OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan i

OLIVEBIOTEQ 2014

5th International Conference for Olive Tree and Oil Production 3‐6 November, 2014 ‐ Amman, Jordan

B O O k O F ABSTRACTS

Editors Dr. Salam Ayoub Dr. Mohammed AlQasem

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Organizing Committee:

Dr. Fawzi Al‐Sheyab ‐ Chair, NCARE, Jordan Dr. Halim Ben Haj Salah ‐ ICARDA Dr. Mohamad Al Ajlouni ‐ AARINENA Dr. Mostafa Qrunfleh ‐ Faculty of Agriculture, University of Jordan, Jordan Dr. Muien Qaryouti ‐ NCARE, Jordan Dr. Salam Ayoub – NCARE, Olive Network, Jordan Dr. Mohammed AlQasem ‐ NCARE, Jordan Dr. Nawaf Freihat ‐ Faculty of Agriculture, Jordan University of Science and Technology Dr. Khalid Al‐Absi, Faculty of Agriculture, Mu’tah University. Dr. Taleb Abu‐Zahra ‐ Faculty of Agricultural Technology, Al‐Balqa' Applied University. Mr. Mohmoud Al Ouran ‐ Jordan Farmers Union, Jordan Eng. Jamal Al Batsh ‐ Ministry of Agriculture, Jordan Eng. Islam Maghaireh ‐ Agricultural Engineers Association, Jordan Eng. Nidal Samain ‐ Jordan Olive Oil Producers Association Ms. Ruba Daghmish ‐ Jordan Olive Products Exporters Association (JOPEA), Jordan

International Advisory Committee:

El Sheyab, F., Jordan Solh, M., ICARDA Chartzoulakis K., NAGREF, Greece Caruso T., University of Palermo, Italy Ismaili ‐ Alaoui M., IAV, Morocco Roussos S., IRD, France Barjol J.L., IOC, Spain Sebastiani L., Scuola Super. Sant' Anna, Italy Dalenda BOUJNAH /AARINENA Olive Network Coordinator, Tunisia

Scientific Committee:

Abdel‐Wali M., Jordan Gargouri K., Tunisia Rapoport H.F., Spain Al‐Absi K., Jordan Gucci R., Italy Roussos S., France Alaoui M., Morocco Kalaitzaki A., Greece Kamoun Rugini E., Italy Aparicio R., Spain N., Tunisia Rusan M., Jordan Ayoub S., Jordan Koubouris G., Greece Sayadi S., Spain Ben Mimoun M., Tunisia Lanza B., Italy Sebastiani L., Italy Caballero J., Spain Lopez‐Lopez A., Spain Sergeeva V., Australia Centritto M., Italy Ozkaya M., Turkey Servili M., Italy Chalak L., Lebanon Ouazzani N., Morocco Shdiefat S., Jordan Chartzoulakis K., Greece Parra‐López C., Spain. Stefanoudaki E., Greece Ferguson L., USA Pereira, José A., Portugal Vita Serman F., Argentina Fernandez E., Spain Perica S., Croatia Xiloyannis C., Italy Fernandez‐Escobar R., Spain Psarras G., Greece Freihat N., Jordan Qrunfleh M. Jordan

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OLIVEBIOTEQ 2014 5th International Conference for Olive Tree and Oil Production

Organizers National Center for Agricultural Research and Extension (NCARE) International Center for Agricultural Research in the Dry Areas (ICARDA) Association of Agriculture Research Institutions in the Near East & North Africa (AARINENA)

In cooperation with Ministry of Agriculture, Jordan

Supporters International Olive Council (IOC) Scientific Research Support Fund, Jordan

Sponsors Jordan Olive Products Exporters Association (JOPEA) Greater Amman Municipality

Conference Secretariat Mrs. Hala Hamati, ICARDA Ms. Hazar Nazzal, AARINENA Mr. Hisham Athamneh, NCARE

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Table of Contents

INTRODUCTORY PRESENTATIONS

What IOC consider people should really need to know about olive oil J. L. Barjol ………………………………………………………………………………………………………………………………...... 2

Olive Sector in Jordan J. Albatsh …………………………………………………………………………………………………….………………….……………...... 3

GENETIC RESOURCES / BREEDING / PROPAGATION

Olive genetic resources: current status of characterization, evaluation and use in breeding programs A. Belaj ………………………………………………………………………………………………………….…………………….……...... 6

Centennial Olive Trees in Lebanon: a Substantial Patrimony L. Chalak, F. Malas, B. Hamadeh, L. Essalouh, B. Khadari ……………………………………………...... 7

Studies on the genetic relationship of wild olive with cultivated varieties and on their resistance to Verticillium wilt in Syria. B. Barhoum, M. Nachit ……………………………………………………………………………………..…………………………… 8

How to select olive varieties for association mapping of agronomic and adaptive traits: towards the setup of Mediterranean network for olive phenotyping. B. Khadari, A. El Bakkali, L. Essalouh, M. Gurbuz, M. Ulas, M. Hakan, O. Cetin, H. Kaya, N. Mete, M. Sahin, L. Sikaoui, A. Moukhli, S. Santoni, E. Costes ……………………………………….…….…………………… 9

Towards the identification of SSR marker linked to vigor in olive tree (Olea europea L.) H. Zaher, B. Boulouha, M. Baaziz, F. Gaboun, S.M. Udupa ……………………….…………………………………… 10

Search for new duel use olive clones in the Central East of Tunisia: Characterization for pomological and chemical traits I. Laaribi, M. Mezghani Aïachi, H. Gouta, M. Ayadi, F. Labidi, M. Mars ………………………………….……. 11

Development of key for identification of seven olive varieties using microsatellite markers M. Iqbal, T. Mehmood, A. Mehmood, S. Jamil ……………………….…………………………………………………….. 12

Exploring and characterizing unknown olive variants R. Mariotti, S. Mousavi, M. Hosseini‐Mazinani, B. Torkzaban, N. Cultrera, S. Pandolfi, J.A. Googlani, L. Baldoni ……………………….………………………………………………………………………………………………… 13 OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan v

The World Olive Collection of Cordoba: 40 years of experience on management of olive genetic resources A. Belaj, J. Hidalgo, G. Beltrán, J. Cano, A. Jiménez, I. Padilla, A. Barceló, G. M. García‐Ruiz, J. Lopez‐Escudero, S. G. Atienza, L. Léon, R. de la Rosa ……………………….…………………………………………… 14

Preliminary Evaluation of Oil Content and Fatty Acids Profile for Olive Varieties Cultivated in Lebanon M. El Riachy, M.Breidi. G. Abou‐Sleymane, L.Chalak ……………………….………………………………………….. 15

Evaluation of some olive cultivars under Kuwait environmental conditions H. Al‐Menaie, O. Al‐Ragam, H. Mahgoub, M. Al‐Hadidi, A. Al‐Shatti and M. Al‐Zalzalah …………… 16

Evaluation of nutrient uptake in different olive cultivars grafted on Gemlik rootstock M. Azimi, M. Ozkaya, H. Çolgecen, H. Buyukkartal ……………………….……………………………………………….. 17

Histological evaluation of graft compatibility in Olea europaea L. scion/rootstock combinations M. Azimi, H. Colgecen, M. Ozkaya, H. Buyukkartal ……………………….……………………………………………… 18

Somatic embryogenesis and plant regeneration from radicles of olive (Olea europaea L.) zygotic embryos, cv. Chemlal. K. Titouh, K. Hadj Moussa, M. Khelifi‐Slaoui, L. Khelifi ……………………….…………………………………………. 19

Varietal diversity of the olive tree in the Chefchaouen region (Northwest of Morocco) M. Ater, H. Barbara1, J. Kassout, M. El Omari ……………………….………………………………………………………… 20

Flowering Phenology in national olive germplasm of Iran A. Zeinanloo, K. Gharibi ……………………….…………………………………………………………………………………………. 21

Assessment of autochthon Tunisian olive varieties by six AFLP markers S. Mnasri, O. Debbabi, M. Ben Saleh, A. Ferchichi ……………………….…………………………………………………. 22

Olive tree architecture as a tool in early selection for short juvenile period S. Hammami, H. Rapoport, M. Ben Mimoun, L. León, R. De La Rosa ……………………….………………… 23

Use of morphological descriptors and principal component analysis for studying the variability of olive descendants issued from controlled pollination I. Laaribi, M. Mezghani Aïachi, F. Labidi, M. Mars ……………………….…………………………………………………. 24

Characterization and performance of selected olive (Olea europaea. L) seedlings issued from a Tunisian breeding program I. Guellaoui, F. Ben Amar, M. Boubaker, M. Ayadi, R. Ayadi, A. Yengui, N. Ben Belgacem ………. 25

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Selection of the best olive genotypes from several crosses among cultivated varieties B. Alfei, A. Paoletti, A. Rosati, A. Santinelli, G. Pannelli ……………………….……………………………………… 26

Chemical and sensory properties of monovarietal olive oils produced by ancient genotypes in Emilia‐Romagna region L. Morrone, A. Rotondi, D. Beghè, T. Ganino, A. Fabbri ……………………….…………..…………………………… 27

Evaluation of pomological traits for some olive cultivars in Tarom Region, Iran S.Vatani, M. Taheri, A. Gholiyan , Z. yousefi, A. A. Zeinanloo ……………………….……………………………… 28

Preliminary results of olive improvement using gamma radiation A. Chaari, M. Maalej, A. Chelli‐Chaabouni, S. Baccari ……………………….…………………………………………. 29

454 pyrosequencing of Olive (Olea europaea L.) transcriptome in response to salinity

P. Kalaitzis, C. Bazakos, M. Manioudaki, E. Sarropoulou ………………………………………………………………. 30

Assessment of variability for three olive varieties in Montenegro B. Lazović, M. Adakalić, D. Bandelj, T. Perović ……………………….………………………………………………………. 31

Genotype effect on embryogenic capacity and plant regeneration from olive (Olea europaea L.) callus, cv. Picual. K. Titouh, L. Khelifi, K. Hadj Moussa, S. Cerezo‐Medina, J.A. Mercado and F. Pliego‐Alfaro ………. 32

Production of olive (Olea europaea L.) rootstocks by in vitro germination of zygotic embryos, cv. Chemlal. K. Hadj Moussa, K. Titouh, M. Khelifi‐Slaoui, L. Khelifi ……………………….…………………………………………. 33

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MOLECULAR BIOLOGY / BIOTECHNOLOGY

Characterization of salinity‐responsive genes in olive (Olea europaea L.) M.T. Sadder, A.M. Zakri, H. Alswalmah, A.A. Alsadon, A.A. Al‐Doss ……………………….……………………. 36

Comparative study of FAD2 gene sequences among some olive cultivars grown in Qom region of Iran M. Sharifani, M. Asheri, A. Yamchi, S. Hosseini Mazinani ……………………….……………………………………. 37

The different adaptations to salt‐stress of two Italian olive cultivars, salt‐sensitive (Leccino) and salt‐tolerant (Frantoio) L. Rossi, A. Francini, A. Minnocci, L. Sebastiani ……………………….……………………………………………………… 38

Transcriptome analysis of seed and mesocarp tissues of olive fruit identifies key genes involved in oil accumulation and fatty acid composition M. Hernández, A. Muñoz‐Mérida, M. Sicardo, O. Trelles, V. Valpuesta, J. Martínez‐Rivas ………… 39

Identifying genes involved in fatty acid synthesis in olive N. Cultrera, R. Mariotti, F. Alagna, V. Sarri, M. Ceccarelli, M. Bellucci, L. Baldoni ……………………….. 40

Hydroxytyrosol – An attractive olive phytochemical produced through metabolic engineering in E. coli F. Ververidis, E. Trantas, Th. Pavlidis, E. Navakoudi, A. Kontaratou, E. Mpalantinaki …………………. 41

Identification of the genes in olive (Olea europaea cv. “Koroneiki”) that may be involved in the biosynthesis of hydroxytyrosol N. Mougiou, E. Trantas, A. Argiriou, F. Ververidis, A.M. Makris, K.E. Vlachonasios …………………….. 42

Response to drought stress of olive (Olea europaea L.) plants transgenic for tobacco osmotin gene. C. Silvestri, V. Cristofori, S. Astolfi, A. Minandri, E. Rugini ……………………….……………………………………. 43

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BIOLOGY / PHYSIOLOGY

Chemical and histological analysis of olive tree tissues to understand seasonal starch distribution E. Cauli, G. Bandino, P. Sedda, M. Mulas, R. Zurru ……………………….………………………………………………. 46

Quantitative behavior of Olea pollen in relation with climatic factors in three regions of Tunisia S. Elhadj Hamda, A. Ben Dhiab, M. Msallem ……………………….……………………………………………………….. 47

Quantitative behavior of Olea pollen in relation with climatic factors in three regions of Tunisia S. Elhadj Hamda, A. Ben Dhiab, M. Msallem ……………………….……………………………………………………… 48

The Estimation of real water needs of olive trees (Olea europaea L. cv. ‘Meski’) conducted in irrigated orchards during flowering S. Ben Mariem, O. Boussadia, F. Ben Mariem, W. Gheriani, N. Boukhrissat, M. Braham …………… 49

Metabolic profiling and gene‐expression analyses in salt stressed olive trees (Olea europea L.) cultivars Leccino and Frantoio L. Rossi, M. Borghi, A. Francini, D. Xie, L. Sebastiani ……………………….……………………………………………… 50

Metabolic profiling and gene‐expression analyses in salt stressed olive trees (Olea europea L.) cultivars Leccino and Frantoio L. Rossi, M. Borghi, A. Francini, D. Xie, L. Sebastiani ……………………….……………………………………………… 51

Effect of Sodium Chloride on olive seedlings under controlled conditions. S. Baccari , A. Chaari Rekhis, A. Chelli‐Chaabouni ……………………….……………………………………………….. 52

Identification of the iridoid synthase involved in oleuropein biosynthesis F. Alagna, F. Geu‐Flores, S. E. O’Connor, L. Baldoni, A. Osbourn ……………………….………………………. 53

Olive hedgerow canopy positions determine inflorescence and flower development E. R. Trentacoste, M. Gómez‐del‐Campo, I. Moreno‐Alías, H. F. Rapoport ……………………….……………. 54

Frost tolerance and recovery ability of eleven olive cultivars in central Italy E.M. Lodolini , B. Alfei, A. Santinelli, T. Cioccolanti, D. Neri ……………………….………………………………….. 55

Use of biosensor for estimating abiotic stresses and technical management of olive groves D. Boujnah, M. Gouiaa, H. Chehab, Z. Mahjoub, S. Lamari, B. Chikhaoui ……………………….……………. 56

Pollen storage and in vitro germination in some olive (Olea europaea L.) cultivars

A.Senbas, M.T. Ozkaya, H. Colgecen ……………………….……………………………………………………………………… 57 OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan ix

Comparison studies among different olive fruit cultivars (Olea europaea L.) reveals differences in expression levels of genes involved in Vitamin E biosynthesis E.C. Georgiadou, G. Koubouris, C. Sergentani, P. Kalaitzis, G.A. Manganaris, V. Fotopoulos …….. 58

Molecular characterization of vitamin E biosynthesis in olive fruit (Olea europaea L.) during three successive growth seasons E.C. Georgiadou, V. Goulas, T. Ntourou, G.A. Manganaris, P. Kalaitzis, V. Fotopoulos ………………. 59

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PLANTING SYSTEMS / ORCHARD MANAGEMENT

Olive planting systems and mechanization J. Tous ……………………….………………………………………………………………………………………………….…………………… 62

Developing mechanical harvesting for California “Black Ripe” table olives (Olea europea cv. ‘Manzanillo) L. Ferguson, J. Miles, S. Garcia, T. Sarcoglu, F. Aranda, F. Jiminez, W. Krueger, E. Ficthner ……….. 63

Characterizing olive tree geometry features using Unmanned Aerial Vehicle (UAV) images J. Torres‐Sanchez,, F. Lopez‐Granados, N. Serrano, O. Arquero, R. Fernandez‐Escobar, J.M. Pena ……………………….………………………………………….……………………….…………………………………………………… 64

Ecosystemic Management Plan of the olive sector in Draa and Tafilalet areas (Morocco) A. Abdellaoui , M. Hemmi, L. Sikaoui, A. El Antari, A. Berahmani, M. Ferrahi ……………………….……… 65

Effect of Planting Density on the Behaviour of Sourani Olive Tree under Kuwaiti Environment O. Al‐Ragam, H. Al‐Menaie, H. Mahgoub, M. Al‐Hadidi, A. Al‐Shatti and M. Al‐Zalzalah ……………. 66

A comparative study of hand‐held harvesting machine with traditional methods used for olive harvesting in Jordan R. Ahmad, S. Ayoub ……………………….………………………………………….……………………….…………………………… 67

Adaptation of different varieties of olive tree to high‐density planting systems O. Arquero, N. Serrano, M. Vinas, M. Lovera ……………………….……………………………………………………… 68

Effect of different pruning methods on olive tree growing A. Gholiyan, A. A. Zeinanloo, Z. yousefi, M. Taheri ……………………….…………………………………………. 69

Effect of lateral mechanical pruning on light interception, vegetative growth, fruit characteristics and fruits yield in a 13 years old high density planting system A. Larbi, I. Dhouibi, M. Msallem ……………………….……………………………………………………………………………. 70

Olive oil characteristics of twelve cultivars produced in a high‐density grove at the Valladolid province (Spain) N. Rodrigues, H. Martín, M. C. Asensio‐S.‐Manzanera, S. Casal, A. Bento, J. A. Pereira ……………… 71

Effect of olive trees density on the production and quality of olive oil from Cv. Arbequina N. Rodrigues, H. Martín, M. C. Asensio‐S.‐Manzanera, S. Casal, A. Bento, J. A. Pereira ……………… 72

Full production olive trees architecture restructuration to continuous harvesting with large straddle machines (over‐row): evaluation of pruning intensity F. Vita Serman, G. Lemole, L. Bueno ……………………….……………………………………………………………………… 73

Measurement of foliar surface of olive orchards using a laser scanner sensor F. Vita Serman, G. Lemole, L. Bueno, C. Soria and M. Munoz ……………………….………………………………. 74 OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan xi

IRRIGATION / STRESS PHYSILOGY / NUTRITION

An environmental‐friendly approach for a productive olive orchard management C. Xiloyannis, A. Palese ……………………….……………………………………………………………………………………………. 76

Olive irrigation management project – Setting the scene V. Nangia, M. Karrou, T. Oweis ……….……………….…………………………………………………………………………….. 77

Water requirement and water‐use efficiency for olive trees under different irrigation systems A. Hijazi, M. Doghoze, N. Jouni, V. Nangia, M. Karrou, T.Oweis ……………………….…………………………… 78

Effect of deficit irrigation on vegetative growth and fruit yield parameters of young olive trees (Olea europaea L.) in semi‐arid area of Morocco L. Sikaoui, A. Bouizgaren, M. Karrou, H. Boulal, B. Boulouha, M. Karama, V. Nangia, T. Oweis 79

Effect of switching from surface to drip irrigation on the performance of mature olive trees in a dry area of Morocco L. Sikaoui, A. El Antari, A. Bouizgaren, M. Karrou, H. Boulal, B.Boulouha, M. Idrissi, V. Nangia, T. Oweis ……………………….……………………………………………………………………………………………………………………. 80

Effect of three irrigation regimes on physiological parameters of new olive orchard under semi‐ arid conditions of Morocco L. Sikaoui, , A. Bouizgaren, M. Karrou, H. Boulal, B. Boulouha, M. Karama, V. Nangia, T. Oweis 81

Impact of different levels of deficit irrigation on olive productivity under drip irrigation system N. Al Jouni, M. Hayek, F. Ayesh, V. Nangia, M. Karrou, T. Oweis ……………………….………………………. 82

Integrated approaches to evaluate the whole‐plant water relations in irrigated and rain fed Olea europaea M. Centritto, G. Marino, C. Cocozza, A. Giovannelli, C. Cantini, R. Tognetti ……………………….……. 83

Irrigating a hedgerow olive orchard to improve production and quality J.E. Fernández, A. Perez‐Martin, M.V. Cuevas, C.M. Rodriguez‐Dominguez, A. Morales‐Sillero, J.M. García, V. Hernandez‐Santana, A. Diaz‐Espejo ……………………….……………………………………………… 84

Responses of olive varieties cultivated under different irrigation regimes M. Aiachi‐Mezghani, C. Masmoudi‐Charfi C ……………………….……………………………………………………….. 85

Irrigation Management of Young Olive Trees in Kuwait A. Al‐Shatti, H. Al‐Menaie, M. Al‐Hadidi, H. Mahgoub, O. Al‐Ragam, and M. Al‐Zalzalah …………… 86

The effect of different irrigation strategies on olive oil (Memecik cv.) antioxidant content and activity D. Sevim, O. Koseoglu, U. Kaya, E. Akkuzu, M. Parlak, N. Pouyafard ……………………….…………………… 87 Effect of deficit irrigation on olive tree performance, yield and oil quality 88 OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan xii

S. Naoum, S. Ayoub, I. Amayreh, M. Diab, M. Ammoush, B. Kawaleet, L. Daoud ……………………..

The timing of water deficit affects oil quality of cv. Frantoio G. Caruso, R. Gucci, C. Gennai, S. Esposto, A. Taticchi, S. Urbani, M. Servili ……………………….……. 89

Olive fruit growth indicators for regulated deficit irrigation scheduling H. Memmi, M.C. Gijón, H.F. Rapoport, J.F.Couceiro, D. Pérez‐López ……………………….………………… 90

Effect of foliar amino acid “Pepton” application on growth, flowering, fruit set and oil quality of olive trees (Olea europaea L., cv. Chetoui) grown under intensive planting system. Ajmi Larbi, Mohamed Ayadi, Samira Yakoubi and Monji Msallem ……………………….……………………… 91

The effect of foliar fertilization on carbohydrates status of 'Chemlali' olive (Olea europaea L.) leaves cultivated under rain‐fed conditions I. Zouari, M. Aïachi‐Mezghani, B. Mechri, F. Labidi, F.Attia, D. Boujneh, H. Chehab, M. Hammami …….……………………………………………………..……………………….………………………………………………….. 92

Development and validation of an irrigation scheduling tool for olives to improve water use efficiency at farm level K. Chartzoulakis, G. Psarras, I. Kasapakis, M. Bertaki ……………………….……………………………………………. 93

Arbuscular mycorrhizal fungus (Glomus deserticola) enhance drought tolerance of olive tree (Olea europaea) W. Khabou, T. Gargouri, S. Kammoun ……………………….…………………………………………………………………. 94

Variability of the floral biology of four olive cultivars grown under two watering regimes in an arid region of Tunisia

M. Aachi‐Mezghani, A. Diab, A. Laroui, I. Zouari, I. Laaribi, F. Labidi, L. Attia, M. Mars …………….. 95

Efficacy of partial root zone drying application on olive tree in arid climate L.Trablesi, S.Maktouf, M.Ghrab, M.Khlifi, N.Soua, K.Gargouri ……………………….…………………………… 96

Physiological responses of young olive (Olea europaea L. cv Ayvalık) trees to water stress N. Pouyafard, E. Akkuzu, U. Kaya ……………………….…………………………………………………………………………. 97

Effect of deficit irrigation strategies on canopy temperature and leaf water potential in olive (Memecik Cv.) U.Kaya, M.Parlak , N.Pouyafard, T.Cakır , M. Gurbuz, G.P.Mengu, E.Akkuzu ……………………….……… 98

Water relations and gas exchanges of olive trees (cv. chetoui) in response to different deficit Irrigation regimes S. Dbara, K. Lahmar, M. Ben Mimoun ……………………….………………………………………………………………….. 99 Ecophysiological response of young olive plants under three irrigation regimes. 100 OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan xiii

M. Gouiaa, F. Zaouay , D. Boujnah ……………………….………………………………………………………………………..

Rainfed olive production as influenced by low quantity of applied water during the first stage of fruit growth R. Razouk, A. Kajji, M. Karrou ……………………….…………………………………………………………………………………. 101

Contribution of the study of the behavior of the olive tree (Olea europaea L.) under different water regimes H. Benjeddou, B. Ben Rouina ……………………….…………………………………………………………………………………. 102

Application of exogenous organic matter promote soil fertility and carbon sequestration Rayda Chaker , Noha Sallemi , Sameh Maktouf , Nabil Soua , Mouna Khilifi , kamel Gargouri 103

Effects of soil management practices on Particulate Organic Matter (POM) fraction involved in soil fertility of rainfed olive orchards A.M. Palese, A. Arous, C. Xiloyannis, G. Celano ……………………….……………………………………………………. 104

Effect of nitrogen and phosphorus fertilizer on the growth of young olive trees in Kuwait H. Al‐Menaie, O. Al‐Ragam, H. Mahgoub, M. Al‐Hadidi, A. Al‐Shatti and M. Al‐Zalzalah …………. 105

Nutritional status of olive orchards in rainfed and irrigated area of Tensift‐Haouz region in Morocco H. Boulal, L. Sikaoui, A. Bouizgaren, M. El Gharous ……………………….……………………………………………. 106

Olive tree nutritional status survey following a non‐typical dry growing season in the area of Chania, Crete, Greece G. Psarras, A. Digalaki, M. Darioti, M. Bertaki ……………………….……………………………………………………….. 107

Role of indigenous weed flora in carbon and main nutrient elements storage in olive orchards in Southern Greece E. Markakis, N. Kourgialas, C. Sergentani, V. Borraccia, L. Mikalef, N. Digalaki, S. Reppas, A. Aggelaki, C. Manolaraki, S. Malliaraki, G. Giakoumaki, K. Giannopolitis, N. Kavroulakis, G. Psarras, G. Koubouris ……………………….……………………………………………………………………………………………… 108

Particulates fallout in the southern Sector of the city of Sfax: monitoring and impact on the soil and plants. The olive tree as an example C. Mbadra, M. Bahloul, K.Gargouri, A. Chafai, F. Ben Abdallah ……………………….……………………………. 109

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PEST AND DISEASE CONTROL

Integrated Pest Management of olive tree in Tunisia M. Ksantini ……………………….………………………………………………………………………………………………………………. 112

Effective, environmentally‐sensitive pest management in olive production V. Sergeeva ……………………….……………………………………………………………………………………………………………… 113

Evaluation of olive fly Bactrocera oleae oviposition preference on Cobrançosa, Madural and Verdeal Transmontana cultivars in Portugal R. Malheiro, S. Casal, L. Pinheiro, P. Baptista, A. Bento, J. A. Pereira ……………………….…………………… 114

New insect pest species recently established on olives groves in Marrakech ‐Tensift – Al Haouz region, Morocco Y. Ouguas, M . Karama, I. Zafzafi , T. Bamhamed ……………………….……………………………………………….. 115

Fruit Fly Net Project: E‐monitoring and pest management design issues of a location–aware system for olive fly and other fruit fly pests M. AlQasem, A. Sciarretta, M. R. Tabilio, M. Miranda‐Chueca, D. Nestel, N. Papadopulos, T. Tsiligiridis ……………………….……………………………………………………..……………………….………………….…………….. 116

Evaluation of susceptibility of the most cultivated olive trees cultivars in Tunisia to Verticillium wilt disease (Verticillium dahlia) M.A. Triki, H. Hassairi, F. Ben Amar, Y. Gharbi, I. Hammami, S. Krid, W. Khabou, A. Rhouma and R. Gdoura ……………………….……………………………………………………..………………………………………………….. 117

Genetic structure of Verticillium dahliae isolates infecting olive tree using AFLP, Pathogenicity and PCR Markers Y. Gharbi, M.A. Triki, F. Daayf, R. Gdoura ……………………….………………………………………………………………. 118

PESTOLIVE: a Mediterranean research project for understanding and managing soil‐borne parasites on olive using historical and ecological approaches T. Mateille, M. Achouri, M. Ater, A. Belaj, G. Besnard, P. Castillo, E. Chapuis, R. De La Rosa, F. De Luca, A.M. D’Onghia, H. El Maraghi, C. El Modafar, A. El Mousadik, A. El Oualkadi, Z. Ferji, N. Horrigue‐Raouani, R.M. Jimenez‐Diaz, M. Kadiri, S. Kallel, B. Khadari, B.B. Landa, L. Leon, M. Montes‐Borrego, A. Moukhli, J.A. Navas‐Cortes, A. Öcal, N. Sasanelli, J. Tavoillot, M.A. Triki, A. Troccoli, E. Tzortzakakis, M. Ulas, N. Vovlas, T. Yaseen ……………………….……………………………………. 119

Bio‐pesticide production by solid‐state fermentation using olive industry by‐products S. Roussos, M.S. Tranier, Q. Carboué, M. Mehjoub, C. Périssol, I. Perraud‐Gaime, R. De la cruz‐Quiroz, C.N. Aguilar, M. Ait Hamza, A. Cherif ……………………….…………………………………………….. 120

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Physical parameters of olive fruits and olive leaves from Portuguese olive varieties and their role in oviposition preference of Bactrocera oleae R. Malheiro, S. Casal, N. Rodrigues, P. Baptista, A. Bento, J. A. Pereira ……………………….………………. 121

Distribution of olive fruit fly in California based on fruit infestations since the 1998 invasion V. Yokoyama ……………………….……………………………………………………..……………..…………………………………….. 122 Study of the duration of attractant action of various attractant solutions used in bait sprays against Bactrocera oleae (Diptera: Tephritidae) Α. Kalaitzaki, E. Malandraki, E. Kapogia, A. Ioannou ……………………….……………………………………… 123

Combined effectiveness of tebuconazole‐trifloxystrobin against infections of Fusicladium oleagineum in comparison to conventional products V. Vizzarri, N. Iannotta , I.Muzzalupo, T. Belfiore. ……………………….………………………………………………… 124

Genetic diversity of Verticillium dahliae from olive trees in Tunisia based on RAMS and IGS‐ RELP Y. Gharbi, M.A. Triki, F. Daayf, R. Gdoura ……………………….………………………………………………………………. 125

Biological control against olive Psyllid Euphyllura olivina by releases of indigenous bug Anthocoris nemoralis N. Gharbi , A. Dibo ……………………….……………………………………………………..………………………………………….. 126

Learning from olive evolution and cultivation to understand the diversity of associated plant‐ parasitic nematodes communities in Morocco N. Ali, E. Chapuis, J. Tavoillot, M. Aït Hamza, Z. Ferji, A. El Mousadik, A. El Oualkadi, G. Besnard, A. El Bakkali, A. Moukhli, B. Khadari, C. El Modafar, M. Ater, T. Mateille ………………… 127

Application of antagonistic bio‐products (Bio‐Zeid and Bio‐Arc) to control Verticillium wilt disease of olive in nursery conditions T. Yaseen, R. Abou Kubaa , A. M. D’Onghia ……………………….……………………………………………………………. 128

Endophytic fungi in olive tree cultivars with different susceptibilities to anthracnose F. Martins, J.A. Pereira, A. Bento, P. Baptista ……………………….………………………………………………………… 129

Phytophthora palmivora: Geographic distribution in different olive tree regions in Morocco and pathogenicity M. Chliyeh, Y. Rhimni, K. Semaoui, A. Ouazzani Touhami, A. Filali‐Maltouf, C. El Modafar, A. Moukhli, A. Oukabli, R. Benkirane, A. Douira ……………………….………………………………………………………… 130

Effect of olive varieties and rearing substrates on plant‐parasitic nematode communities in southern Morocco olive nurseries M. Ait Hamza, Z. Ferji, N. Ali, H. Tazi, J. Tavoillot, A. Moukhli, H. Lakhtar, S. Roussos, H. Boubaker, A. El Mousadik and T. Mateille ……………………….…………………………………………………………….. 131

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Comparative effect between biopesticides and biofertilizers on bioagressors of Olea europaea L. Brahimi, H. Halladj, Z. Djazouli ……………………….……………………………………………………..…………………….. 132

First report of Botryosphaeria dothidea and Diplodia seriata associated with olive tree wood lesions in Algeria F. Ammad, S. Benarous ……………………….……………………………………………………..……………………….…………… 133

Biological control of Colletotrichum species associated with olive anthracnose in Tunisia M. Chattaoui, M. Msallem, A. Rhouma ……………………….…………………………………………………………………. 134

Current status of Olive Weevil Rhynchites cribripennis Desbrochers (Coleoptera: Attelabidae) in Montenegro T. Perovic, S. Hrncic, M. Cizmovic ……………………..……………………….………………………………………………….. 135

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OLIVE OIL QUALITY/ HEALTH / TABLE OLIVE

Olive Oil Quality and Health A. Kiritsakis, D. Gerasopoulos, E. L. Iorio and K. Kiritsakis ……………………….………………………………….. 138

Which of the factors: cultivar, extraction systems or storage conditions is determinant on the variation of olive oil physicochemical and the sensory profiles? K. Ben‐Hassine, D. Malouche, K. Talmoudi, T. Ameni, H. Sebei, M. Hamami, N. Grati‐Kamoun 139

Potential usefulness of a colorimetric test to measure olive oil peroxidation E. L. Iorio, A. Kiritsakis, M. Carratelli ……………………….………………………………………………………………………. 140

Fatty Acid Alkyl Esters in Extra Virgin Olive Oil: an Evolving Parameter R. B. Gómez‐Coca, G. D. Fernandes, M. C. Pérez‐Camino, W. Moreda ……………………….………………. 141

Hepatoprotective activity of oleocanthal extracted from olive oil amurca in the rat S. Janakat, A. Al Amour ……………………….……………………………………………………..……………………….…………… 142

Effect of cultivar and climatic conditions on olive (Olea europaea L.) oil fatty acid composition A.A. Zeinanloo ……………………….…………………………..……………………….………………………………………………….. 143

Role of chemometric techniques in the typicality determination of virgin olive oil from Moulay Driss Zarhoun (Volubilis)/ Morocco S. Rouas1, M. Rahmani, A. Antari, E. Baamal, N. Maata ……………………….………………………………………… 144

Deodorized olive oils: from fraud to nutrition R. Aparicio‐Ruiz, D.L. García González, I. Romero, C. Oliver, R. Aparicio, M.T. Morales ……………… 145

Olive Oil: An Exceptional Functional Lipid Food Protecting Our Body From Oxidative Stress A. Kiritsakis, E. Iorio, F. Shahidi, D. Gerasopoulos, K. Kiritsakis ……………………….……………………………. 146

Reduction of acrylamide in potato chips using olive oil as frying mean. Immunoassay vs LC‐ MS/MS techniques for acrylamide determination. G. Siragakis, G.Miliadis, A.Vrettakou ……………………….…………………………………………………………………….. 147

The effect of chlorophyll pigments and squalene addition on refined olive oil oxidative stability G. Rigane, R. Ben Salem ……………………….………………………………………………………………………………………….. 148

Healthy and sensory properties in virgin olive oil: new approach to extraction process A. Taticchi, S. Esposto, R. Selvaggini, S. Urbani, B. Sordini, M. Servili ……………………….………………….. 149

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Comparison of olive oils from autochthonous and foreign cultivars of the region of Qom, Iran M. Sharifani, M. Asheri, A.Yamchi and S. M. Hosseini Mazinani ……………………….………………………….. 150

The evaluation of Turkish virgin olive oil fatty acid profiles by Chemometrics during different ripening index and storage conditions O. Koseoglu, M. K. Unal, D. Sevim, D.ozdemir ……………………….……………………………………………………. 151

Chemical composition and antibacterial activity of two different oils of Olea europea L. A. Chaker, H. Boukhebti, M. Ramdhani, S. Dahamna ……………………….…………………………………………… 152

Steryl glucosides composition in monovarietal extra virgin olive oils R. Cruz‐Hidalgo, R. B. Gómez‐Coca, G. D. Fernandes, M. C. Pérez‐Camino, W. Moreda ………….. 153

Changes in virgin olive oil quality characteristics due to heating in comparison with vegetable oils A. Yorulmaz ……………………….……………………………………………………..……………………….…………………………….. 154

Preliminary results of olive zoning study in the province of Sassari: the peculiarities of cv Bosana oils produced in different growing areas L. Morrone, A.Rotondi,N. Di Virgilio,B.Alfei, C.Cantini, E.Vagnoni, P.Duce ……………………….………. 155

Virgin olive oil carotenoid fraction improvement by addition of microalgae extracts of Scenedesmus almeriensis P. Limon, R. Malheiro, G. Acién‐Fernández, J. M. Fernández‐Sevilla, S. Casal, N. Rodrigues, R. Cruz, R. Bermejo, J. A. Pereira ……………………….………………………………………………………………………………… 156

Effect of olive oil on lipid profile in female climacteric ‐ Research project in progress F. A. conte, L. B. Franz, O. B. Oliveira ……………………….………………………………………………….…………………. 157

Effect of Olea Europaea L. folium ethanolic extract in rats submitted to LPS M. Ramos, O. B. Oliveira, C. Rasvickas, A. Pereira , N. Schor ……………………….………………………………… 158

Nutritional value and health benefits of table olives F. Gungor, A. Yildırım ……………………….………………..……………………….………………………………………………….. 159

Comparison of single strain starter culture and a selected inoculum enrichment in the processing of natural table olives M. Campus, P. Sedda, E. Cauli, R. Comunian, A. Paba, E. Daga, S. Schirru, R. Zurru, G. Bandino 160

Natural fermented table olives from Trás‐os‐Montes region as a source of bioactive compounds with antioxidant and antimicrobial properties R. Malheiro, P. Mendes, A. Bento, J. A. Pereira ……………………….…………………………………………………….. 161

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OLIVE MILL WASTE MANAGEMENT

Alternative technologies for olive mill wastewater management with emphasis on soil application K. Chartzoulakis, G. Psarras, N. Kalogerakis, F. Santori ……………………….………………………………………… 164

Agronomic and environmental valorization of olive mill wastewaters A. Abichou, L. Mohamed ……………………….……………………………………………………..……………………….……….. 165

Impact of olive mill wastewater treated by different techniques on seed germination of Barley M. Rusan, A. Balasmeh, S. Zuraiqi, M. Bashabsheh ………………………….…………………………………………. 166

The different points of view of olive mill wastewater in Turkey: A result of New Ecological Paradigm R. Tunalioglu, R.Yildirim……………………….…………………………………………………………………………………………… 167

Valorization of olive husk into valuable organic amendment: does initial C/N ratio affect the quality of the produced compost? K. Azim , S. Roussos , C. Périssol, I. Thami Alami , B. Soudi ……………………….……………………………………. 168

Coagulation/flocculation followed by solar Fenton oxidation at a pilot scale for the purification of olive mill wastewater I. Michael, A. Panagi, L. A. Ioannou, Z. Frontistis, D. Fatta‐Kassinos ……………………….……………………. 169

Production of extracellular lignocellulolytic enzyme by Streptomyces Sp grown on olive pomace L. Medouni‐Haroune, M. Kecha, F. Zaidi, S. Roussos, V. Desseau ………………….…….…………………….. 170

Olive mill wastewater agronomic application on olive tree (Olea europaea L.) in the South of Tunisia: Some biochemical and nutritional traits S. Magdich, C. Ben Ahmed, B. Ben Rouina and E. Ammar …………………….….…………………………………… 171

Influence of olive mill wastewater applied at different quantities on an alkaline soil fertility R. Razouk, K. Oubella ……………………….……………………………………………………………………………………………… 172

Performance of Jet‐Loop reactors with ultrafiltration membrane system (JACTO.MBR) for olive mill wastewaters biotreatment B. Ribeiro, I. Torrado, L. Baeta‐Hall, A. Amer, M. Rusan, A. Eusébio ……………………….…………………. 173

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ECONOMICS / MARKETING

Olive Oil Promotion: A must for the future. M. Rappou ……………………….……………………………………………………..……………………….………………………………. 176

Performance of the olive trees and economic profitability of olive production in the Haouz region in Morocco: effect of drip irrigation with full and deficit water regimes compared to flooding method A. Ait Hmida , L. Sikaoui , M. Karrou, V. Nangia ……………………….…………………………………………………….. 177

Estimating price and income elasticity of olive oil demand in Libya for the period 1980‐2010 K. Elbeydi1 , A. Hamuda2 178

Preference mapping of Extra Virgin olive oil with variety, extraction systems and analytical parameters: Multiple Factor Analysis and Principal Component Analysis K. Talmoudi, K. Ben‐Hassine, A. Taamalli, M. Hamami, N. Grati‐Kamoun, D. Malouche ……………. 179

Olive production in Algeria: situation and perspective B. Lotmani, A. Berkani, D. Labdaoui ……………….……….……………………………………………………………………… 180

The diversity of the Italian monovarietal extra virgin olive oils: from trained panel to consumer experience B. Alfei, M. Magli, A. Rotondi, L. Morrone ……………………….…………………………………………………………… 181

Analysis of efficiency in organic and conventional Italian olive farms using F.A.D.N. dataset N. Galluzzo ……………………….………………………………………………………….………………………………………………….. 182

Olive growing in Morocco: Competitive agribusiness M. Ismaili Alaoui ……………………….……………………………………………………..……………………….……………………… 183

Factors affecting consumers' knowledge of olive oil Protected Designation of Origin label Y. Erraach, S. Sayadi, C. Parra‐López ……………………….………………………………………………………………….... 184

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 1

INTRODUCTORY PRESENTATIONS

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 2

What IOC consider people should really need to know about olive oil

J. L. Barjol

International Olive Council (IOC), Principe de Vergara, 154, 28002 Madrid, Spain

Since 1959, the IOC is the international, intergovernmental forum for debating olive and olive oil issues. Member governments enter into a commitment to apply IOC standards in their international trade. Nowadays, the IOC has 17 Members that account for 97% of world olive oil production and 96% of world exports. World consumption has almost doubled in 25 years. At present, (except this campaign 2014/15 that will be lower) around 3 million tonnes of olive oil are produced in the world, of which 0.8 million tonnes go into international trade (intra‐European Union trade excluded). More than 80% of the world’s olive oil is produced by five countries: Spain (45% share of the total, except this campaign 2014/15), followed by Italy and then Greece, Tunisia and Turkey. The top importers are the United States, the European Union (excluding intra‐ European Union trade), Brazil (newcomer), Japan, China (newcomer), Canada and Australia. These are followed by a long list of other countries since olive oil is now a globalised product. The IOC is mandated to fix the designations and definitions of olive oil to be used by member governments in their international trade. A clear distinction has to be made between the family of virgin olive oils (extra virgin, virgin and ordinary virgin plus lampante virgin which is not fit for direct human consumption) and olive oil (blend of refined lampante virgin olive oil with extra virgin or virgin olive oil) and olive pomace oil (blend of refined crude pomace oil extracted from olive pomace, the by‐product of virgin olive oil extraction, with extra virgin or virgin olive oil). IOC standards fix purity criteria to guarantee that the oil has been obtained solely from olives and quality criteria (including sensory evaluation performed by a recognised panel of trained tasters) to classify the different types of virgin olive oils. Methods and limits are also specified in the standards alongside the criteria. All are regularly updated on the basis of scientific research validated by scientific peers as well as in ring tests to make sure there are no problems due to the geographical or varietal origin of oils. Virgin olive oils, olive oil and olive pomace oil are predominantly made up of monounsaturated fatty acids. Their positive impact on health is well documented (especially in preventing cardiovascular disease and decreasing LDL‐cholesterol without decreasing HDL‐cholesterol) and justifies the issue of a qualified health claim. The minor components (antioxidants and vitamins) present in virgin olive oils (and to a certain extent in olive oil and olive pomace oil because they are blended with virgin olive oils) have a high biological value. They help to prevent various diseases (cancer, diabetes, high blood pressure …), combat ageing and protect from memory decline and neurodegenerative diseases (Alzheimer’s, Parkinson’s).

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Olive Sector in Jordan

J. Albatsh

Ministry of Agriculture, Jordan

The cultivation of olive trees (Olea europaea L.) started in the Mediterranean basin some 6000 years ago . Jordan can be considered one of the homelands and natural habitat of cultivated olives. The olive sector is one of the most important components of the Jordanian agricultural sector. The area planted with olive trees is about 125986 hectare, representing about 77% of the total area planted with fruit trees, and 44% of the actually cultivated area in Jordan in 2013. The sector also constitutes a pillar to socio‐economic development in the country, since it is a major tool for poverty and unemployment alleviation. Thus, olive farming is an important job creator and income source for 80 thousand Jordanian families. Jordan investments in olives sector is estimated more than one billion JD including the value of cultivated lands and related industries. Olive's tree is mainly cultivated within two areas in Jordan, first: the western highlands (rain fed) crossing Jordan from North to South and second: the Northern Western deserts area (irrigated). According to MOA, 2013 statistics around 77.3 % of the total area of olives is rain fed area, where 22.7% of the total area is under permanent irrigation. In addition, there are 87% of cultivated trees are bearing and some 80% of its production is converted into oil. The Jordanian olives oil is characterized by a natural flavor and excellent chemical composition compared to other oils over the world. There is potentiality for producing best quality of virgin olive oil in both rain fed and irrigated areas. The major strengths of the Jordanian Olive Oil Cluster are: good indigenous olive varieties with differentiated characteristics, some good agricultural practices like olive hand picking, and an acceptable modem oil producing infrastructure, based on continuous flow system.

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Genetic Resources Breeding

Propagation OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 6

Olive genetic resources: current status of characterisation, evaluation and use in breeding programs

A. Belaj

IFAPA Centre “Alameda del Obispo”, Córdoba, Spain

Unlike most other fruit species, as a consequence of tree longevity and lack of turnover with new genotypes, the genetic patrimony of olive is very large. More than 1,200 cultivars maintained ex situ in 79 international and national repositories, are still actively cultivated. In addition, in situ conservation of ancient olive trees has recently been considered as an alternative approach to preserve this unexploited reservoir of genetic diversity. Wild relatives and related subspecies might also be very interesting source of genes for some agronomic traits as resistance to stresses. In spite of this rich genetic biodiversity, the knowledge on olive genetic is still limited. In this paper we report the efforts to characterize, identify and exploit the species genetic resources. Up to date, the application of molecular tools has been mainly related to the analysis of genetic variability in Olea europaea complex and for the olive oil origin traceability. While, the recent generation and application of genomic tools can be of great help for understanding the molecular basis of fruit and oil quality and that of traits of agronomical importance looking for cultivars with more desirable phenotypes for any of these traits. However, due to the long period of time needed and to the difficulties of field trials, there is a certain gap between the molecular and phenotypic evaluation of this diversity. And there is a general need of comparative cultivar field trials in Mediterranean countries. Finally, wild olives are also being characterized at molecular and phenotypic level and the possibility to introgress new and superior alleles into cultivated varieties is under exploration.

Keywords: Olea europaea, genetic patrimony, olive cultivars, identification, evaluation, olasters

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 7

Centennial Olive Trees in Lebanon: a Substantial Patrimony

L. Chalak1, F. Malas1, B. Hamadeh2, L. Essalouh3, B. Khadari4,5

1The Lebanese University, Faculty of Agronomy Sciences, Dekwaneh, Beirut, Lebanon. 2Lebanese Agricultural Research Institute, Fanar Station, Beirut, Lebanon. 3Montpellier SupAgro, UMR 1334 AGAP, 34398 Montpellier, France. 4INRA, UMR 1334 AGAP, F‐34398 Montpellier, France. 5Conservatoire Botanique National Méditerranéen (CBNMED), UMR 1334 AGAP, 34398 Montpellier, France.

This study aimed to assess the centennial olive trees growing across Lebanon, with the perspective of conservation of the ancient germplasm. The survey indicated the existence of numerous centennial olive trees distributed in different agro‐climatic areas, from 80 to 1350 meters altitude across the country. Centennial olives were found in large size orchards and scattered as well as in young orchards, road hedges and gardens for ornamental purposes. Yet, no reliable information is available regarding the age of the centennials, but they can considered as 500 to 1000 years old. Among these, only six centennials located in Bcheale village in the north of the country at 1000 meters altitude are considered as “millenials” or “monumentals” by the Ministry of Tourism, while the remaining ones widespread across the countries are still ignored. As to the preliminary morphological characteristics of the trees conducted on 292 centennials spread in 48 orchards, a large variability was recorded for foot, trunk and central cavity sizes. Principal component analysis showed that foot and trunk circumferences as well as central cavity diameter were the most discriminating descriptors. Most of the 48 orchards were clustered together in one pool sharing similar traits while few others are found out. The most outstanding orchards were located in 10 locations distributed across Lebanon. At the tree level, six single trees located in North and Mount Lebanon were well differentiated by their large sized foot, trunk and central cavity diameter. These centennials should be further characterized using morphological and agronomical descriptors in order to understand their performance through time and to valorize them in selection programs.

Keywords: Olea europaea L., centennials, Lebanon, distribution, trees characteristics.

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Studies on the genetic relationship of wild olive with cultivated varieties and on their resistance to Verticillium wilt in Syria.

B. Barhoum1 , M. Nachit2

1Agricultural Research center at .Latakia, Syria 2ICARDA, P.O.Box 6299, Rabat‐Instituts, Morocco

The Olive tree (Olea europaea) belonging to the family Oleaceae is grown on 10. 5 million hectares; where the Mediterranean basin represents more than 97 % of the world cultivation. The olive tree is originated from the Eastern Mediterranean and moved historically to the Western Mediterranean region. In Syria, the olive tree covers an area close to 650 thousand hectares with a number of trees approaching 100 million trees. The Olive tree is divided into two groups: the wild (O. europaea subsp. europaea var. sylvestris) and the cultivated group (O. europaea subsp. europaea var. europaea). One of the most important diseases affecting olive tree (Olea europaea) is Verticillium wilt; it affects the commercial orchards and landscape plantings; and is caused by soil‐borne fungi Verticillium dahliae . This study was conducted to identify the resistance to Verticillium wilt in 76 wild olive accessions collected from different agro‐ecologies in Syria. In addition, the most cultivated olive cultivars in this country (Sorani and Zaity) were also tested for the resistance. The results showed high level of resistance to Verticillium wilt diseases in the wild olive accessions. The principal components analysis for the morphological characters showed the two cultivars' clustering differently with different wild olive accessions. In addition, 60 SSR were used for genotyping the wild and cultivated olive; the results showed that the olive cultivars were closely related to some of the wild olive accessions collected at the areas of Fakro, Aljarras, and Sighata. This indicates very clearly the possibility that the cultivated olive originated from the wild olive accessions of these 3 sites. These sites are located in the North‐western regions of Syria. These findings are of importance to the olive tree breeding to improve the resistance to Verticillium wilt from the wild olive types, particularly by using the accessions originating from the North‐western areas of Syria.

Keywords: Olive, olea europea, sylvestris, Vericillium wilt, resistance

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How to select olive varieties for association mapping of agronomic and adaptive traits: towards the setup of Mediterranean network for olive phenotyping.

B. Khadari1,6, A. El Bakkali5, L. Essalouh2, M. Gurbuz3, M. Ulas3, M. Hakan3, O. Cetin3, H. Kaya3, N. Mete3, M. Sahin3, L. Sikaoui4, A. Moukhli4, S. Santoni1, E. Costes1

1INRA, UMR 1334 Amélioration Génétique et Adaptation des Plantes (AGAP), F‐34070 Montpellier, France. 2Montpellier SupAgro, UMR 1334 AGAP, F‐34070 Montpellier, France 3Olive Research Station, 35100 Bornova, Izmir, Turkey. 4INRA, CRRA‐Marrakech, UR Amélioration des Plantes, B.P. 533 Menara, Marrakech, Morocco. 5INRA, CRRA‐Meknès, UR Amélioration des Plantes et Conservation des Ressources Phytogénétiques (APCRPG), B.P. 578, Meknès, Morocco. 6CBNMed, UMR 1334 AGAP, F‐34398 Montpellier, France.

Despite the socio‐economic importance of oil olive and the need of olive breeding, genetic studies on agronomic and adaptive traits are restricted to few biparental populations limiting the efficiency of QTL (quantitative trait loci) mapping strategy. Association mapping based on a diversified collection of olive germplasm can be proposed as a complementary strategy. Here, we aimed at defining an olive collection to efficiently link gene variation to phenotype traits taking advantage from the worldwide olive germplasm bank of Marrakech (586 accessions from 14 Mediterranean countries) and Izmir (85 accessions). These 671 accessions were characterized using 24 nuclear SSRs and cpDNA markers and were classified into 535 distinct genetic profiles. We detected 476 nuclear alleles and 17 cpDNA haplotypes and classified this worldwide collection into east, centre and west Mediterranean gene pools. Combining two sampling methods to maximize the capture of diversity and genetic distance, we proposed two core collections of 55 and 118 accessions including all nuclear SSR alleles, cpDNA haplotypes. These cores collections include varieties considered as the most important in Mediterranean olive producing countries and display a limited genetic structure between east and west/center gene pools. Selected olive accessions will be set up as randomized experimental trials in several sites such as INRA Montpellier, INRA Marrakech and ORS Izmir for phenotyping agronomic and adaptive traits. Hence, the defined core collections will constitute good candidates for exploring the largest variability known in distinct environmental conditions.

Keywords: Olea europaea L., domestication, genetic structure, core collection, association mapping, phenotyping

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 10

Towards the identification of SSR marker linked to vigor in olive tree (Olea europea L.)

H. Zaher1, 2, B. Boulouha1, M. Baaziz2, F. Gaboun3 and S.M. Udupa4

1INRA Marrakech, Morocco 2Faculty of sciences Semlalia, Marrakech, Morocco 3INRA Rabat, Morocco 4INRA/ICARDA cooperation, Rabat, Morocco

In this study, a progeny resulting from a cross between "Menara" and "Arbequina" varieties was characterized by both morphological and SSR markers (microsatellites). The technique Bulk Segregant analysis (BSA) was used to establish linkage between molecular markers and vigor parameters (tree height, perimeter of trunk diameter and canopy circumference). Thirty SSR primers were used to compare the parents and bulks of individuals of different vigor. Eleven SSR primers were selected for their ability to detect polymorphism between the two parents and bulks. Results of morphological analysis showed highly significant genotypic differences for all measured parameters. Principal component analysis revealed highly significant phenotypic variation where the first two axes represented 85.32 % of the total variation. For association analysis, results obtained by the General Linear Model showed that some of the used SSR markers are significantly associated with the phenotypic traits studied. Two SSR markers, DCA9 (p <0.05) and GPAU59 (p <0.05), were found significantly associated with tree height. Also PA2 (p <0.05) and GPAU59 (p <0.05) were associated to the perimeter of trunk diameter. Finally GPAU59, DCA9 and DCA18 were linked with the canopy circumference. These results are of great importance and incite to use these primers association analysis in characterizing our olive progeny.

Keywords: Olea europea, morphological characterization, molecular marker, SSR, association analysis

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Search for new duel use olive clones in the Central East of Tunisia: Characterization for pomological and chemical traits

I. Laaribi1,2, M. Mezghani Aïachi1, H. Gouta1, M. Ayadi3, F. Labidi1, M. Mars2

1Institut de l’Olivier, station spécialisée de Sousse, BP 14, 4061, Ibn Khaldoun, Tunisia. 2U.R. Agrobiodiversité, Département des Sciences Horticoles, Institut Supérieur Agronomique Chott Mariem Sousse 4042, Tunisia. 3Institut de l’Olivier, BP 1087, 3000 Sfax, Tunisia .

The region of Sousse ‘Capital of the Tunisian Sahel’ is subjected to a high risk of olive genetic erosion due to the rapid and uncontrolled urbanization. Moreover, this region is a historical location of olive biodiversity which has been relatively neglected in the literature sources of olive characterization. In order to evaluate and exploit olive diversity in this region, a prospecting effort was carried out in many old olive groves. In the present work, we performed pomological and chemical traits of ten accessions selected for their fruit characteristics. Results showed that all studied accessions have medium fruit weight that ranged from 2.13g to 3.57g. Flesh to stone ratio and flesh percentage varied from 4.86 to 7.97 and from 82.94 % to 88.82 % respectively. Olive oil content was average to high for most tested accessions, varying from 14.92% to 29.79%. Regarding fatty acid composition the majority of the tested olive oils were classified as extra‐ virgin oils due to their high oleic acids (from 55.38% to 76.39%) and low palmitic and linolenic acids contents. Analysis of polyphenols and tocopherols showed a significant variation between samples; they varied respectively from 42.2 ppm to 943.1 ppm and from 149.5 ppm to 534.7 ppm. These findings are of great interest while they underline the importance of chemical and molecular analysis to enhance effort to preserve such valuable germplasm and to protect many specimens that can be valorized as oil and table olive and enrich a patrimony threaten by genetic erosion.

Keywords: Olea europaea, Sousse region, morphological traits, chemical traits, oil‐table olive.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 12

Development of key for identification of seven olive varieties using microsatellite markers

M. Iqbal, T. Mehmood2, A. Mehmood, S. Jamil

1Director Agricultural Biotechnology Research Institute, AARI, Faisalabad, 2Director Agricultural Biotechnology Research Institute, AARI, Faisalabad, Director 3Barani Agricultural Research Institute, Chakwal. 4Director Cotton Research Institute, AARI, Faisalabad.

The present study was carried out to develop a standardize reference, based on DNA fingerprints for the identification of seven olive cultivars (Sohawa selection, Manzanilla, Bulkasar selection, Chugtai selection, Arbequina, Arbosana and Koroneikai) using SSR markers for their ability to discriminate between genotypes and to verify cultivar identity using database of DNA fingerprints. These genotypes have been planted at Barani Agricultural Research Institute (BARI), Chakwal, Punjab, Pakistan, introduced from abroad or collected from farmer field introduced by the desirous tourists or pilgrims visiting Syria, Iraq and Iran. The BARI is involved in vegetative propagation of these olive varieties and provides nursery plants to the farmer for commercial orchid planting. The aim of this study was to develop molecular identification key of these varieties to identify at nursery level and quantification of genetic similarities among these varieties. Simple Sequence Repeat (SSR) molecular marker system was applied for molecular identification and genetic diversity of these olive genotypes. Among the 58 surveyed markers 51 were polymorphic and percentage of polymorphism was 87.9%. The 51 polymorphic primers produced 551 alleles out of which 19 were monomorphic and 532 were polymorphic. The number of alleles per SSR varied from 1‐23 with an average of 10.8 alleles per locus. Two SSR primers (OR‐2 & UDO‐26 or OR‐2 & GAPU‐71A or OR‐2 & UDO‐42 or OR‐2 & DCA‐7) used in various combination can distinguish all the seven olive varieties under study. Three SSR primer pair, i.e., UDO‐3, UDO‐25 and DCA‐4 can also discriminate all the seven olive varieties from each other. Variety specific alleles were also found in all the varieties except Bulkasar selection and Arbosana. Statistical approach for primer efficiency, i.e., Confusion probability (Cj), Discriminating power (Dj) and Discriminating power (DL) when N tends towards infinity also confirm these findings. Clustering analysis yielded seven different clusters when drawn on 70% similarity as base line. Genetic similarity values ranged from 0.545 to 0.668.

Keywords: Olea europae, Simple Sequence Repeat, diversity, variety identification.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 13

Exploring and characterizing unknown olive variants

R. Mariotti1, S. Mousavi1, M. Hosseini‐Mazinani2, B. Torkzaban2, N. Cultrera1, S. Pandolfi1, J.A. Googlani1, L. Baldoni1

1Institute of Biosciences and Bioresources (IBBR), CNR, 06128 Perugia, Italy 2National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran

It has been recently demonstrated that the Iranian olive germplasm, even if poorly explored in the past, represents an invaluable new source of variation for olive production and breeding. Olive tree samples, occurring as few or even single trees, abandoned or living under wild or semi‐natural growing status, mostly under extreme climate and soil conditions, have been collected from different provinces, analyzed at molecular level and phenotypically characterized. They certainly represent a broad repertoire of olive ecotypes, holding a high potential value not only for characters such as climate and soil stress resistance, but also for their high fruit quality (high oil content, big fruit size, etc.). When analyzing the OeACP2 gene, previously demonstrated to contribute to the fatty acid synthesis in olive fruit mesocarp, it resulted that some Iranian ecotypes carry unique allele combinations. Data on rooting ability, recorded on 100 ecotypes, showed that most of them had more than 85% of rooting percentage. The physiological evaluation of cold resistance, performed on a subset of Iranian ecotypes, demonstrated a good level of resistance when compared to outstanding Mediterranean cultivars. Finally, experiments to evaluate the level and type of salt tolerance are currently ongoing on seedlings and clonally propagated ecotypes collected from salty soils.

Keywords: Olea europaea, olive ecotypes, SSRs, SNPs, salt resistance, rooting

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 14

The World Olive Collection of Cordoba: 40 years of experience on management of olive genetic resources

A. Belaj1, J. Hidalgo1, G. Beltrán2; J. Cano2, A. Jiménez2, I.Padilla3, A. Barceló3, G. M. García‐Ruiz1, J. Lopez‐Escudero4, S. G. Atienza5, L. Léon1, R. de la Rosa1

1IFAPA Centre “Alameda del Obispo”, Córdoba, Spain 2IFAPA Centre “Venta del Llano”, Mengibar, Jaén, Spain 3IFAPA Centre de Churriana, Málaga, Spain 4Departamento de Agronomía, ETSIAM, Universidad de Córdoba, Spain 5Instituto de Agricultura Sostenible (IAS). CSIC. Córdoba. Spain

Germplasm collections are basic tools for conservation, characterization and efficient use of olive genetic resources. In this sense, the establishment in the 70s of the World Olive Collection of Córdoba, Spain, represented the first international attempt of conservation and management of the olive germplasm. Nowadays this Collection counts 885 accessions from 24 countries, most of them from Mediterranean Basin. In order to ensure the conservation of this rich patrimony, a security copy of this collection is being established at IFAPA Centre “Venta del Llano”, Mengibar, Jaén. At the same time, in vitro propagation of olive cultivars is also being carried out. The identification of the olive cultivars maintained in this collection is an important ongoing task. Different types of markers such as RAPDs, AFLPs, SSRs, and more recently SNPs and DArTs have been successfully used for such purpose revealing thus synonymy and homonymy cases, mislabelling and propagation errors in the collection. These markers have evidenced high levels of the diversity in the olive germplasm. The olive cultivars maintained in the World Olive collection have systematically been evaluated for several pomological traits related to vigour, phenology, production, fruit quality and for their resistance or susceptibility to Verticillium dahliae Kleb and other diseases. In addition, these cultivars have been evaluated for their olive oil quality and composition as well as sensorial characteristics. All the above mentioned evaluations have also revealed high levels of variability among the olive cultivars. In spite of the great importance of such evaluations, they only represent the first step to determine the most interesting cultivars for their use as potential parents in future crosses of the olive breeding program as well as for the establishment of comparative trials in different agro‐climatic conditions. In this sense, some selected olive cultivars of this collection are being evaluated in 7 different locations of Andalusia, Southern Spain,). The activities carried out in the World Olive Collection of Córdoba are fruit of collaboration between different research teams and scientific institutions.

Key words: Olea europaea, germplam bank, olive cultivars, identification, evaluation

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 15

Preliminary Evaluation of Oil Content and Fatty Acids Profile for Olive Varieties Cultivated in Lebanon

M. El Riachy1, M.Breidi1. G. Abou‐Sleymane2, L.Chalak3

1Olive Oil Laboratory, Lebanese Agricultural Research Institute (LARI), Tal Amara, Lebanon 2Faculty of Health Science, American University of Science and Technology (AUST), Achrafieh, Lebanon. 3Faculty of Agronomy, The Lebanese University (LU), Dekwaneh, Beirut, Lebanon.

Fatty acids (FAs) composition represents critical aspect that affects virgin olive oil (VOO) quality, as the balance of concentrations of saturated, monounsaturated and polyunsaturated FAs has a direct impact on oil stability and health benefits. Besides, oil content represents an indicator of the oil yielded from each variety. The present work aimed at characterizing the oils of 12 olive varieties growing in different agro‐climatic conditions of Lebanon. Olive fruits were harvested in autumn 2013 and olive oils were extracted using an Abencor system. Oil content in humid and dry matter of the crushed olives was determined by using Soxhlet method. The percent of esterified FA was determined by using gas chromatography with flame ionization detection (GC– FID). The results showed no qualitative differences in FAs profile from different varieties. However, quantitative differences were observed mainly in case of major FAs, namely: oleic acid ranged between 64.88% (Baladi Ijd Ibrine) and 73. 75% (Bou Chawkeh Fakehe); Palmitic acid between 10.42% (Bou Chawkeh Fakehe) and 15.53% (Nabali Kfarchakhna); linoleic acid between 7.76% (Roumani Qana) and 15.10% (Baladi Ijd Ibrine); and stearic acid between 1.86% (Bou Chawkeh Fakehe) and 4.49% (Soury Iaal). As to the oil content in dry matter, Baladi Ijd Ibrine showed the higher content (54.27%) and Baladi North the lowest (23.84%). This preliminary characterization indicates an important variability among oils extracted from different varieties; it should be complemented by a more complete characterization based on fruit harvesting and oil characterization at different ripening stages.

Keywords: Olea europaea, local varieties, olive fruits, virgin olive oil, fatty acids, oil content

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 16

Evaluation of some olive cultivars under Kuwait environmental conditions

H. Al‐Menaie, O. Al‐Ragam, H. Mahgoub, M. Al‐Hadidi, A. Al‐Shatti and M. Al‐Zalzalah

Environment and Life Sciences Research Center, Kuwait Institute for Scientific Research, PO Box 24885, Safat 13109, Kuwait, Tel: (+965) 24989806, Fax: (+965) 24989809

Olea europaea, a native plant of the Mediterranean region became a cynosure among agronomists worldwide, owing to its health beneficial properties and tolerance to harsh environmental conditions, along with its economic importance. The prevailing climatic and environmental conditions constitute one of the major factors affecting the productivity of different olive cultivars. Hence, an adequate knowledge of the best olive cultivars in the arid area is mandatory to efficiently utilize them for greenery development, desertification control, table olive, and olive oil production. As a preliminary effort, twenty‐ six olive cultivars were subjected to performance evaluation in coastal sites (Salmiya) of Kuwait. Various growth parameters such as plant height, stem girth, number of shoots, and length of shoots were clearly examined. It was found that except the cultivar Shauki, all other cultivars under observation survived under Kuwait environmental conditions. Moreover, all the varieties flowered and produced fruits, indicative of their ability to reproduce under harsh climatic conditions of Kuwait. This study on the suitability of various olive cultivars under Kuwait harsh climatic conditions outlined the superior performance of certain cultivars such as Picual, Picholine, Pendulino, Coratina, Arbequina, and Frantoio in the coastal sites of Kuwait.

Keywords: Olea europaea, horticultural practices, Kuwait.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 17

Evaluation of nutrient uptake in different olive cultivars grafted on Gemlik rootstock

M. Azimi1, M. Ozkaya1, H. Çolgecen2, H. Buyukkartal3

1Ankara University, Faculty of Agriculture, Department of Horticulture, 06110 Diskapi Ankara, Turkey 2Bülent Ecevit University, Faculty of Arts and Science, Department of Biology, 67100 Incivez, Zonguldak, Turkey 3Ankara University, Faculty of Science, Department of Biology, 06100 Tandoğan, Ankara, Turkey

The olive (Olea europaea L.) varieties are cultivated by grafted or self‐rooted plants. Gemlik cvs is an easy‐to‐root variety so can be used as a clonal rootstocks. However there could be a grafting incompability problems between cvs. So there could be some problems in vascular system. Self‐ rooted Ayvalik (A) and Gemlik (G) cvs and six grafting (T bud) combinations; Ayvalik/Gemlik (A/G), Domat/Gemlik (D/G), Gemlik/Gemlik (G/G), Memecik/Gemlik (M/G), Nizip Yaglik/Gemlik (N/G) and Sari Ulak/Gemlik (S/G) were used as plant materials. These plants had been grown in the greenhouse for the first two years and planted to the pots include a compost of soil:sand:manure (1:1:1). According to the plant nutrient analysis it was observed that there were significant differences (p<0.05) in self‐rooted cvs and grafted cvs. In evaluation of average data in all macro and micro elements, it was observed that Gemlik and Gemlik/Gemlik have higher amounts of plant nutrients than Ayvalik and other grafting combinations. However the lowest amount of nutrients were observed in Ayvalik/Gemlik and Domat/Gemlik. According to cluster analysis for all nutrients, Ayvalik/Gemlik and Domat/Gemlik were located in same group which has lowest amount of nutrients. On the other hand other grafting combinations with Ayvalik and Gemlik self‐ rooted cvs were located in another group. According to the results for mean value and cluster analysis, there is a problem in grafting region in both Ayvalik/Gemlik and Domat/Gemlik cvs. As a result, may be there are some problems in proliferation of cambium cells and formation of vascular system.

Keywords: Olea europaea, graft combination, Ayvalik, Domat, nutrient uptake

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 18

Histological evaluation of graft compatibility in Olea europaea L. scion/rootstock combinations

M. Azimi1, H. Colgecen2, M. Ozkaya1, H. Buyukkartal3

1Ankara University, Faculty of Agriculture, Department of Horticulture, 06110 Diskapi Ankara, Turkey 2Bülent Ecevit University, Faculty of Arts and Science, Department of Biology, 67100 Incivez, Zonguldak, Turkey 3Ankara University, Faculty of Science, Department of Biology, 06100 Tandoğan, Ankara, Turkey

The plant material in this study consisted of cultivars ‘Ayvalik’, ‘Gemlik’, ‘Domat’, ‘Memecik’, ‘Nizip Yaglik’ and ‘Sari Ulak’. These cultivars were grafted (T budding) onto one year‐old ‘Gemlik’ rootstock propagated by cuttings. The grafted plants samples were selected 3, 6 and 12 months after grafting, then were protected using formalin aceto alcohol (FAA) solution by application at 1 cm above and below the graft. The grafted samples were sectioned using a microtome to obtain 30 µm‐thick sections for microscopy and investigated with light microscope. In all graft combinations dark brown necrotic regions along the lines of the graft in the callus tissue borders were also observed. Phenolic compounds causing necrosis in the graft zone were also identified. Results showed that in this study reports the extent of callus growth, cambial differentiation, cambial continuity, sclerenchyma tissues and formation of vascular tissues of these olive cultivars in applied grafts. A good callus growth was observed in the cambial development of vascular tissues in these samples. The scions of the ‘Gemlik’, ‘Memecik’ and ‘Sari Ulak’ cultivars were compatible when grafted onto 'Gemlik' rootstock. Conversely, the scions of ‘Ayvalik’, ‘Domat’ and ‘Nizip Yaglik’ grafted onto 'Gemlik' rootstock induced limited callus and cambial development, sclerenchyma tissues and vascular tissues. Moreover, in air pockets zones of grafts founded callus tissues after 6 and 12 months of grafts procedure, resulting in weak grafts.

Keywords: Olea europaea, T budding, Domat, Ayvalik, cambial differentiation.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 19

Somatic embryogenesis and plant regeneration from radicles of olive (Olea europaea L.) zygotic embryos, cv. Chemlal.

K. Titouh1,2, K. Hadj Moussa2, M. Khelifi‐Slaoui2 and L. Khelifi2

1National Institute of Agronomic Research (INRAA), Centre of Béjaïa, Algeria. 2High National School of Agronomy (ENSA), Laboratory of Genetic Resources and Biotechnologies (LRGB), Algiers, Algeria.

The olive tree (Olea europaea L.) is one of the most important fruit crops in the Mediterranean region. However, this species is attacked by several diseases and pathogens causing considerable losses in yield. Genetic control is the solution of choice for these problems. Nevertheless, olive improvement by classical methods is difficult due its long juvenile period. Therefore, the use of biotechnological tools could be of great importance for the improvement of this species. However, such techniques require the use of plant material with good quality as well as the establishment of an effective approach of regeneration. Thus, the aim of our work is to establish a protocol of regeneration via somatic embryogenesis from radicles of zygotic embryos of the principal cultivar in Algeria ‘Chemlal’. Callogenesis induction was achieved in olive medium (OM) (Rugini, 1984) or MS solid medium (Murashige and Skoog, 1962) supplemented with kinetin combined with 2,4‐dichloro‐phenoxy‐acetic acid (2,4‐D), indole‐3‐acetic acid (IAA), naphthalene‐ acetic acid (NAA) or indole‐3‐butyric acid (IBA) or in presence of IBA combined with 6‐ benzyladenine (BAP), kinetin or zeatin. After 3 weeks of culture in complete darkness, explants were transferred to the same media without cytokinin and with lower concentration of auxin. After two months of culture, the obtained calli were maintained on olive cyclic embryogenesis solid medium (ECO) (Pérez‐Barranco et al., 2007) with monthly subculture until the appearance of embryogenic structures. Our results indicate that the induction of callus from radicles of zygotic embryos was strongly influenced by hormonal balance applied, harvest period of olives and the culture conditions. In fact, more embryogenic calli were obtained after induction in presence of IBA or NAA combined with zeatin. In addition, OM medium was more effective to obtain embryogenic calli than MS medium. Somatic embryos (SE) differentiation and maturation were achieved on ECO solid medium supplemented with 1 g l‐1 activated charcoal for two months. SE germination carried out on a modified MS solid medium allowed to obtain plantlets without any phenotypic anomalies.

Keywords: Callogenesis, somatic embryogenesis, olive, radicle, In vitro regeneration.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 20

Varietal diversity of the olive tree in the Chefchaouen region (Northwest of Morocco)

M. Ater1, H. Barbara1, J. Kassout1, M. El Omari2

1Faculté des Sciences de Tétouan. Avenue Khénifra. BP 2062. 93030. Tétouan. Morocco. 2Direction Provinciale de l’Agriculture. Chefchaouen. Morocco.

The Chefchaouen region (northwestern Morocco) is a representative area of rural landscapes of the Rif mountains and traditional agroecosystems. The forest area is large and covers 35% of the territory. Agricultural Area (SAU) corresponds to 40.6% of the territory where the tree dominates with 77% of the SAU. The cultivation of olive trees is important and occupies 26.5% of the SAU of the region. Surveys were conducted with farmers in 15 villages (dchars) of the study area. The results show a varietal diversity in olive particular through the continued cultivation of local varieties. However, it should be noted that local varieties are used by 44 % of farmers surveyed, while selected varieties (Haouzia and Mnara essentially) are on the increase and are used by 64.5% of farmers. The Zeitoun Beldi variety which corresponds to the " Moroccan Picholine " is the dominant variety with a frequency of 0.79. The other varieties are relatively rare as Bouchouk (0.35 ), Meslal (0.17), Homrani (0.9), Semlal (0.2) and Khoubzi (rare). Generally, these varieties are propagated by grafting on oleaster and rarely by cuttings. This diversity is maintained despite massive introductions of selected varieties programs. Note that the rest of the Moroccan territory, varietal diversity of the olive tree is smaller and the « Moroccan Picholine » is the dominant variety. Spontaneous form (oleaster) is integrated into the operation and management of traditional orchards. This is explained by the traditional practices of farmers commonly use not only for the grafting of local varieties of olive trees but also for the production of oil. Socio‐ economic transformations underway disrupted the practice of polyculture and threaten the maintenance of agrodiversity.

Keywords: Olivier, varietal diversity, Oleaster, Agrodiversity, Chefchaouen

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 21

Flowering Phenology in national olive germplasm of Iran

A. Zeinanloo1 and K. Gharibi2

1Horticulture dept , Seed and plant improvement institute Karj , Iran 2Agriculture insurance foundation, Tehran, Iran

More than 100 local olive genotypes were collected from different provinces of Iran and this collection was established in Augmented design with 4 olive cultivar (‘Roghani’, ‘Zard’, ‘Koroneiki’ and ‘Conservolia’) as control in olive research station of Tarom in 2005. In this research investigated phenology of flowering of many of these genotypes. Traits investigated in this study were the time of inflorescence buds start to swell, growth and development of inflorescences, flower opening, full bloom and petal fall. The result showed that the time of emergence of inflorescence was related to cultivar or genotype. The cultivars of ‘Roghani’ and ‘Koroneiki’ were the earliest and of ‘Kh11’ and ‘Bash’ had the latest start time of inflorescence buds swelling. Among the genotypes the Flower opening time was different and the genotype of ‘Bash’ had the latest flower opening. The genotype of Ds7 had earliest petal fall and the latest were ‘Derak’ and ‘Bash’. Flowering period among genotypes was 10 days. The Comparison of means indicated that the most perfect flower were belong to genotype of ‘Qg28’ with65% and cultivar ‘Koroneiki’ with 60.7. The genotype of NO12 with 3% had the lowest amount of perfect flowers.

Keywords: Olive, phenology, flowering, genotypes

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 22

Assessment of autochthon Tunisian olive varieties by six AFLP markers S. Mnasri1, O. Debbabi1, M. Ben Saleh3 and A. Ferchichi 2

1National Gene Bank of Tunisia, Street Yesser Arafet, 1080, Tunis, Tunisia 2National Institute of Agronomy of Tunisia, 43 Street Charles Nicolle, 1082‐Tunis 3Institute of Arid Regions of Gabes, 6051 Nahal Gabes Tunisia

In Tunisia, little is known about the existing olive patrimony. The number of local cultivars is probably underestimated because of the scarce information on Tunisian olive biodiversity. The aims of this work were to make a molecular characterization of thirty local cultivars with the use of six different AFLP primer combinations. Auto radiographs revealed 92 polymorphic markers in a total of 237 detected fragments. A set of redundant marker patterns was identified and deleted from the binary data matrix; data analysis demonstrated a high degree of polymorphism with an average of 38.8%. The analysis of AFLP profiles found in our set of olive cultivars showed a wide genetic diversity among olive germplasm. The UPGMA cluster analyses using Jaccard’s index and the Principal coordinate analysis (PCO) revealed that the genetic diversity was predominantly structured according to the qualitative morphological parameters of the fruit and the endocarp. The data obtained can be used for the varietal survey and construction of a database of autochthon olive varieties in Tunisia and providing also additional information that could form the basis for the national design of breeding programs.

Key words: AFLP, autochthon varieties, genetic diversity, olive.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 23

Olive tree architecture as a tool in early selection for short juvenile period

S. Hammami1,3, H. Rapoport1, M. Ben Mimoun3, L. León2 , R. De La Rosa2

1Instituto de Agricultura Sostenible (CSIC), Avda. Menéndez Pidal s/n, 14004 Córdoba. Spain. 2IFAPA Centro “Alameda del Obispo”. Avda. Menéndez Pidal s/n, 14004 Córdoba, Spain 3Institut National Agronomique de Tunisie, 43 av. Charles Nicolle, 1082 Tunis, Tunisia

A critical step for olive breeding is the early elimination of seedlings with a long juvenile period. Basic parameters of architecture, such as seedling height have been strongly correlated with juvenile period duration in the olive tree. In other fruit trees, such as apple and pear, additional, more detailed growth‐habit parameters have also been associated with the transition to flowering. The purpose of this study is to evaluate relationship between juvenile period duration and more detailed qualitative and quantitative olive tree architecture parameters. We evaluated a population of 860 seedlings from six different crosses of seven parental cultivars and open pollination. The seedlings were grown in a greenhouse under optimum conditions and without any pruning. At 11 months, just prior to transplanting to the field, the number of the lateral shoots (quantitative parameter) was recorded and the branching rhythm (qualitative parameter) was described. All defined branching rhythms (continuous, rhythmic, diffuse) were present in varying degrees in the different crosses, but the “diffuse” rhythm, in which there is no regular pattern of which nodes form lateral branches, was clearly dominant. Two years after transplanting, only plants with "diffuse" branching rhythm produced flowers. The following year, both “diffuse” and “continuous” branching rhythms showed the highest percentage of flowering plants. In addition, plants which were flowering by this year had produced significantly more lateral shoots than those still juvenile.. Those studies will aid in the development of early selection criteria for short juvenile period in olive breeding programmes. Furthermore, the implication of their use for the description and selection of growth habit is discussed.

Keywords: growth habit, olive seedling, lateral shoot, branching rhythm.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 24

Use of morphological descriptors and principal component analysis for studying the variability of olive descendants issued from controlled pollination

I. Laaribi1,2, M. Mezghani Aïachi1, F. Labidi1, M. Mars2

1Institut de l’Olivier, station spécialisée de Sousse, BP 14, 4061, Ibn Khaldoun, Tunisia. 2U.R. Agrobiodiversité, Département des Sciences Horticoles, Institut Supérieur Agronomique Chott Mariem Sousse 4042, Tunisia

Morphological descriptors are a preliminary identification tool to exploit in breeding programs and also represent the first choice used for describing and classifying genotypes. In this study, one hundred twenty four descendants issued from thirty nine genetic combinations, were investigated and characterized by combining twenty six quantitative and qualitative morphological descriptors. In order to evaluate the phenotypic variability of the genetic material and the discriminate power of the descriptors, cluster analysis and principal component analysis were used. Correlations between tree, leaf, fruit and endocarp characteristics were also studied. The first three principal compounds (PC1, PC2 and PC3) accounted for 26.2, 17.2 and 9.8% of the total variance respectively and accumulated 43.8% of the variability. The PC1 was correlated positively with fruit and endocarp size and negatively with leaf and tree dimensions. The PC2 was due to the contribution of fruit and endocarp shape. The PC3 was correlated to the leaf shape and endocarp grooves parameters. The most discriminate descriptors, are respectively: leaf width, endocarp shape, leaf size, endocarp weight, endocarp width, endocarp length, fruit length, fruit shape, fruit symmetry and fruit width. UPGMA dendrogram allowed to group descendants into five main clusters according to their morphological similarities. No clear association between genetic combinations and morphological similarities was founded. Molecular markers should be considered in a future work to provide a more complete understanding of the morphological diversity.

Keywords: Olea europaea, morphological descriptors, controlled pollination, discrimination power.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 25

Characterization and performance of selected olive (Olea europaea. L) seedlings issued from a Tunisian breeding program

I. Guellaoui1, F. Ben Amar1, M. Boubaker2, M. Ayadi1, R. Ayadi1, A. Yengui1, N. Ben Belgacem1

1Institut de l’Olivier, Sfax, Tunisia 2Institut Supérieur Agronomique de Chott‐Mariem, Sousse, Tunisia

The Tunisian olive sector is dominated by two main oil varieties, Chetoui in the north and Chemlali Sfax in the centre and the south of Tunisia. Chemlali Sfax is highly productive and well adapted to the arid conditions. However, its oil contains high level of palmitic acid and relatively low level of oleic acid. Therefore, crosses of Chemlali Sfax with Tunisian and foreign varieties have been undertaken. A preliminary selection of hybrids on the basis of the fatty acid composition has been made which are planted in an irrigated and intensive orchard in the Sfax region (Tunisia) since 2005. In this study, we proposed to characterize seven hybrids on morphological, agronomic and technological plans. The morphological parameters concerned the leaf, the fruit and the pit. Agronomic characterization deals with the fruit weight, the flesh/pit ratio, the maturity index (November) and the olive production per tree. Technological analysis was done for acid composition. The follow‐up has concerned three trees by hybrid during three years. The morphological characterization has shown differences and similarities between the hybrids for most of the parameters. Each hybrid was characterized by specific parameters mainly at the level of the fruit and the pit. The agronomic characterization showed an important variability. The average fresh fruit weight varies from 1.27 to 3 g and the flesh/pit ratio from 2.1 to 8.5. The maturity index in November (scale from 0 to 7) varies widely from 1 to 4.65. The olive production for the period 2007‐2013 was between 0.34 and 6.86 kg per tree with high level of alternation bearing. Technological characterization revealed an important gain for acid composition in comparison with the original variety. So, the rate of palmitic acid does not exceed 15 % and that of the oleic acid is higher than 70 %. This work has allowed us to characterize the pre‐selected hybrids and will greatly help to make a final selection of new olive varieties in Tunisia in few years.

Keywords: Chemlali Sfax, cross pollination, morphology, agronomy, technology, characterization.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 26

Selection of the best olive genotypes from several crosses among cultivated varieties

B. Alfei1, A. Paoletti2, A. Rosati2, A. Santinelli1, G. Pannelli2

1Agenzia per i Servizi nel Settore Agroalimentare della regione Marche, ASSAM, Ancona, Italy 2Consiglio per la Ricerca e la sperimentazione in Agricoltura, Centro di ricerca per l'olivicoltura e l'industria olearia (CRA‐OLI), Via Nursina 2, 06049 Spoleto, PG, Italy

Sixty four genotypes obtained from a breeding program initiated 60 years ago in central Italy by crossing six olive varieties (Ascolana tenera, Carboncella, Dritta, Gentile di Chieti, Intosso e Leccino), with the aim to select genotypes with superior traits, were evaluated in this study. Both vegetative and productive parameters were examined for these genotypes in a field in Carassai and another one near Jesi. Ten genotypes were selected on the basis of their yield. Here we report on the characteristics of trees (vigour, habitus, age of first bearing, total cumulative yield), fruits (weight, oil content, evolution of fruit coloration over time, pulp firmness), oil (sensory parameters, fatty acid composition, phenols content), susceptibility to biotic and abiotic stresses (cold, peacock eye), and suitability to different harvesting methods (combs, trunk shakers, over‐ the‐row harvester). In addition to the highest yielding genotypes, one genotype was selected for a high yield efficiency (yield per unit of trunk cross sectional area). The genotype with the highest cumulative yield, considering both fields, had the lowest polyphenols and oleic acid content. The selected genotypes represent promising material to be further tested, comparing their yield and quality performance with that of parents and other cultivars, before a final judgment can be made.

Keywords: cross‐breeding, genetic improvement, productivity, yield efficiency, oil quality

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Chemical and sensory properties of monovarietal olive oils produced by ancient genotypes in Emilia‐Romagna region

L. Morrone1, A. Rotondi1, D. Beghè2, T. Ganino2, A. Fabbri2

1IBIMET‐CNR Via Gobetti 101, 40129 Bologna, Italy 2Dipartimento di Scienze degli Alimenti, Università degli Studi di Parma, Viale delle Scienze, 59/A 43100 Parma, Italy

Olive trees (Olea europaea L.) growing in the Emilia territory (Italy) and assessed by SSR markers are characterized by an interesting genetic diversity in the studied genotypes. During genotypes selection the chemical and sensory characterization of monovarietal olive oils produced by the studied accessions represents a key factor. During several consecutive crop seasons the products of the most interesting accessions, located in the four western provinces of the Emilia‐Romagna region, were processed using a low‐scale mill. Chemical and sensory properties of monovarietal olive oils were evaluated according to EC Reg. 1513/2001 and 796/02. Some oils were distinguished for their elevated contents in total phenols, other samples were also interesting by their high contents in oleic acid, above 80%. Sensory analysis emphasized the aromatic peculiarities of some monovarietal oils, characterized by a relevant intensity of fruitiness, bitterness and pungency. Moreover oils were singled out for their particular flavours (almond, tomato, artichoke and green notes), ascribable to the different genetic matrix of the mother plants.

Keywords: monovarietal olive oil, genetic matrix, SSR, phenols, sensory analysis, Olea europaea

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Evaluation of pomological traits for some olive cultivars in Tarom Region, Iran

S.Vatani1, M. Taheri2, A. Gholiyan1 , Z. yousefi3, A. A. Zeinanloo4

1Department of Horticulture, Rasht Branch, Islamic Azad University, Rasht, Iran. 2Agricultural and Natural Resources Research Center of Zanjan province, Zanjan, Ira 3Agricultural and Natural Resources Research Center of Guilan , Rasht, Iran 4Department, Horticulture. Seed and Plant Improvement research Institute, Karaj, Iran.

Olive (Olea europaea L.) is one of the most important horticultural crops in Zanjan province. The aim of this study conducted in 2012 was to investigate the important characteristics of flowers and fruits for four commercial olive cultivars growing in Tarom region: Cailletier, Mission, Manzanilla and Conservolia. Some vegetative and reproductive traits were measured and then analyzed and evaluated by SAS program.Variance analysis showed that there was a significant difference between the varieties with respect to fruit‐set percentage, yield, fruit weight, fruit flesh percentage, flesh to stone ratio, dry matter oil percentage. Manzanilla cultivar had the highest yield and dry matter fruit oil percentage; whereas Conservollia had the highest percentage of fruit‐set, flesh percentage, flesh to stone ratio and fruit weight among the studied varieties. Cailletier cultivar showed the lowest values.

Key words: olive, cultivar, yield, oil percent, Iran

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Preliminary results of olive improvement using gamma radiation

A. Chaari1, M. Maalej2, A. Chelli‐Chaabouni3, S. Baccari1

1Laboratoire de l’amélioration et la protection des ressources génétiques de l’olivier, Institut de l’olivier ‐ Université de Sfax, Tunisie. 2Laboratoire de l’amélioration des ressources génétiques, Faculté des Sciences de Sfax ‐ Université de Sfax, Tunisie. 3Laboratoire de l’amélioration et la protection des ressources génétiques de l’olivier, Institut national de Recherche Agronomique de Tunis ‐ Université de Carthage, Tunisie.

In light with the fast and severe climate change, olive genetic improvement is crucial to make available ecotypes that respond to current and future needs of the Tunisian farmers. Application of Gamma rays on olive may be at the origin of obtaining new interesting cultivars. Thus, in vitro vegetative explants (axillaries buds) of Chemlali Sfax variety have been subject to a mutagenic treatment by gamma rays at increasing doses (0.5, 10, 15, 20, 25, 30, 40 and 50 Grays). After 8 weeks of in vitro culture, results showed that doses of 5 ,10 and 15 Gray allowed the regeneration of explants with the highest viability rates respectively 95, 90 and 90 %. After three subcultures in MM medium, shootlets with an average length of 2 to 3 cm with 3 to 4 nodes were regenerated. Subsequent rooting in ex vitro conditions resulted in thirty well hardened plants which were transferred in the field. After 2 years, seven olive trees are in bloom.

Keyswords: Olive, gamma radiation, in vitro culture

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454 pyrosequencing of Olive (Olea europaea L.) transcriptome in response to salinity

P. Kalaitzis1, C. Bazakos1,2, M. Manioudaki1, E. Sarropoulou3

1Department of Horticultural Genetics and Biotechnology, Mediterranean Agronomic Institute of Chania (MAICh), Crete, Greece. 2Department of Horticulture, Aristotle University of Thessaloniki, Thessaloniki, Greece. 3Institute of Marine Biology and Genetics, Hellenic Centre for Marine Research, Heraklion, Crete, Greece.

The continuous expansion of olive cultivation in irrigated lands and the increase in the utilization of low quality, mostly saline, water for irrigation during the summer period slowly leads to highly saline soil causing severe problems to olive cultivation. Our knowledge of the response of olive trees to salinity is restricted to physiological level where numerous studies are available. The up to day lack of Olea europaea reference genome sequence prevents contemporary approaches towards the identification and characterization of regulatory genes. Therefore high‐throughput transcriptome sequencing is emerging as a rapid, efficient and attractive alternative to microarrays approaches for gene expression studies. Towards this direction, we used Roche/454 GS FLX pyrosequencing platform to investigate the molecular response of the salt‐tolerant olive cultivar, Kalamon, in order to characterize the transcriptome profile of roots and leaves and identify differentially expressed ESTs that are related to salt tolerance response. A total number of 28270 unique transcripts were identified of which 35% are annotated. This 454‐based high throughput transcriptome analysis resulted in the identification of salt response related transcripts while a proportion of the differentially expressed transcripts were also annotated providing tools for further molecular studies in olive.

Keywords: Olea europaea L., transcriptome, salinity stress, 454 pyrosequencing

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Assessment of variability for three olive varieties in Montenegro

B. Lazović1, M. Adakalić1, D. Bandelj2,3, T. Perović1

1University of Montenegro, Biotechnical Faculty, Center for Subtropical Cultures, Topolica bb, 85000 Bar, Montenegro 2University of Primorska, Science and Research Centre, Institute of Olive culture, Garibaldijeva 1, SI‐6000 Koper, Slovenia 3University of Primorska, Faculty of Mathematics, Natural Sciences and Information Technologies, Glagoljaška 8, SI‐6000 Koper, Slovenia

Montenegro is a small country in the Adriatic area with long tradition of olive growing. During the long period of olive cultivation a large number of varieties have been developed. Olive plant material was propagated mainly by vegetative parts of the tree, the 'tuber sprouts'. In this study, we compared the capacity of morphological and molecular markers for assessing the varietal identity and connection of selected clones / accessions of three local varieties: Crnica (9), Lumbardeška (4) and Barkinja (3). A study of morphological parameters showed differences between the tested clones, especially for degree of imperfect flowers (30.31 to 77.10%), fruit weight (2.89 to 5.36 g), stone weight (0.58 to 0.80 g) and oil content in dry matter (19.08 to 37.95%). The UPGMA analysis of 16 morphological traits showed variability and inter connection among the accessions of the three varieties. Analysis with nine SSR markers showed higher efficiency in the characterization of close individuals developed through clonal selection. The average number of allels per locus was 3.89, ranged from 2 to 5. Observed heterozygosity (He) ranged from 0.21 to 1 with a mean of 0.90. The dendrogram based on SSR markers revealed variability within Crnica variety and presence of synonymy in Barkinja and Lumbardeška varieties.

Keywords: olive, morphological characteristics, SSR markers, Montenegro

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Genotype effect on embryogenic capacity and plant regeneration from olive (Olea europaea L.) callus, cv. Picual.

K. Titouh1,2, L. Khelifi2, K. Hadj Moussa2, S. Cerezo‐Medina3,4, J.A. Mercado4 and F. Pliego‐Alfaro4

1National Institute of Agronomic Research (INRAA), Centre of Béjaïa, Algeria. 2High National School of Agronomy (ENSA), Laboratory of Genetic Resources and Biotechnologies (LRGB), Algiers, Algeria. 3Instituto de Hortifruticultura Subtropical y Mediterránea “La Mayora”, Consejo Superior de Investigaciones Científicas‐Universidad de Málaga (IHSM‐CSIC‐UMA). 4Departamento de Biología Vegetal, Universidad de Málaga, 29071 Málaga, Spain.

The olive tree (Olea europaea L.) is one of the most important crops in the Mediterranean area. Olive improvement by biotechnological tools is very difficult due to its recalcitrance to in vitro regeneration. In fact, the selection of an embryogenic line with an adequate regeneration capacity is of key importance for in vitro breeding. In this study, the embryogenic potential of several olive embryogenic lines, referred to the capacity to produce somatic embryos followed by plant conversion, was evaluated. Embryogenic lines were initiated from radicles of zygotic embryos of cv. ‘Picual’ and maintained in ECO solid medium. The eleven lines tested showed a good proliferation rate in solid medium, with increments on fresh weight in the range 0.3‐1.0 g per subculture. For embryo maturation, embryogenic callus was cultured for 4 weeks in liquid ECO medium and later transferred to solid ECO medium with activated charcoal. Three of the lines did not yield mature somatic embryos while in the rest of lines, the percentage of globular embryos giving rise to mature embryos varied between 1.7 and 17.4%. Embryo conversion was also highly variable among the different embryogenic lines. Moreover, the number of plants recovered from mature embryo ranged between 0.1 and 1.2, although plants could not be recovered from some lines. The results obtained showed that the embryogenic potential varies considerably among the embryogenic lines, and therefore, a screening of different genotypes should be performed prior to select a material for in vitro breeding.

Keywords: Embryogenic capacity, In vitro regeneration, Olive, Somatic embryogenesis

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Production of olive (Olea europaea L.) rootstocks by in vitro germination of zygotic embryos, cv. Chemlal.

K. Hadj Moussa1, K. Titouh1,2, M. Khelifi‐Slaoui1 , L. Khelifi1

1High National School of Agronomy (ENSA), Laboratory of Genetic Resources and Biotechnologies (LRGB), Algiers, Algeria. 2National Institute of Agronomic Research (INRAA), Centre of Béjaïa, Algeria.

The olive tree (Olea europaea L.) is the first fruit crop in Algeria which multiplies vegetatively by grafting or rooting of semi‐hardwood cuttings. However, this type of multiplication requires a long time and a large space. Our work aims to produce in a short time and limited space rootstocks after germination of olive zygotic embryos of the main cultivar in Algeria, Chemlal. Germination was achieved on three different culture media: olive medium (OM) (Rugini, 1984), MS medium (Murashige and Skoog, 1962) or Driver and Kuniyuki modified medium (DKW) (Revilla et al., 1996) supplemented or no with 0.01 mg l‐1 IBA and 1 mg l‐1 BAP. Our results indicate that the presence of growth regulators significantly improved germination which begun since the first week of culture, contrary to the media without hormones. Similarly, the size, the average number of leaves and the average number of micro‐cuttings recovered per obtained plant were higher in the presence of hormones either on the germination medium or the multiplication one. However, the length and the average number of emitted roots were affected by the presence of growth regulators. In fact, OM medium allowed a good germination and better seedling development while DKW was more beneficial for the micropropagation of derived plantlets. Furthermore, the insertion level of the micro‐cutting within plant strongly influenced the in vitro propagation. Therefore, the best results were obtained with median micro‐cuttings resulting to good plantlets without phenotypic anomalies.

Keywords: In vitro germination, Micropropagation, Olive, Rootstock, Zygotic embryo.

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MOLECULAR BIOLOGY BIOTECHNOLOGY

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Characterization of salinity‐responsive genes in olive (Olea europaea L.)

M.T. Sadder1,2,, A.M. Zakri2, H. Alswalmah2, A.A. Alsadon2, A.A. Al‐Doss2

1Plant Biotechnology Lab, Department of Horticulture and Crop Science, Faculty of Agriculture, The University of Jordan, Amman, 11942, Jordan 2Department of Plant Production, College of Food and Agricultural Sciences, King Saud University, Riyadh 11451, Saudi Arabia

Low quality water and soil salinization are increasingly becoming limiting factors for world food production. Novel salinity‐responsive genes (SRGs) were characterized in olives, namely, monooxygenase 1 (OeMO1), cation calcium exchanger 1 (OeCCX1), salt tolerance protein (OeSTO), proteolipid membrane potential modulator (OePMP3), universal stress protein (OeUSP2), adaptor protein complex 4 medium mu4 subunit (OeAP‐4), WRKY1 transcription factor (OeWRKY1) and potassium transporter 2 (OeKT2). Sequence analysis and protein structure were described and discussed. Full length olive SRGs were translated and utilized for phylogenetic analysis. Two major clusters were resolved for monooxygenase. OeMO1 was grouped in one cluster, which was subdivided into two major clades (monocots and dicots). On the other hand, OeSTO, OePMP3 and OeKT2 proteins did not cluster with any plant homolog. However, both OeUSP2 and OeAP‐4 were found to be clustered with their corresponding tomato homologs with relatively good bootstrap values of 74% and 65%, respectively. The expression of olive SRGs was investigated using quantitative real‐time PCR calibrated to actin1 expression. Two olive cultivars were utilized; 'Nabali' (moderately tolerant) and 'Picual' (tolerant). Six salinity stress levels were applied to in vitro plantlets (0, 30, 60, 90, 120 and 150 mM NaCl). The differences in photosynthetic activity between applied stress levels were not significant. However, tangible differential gene expression was prominent. Several olive SRGs were found to be up‐regulated at 60 mM NaCl in both cultivars, however, they were more prominent in 'Nabali'. On the other hand, SRGs showed higher fold increase in expression at 90 mM NaCl in 'Picual' than in 'Nabali'. Identifying essential genetic factors of salinity‐stress tolerance would be helpful in future olive selection of new tolerant lines and in crop management.

Keywords: Abiotic stress, 'Nabali', olives, 'Picual', salt, genes.

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Comparative study of FAD2 gene sequences among some olive cultivars grown in Qom region of Iran

M. Sharifani1, M. Asheri1, A. Yamchi1 and S. Hosseini Mazinani2

1Faculty of crop Production, Gorgan University of Agricultural Sciences and Natural Resources, Gorgan, Iran 2National Institute of Genetic Engineering and Biotechnology, Tehran, Iran

Fatty acid desaturases are enzymes capable of modifying carbon‐carbon bonds within fatty acids. All higher plants contain one or more oleic acid desaturases that catalyze insertion of a double bond between carbons 12 and 13 of oleic acid to produce linoleic acid, an unsaturated fatty acid. Fatty Acid desaturase 2 (FAD2) genes’ encoding enzymes convert oleic acid to linoleic acid. The higher the amount of oleic acid, the better the olive oil quality as long as it falls within the range permitted by IOC. In this study, five olive cultivars were chosen according to their fatty acid profiles with different range of oleic acid (Mari, Shengeh, Arbequina, Beledy, and Koroneiki), with the aim of identifying differences in FAD2‐2 gene sequences of local and imported varieties. Forward and Reverse degenerative primers were designed for FAD2‐2 gene using OLIGO ver 5.0 software. DNA was extracted from olive leaves with the CTAB method. PCR was applied to amplify FAD2‐2 gene after a gradient PCR approach was conducted. The PCR products were sequenced and the sequences were aligned together. Then alignment of each of these with the NCBI reference accession was conducted using Multalin and mega softwares. Alignment of the Koroneiki cultivar sequence with the sequences of the other four cultivars revealed different percentages of identity, such that. Mari showed 95%, Arbequina 96%, Beledy 93% and Shengeh 91% identity with the Koroneiki sequence. Furthermore, alignment of Arbequina, Koroneiki, Shengeh, Mari and Beledy sequences with FAD2 cDNA clone from olive cultivar Picual (Accession No. AY733077) registered in NCBI showed 92%, 90%, 87%, 85%, and 84% identity, respectively. Relatively low percentages of shared identity could possibly be the result of comparing the sequences directly from PCR products with cDNA clone without intron parts in NCBI. Results of aligning FAD2 gene sequences implied that there are considerable differences in sequences of FAD2 genes among the studied cultivars.

Keywords: FAD2, gene sequencing, olive oil, oleic acid

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The different adaptations to salt‐stress of two Italian olive cultivars, salt‐sensitive (Leccino) and salt‐tolerant (Frantoio)

L. Rossi1,2, A. Francini1, A. Minnocci1, L. Sebastiani1

1Institute of Life Sciences, Scuola Superiore Sant’Anna, I‐56127 Pisa, Italy 2Department of Plant Biology, North Carolina State University, Raleigh, NC‐27695, USA

The development of the soil salinity is one of the major problems of the arid zones. Olive trees are salt‐stress tolerant, in a cultivar‐related manner, but salinity tolerance mechanisms are not clear. We have studied root adaptations to salinity analyzing the changes of its apoplastic barriers, the ions accumulation and Na+ cell‐specific distribution in one salt‐sensitive (Leccino) and one salt‐tolerant (Frantoio) cultivar. We found that tolerant cultivar retained salt in root, while sensitive cultivar accumulated in leaves. Salt stress influenced also the development of the apoplastic barriers in the roots of both the two cultivars. Na+ cell concentrations were significantly different between genotypes and cell type, showing a gradient concentration from exodermis to stele, stronger in Leccino than in Frantoio. This study demonstrate how salt tolerant and sensitive olive cultivars partially avoid salt intrusion creating barriers at root levels in response to salt stress, but this not result sufficient to completely avoid salt translocation in the shoot.

Keywords: Olea europaea, aeroponic, apoplastic barriers, cryo‐SEM, Na+, X‐ray microanalysis

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Transcriptome analysis of seed and mesocarp tissues of olive fruit identifies key genes involved in oil accumulation and fatty acid composition

M. Hernández1, A. Muñoz‐Mérida2, M. Sicardo1, O. Trelles2, V. Valpuesta3, J. Martínez‐Rivas1

1Department of Biochemistry and Molecular Biology of Plant Products, Instituto de la Grasa (IG‐ CSIC), 41012 Sevilla, Spain. 2Integrated Bioinformatics, National Institute of Bioinformatics, University of Málaga, 29071 Málaga, Spain. 3Department of Molecular Biology and Biochemistry, Instituto de Hortofruticultura Subtropical y Mediterránea, University of Málaga (IHSM‐UMA‐CSIC), 29071 Málaga, Spain.

Olive breeding programs are focused on selecting for different traits such as oil content or oil characteristics including fatty acid, phenolic and volatile composition to suit new markets. Understanding the molecular basis of these characteristics and improving the efficiency of such breeding programs require the development of genomic information and tools, which are still scarce in olive. Within the project OLEAGEN, we have applied Sanger and 454 pyrosequencing technologies to generate close to two million reads from twelve cDNA libraries obtained from the Picual, Arbequina and Lechin de Sevilla cultivars, plus seedlings from a segregating progeny of a Picual x Arbequina cross. The libraries include fruit mesocarp and seeds at three relevant developmental stages, young stems and leaves, active juvenile and adult buds as well as dormant buds, and juvenile and adult roots. The reads were assembled into 81,020 unigenes with an average size of 496 bases, and their functional annotation was carried out.Digital expression analysis of the three non‐normalized 454 cDNA collections corresponding to seeds and mesocarp tissue at two different developmental stages from Picual cultivar has been performed. Sequences related to sugar and lipid metabolism have been further annotated on the basis of Arabidopsis proteins since they closely matched with Arabidopsis orthologs. The ESTs counts assigned to each gene have been expressed as ESTs/100 000 ESTs and this value has been used as a measure to compare relative gene expression in different olive fruit tissues. Genes corresponding to the metabolic pathways for fatty acid biosynthesis, triacylglycerol assembly, biosynthesis of plastidial lipids, and carbon supply for fatty acid and glycerolipid biosynthesis that exhibit differential expression between the studied tissues have been identified. Their involvement in the regulation of the mentioned metabolic pathways will be discussed.

Keywords: Olea europaea, transcriptome, pyrosequencing, digital expression analysis, oil accumulation, fatty acid biosynthesis

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Identifying genes involved in fatty acid synthesis in olive

N. Cultrera1, R. Mariotti1, F. Alagna1, V. Sarri2, M. Ceccarelli2, M. Bellucci1, L. Baldoni1

1CNR ‐ Institute of Biosciences and Bioresources (IBBR), 06128 Perugia, Italy 2University of Perugia, Department of Chemistry, Biology and Biotechnology, 06123 Perugia, Italy

Olive accumulates oil triacylglycerols (TAGs) in the fruit mesocarp and the study of the genetic determinants affecting TAG biosynthesis, including the early steps of fatty acid (FA) elongation, is extremely important to clarify the mechanisms that regulate or restrict this pathway. We have identified and characterized four main gene families involved in FA elongation and desaturation: Acyl Carrier Protein (ACP), β‐ketoacyl‐ACP synthase I (KAS I) and II (KAS II), and stearoyl‐ACP‐ desaturase (SAD). ACP is the central cofactor protein for plastidial FA synthesis, KAS I catalyzes the substrate C4:0‐ACP to produce C16:0‐ACP, KAS II conduces the final elongation from C16:0‐ ACP to C18:0‐ACP and SAD desaturates stearoyl‐ACP to oleoyl‐ACP in the plastid. Three OeACP genes have been identified at genomic and cDNA level and numerous haplotypes have been distinguished within olive cultivars. Their expression has been characterized in different olive tissues and an OeACP cDNA has been overexpressed in tobacco plants, in order to alter the levels of ACP and analyze the possible consequences on lipid biosynthesis. The phenotypic analysis of tobacco transformants expressing high amounts of olive ACP in different tissues is currently in progress. The analysis of preliminary olive genomic sequence data and fruit EST (Expressed Sequence Tags) collections have allowed to identify three OeKASI copies carrying seven exons. Their sequence has been characterized, many polymorphisms have been identified and expression analyses carried out on RNA of fruit mesocarp accumulating oil allowed to identify the most strong candidates involved in the FA synthesis of olive oil. The same approach has been used to identify two OeSAD variants, respectively carrying two and three exons and four OeKASII copies, all carrying 12 exons. The identification of haplotype variants and the analysis of their expression in oil‐accumulating tissues is currently in progress.

Keywords: Olea europaea, oil synthesis, ACP, KAS, SAD, functional genomics

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Hydroxytyrosol – An attractive olive phytochemical produced through metabolic engineering in E. coli

F. Ververidis, E. Trantas, Th. Pavlidis, E. Navakoudi, A. Kontaratou, E. Mpalantinaki

Plant Biochemistry and Biotechnology Laboratory (PBBL), Department of Agriculture, School of Agriculture, Food and Nutrition, Technological & Educational Institute (TEI) of Crete, P.O. BOX 1939, GR 710 04, Heraklion – Crete, Greece

Hydroxytyrosol (HT) along with tyrosol are the most abundant phenethyl alcohols in olive oil, as they are biosynthesized in olive fruit. HT contributes most to its qualitative characteristics and was recently approved as a part of “olive oil healthy polyphenols”, by EC Regulation 432/2012. HT has been associated with significant pharmacological properties; it possess high capacity for free radical scavenging, can reduce the risk of cancer, may have critical effects on the formation and maintenance of bones, can be used as an effective remedy in the treatment of osteoporosis symptoms, and possess antimicrobial and antiviral activity. The content of HT and tyrosol in olive fruit, leaves and olive pulp varies significantly among the cultivars and in general decreases during fruit development and maturation. However, in some varieties there is an increase in HT content which drops again before full maturation, showing some specificity for certain cultivars that have high phenolic content. This biological incident might be related to the fact that there is an underlying structural connection between phenylethanoids biosynthesized in olive tissues, as they contain a hydroxytyrosol or a tyrosol moiety as part of their structure, which during ripening process breakdown to their building molecules. Recent research data have shown that HT biosynthesis in olive tissues has been presumably derived from tyrosine, which proceeds through DOPA and dopamine to hydroxytyrosol. Moreover, HT and tyrosol appear to be also degradation products during processing of olives, particularly in the mechanical production of olive oil. Thus, they are also present in olive mill waste waters (OMWWs), as hydrolysis products of phenylethanoids that are non‐soluble in olive oil. As a result, OMWWs contain the majority of hydroxytyrosol of the olive fruit compared to the olive oil. In this contribution, we present a heterologous microbial expression system metabolically engineered to bio‐convert precursor molecules deriving from primary metabolism into HT. A novel, artificial biosynthetic pathway for HT production in Escherichia coli was developed enabling its production directly from L‐glucose or L‐tyrosine, thus mimicking the putative native pathway in olive tissues. This pathway involves two heterologous enzymes for the conversion of L‐tyrosine into HT with similar intermediate products. Additionally, by using engineered E. coli strains capable of overproducing tyrosine, we managed to produce 1.7mM of HT from L‐ glucose in the culture medium after 24hrs growth. Our results indicate that there is considerable space for strain optimization to increasing heterologous HT production efficiency in E. coli.

Keywords: Olea europaea, Hydroxytyrosol, metabolic engineering, Escherichia coli

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Identification of the genes in olive (Olea europaea cv. “Koroneiki”) that may be involved in the biosynthesis of hydroxytyrosol

N. Mougiou1, 2, E. Trantas3, A. Argiriou2, F. Ververidis3, A.M. Makris2, K.E. Vlachonasios1

1Department of Botany, School of Biology, Faculty of Science, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece 2Institute of Applied BioSciences, CERTH, 6th km Charilaou‐ Thermis, 57001 Thermi Thessaloniki, Greece 3Plant Biochemistry and Biotechnology Laboratory (PBBL), Department of Agriculture, School of Agriculture, Food and Nutrition, Technological & Educational Institute (TEI) of Crete, 71004 Heraklion, Greece

Hydroxytyrosol (HT), a derivative of oleuropein, is a phenolic compound with high antioxidant activity present in olive fruits. HT and oleuropein consumption through olive oil and fruits has been correlated with health benefits in humans. Moreover HT has antibacterial activity against various plant pathogenic bacteria. The HT biosynthesis pathway has not yet been clarified. Our goal is to identify the genes that are involved in olive HT biosynthesis. We used RNA‐sequencing of the “Koroneiki” fruit at 30 days after fruit set. The transcriptome analysis data revealed 76272 unigenes (unique transcripts), 53864 of which were functionally annotated in databases. The 4 proposed genes for HT biosynthesis, tyrosine decarboxylase (TDC), monoamine oxidase (MAO), phenylacetaldehyde reductase (PAR) or alcohol dehydrogenase (ALDH) and tyrosinase (TYR) were identified through bioinformatics analysis. The olive transcriptome bears 12 TDC, 6 MAO, 9 ALDH and 6 TYR partial transcripts. For each gene the transcript was chosen that had an intact open reading frame and the highest homology with translated transcripts in other olive cultivars. For instance, the TYR protein from “Koroneiki” has 99% homology with “Coratina” and “Dolce d’ Andria” cultivars, MAO has 88% homology with “Tendellone”, TDC has 68% with “Coratina” and ALDH has 99% homology with “Carolea”. These genes will be used in our effort to reconstitute the HT biosynthesis pathway in other organisms.

Keywords: Olea europaea, Koroneiki, Hydroxytyrosol biosynthesis, RNA‐sequencing

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Response to drought stress of olive (Olea europaea L.) plants transgenic for tobacco osmotin gene.

C. Silvestri, V. Cristofori, S. Astolfi, A. Minandri, E. Rugini

Department of Agriculture, Forestry, Nature and Energy (DAFNE), Tuscia University, 01100 Viterbo, Italy

Olive is a Mediterranean species well adapted to grow in arid and semi‐arid areas, where the plants are often subjected to water deficiency. In the present work we studied the in vitro response of transgenic olive plantlets to drought‐stress conditions. The transgenic plantlets were initially obtained by agrobacterium‐mediated transformation of Canino cultivar for the tobacco‐ osmotin gene. In vitro drought‐stress conditions were achieved by adding different concentrations (0, 1, 2 and 4%) of polyethylene glycol (PEG‐8000) to the proliferation olive medium. The osmotin transgenic shootlets showed higher tolerance to PEG‐induced water deficiency as compared to their un‐transformed counterparts. The transgenic shootlets showed a normal growth rate in the presence of 2 and 4% of PEG and furthermore, there was no difference with respect to the un‐treated shootlets. On the contrary, the exposure of un‐ transformed shootlets to PEG increased concentrations led to severely damaged leaves. Among the biochemical parameters investigated, the enhanced tolerance to PEG‐induced stress was also accompanied by higher proline accumulation in the tissues, indicating that the transgenic material that is over‐expressing the tobacco‐osmotin gene is able to better tolerate the in vitro drought‐stress conditions. In addition preliminary observations in pots and field trials showed that the transgenic plants, contrary to un‐transformed counterparts, grew poorly under irrigation, showing stunted growth and randomly leaf drop in the young shoots. The observations resulted incomplete due to the lack of renewal of to field trial authorization by the Italian Ministry of Environmental.

Keywords: Olea europaea L., drought‐stress tolerance, osmotin‐transgenic olive, proline accumulation, plant growth. OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 44

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BIOLOGY PHYSIOLOGY

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Chemical and histological analysis of olive tree tissues to understand seasonal starch distribution

E. Cauli1, G. Bandino1, P. Sedda1, M. Mulas2, R. Zurru1

1Department for Fruit and Forestry Research of AGRIS‐Sardegna, I‐09123 Cagliari, Italy 2Department of Nature and Land Sciences, University of Sassari, I‐07100 Sassari, Italy

One of the main problems of the olive industry is the irregular or alternate fruit bearing. The study of carbohydrate reserve distribution on tree may provide an opportunity to better understanding the influence of leaf activity and wood starch distribution on fruit yield. Starch reserve distribution on wood tissues and in leaves of 18‐year‐old trees of ‘Tonda di Cagliari’ olive cultivar were observed from February 2011 to March 2012 in an orchard located in the South of Sardinia (Italy). Six trees were considered as off‐plants (not bearing fruits) and six as on‐plants (abundant fruit yield). Young wood of main roots showed the highest starch content, ranging from 5 to 15% of the dry weight (DW). In the external woody rings of the trunk collar starch was between 3.5 to 7.5% of DW, in the upper part of the trunk ranged from 0.5 to 5.0% of DW, and in main branches between 1.0 and 4.5% of DW. Starch content of wood of the trunk and of the main branches was significantly lower in on‐trees with respect to off‐trees. Fruit bearing one‐ year‐old branches showed an initial starch content ranging from 2.5 to 4.5% of DW, with significantly higher values in on‐trees. This contend decreased sharply in the summer‐autumn season and subsequently increased in winter. Leaves formed in the previous spring have a starch content between 1.5 and 5.5% from February to June 2011, with a big decrease in July, while leaves formed in the spring of the 2011 showed low starch content in the first months of life until the subsequent winter season. Starch reserves of trunk and main branches seemed to have a key role to support the growing and ripening fruits.

Keywords: Olea europaea, yield behavior, alternate fruit bearing, carbohydrates, nutritional reserves.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 47

Quantitative behavior of Olea pollen in relation with climatic factors in three regions of Tunisia

S. Elhadj Hamda1, A. Ben Dhiab2, M. Msallem1

1Institut de l’olivier, BP 286, Cité Elmahrajène 1002 Tunis, Tunisie 2Institut de l’olivier, BP14, Sousse, Sousse

Meteorological parameters are the main factors affecting atmospheric olive pollen dispersion and pollination progress. Different aeropalynological methods were developed allowing monitoring atmospheric pollen content. The aim of this study is to monitor the pollen emission in conjunction with meteorological factors in three regions of Tunisia, Mornag (36°39’N; 10°26’E) in the Northern part, Jemmel (35°38’N; 10°45’E) in the Central East part and Chaal (34°19’N; 10°04’E) in the Southern part using Hirst‐Type spors traps. The pollen numerical data analyzes showed a similar pollen index recorded in two bounded regions Mornag and Chaal. The flowering of these regions was characterized by a long period reached 45 and 50 days respectively with two peaks pollen emission reflecting staggered flowering. However, in Jemmel flowering lasted only 29 days with the lowest pollen index and only one peak pollen emission indicating grouped flowering process. The combination between biological parameter in terms of pollen and meteorological data showed a positive correlation between daily mean temperature, wind speed and pollen dispersion in the atmosphere which is blocked in presence of rainfall and high hygrometric degrees of the air reflected by low atmospheric pollen concentrations.

Keywords: Aéropalynology, olive pollen emissions, airborne pollen counts, meteorological parameters.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 48

A de novo trascriptome approach to identify the gene expression in the development of the pollen tube in olive (Olea europaea L.)

D. Iaria, A. Chiappetta, V. Vizzarri, I. Muzzalup

1Olive growing and oil industry research centre (CRA‐OLI), 87036, Rende (CS), Italy 2University of Calabria (DiBEST), 87036, Arcavacata di Rende (CS), Italy

Olive (Olea europaea L.) is a wind‐pollinated, allogamous species that is generally not considered to be self‐compatible. However, the processes controlling self‐incompatibility are still unclear and the molecular basis that underlie this process are still not fully characterized. These topics are regarded as important both for fundamental studies of the control of fertility and reproduction in plants and also as an attractive model system for the investigation of polarized tip growth, cell–cell interactions, and signal transduction. In order to determine compatibility relationships, using modern next‐generation sequencing techniques and a de novo trascriptome assembly strategy we show that pollen tubes from different olive plants, that have grown in vitro in a medium containing its own pistil and in combination pollen/pistil from self‐sterile and self‐fertile cultivars, have a distinct gene expression profile and many of the differentially expressed sequences between the samples fall within gene families involved in the development of the pollen tube, as lipase, carboxylesterase, pectinesterase, pectin methylesterase, callose synthase. Moreover different genes involved in signal transduction, transcription and growth are overrepresented. The analysis also allowed us to identify members in actin, actin depolymerization factor and fimbrin gene family and member of the Ca2+ binding gene family related to the development and polarization of pollen apical tip. The whole transcriptomic analysis, through the identification of the differentially expressed transcripts set and an extended functional annotation analysis, will lead to a better understanding of the mechanisms of pollen germination and pollen tube growth in olive.

Keywords: next‐generation sequencing, pollen germination, gene expression, olive fertility.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 49

The Estimation of real water needs of olive trees (Olea europaea L. cv. ‘Meski’) conducted in irrigated orchards during flowering

S. Ben Mariem1, O. Boussadia2, F. Ben Mariem2, W. Gheriani2, N. Boukhrissat2, M. Braham2

1Higher Agronomic Institute of Chott Mariem, Sousse, Tunisia 2Olive Institute – Sousse, Tunisia

In Tunisia, the majority of olive irrigated orchards recently installed, are localized in arid and semi‐ arid regions, where trees are frequently subjected to high temperature and scarcity of water. This implies the adoption of reasonable management of irrigation based on the knowledge of water requirements in order to economize and rationalize the water use. In this context, this study focuses on the estimation of real water needs and the assessment of those needs by controlling the ecophysiological behavior and the water status in the table olive (cv. ‘Meski’) conducted in an irrigated orchard in the semi‐arid climate in Tunisia. To do this, a calculation of water requirements and application was performed by two methods, one is climatic (water balance) and the second is physiologic (xylem sap flow). The experiment was installed in an orchard irrigated by drip system in the region of sidi El Heni (Sousse, Tunisia). The doses applied was: T0 ; irrigation with a dose of 100 % crop evapotranspiration (ETc), T1 ; irrigation with a dose of 100 % xylem sap flow with black plastic mulching, T2 ; irrigation with a dose of 100 % xylem sap flow without black plastic mulching. During this essay, we could quantify the right amount of irrigation converted to percentage of Etc from ‘cluster formation’ until the first stage of fruit growth. This percentage varies from 19 % to 10 % ETc according to the phenology of the olive tree.

Keyword: Olea europaea, water needs, xylem sap flow, ecophysiological parameters, water status, flowering.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 50

Metabolic profiling and gene‐expression analyses in salt stressed olive trees (Olea europea L.) cultivars Leccino and Frantoio

L. Rossi1,2, M. Borghi2, A. Francini1, D. Xie2, L. Sebastiani1

1Institute of Life Sciences, Scuola Superiore Sant’Anna, I‐56127 Pisa, Italy 2Department of Plant Biology, North Carolina State University, Raleigh, NC‐27695, USA

Olea europaea L. (olive tree) is an important crop in the Mediterranean Basin, which produces 95% of the world total fruits and oil (approximately 11 million tons), and lends important economic and dietetic benefits to the people of that region. However, the yield of olives and olive oil could be compromise because of climate changes in the areas of its major cultivation. Sporadic irrigation with sea water is currently under evaluation as a viable method to overcome prolonged periods of drought. Preliminary results of these experiments have shown that olive trees can tolerate a moderate salt stress induced by irrigation with sea water, which helps to maintain constant the average annual production. Given the importance that metabolites with nutraceutical properties have for the human health, the major aim of this research is to investigate the added nutraceutical value of olives from plants exposed to irrigation with salty water. Results reveal that olive leaves, stem and roots grown under salt‐stress increased content of metabolites with nutraceutical properties (Kaempferol, Luteolin, Quercetin). In conclusion salt irrigation enhances the nutraceutical properties of olive leaves. That last achievement represents a milestone in metabolomic studies because is the first one conducted on olive tree leaves, that are an important by‐product (after the fruits and its oil) with an economic relevance in terms of olive tea leaf.

Keywords: Olea europaea, leaves, nutraceutical properties.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 51

Chilling and heat requirements of different olive varieties (Olea europaea L.) grown in the south of Tunisia

I. Zouari1,2, M. Aïachi‐Mezghani2, A. El‐Aroui1, F. Labidi2, M. Mars1

1Institut Supérieur Agronomique de Chott Mariem, B.P : 47, 2042 Chott Mariem, Sousse, Tunisia 2Laboratoire d’amélioration de la productivité de l’olivier et de la qualité des ses produits, Institut de l’Olivier, Unité Spécialisée de Sousse, Rue Ibn Khaldoun, B.P.: 14, 4061 Sousse, Tunisia

Tunisian olive oil production is fluctuating both in quantity and quality. Thus intensification program is important to improve yield of the local varieties and introduction of new varieties which their climatic adaptation degree is not yet well established. Winter chilling is required to release previously initiated flower buds from dormancy and enhance flowering after receiving the required heat requirements. However, information concerning chilling and heat requirements of varieties is generally poor. In this context, a study was carried out on local and foreign cultivars grown in south of Tunisia to calculate the chilling requirements for breaking of dormancy and heat requirements for flowering during the period 2002‐2012. Heat Unit was expressed in Growing Degree Day (GDD). Chilling accumulation was equal to the sum of the chilling hours less than 7.2°C. Results indicated a large inter‐annual variability. Chilling ranged from 70 hours (2005‐2006) to 229 hours (2007‐2008). Heat accumulation varied between 197 GDD (2012) and 322 GDD (2010). For the flowering, differences between varieties were noted during 2010‐2011. Koroneiki showed a relatively early entry into bloom (05 of April) just after accumulation of 87.5 GDD comparing to the other varieties which presented a beginning of bloom after accumulation of 247.33 GDD (30 of April). For the year 2011‐2012, the variety Meski showed an early bloom after the accumulation of 118.10 GDD (12 of April) while the other cultivars entered to bloom after accumulation of GDD ranging between 169.45 and 267.6. These results indicated that chilling requirement is highly correlated with heat requirements for flowering of each variety and varied according the climatic conditions.

Keywords: Olive tree, Cultivar, Chilling hours, Growing Degree Day

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 52

Effect of Sodium Chloride on olive seedlings under controlled conditions.

S. Baccari1 , A. Chaari Rekhis1, A. Chelli‐Chaabouni2

1Institut de l’olivier .Université de Sfax. BP 1087. 3000 Sfax .Tunisia. 2Institut National de Recherche Agronomique de Tunis. Université de Carthage. Hedi Karray Street. 3049 Ariana. Tunisia

Salinity is a one of the major environmental limiting factors in plants growth and productivity. The effect of salt stress on olive (O. europaea L.) plantlets was evaluated under controlled conditions. One‐year‐old seedlings from Chemlali variety were irrigated twice weekly, in turns, with a nutrient solution and distilled water supplemented with 75 or 150 mM of NaCl. After 3 months, the results achieved show that the NaCl caused a significant decrease in the stem elongation, in the axillary shoots formation, in the number of leaves as well as in the dry weight of plantlets. In addition, the fresh weight of aerial part was more affected by the salt than roots system. Symptoms of toxicity such as falling leaves and chlorosis were detected from the first month. Mineral analysis showed a significant increase in the sodium content (Na+) and chloride (Cl‐). Na+ concentrations were comparable in both aerial and root parts, while the Cl‐ concentrations were higher in the aerial part in seedlings treated with 75 mM NaCl. Among olive plantlets treated with 150 mM of NaCl, the accumulation of chlorides was comparable in both aerial and root parts. Sodium Chloride also has an effect on Potassium concentration. A significant decrease was recorded in the aerial and root parts for Potassium concentrations with the exception of seedlings treated with 150 mM NaCl which maintained a K+ concentration comparable to the control. However, potassium levels were higher in the aerial part.

Keyswords: olive, Sodium Chloride, toxicity, Potassium.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 53

Identification of the iridoid synthase involved in oleuropein biosynthesis

F. Alagna1, 2, F. Geu‐Flores1, S. E. O’Connor1, L. Baldoni2, A. Osbourn1

1Department of Metabolic Biology, John Innes Centre, Norwich NR4 7UH, United Kingdom 2Institute of Biosciences and Bioresources (IBBR), CNR, 06128 Perugia, Italy

In recent years, interest in the therapeutic benefits of olive oil and olive fruits consumption has grown considerably, due to the presence of beneficial bioactive compounds. From the pharmacological viewpoint the secoiridoids (mainly oleuropein) are the most active substances of olive oil. Oleuropein has shown several pharmacological properties, exerting antioxidant, anti‐ inflammatory, anti‐atherogenic, anti‐cancer, anti‐microbial and anti‐viral effects, and, for these reasons, it is commercially available as food supplement. Despite the importance of this compound, the genes responsible for its biosynthesis in olive are still unknown. The aim of our work was to identify the genes involved in secoiridoid biosynthesis and to characterise the enzymes that they encode. Putative genes were identified by mining transcriptomicand genomic data. Furthermore, the olive iridoid synthase (OeISY), which catalyses the cyclization of 8‐ oxogeranial to generate the iridoid ring scaffold of secoiridoids, has been identified by homology with the corresponding gene involved in the indole alkaloid pathway in Catharanthus roseus. The gene was expressed in E. coli and the purified protein was used for biochemical assays. In vitro assays confirmed its predicted function, nepetalactol and its open dialdehyde forms being obtained as products. These data, together with the OeISY expression pattern analysis, which was in accordance with the oleuropein content in olive tissues, strongly suggest that OeISY is the enzyme which synthesises the monoterpene scaffold of oleuropein. Other three genes showing high similarity to OeISY were also cloned and the products biochemically characterized. Even if their physiological role still remains unknown, our results shed some light on their putative function. The discovery of genes involved in secoiridoid biosynthesis is a first step toward increasing the dietary value of olive fruits and oil and improving the extraction of their bioactive components. It also offers opportunities for metabolic engineering in microbial or plant hosts for their large scale production by the pharmaceutical or herbal industries.

Keywords: Olea europaea, secoiridoid, secondary metabolites, olive

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 54

Olive hedgerow canopy positions determine inflorescence and flower development

E. R. Trentacoste1,2, M. Gómez‐del‐Campo2, I. Moreno‐Alías3, H. F. Rapoport3

1INTA EEA Junín, Mendoza, Argentina. 2Dpto Producción vegetal: Fitotecnia. Univ. Politécnica de Madrid, Spain 3Instituto de Agricultura Sostenible, C.S.I.C., P.O. Box 4084, 14080 Córdoba, Spain

Productivity of the olive tree is highly responsive to radiation and temperature conditions, and understanding the responses of the different developmental processes that determine yield components is essential for orchard design and management. Inflorescence and floral development are key initial stages for the formation of fruits. In the new intensive hedgerow orchards, radiation and temperature are highly modified by orientation and height within the canopy. In a N‐S (North‐South) oriented hedgerow (cultivar Arbequina) located in Toledo (Spain), inflorescences were sampled from both sides (faces) of the hedgerow canopy, at two heights: 0‐ 0.4 and 1.6‐2.0 m above the soil. Floral was quality determined at different levels of morphogenetic organization, from the inflorescence, to flower, ovary and ovule. Temperature and radiation were measured in the sampling positions and compared with model predictions. Developmental effects which potentially affect fruit production were observed particularly at the higher levels of organization. Inflorescence structure and flower number both showed tendencies depending on height within the canopy, with greatest values for both parameters in the high position of the eastern face. Pistil abortion was highest for the lower positions of both east and west faces, with highest number and percent of perfect flowers per inflorescence again in the upper position of the eastern canopy face. Flower quality, observed by ovary size and ovule development in histological preparations, were found to be highly conservative inside the hedgerow. Final fruit number most correlated with inflorescence total flower number. Overall, the results emphasize the importance of both irradiance and temperatures at different heights and orientations within the hedgerow, on assimilate partitioning to olive inflorescence and floral structures.

Keywords: Olea europaea, pistil abortion, ovule development, flower number, ovary size

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 55

Frost tolerance and recovery ability of eleven olive cultivars in central Italy

E.M. Lodolini1 , B. Alfei2, A. Santinelli2,, T. Cioccolanti1, D. Neri1

1Dipartimento di Scienze Agrarie, Alimentari e Ambientali, UNIVPM, 60131 Ancona, Italy 2Agenzia Servizi Settore Agroalimentare delle Marche, ASSAM, 60131 Ancona, Italy

Olive is an evergreen species that can be subjected to frost damages when low temperatures or cold returns appear in winter and spring, respectively. Frost conditions occur quite frequently in central Italy and symptoms on the tree can range from shoot tip burns and defoliation up to bark split on branches or trunk in case of intense injury. The selection of the less susceptible olive cultivars represents the most effective method to avoid frost damage. The objective of the present study was to assess the level of frost damage in 11 olive cultivars after a freezing event occurred in 2012 in Marche Region (central Italy) and their recovery ability during the following growing season. The studied cultivars were Nebbia del Menocchia, Rosciola, Sarganella, Capolga, Oliva Grossa, Piantone di Mogliano and Piantone di Falerone locally diffused in the Marche Region; FS17, Don Carlo and Giulia, nationally spread; and Arbequina internationally used for super‐high density olive orchards. At the moment of the freezing event (February 2012), the trees were three years old and a visual index from 0 (no injuries) to 4 (very intense injuries) was used to evaluate the level of frost damage. During the following growing season, the recovery ability of the cultivars (re‐sprouting in the apical, central and basal portion of the tree) was studied using a visual index from 0 to 3. Results confirmed a different frost tolerance and recovery ability of the studied cultivars. In particular, Arbequina, FS17, Giulia, Sarganella, Oliva Grossa, Piantone di Mogliano and Piantone di Falerone showed a high frost susceptibility reporting intense damages on primary branches and trunk (visual index for injury level above 3); Don Carlo, Capolga and Nebbia presented an intermediate sensitivity showing damages mainly on 2‐ and 3‐year‐old branches (values included between 2 and 3), while Rosciola showed a high frost tolerance with injuries reported only on 1‐year‐old shoots (average level of damage of 1.7). Vegetative re‐sprouting during the season was good and homogeneous on the whole tree for Rosciola and Don Carlo, while it was concentrated mainly in the trunk and in the primary branches of the medium‐basal portion of the tree for the other cultivars. This study allowed the characterization of local, national and international olive cultivars in respect to the frost tolerance and supplied helpful evidence about their recovery ability. Such information can be used to select the most suitable varieties to be planted in cold areas.

Keywords: Olea europaea, Rosciola, FS17, Arbequina, visual index, vegetative re‐sprouting.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 56

Use of biosensor for estimating abiotic stresses and technical management of olive groves

D. Boujnah1, M. Gouiaa2, H. Chehab1, Z. Mahjoub1, S. Lamari1, B. Chikhaoui1

1Instuitut de l’Olivier, Rue Ibn Khaldoun , BP: 14‐ 4061 Sousse ,Tunisie. 2 Institut national Agronomique de Chott Mariem, Tunisie

The olive tree is a principal element of the agro Mediterranean system. It is characterized by its flexibility adaptation. The repositioning of the varieties according to the evolution of the bioclimatic context is one of the adaptation possibilities of the cultivated species to the climate changes. This option would be however conditional by a technological support to identify abiotic stress at the plant level and the optimization of the technical management of olive groves as well. In this study we report on the use of biological sensors to identify the abiotic stress and the tree behaviour after the application of some cultural practices to mitigate the stress recorded (pruning and providing additional water using hydroabosrbant). The experiment is conducted on farm and under rain fed. The results show that the effect of hydroabosrbant on the turgor potential of the olive tree is not immediate. From February, following the lack of rainfall a clear difference of evolution curves appear. In fact, we note the onset of water stress in the control treatment which is reflected by an increase of the Pp peak value at noon on the following day this indicate that the water loss is not compensated by an appropriate uptake of water by the roots, thus leading to an increased turgor pressure loss. In trees with water‐absorbing injected into the soil leaf turgor is better. The method that we tested show that (the leaf patch clamp pressure probe) enable real‐time monitoring of plant water status under field conditions, and are non‐invasive measuring method that can elucidate short‐and long‐term, temporal and spatial dynamics of leaf water status of trees in dependence on microclimate and soil factors.

Keywords: olea europaea, water stress , hydroabsorbant, cultural practices

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 57

Pollen storage and in vitro germination in some olive (Olea europaea L.) cultivars A.Senbas1, M.T. Ozkaya1, H. Colgecen2,

1Ankara Univ., Fac. of Agriculture, Dept. of Horticulture, Ankara, Turkey 2 Bulent Ecevit Univ., Fac. of Science and Literature, Dept. of Botany, Zonguldag, Turkey

Since there are some insufficient pollination problems in olive because of rain and high humidity during flowering, growers have to apply supplemental pollen. In order to achieve success in supplemented pollination, there are some necessities such as viability of pollen grains of pollinator cultivars. The most important criteria for keeping viabile of pollen are; storage conditions and temperature The pollen grains have been stored in 3 different temperatures (+4OC, ‐20OC, ‐80OC) ‘Gemlik’ and ‘Domat’ are the most important table olive cultivars. ‘Gemlik’ is cultivated almost everywhere in Turkey and advised as a pollinator for olive orchards. The pollen of ‘Gemlik’, ‘Domat’ and two clones of ‘Gemlik’ cvs; GE119 and GK138 were used for in vitro germination experiment. 23 different media were used for in vitro germination. The pollen grains were germinated in optimum media (water, agar, sucrose, boric acid, calcium chlorite).The highest pollen germination was observed in ‘Gemlik’ cvs., lowest was GK138. Germination rate were; Gemlik 45%, Domat 44%, GE119 42% and GK138 27%. When the pollen of ‘Gemlik’ and GE119 were stored 35 days at ‐20oC the germination rate were 41% and 40%, respectively. So pollen of ‘Gemlik’ cvs can be harvested, stored in the fridge for one month and used for supplemented pollination in the same season.

Keywords: Gemlik, Domat, pollen viability, pollen germination, pollen storage, Olea europaea L.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 58

Comparison studies among different olive fruit cultivars (Olea europaea L.) reveals differences in expression levels of genes involved in Vitamin E biosynthesis

E.C. Georgiadou1,2,3, G. Koubouris3, C. Sergentani3, P. Kalaitzis2, G.A. Manganaris1, V. Fotopoulos1

1Department of Agricultural Sciences, Biotechnology & Food Science, Cyprus University of Technology, 3603 Lemesos, Cyprus 2Department of Horticultural Genetics and Biotechnology, Mediterranean Agronomic Institute of Chania (MAICh), Alsyllio Agrokepiou, 73100 Chania, Greece 3ELGO DEMETER, NAGREF – Institute for Olive Tree and Subtropical Plants, Agrokipion, 73100 Chania, Greece.

Tocopherols and tocotrienols are lipid‐soluble molecules represented by four different forms (α, β, γ and δ) that belong to the vitamin E group of compounds. This study compares the oil content and the gene expression of vitamin E in the fruit flesh of olive cultivar ‘Koroneiki’ with four other common Greek cultivars (Kalamata’, ‘Mavrelia’, ‘Kalokairida’ and ‘Lianolia Kerkyras’) based on skin color (green‐yellow, yellow‐purple, purple, and black), and weight of black fruit [1: Typical fruit weight for each cultivar, 2: Fruit with weight at least 10% lower than #1 (small fruit), 3: Fruit with weight at least 10% higher than #1 (large fruit)]. Thirty‐ to forty‐year‐old irrigated trees of the examined cultivars were selected based on growth and fruit yield uniformity. Representative healthy fruit samples (1‐2 kg) were harvested in triplicate around the canopy at human height between the beginning of November and the middle of December from 3‐4 olive trees for each cultivar according to the maturity stage of each cultivar. Morphological measurements were conducted according to UPOV (1999) TG/099/3. The oil content was determined by Soxhlet extraction and was expressed as a percentage of dry and fresh olive paste weight, while a fruit flesh subsample was stored at ‐80oC for further analyses. The cvs ‘Koroneiki’ and ‘Mavrelia’ had the lowest fruit weight and diameter but they had the highest oil content as a percentage of dry and fresh olive paste weight compared with to the other cultivars. For the molecular analysis of tocochromanol biosynthesis in fruit flesh, real‐time RT‐PCR (qRT‐PCR) was performed using ubiquitin (UBQ2) as a reference gene. Gene expression studies showed a general down‐regulation pattern of tocopherols and tocotrienols biosynthetic genes in all cultivars examined, with the exception of 4‐hydroxyphenylpyruvate dioxygenase (HPPD) in cv. ‘Lianolia Kerkyras’ and the genes HPPD and vitamin E2 (VTE2) (on skin color yellow‐purple) in cv. ‘Kalamata’.

Keywords: Olea europaea, oil content, morphological measurements, vitamin E

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 59

Molecular characterization of vitamin E biosynthesis in olive fruit (Olea europaea L.) during three successive growth seasons

E.C. Georgiadou1,2, V. Goulas1, T. Ntourou2, G.A. Manganaris1, P. Kalaitzis2, V. Fotopoulos1

1Department of Agricultural Sciences, Biotechnology & Food Science, Cyprus University of Technology, 3603 Lemesos, Cyprus 2Department of Horticultural Genetics and Biotechnology, Mediterranean Agronomic Institute of Chania, Alsyllio Agrokepiou, 73100 Chania, Greece

Vitamin E is a general term used to describe a group of eight fat‐soluble compounds known as tocochromanols. These eight forms of vitamin E are divided into two groups with four forms each, namely α‐, β‐, γ‐, and δ‐tocopherols and tocotrienols, respectively. Vitamin E has strong antioxidant capacity and is found in high concentrations in olive fruits, with previously demonstrated protective effect against cancer, diabetes, cardiovascular and neurological diseases. The present study focuses on the molecular characterization of vitamin E biosynthesis during eight developmental stages (12‐30 weeks after flowering) of cv. ‘Koroneiki’ during three consecutive growth seasons. Fruit material was collected from the olive grove of the Mediterranean Agronomic Institute of Chania, Crete starting in the middle of July and ending the beginning of December each year. Gene expression levels were examined using real‐time RT‐PCR (qRT‐PCR) with the ubiquitin (UBQ2) gene as housekeeping reference gene. During the first year, a general down‐regulation pattern was observed in tocopherols and tocotrienols biosynthetic genes in fruit flesh. In contrast, a significant up‐regulation was observed in specific genes [(Geranylgeranyl reductase, 4‐hydroxyphenylpyruvate dioxygenase (HPPD) and vitamin E4 (VTE4)] during the second year. In the third year the expression of the tocopherols and tocotrienols biosynthetic genes was up‐regulated [with the exception of the gene phytol kinase (VTE5)]. The observed changes in gene expression patterns can be attributed to differences in weather conditions in each one of the three years.

Keywords: Olea europaea, fruit developmental stages, gene expression, tocopherols, tocotrienols, tocochromanols OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 60

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 61

PLANTING SYSTEMS ORCHARD MANAGEMENT

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 62

Olive planting systems and mechanization

J. Tous

EMP Agrícola. C/ Sant Antoni, 44. 43480 Vila‐seca (Tarragona), Spain.

The productivity of the traditional olive orchards is relatively low and the crop costs are very high (harvesting, pruning, etc.). Since the 70‐80s, the increase of the olive surface has been associated to an intensification of the orchards and several authors recommend to use higher densities (HD), about 200‐500 trees/ha, with drip irrigation, designed for harvesting with trunk shakers, and with higher yields and low‐medium production costs. Finally, at the beginning of the 90s, a new type of olive orchards (super‐high density hedgerow, SHD) appeared in Catalonia (NE of Spain), with densities ranging between 1,500 and 2,500 trees/ha. Later they were introduced into other Spanish regions (Andalusia, Aragon, etc) and other countries (Portugal, Chile, California, etc.). This system facilitates the use of continuous mechanical straddle harvesters and the achievement of higher yields within a few years after planting. To improve the efficiency of the harvester, vigour must be managed to limit tree size while maintaining high productivity. However, there are few cultivars adapted to this system and their management is more difficult than the intensive models. A comparative studies on the economic viability of high and super‐high density olive orchards in Spain, indicate that HD economic ratios are more profitable than the SHD ratios. However, the latter could be the most sustainable option in large orchards with short term investments, mainly due to the full harvest mechanization and reduced labor requirements. Finally, the planting model choice in each region depends on different factors, such as ecological conditions, orchard size, manpower availability and the economic life investment.

Keywords: Planting models, cultivars, yields, mechanical harvesting, production cost.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 63

Developing mechanical harvesting for California “Black Ripe” table olives (Olea europea cv. ‘Manzanillo)

L. Ferguson1, J. Miles2, S. Garcia3, T. Sarcoglu4, F. Aranda5, F. Jiminez3, W. Krueger6, E. Ficthner6

1Department of Plant Sciences, University of California Davis, Davis CA, USA 2Department of Biological & Agricultural Engineering, University of California Davis, Davis, CA, USA 3Department of Rural Engineering, University of Cordoba, Cordoba, Spain 4Department of Biosystems Engineering, Faculty of Agriculture, Aydin, Turkey 5Institute of Agricultural Research and training, Andalusia, Granada, Spain 6University of California Farm Advisors, Glenn and Tulare Counties, California USA

California’s major table olive cultivar, the Olea europea cv. ‘Manzanillo’ is harvested physiologically immature and oxidized from green to black in processing. In recent years the hand harvesting costs have ranged from 40% to 75% of gross return while prices per ton have stagnated. Recent legislation has decreased the available labor pool, there is competition for harvesters from the table grape industry, and the dwindling labor pool is aging. The combination of rising labor prices, less labor and stagnant per ton has reduced the California table olive industry to less than 9,000 hectares. In a 2013 trial a prototype canopy contact harvester removed 92% of an 11.88 Mg.ha‐1 mechanically pruned crop, versus 81% for a 27.12 Mg.ha‐1 hand pruned control crop in a 19‐year‐old, 4 x 8‐m grove with 335 trees.ha‐1 adapted for mechanical harvesting with 6 years of mechanical topping and hedging. Approximately 85% of the hand‐ pruned olives were cannable olives versus 86% of the mechanically pruned olives. While there was no significant difference in value per ton between the two pruning treatments the mechanical pruning produced a higher percentage of extra large olives versus the more desirable medium size olives. Over the 6 years of the mechanical pruning trial the mechanically pruned trees averaged an insignificantly different annual 9.41 Mg.ha‐1 versus 11.88 Mg.ha‐1 for hand‐ pruned trees. Again in 2013 the canopy contact harvester achieved 81% harvester efficiency with a 13 Mg.ha‐1 crop in a 12‐year‐old, 3.7 x 5.5‐m, 487 trees.ha‐1, mechanically pruned hedgerow grove versus 80% efficiency for an 11.44 Mg.ha‐1 crop with hand pruned hedgerow trees. Similarly, pruning treatments produced no significant differences in the percentage of cannable olives, fruit size distribution, or value per ton. In this 3‐year trial hand and mechanical pruning were used to produce a hedgerow; the hand pruned trees have averaged an insignificantly different 8.94 Mg.ha‐1 versus 8.29 Mg.ha‐1 for mechanically pruned trees.

Keywords: mechanical pruning, canopy contact harvester.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 64

Characterizing olive tree geometry features using Unmanned Aerial Vehicle (UAV) images

J. Torres‐Sanchez1,, F. Lopez‐Granados1, N. Serrano2, O. Arquero2, R. Fernandez‐Escobar3, J.M. Pena1

1Department of Crop Protection, IAS‐CSIC, 14080 Córdoba, Spain 2IFAPA Alameda del Obispo, Avda Menendez Pidal s/n 14071, Córdoba, Spain 3Department of Agronomy, UCO, 14080 Córdoba, Spain

Olive tree geometric features such as height, diameter, area and volume are used by olive researchers and producers for inventory and crop monitoring. Tree height and diameter are traditionally measured after an intensive field work and, subsequently, volume and surface trees are estimated from these measures by using equations that treat the trees as geometric solids, which may produce inconsistent results. In this work, we present an innovative automatic method for olive tree characterization based on two phases: 1) close range photogrammetry from Unmanned Aerial Vehicles (UAV) and 2) use of object‐ based image analysis (OBIA) techniques. In the first phase, a multispectral camera onboard a multirotor UAV was used to acquire a set of images with high overlapping over an olive orchard, and a Digital Surface Model (DSM) was automatically generated by stereoscopy. In the second phase, the DSM and the orthomosaic generated with these images were analyzed with OBIA methods in order to detect the single trees and measure their geometric features. The presented methodology was tested in two olive orchards in southern Spain. 100% of the olive crowns were reconstructed by the photogrammetric software, and 100% of the olives trees were detected by the OBIA algorithm. Area, length, width, maximum height and volume of the olive trees were calculated and all these data were automatically georeferenced to create maps representing the spatial variability of the crop. The presented methodology allowed an accurate counting of the olive trees, which is very useful to orchard inventory. It also allowed a notably reduction in time and labor of measuring tasks for orchard monitoring, making possible to remotely measure several geometric features. Another important point is that this method could be adapted to other similar woody crops.

Keywords: Olea europaea, stereoscopy, tree morphology, remote sensing, OBIA

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 65

Ecosystemic Management Plan of the olive sector in Draa and Tafilalet areas (Morocco)

A. Abdellaoui1 , M. Hemmi2, L. Sikaoui3, A. El Antari3, A. Berahmani4 and M. Ferrahi4

1ORMVA Ouarzazate, Morocco 2ORMVA Tafilalet, Morocco 3INRA, CRRA Marrakech, BP 533 Morocco 4CRPII, Ifrane, Morocco

Ecosystemic Management Plan (EMP) of the olive sector in Draa and Tafilalet areas (South Eastern of Morocco) is based on potentials and constraints of the biophysical and socio‐economic status, their spatial‐temporal dynamics and their interrelations. This new approach has the advantage of considering in their globosity ecosystems that support agricultural activity. This approach is thus a tool allowed to capitalize on knowledge, to ensure consistency of data, the networking of actors from different links in the system and to generate interest and adherence of all stakeholders. In addition it allows us to create a database which can be updated and enriched progressively by different stakeholders, to have spatially vision of all links in the value chain and to have a dynamic overall vision of ecosystems underlying agricultural production and reduce the impact of chain on the environment. The work is accomplished in three phases. The preparatory phase is the first step for identifying the territory of action with its area of influence, collect and spatialization of information, the collection of biophysical, socio‐economic and legal data, and planning tools of the sector at different scales. The Diagnosis phase permitted to develop six maps which are: production base; production diagnosis, water resources, pressures, olive processing, marketing and planning tools. The participatory planning phase was conducted as participatory workshops with stakeholders consisted of the identification of actions to overcome constraints identified during the diagnostic phase. These actions are spatially related on four maps: production, processing, marketing and preservation of the environment. The main results show that olive carries great pressure on the oasis environment. Actions are proposed to mitigate these pressures.

Keywords: olive, ecosystem plan, constraints, environment preservation actions.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 66

Effect of Planting Density on the Behavior of Sourani Olive Tree under Kuwaiti Environment

O. Al‐Ragam, H. Al‐Menaie, H. Mahgoub, M. Al‐Hadidi, A. Al‐Shatti and M. Al‐Zalzalah

Kuwait Institute for Scientific Research, P.O. Box 24885, Safat 13109, Kuwait

It is a known fact that planting systems/density and canopy management are integrated techniques aimed at maximizing productivity and production efficiency in olive growing. In this regard, it is important to note that the planting density, besides affecting the growth of the trees, has an influence on time required, having full production in terms of yield per hectare, microclimate conditions, use of soil resources, landscape appearance, and contrast to wind. These aspects are assumed to be of particular importance in countries like Kuwait, where olive is studied for both commercial (for the production of olives/oil) and environmental/beautification (greenery and, desertification control) purposes. Hence, one‐year old acclimatized plants of the Sourani variety were studied to evaluate different planting densities (6 x 6 m, 6 x 3 m, and 6 x 1.5 m) and training systems for commercial and environmental purposes. The planting density strongly affected the tree growth.

Keywords: Olive tree, density, canopy management

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 67

A comparative study of hand‐held harvesting machine with traditional methods used for olive harvesting in Jordan

R. Ahmad , S. Ayoub

Olive Research Department, National center for Agriculture Research and Extension, Jordan.

This study was carried out for two seasons (2011‐2013) to evaluate the effectiveness of hand held machine and to compare it with traditional methods for harvesting of two olive cultivars. This experiment was performed at Raba Research Station, Karak, Jordan, using two olive cultivars "Nabali Baladi" and "Nabali Mohasan" which were about 15 years old. Three methods of harvesting were used (hand harvesting ‐the most traditional and familiar method in Jordan, pneumatic comb machine and plastic combs. Result revealed that mechanical harvesting increased significantly harvesting productivity (the weight of harvested fruits by one worker /time spend by worker for harvesting) by five to six times for both cultivars "Nabali Baladi" and "Nabali Mohasan" when compared with traditional method (hand harvesting), and increased by two times when compared with harvesting with plastic combs for both cultivars. The result of harvested fruits was more than 95% using hand harvesting and ranged from 86% ‐ 88% using pneumatic comb machine for both cultivars and the differences was significant between the two harvesting methods. Percentage of detached leaves (number of detached leaves / total leaves numbers / 100) was about 9‐15% with mechanical harvesting by hand‐ held shaker in the two cultivars and from 4‐5% with manual harvesting methods. The percentage of injured fruits was less than 2% in "Nabali Baladi" and "Nabali Mohasan" in the two seasons, and it increased to 6‐ 7% with mechanical harvesting by pneumatic comb machine. Mechanical harvesting using hand‐ held pneumatic comb in olive cultivars "Nabali Baladi" and "Nabali Mohasan" greatly improved the time of harvesting efficiency and reduce the cost of labors.

Keywords: Olea europaea L., "Nabali Baladi", "Nabali Mohasan", mechanical harvesting, pneumatic comb, harvesting productivity

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 68

Adaptation of different varieties of olive tree to high‐density planting systems

O. Arquero, N. Serrano, M. Vinas, M. Lovera

IFAPA Alameda del Obispo de Córdoba. Junta de Andalucía. Spain

Spain is the country with the largest surface of olive trees in the world. Currently olive tree surface is over 2,584,000 ha, most of it covered by traditional design plantations i.e. plantation frameworks going from 10x10 m to 16x16 m and plants aged from fifty to several hundred years old. These factors have resulted in a difficult mechanization of plantations and very low production yields. In the nineties new plantations systems started with higher plants density and hedgerow planting allowing a greater mechanization of different cultivation works. Most of these plantation systems were established with “Arbequina” as only single cultivar. This cultivar has a very good adaptation to hedgerow planting, is very productive, early and with a drooping growth habit. The objective of this work is the characterization of 17 olive trees varieties with hedgerow plantation design and high density of plants. Chosen varieties are the main Spanish varieties, other producing countries varieties, and varieties coming from enhancement programs of Córdoba University in Spain. Measured values were: vigour, growth habit, branching degree and production yields data. Up to now have been identified a set of varieties that can be adapted to hedgerow planting, most significant are “Arbequina”, “Chiquitita”, “Arbosana”, etc. On the other hand, some varieties were found too vigorous and little early to be established in this plantation design.

Keyworks: olive tree plantation, olive varieties, high density.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 69

Effect of different pruning methods on olive tree growing

A. Gholiyan1, A. A. Zeinanloo2, Z. yousefi3, M. Taheri4

1Department of Horticulture, Rasht Branch, Islamic Azad University, Rasht, Iran. 2Seed and Plant Improvement Institute, Horticulture department, Karaj, Iran. 3Agricultural and Natural Resources Research Center of Guilan, Rasht, Iran. 4Agricultural and Natural Resources Research Center of Zanjan, Zanjan, Iran

There are 5000 hectares of old olive orchards in Iran. Due to a lack of experience with the principles of olive rejuvenation pruning, fruit production of these orchards has reduced and their maintenance is not economical. There are many latent buds in the older parts of olive tree wood, that if they were stimulated by a proper pruning could be able to grow like common vegetative buds and where necessary, they can revive the tree. This research was performed in a traditional olive orchard, located in region of Tarom in Zanjan province from February 2012 to July 2013. The Experiment conducted as randomized complete block design with pruning techniques at three levels (continuous, periodic, complete topping the trees) with three replications. The results of the first and second year showed that the two methods of olive pruning (periodic and continuous) significantly have more new shoot growth length and weight than complete topping. Percentage of nitrogen in leaves of sucker of continuous pruning was higher than the other methods. According to the results, for rejuvenation of old olive orchards, continuous and periodic methods were suitable.

Keywords: rejuvenation pruning, periodic, continuous, topping, old olive tree

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 70

Effect of lateral mechanical pruning on light interception, vegetative growth, fruit characteristics and fruits yield in a 13 years old high density planting system

A. Larbi, I. Dhouibi, M. Msallem

Institut de l’Olivier, BP 286 , 1082, Tunis, Tunisia

Among the objectives of an olive high density planting system is to reach a high degree of mechanization in order to reduce the production costs and then to increase the grower incomes. The decrease of pruning costs is the most important factor to increase benefits per ha. The lateral mechanical pruning instead of the manual pruning can be used since the third year after planting once the vegetative structure of the canopy is formed. The main objective of the present study is to assess the impact of lateral mechanical pruning applied to one side of the canopy at 30 cm from the tree axis, on light interception, vegetative growth, fruit size, maturity index, oil content and yield. In this context, an experiment was carried out in a commercial orchard (Arbosana) located in Mornag (north of Tunisia). Trees were planted since 2000 with a density of 1250 trees/ha. Trees were divided into two layers, the bottom (0‐1m) and the medium layer (1‐2 m). It must be pointed out that control and lateral pruned trees were topped at 2.20 m. Results obtained here indicated that the application of lateral pruning increased light interception in both bottom and top layer by 5 and 8%, respectively when compared to the control trees. Vegetative growth increased significantly in the pruned side of trees when compared to the other side that has not been pruned and to the control trees. Indeed, vegetative growth increase was about 7 and 5.5 cm in the pruned side in both bottom and the top layers, respectively while the increase was only about 3.6 and 3 cm in the same layers in the control trees. Fruit weight average was also higher in pruned trees in both top and bottom layers when compared to the controls trees, although this increase was more pronounced in the bottom layer comparing to the top. The lateral pruning did not affect olive yield being approximately the same in pruned and control trees. Finally, the lateral pruning increased significantly fruit maturity index and fruit oil content when compared to control trees, although this increase was significant in the bottom layer.

Keywords: lateral pruning, yield, vegetative growth and light interception

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 71

Olive oil characteristics of twelve cultivars produced in a high‐density grove at the Valladolid province (Spain)

N. Rodrigues1, H. Martín2, M. C. Asensio‐S.‐Manzanera2, S. Casal3, A. Bento1, J. A. Pereira1

1Mountain Research Centre (CIMO), School of Agriculture, Polytechnic Institute of Bragança, Campus de Santa Apolónia, Apartado 1172, 5301‐855 Bragança, Portugal 2Instituto Tecnológico Agrario de Castilla y León, Ctra. de Burgos Km.119, 47071 Valladolid, Spain 3REQUIMTE/Laboratory of Bromatology and Hydrology, Faculty of Pharmcacy, Porto University, Rua de Jorge Viterbo Ferreira, 228, 4050‐313 Porto, Portugal

In the Mediterranean region olive crop is one of the agricultural activities with higher social‐ economic and cultural relevance. In the last decades, oliviculture expanded to regions where olives are not usually produced, like Valladolid province (Castilla y Léon, Spain). In the present work, we intend to evaluate the olive oils characteristics of 12 olive varieties (Cvs. Arbequina, Arbosana, Arroniz, Cornicabra, Frantoio, Hojiblanca, Manzanilla, Picual, Picudo, Redondilla, Royuela and Zorzal) produced in a high‐density olive grove at the province of Valladolid (Spain). Characterization occurred in two consecutive crops season (2012/13, and 2013/14) and the quality parameters (free acidity, peroxide value, UV analysis, sensory analysis), chemical composition (fatty acids profile, and tocopherols content), oxidative stability (by Rancimat method) and antioxidant activity (ABTS, DPPH and total reducing capacity methods) were evaluated. The results indicated that all the obtained olive oils were classified as extra virgin olive oils (EVOO’s), with low levels of acidity (between 0.2‐0.3%), peroxide value (between 1.9 mEq.O2/kg in Cv. Arroniz and 7.1 mEq.O2/kg in Cv. Picual), K232 (between 0.73 in Cv. Arbosana and 1.70 in Cv. Zorzal) and K270 (between 0.07 in Cv. Arroniz and 0.22 in Cv. Frantoio). Oleic acid was the most abundant (varying between 76.9 and 82.8% respectively in Cvs. Picual and Arbequina), followed by palmitic acid (7.0 and 11.6 corresponding to Cvs. Frantoio and Redondilla, respectively) and linoleic acid (3.6 and 10.4% in Cvs. Cornicabra and Zorzal, respectively). Total tocopherols content ranged from 165.7 to 341.7 mg kg‐1, reporting Cv. Royuela higher content, being α‐tocopherol the most abundant. Oxidative stability ranged between 7.46 and 28.55 h reporting Cv. Cornicabra higher stability. Cvs. Frantoio and Hojiblanca reported higher antioxidant activity for DPPH and ABTS methods respectively. Whilst Cv. Royuela showed possessing higher total reducing capacity.

Keywords: Valladolid, olive oil quality, chemical composition, oxidative stability, antioxidant activity.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 72

Effect of olive trees density on the production and quality of olive oil from Cv. Arbequina

N. Rodrigues1, H. Martín2, M. C. Asensio‐S.‐Manzanera2, S. Casal3, A. Bento1, J. A. Pereira1

1Mountain Research Centre (CIMO), School of Agriculture, Polytechnic Institute of Bragança, Campus de Santa Apolónia, Apartado 1172, 5301‐855 Bragança, Portugal 2Instituto Tecnológico Agrario de Castilla y León, Ctra. de Burgos Km.119, 47071 Valladolid, Spain 3REQUIMTE/Laboratory of Bromatology and Hydrology, Faculty of Pharmcacy, Porto University, Rua de Jorge Viterbo Ferreira, 228, 4050‐313 Porto, Portugal

In recent years, has increased sharply the number of olive trees in non‐traditional areas. The number of plants per hectare, is one of the determining factors in their productivity. In this sense, in the present work we intended to study the effect of planting density of Cv. Arbequina, in Valladolid, a non‐traditional olive producing region, on olives production and oil quality from the first years of production. Thus, we studied whether different distances on the line, 1, 1.25, 1.5, 2, 3 and 3.5 m, influences the olive production, the olive oil quality parameters (free acidity, peroxide value, UV analysis), the oils composition (fatty acids profile, tocopherols content), stability (by Rancimat method), and antioxidant activity (ABTS, DPPH, and total reducing capacity methods). The results obtained indicate that all the olive oils obtained at different densities were classified as extra virgin olive oils (EVOO), with low acidity values (0.2 ‐ 0.3%), peroxide value (2.1 ‐ 7.1 mEq.O2/Kg), K232 (0.71 ‐ 1.78) and K270 (0.08 ‐ 0.20). Regarding fatty acids composition, oleic acid was the major fatty acid (between 78.6 and 82.9%), followed by palmitic acid (8.8 – 10.6) and linoleic acid (4.5 ‐ 6.0%). Total tocopherols content ranged between 242.8 and 296.5 mg kg‐ 1, without significant differences among the plant densities studied. On oxidative stability values ranged between 9.6 and 25.2 h. Antiradical potential varied between 27.5 and 95.0% for DPPH method, and 93.0 and 95.0% for ABTS method. Total reducing capacity values ranged from 148.0 and 226.3 mg of cafeic acid per kg of olive oil. From the results obtained we can conclude that in the two first crop production years, planting density don’t interfere in the quality, composition and antioxidant capacity of Cv. Arbequina olives oils. By other hand, the effect of year is a major factor of variance, mainly due to olives production and maturation index at harvest moment.

Keywords: planting density, Cv. Arbequina, quality, chemical composition, oxidative stability, antioxidant activity.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 73

Full production olive trees architecture restructuration to continuous harvesting with large straddle machines (over‐row): evaluation of pruning intensity

F. Vita Serman1, G. Lemole1, L. Bueno1

1Instituto Nacional de Tecnología Agropecuaria (INTA) EEA San Juan. Departamento de Agronomía, Facultad de Ingeniería. Universidad Nacional de San Juan. Argentina.

New modern orchards have developed a deep intensification of the olive production system including high density (HD) designs with hedgerows structures and mechanical harvesting. However, most of the HD olive orchards, planted during the 90's in Argentina, were not considered to be harvested by large straddle machines (Colossus), making its work very inefficiently or even impossible. An experiment was developed in a 15 year‐old, high density (6.5 x 3.5 m) commercial orchard, located in San Juan, Argentina (31°55'06.7" S 68°18'56.2" W). In first two seasons (2011‐12 and 2012‐13), branches removal intensity was applied in order to adapt the orchard structure to a continuous mechanical harvesting with over‐row machines. The productive and vegetative response was then evaluated. Pruning treatments consisted of: manual‐selective intensive pruning (PMaI) (removing all the not‐well‐oriented branches to fit a suitable hedgerow width); manual‐selective semi‐intensive pruning (PMaS) (two consecutive years removing all the not‐well‐oriented branches); intensive mechanical pruning (PMeI) with lateral hedging at both sides of the row to fit a suitable hedgerow width; and a control treatment (C) with no pruning. Pruning material weight was higher on PMaI (55 Kg.plant‐1) than in PMaS (24 Kg.plant‐1) and PMeI (24.5 Kg.plant‐1) for the first year. In the second winter (2012‐13), 86.2 Kg.plant‐1 were extracted from PMaS. The canopy volume and interception of solar radiation was significantly reduced in PMaI and PMeI after the first pruning year and in PMaS after the second year, over the control. Changes in hedgerows structure modified the canopy surface area (SA) with important reductions on pruned treatments (PMaI= 10,824 m2.ha‐1; PMeI= 11,369 m2.ha‐1; PMaS=14,542 m2.ha‐1), over the control (C=17,384.4). This led to a significant reduction on fruit yield in the first growing season (PMaI= 50.8%; PMeI= 42.2%; PMaS=21.7%). In the second season, the SA was quickly recovered on PMaI and PMeI and there were not significant differences on yield production over the control. However, the SA in the second pruning year was highly reduced in PMaS, thus the fruit yield was severely affected. According to these first two‐ season results, both mechanical and manual‐selective intensive pruning ar e recommended to modify and adapt the hedgerow structure to continuous mechanical harvesting with large straddle machines (over‐row).

Keywords: Olea europaea L, Colossus, mechanical harvesting, pruning.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 74

Measurement of foliar surface of olive orchards using a laser scanner sensor

F. Vita Serman1,2,3, G. Lemole1,2, L. Bueno1,2, C. Soria4 and M. Munoz4

1Instituto Nacional de Tecnología Agropecuaria (INTA) EEA San Juan. 2Departamento de Agronomía, Facultad de Ingeniería. Universidad Nacional de San Juan. Argentina. 3Departamento de Ciencias Básicas y Tecnológicas. Universidad Nacional de Chilecito 4 Instituto de Automática‐CONICET, Facultad de Ingeniería, Universidad Nacional de San Juan

The foliar surface describes the geometric shape of the canopy and indicates photosynthetic ability and biomass. The laser scanner is a portable scanning Lidar (light detection and ranging) instrument utilized to obtain 3‐D structural properties of plants. In recent years, Lidar sensors have been widely used for the measurement of environmental parameters, particularly for the characterization of forest and agricultural systems. For agricultural applications, was proposed a methodology for managing and computing laser sensor data to obtain several parameters related to the geometric characteristics of olives trees (height, surface, volume) and to some properties that define the structural characteristics (foliage density and foliage distribution). Some experiments have been carrying out with a lidar Sick LMS291 mounted on an autonomous all terrain vehicle in an olive plantation. As conclusion, the laser‐based measurement system described here proved to be a valuable tool for the measurement of the physical and structural characteristics of olives, such as tree volume, foliar surface, leaf area density and LAI. The ability of laser sensors for measuring, in a rapid and non‐destructive manner, crop leaf area, tree‐row volume and other crop parameters, makes this system a promising tool to be used as support for the decision making related to the optimization of olives management practices.

Keywords: foliar surface, olive tree structure, canopy architecture, Lidar

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 75

IRRIGATION STRESS PHYSILOGY NUTRITION

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 76

An environmental‐friendly approach for a productive olive orchard management

C. Xiloyannis, A. Palese

Dipartimento delle Culture Europee e del Mediterraneo: Architettura, Ambiente, Patrimoni Culturali (DiCEM) – Università degli Studi della Basilicata, via San Rocco, 3 – 75100 Matera (Italy)

The productive performance of an olive orchard is strongly affected by the grower capacity in choosing, combining, and properly performing the several agronomical practices, such as soil management, fertilization, irrigation, also taking into account their environmental impact. Such expertise is taken into consideration by the recent agricultural European Union policy and at worldwide level, in order to support and encourage the use of production technologies aimed to preserve natural resources (soil, air and water), to control environmental pollution (greenhouse gas emissions, nitrates, pesticides), and to assure product quality (from hygienic, nutritional, and organoleptic point of views). As shown by the recent scientific literature on this topic, an environmental‐friendly management in olive orchards can determine an optimal plant nutritional equilibrium, avoid nutrients accumulation and leaching risks, improve irrigation efficiency, prevent soil erosion and root asphyxia, fix atmospheric CO2, etc. The application of agricultural techniques with low negative environmental impacts allows also to guarantee top yields of high quality. Some practical suggestions for a sustainable soil management and a proper irrigation and fertilization of olive orchards are presented in this paper taking into account the current and urgent environmental emergencies and what are the production expectations of olive growers.

Keywords: sustainability, organic matter recycling, residues quality, soil nutrient synchronization, irrigation efficiency.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 77

Olive irrigation management project – Setting the scene

V. Nangia, M. Karrou, T. Oweis

International Center for Agricultural Research in the Dry Areas (Jordan/Morocco)

The key success factors of the olive oil market are the enhancement and stabilization of olive yields, improvement of olive oil quality and reduction of production costs. There is great potential for water saving in olive production in Morocco and Syria. This presentation is to set the scene for the following six presentations on a study on olive irrigation management being implemented in Morocco and Syria. The main purpose of the four year (2010‐2014) study is to enhance crop yields of smallholder olive farmers in two Mediterranean countries ‐ Morocco and Syria, through the optimization of water management practices applied to olive cultivation. Due to the sufficiency of rainfall in Syria for growing olives, the study investigates the effect of supplemental irrigation in increasing and stabilizing yield, whereas in Morocco the low rainfall allows only irrigated olive cultivation and so the study investigates the effects of deficit irrigation in reducing water consumption without adversely affecting the olive yield. Results indicate that adding 230 mm (a 100% of crop water requirement) supplemental irrigation led to as much as 175% increase in olive yields compared to rainfed olive in Syria, and in Morocco, adding water by drip irrigation method at 70% of ETc led to 63‐77% saving in water compared to flood irrigation along with 13‐ 43% increase in yields.

Keywords: Drip irrigation, deficit irrigation, Morocco, supplemental irrigation, Syria, water productivity

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 78

Water requirement and water‐use efficiency for olive trees under different irrigation systems

A. Hijazi1, M. Doghoze1, N. Jouni1, V. Nangia2, M. Karrou3, T.Oweis2

1 General Commission for Scientific Agricultural Research (GCSAR), Syria 2 ICARDA, Amman, Jordan 3 ICARDA, Rabat, Morocco

The research was carried out in Nashabia agricultural research station in rural Damascus province, which belongs to General Commission for Scientific Agricultural Research (GCSAR), it is located 16 km east of Damascus city. The research aimed to determine Olive water requirement and irrigation frequency under different irrigation techniques according to the phonological stages, and to find out crop coefficient (Kc), also studying irrigation efficiency and water use efficiency (WUE). Experimental design included four treatments: 1st is drip irrigation system (6 drippers/tree, with discharge of 8 L/h for each), 2nd is mini‐sprinkler irrigation system (2 mini‐ sprinklers/tree, with discharge of 55 L/h for each), while 3rd is surface irrigation system by boarder stripes (100*3 m, with a discharge of 2 L/Sec/m on the head of the stripe), and finally 4th is surface irrigation system by soil rings which considered as a control (with a diameter of soil ring around tree of 4.5 m, which receives irrigation water from a channel on the top of the field), neutron probe was used to follow up soil moisture changes, and irrigation was applied when soil moisture declines to 80% of field capacity, also climatic data from agro‐meteorological station were used for getting ET0 values. Studied cultivar was Qaisi, with a trees spacing of 7*8 m. As a result, drip irrigation was superior to other irrigation systems in the domain of fruit yield, with 8532 kg/ha compared with 7155 kg/ha for the control, where the difference between drip and mini‐sprinkler treatments was not significant, with 8384 kg/ha for the mini‐sprinkler treatment. Also drip irrigation treatment was superior to other treatments, in the domain of water use 3 3 efficiency with 2.36 kg/m , compared to 1.30 kg/m for the control. As for crop coefficient Kc , it was found that the lowest values were at beginning and the end of season, where the highest values were in both of June and July months, which corresponds with the physiologic maturity and colouring of fruits. Finally it was clear that applying drip irrigation system in olive trees irrigation, can save 34.4% of total applied irrigation water, and increase fruit yield by 19.2%, when compared with surface irrigation system by soil rings (control).

Keywords: Deficit irrigation, drip irrigation, water requirement, water use efficiency, mini‐sprinkler, surface irrigation, olive.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 79

Effect of deficit irrigation on vegetative growth and fruit yield parameters of young olive trees (Olea europaea L.) in semi‐arid area of Morocco

L. Sikaoui1, A. Bouizgaren1, M. Karrou2, H. Boulal2, B. Boulouha1, M. Karama1, V. Nangia3, T. Oweis3

1INRA, CRRA Marrakech, BP 533 Morocco 2ICARDA, Rabat, Morocco 3ICARDA, Amman, Jordan

The availability of irrigation water in Morocco is an issue of growing concern, particularly in semi‐ arid areas which are characterized by high evaporative demand (more than 1500 mm/ year) and irregular and low rainfall (200‐300 mm/year), that increased the demand for irrigation of olive trees. The objective of this study was to evaluate the effect of deficit irrigation on vegetative growth and fruit yield parameters of young olive trees. The trial carried out at Sâada experiment station INRA in Marrakech region, where the average yearly cumulated rainfall is less than 200 mm. The olive orchard was planted on December, 2010. The trees were exposed to three irrigation treatments, T1: Full drip irrigation (100% ETc), T2: Deficit drip irrigation (70% ETc and T3: flood irrigation using the farmer’s technique. The amount of irrigation applied between January and November 2013 was 538 mm in flooded plots against 116 mm in the case of 100% ETc and 81 mm for 70% ETc. The results showed that the trunk perimeter was affected by irrigation treatments with the highest mean values were obtained with T2 treatment, although it was not significantly different than T1 treatment. The average canopy diameter also varied significantly among treatments with the highest values were obtained with T1 treatment (137.2 cm), which did not differ significantly from deficit drip irrigation treatment T2 (133.9 cm). Significant variations were noted among treatments for canopy volume, and the values were obtained by treatments T1 (1.22 m3) and T2 (1.24 m3) which were significantly higher than that of treatment T3 (0.71 m3). The effect of irrigation regime was significant on the fruit set rate and the high values were achieved by deficit drip irrigation (62%), followed by full drip irrigation (60%) and then flood irrigation (40%). The olive fruit yield showed significant differences between the three treatments. The highest yield was recorded by T1 (146 kg/ha) which does not differ significantly from T2 (117.6 kg/ha); the lowest yield was obtained by T3 (64.2 kg/ha). Moreover, the difference between the three treatments for olive fruit weight was significant and the values were the highest under T1 (3.42 g) and T2 (3.36 g) and lowest under T3 (3.38 g). However, the oil content was not affected by the treatments tested. From this study, we can conclude that deficit irrigation (70% ETc) saves irrigation water without affecting negatively the growth and yield of young olive trees.

Keywords: Young olive trees, irrigation regime, growth, yield.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 80

Effect of switching from surface to drip irrigation on the performance of mature olive trees in a dry area of Morocco

L. Sikaoui1, A. El Antari1, A. Bouizgaren1, M. Karrou2, H. Boulal2, B.Boulouha1, M. Idrissi1, V. Nangia3, T. Oweis3

1INRA, CRRA Marrakech, BP 533 Morocco 2 ICARDA, Rabat, Morocco; 3ICARDA, Amman, Jordan

In Morocco, the major problem that limits olive production is drought and the scarcity of water for irrigation in the southern part of Morocco. Rainfall in the area does not exceed 240 mm on average. Consequently, olive orchards are irrigated throughout the year to ensure an economic yield. However, a large proportion of orchards area is irrigated by a traditional inefficient surface irrigation system leading to high water losses. When this technique is practiced, the water applications are often excessive and irrigation scheduling is not based on the requirements of the crop. The objective of this study is to compare the performance of 36 years‐old olive trees under three irrigation regimes (100% ETc and 70% ETc under drip irrigation system and flooded surface irrigation) and two tillage treatments (deep and minimum tillage). In total four treatments are tested: T1 (100% ETc x Minimum tillage), T2 (100% ETc x Deep tillage), T3 (70% ETc x Minimum tillage), T4 (70% ETc x Deep tillage) compared to T5 (Flood irrigation farmers’ technique). The switching from conventional to drip irrigation was made on December, 2010. The average amounts of irrigation water applied in 2012 and 2013 were 936 mm for flooding treatment, 524 mm for 100% ETc and 367 mm for 70% ETc. The results showed significant effect of the treatments on olive yield. The high values were achieved under T4 treatment with 7.9 t/ha and 9.3 t/ha in 2012 and 2013, respectively. The low values were obtained under T5 treatment with 4.5 t/ha and 7.9 t/ha in 2012 and 2013, respectively. Significant variations were also shown among irrigation treatments for the shoot growth rate. The drip irrigation induced higher values of this parameter (16.9%) in comparison with flood irrigation (11.1%). We noted a significant difference between deep and minimum tillage; but with the same tillage, we did not observe any significant difference between full and deficit irrigation. Average olive weight was higher under drip irrigation (4.96 g/olive) compared to flood irrigation (4.48 g/olive). The comparison of tillage treatments under drip irrigation showed that the highest average olive weight was obtained by deep tillage (5.16 g/olive). In conclusion switching mature olive orchard from surface to drip irrigation requires a deep tillage that improves water use by the trees, while the combination of deficit irrigation (70% ETc and deep plowing increases yield.

Keywords: olive, switching, flood irrigation, drip irrigation

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 81

Effect of three irrigation regimes on physiological parameters of new olive orchard under semi‐arid conditions of Morocco

L. Sikaoui1, , A. Bouizgaren1, M. Karrou2, H. Boulal2, B. Boulouha1, M. Karama1, V. Nangia3, T. Oweis3

1INRA, CRRA Marrakech, BP 533 Morocco 2ICARDA, Rabat, Morocco 3 ICARDA, Amman, Jordan

Flood irrigation, which is the predominant technique practiced by farmers in Morocco, is a source of excessive and inefficient use of irrigation water. The objective of this study is to evaluate the effect of deficit irrigation on physiological processes of young olive trees. A trial was carried out at Sâada experiment station in Marrakech region. The climate is a typical Mediterranean semi‐ arid with a hot and dry summer. The average yearly cumulated rainfall is less than 200 mm. The olive orchard was planted on December, 2010. Trees in orchards had a spacing of 8 m x 8 m. The trees were exposed to three irrigation treatments, T1: Full drip irrigation corresponding to 100% ETc, T2: Deficit drip irrigation (70% ETc) and T3: flood irrigation using the farmer’s technique (with 6 to 7 irrigations). The ETc was calculated by the following equation: ETc= ET0 x Kc x Kr/Ne, where ETo is the reference evapotranspiration (Penman Monteith), Kc is the crop coefficient for olive tree, Kr is the coefficient to correct for incomplete cover and Ne is the efficiency of irrigation network. The amount of irrigation applied between January and November 2013 was 538 mm in T3 treatment against 116 mm for T1 and 81 mm for T2. The physiological parameters were measured, during the third year after planting, at flowering, fruiting and fruit set stages. Results showed that at the three phenological stages, all the parameters varied significantly with treatments. The lowest values of leaf relative water content were obtained under T3 treatment, while no significant differences were recorded between T1 and T3 treatments. For stomatal conductance, the highest and lowest values were recorded under T1 and T3 regimes, respectively. The maximum values of total soluble carbohydrates were obtained with T3 treatment and the minimum ones with T1 treatment which did not differ significantly from T2 treatment. Leaf protein content was significantly lower under drip irrigation compared to surface irrigation; no significant differences were noted between full and deficit irrigation. Deficit irrigation (T3) did not affect significantly leaf proline content compared to full drip irrigation. The high values of this parameter were obtained under surface irrigation treatment. Significant decrease in leaf total chlorophyll content was found under T3 treatment in comparison with T1 and T2 treatments. The highest and lowest values of leaf total mineral content were obtained by T1 and T3 treatments respectively. From this study, we can conclude that deficit irrigation did not affect negatively the physiological parameters measured on young olive trees; however, surface irrigation technique expose the plants to water stress despite the high amount of water applied.

Keywords: olive, water stress, deficit irrigation, physiological parameters

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 82

Impact of different levels of deficit irrigation on olive productivity under drip irrigation system

N. Al Jouni1, M. Hayek1, F. Ayesh1, V. Nangia2, M. Karrou3, T. Oweis2

1General Commission for Scientific Agricultural Research (GCSAR), Syria 2 ICARDA, Amman, Jordan 3 ICARDA, Rabat, Morocco.

The research was carried out in two sites in Dara'a province, the 1st was in Jillin agricultural research station, which belongs to General Commission for Scientific Agricultural Research (GCSAR), it is located 25 km North of Dara'a city, While 2nd was in Al Asha'ari area, in a field belongs to a private farmer, it is located 16 km North West of Dara'a city. The research aimed to determine Olive water requirement at different levels of deficit Irrigation, and to find out irrigation efficiency and water‐use efficiency (WUE), also studying the relationship between yield and water, according to different treatments. Experimental design included three water treatments: 1st is given 100% of crop water requirement, 2nd is given 50% of crop water requirement, while 3rd is a rainfed treatment (control), drip irrigation system was applied, irrigations were applied when soil moisture declines to 50% of available water, soil moisture was followed up by using neutron probe, and climatic data from agro‐meteorological station were used for getting ET0 values, fertigation was applied by using Dosatron‐type fertilizer injector. Studied cultivars in the 1st site were Sorani, Jluott, with a trees age of 30 years, and a spacing of 8 x 8 m, while studied cultivar in the 2nd site was Sorani only, with a trees age of 46 years, and a spacing of 10 x 10 m. As a result, it was found that applying deficit irrigation (50%) of water requirement on olive trees, when compared with full irrigation, can save about 1500 m3/ha, which means saving millions of cubic meters at country level, also it can cause a fruit yield reduction, between 1200 and 2000 kg/ha, where this loss can be endured under water irrigation shortage and the high power costs. The loss in the oil yield can be neglected, with a values ranged between 50 – 250 kg/ha, where it was found that oil ratio increases under deficit irrigation and rainfed conditions. Finally it was clear that applying deficit irrigation (50%) of water requirement on olive trees, can create a significant fruit yield increase with 3500 – 5400 kg/ha when compared with rainfed trees, when under conditions of irrigation water resources availability.

Keywords: deficit irrigation, drip irrigation, rainfed, water use efficiency, olive, ET0, fertigation

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 83

Integrated approaches to evaluate the whole‐plant water relations in irrigated and rainfed Olea europaea

M. Centritto1, G. Marino2, C. Cocozza3, A. Giovannelli1, C. Cantini1, R. Tognetti3

1CNR‐IVALSA, via Madonna del Piano 10, I‐50019 Sesto Fiorentino (FI), Italy 2CNR‐IPSP, via Madonna del Piano 10, I‐50019 Sesto Fiorentino (FI), Italy 3DIBT, Università del Molise, I‐86090 Pesche (IS), Italy

The major need for development of irrigation is to save substantial water through improved irrigation management and increased water productivity. The development of new strategies in water saving requires improved determination of whole‐plant water relations. In this sense, a better knowledge on water transport in trees can provide practical information, through simultaneous measurements of whole tree water loss. This work presents an integrated approach to evaluate the whole plant water status through the use of canopy spectral reflectance indices, sap flow and stem radius variations in rainfed and irrigated mature olive (Olea europaea L.) trees in field conditions in central Italy. Continuous stem water relations, sap flux and radial variation, and soil moisture were monitored for detecting the early plant response to water deficit, whilst canopy reflectance indices were used for detecting plant functioning in response to seasonal drought. On a daily basis, sap flux density showed an opposite trend with respect to patterns of vapour pressure deficit and evapotranspiration (ET0) in spring, corresponded to the course of ET0 in summer and lagged behind ET0 in autumn. Maximum daily shrinkage was correlated with vapour pressure deficit (R2 = 0.50, in rainfed; R2 = 0.51, in irrigated plants), whereas it was not significantly related to ET0. Conversely, cumulative sap flow showed a good 2 2 relationship with ET0 (R = 0.60, in rainfed; R = 0.51, in irrigated plants) and scaled exponentially with maximum daily shrinkage (R2 = 0.56, in rainfed; R2 = 0.61, in irrigated plants), indicating consistent radial transfer of water between xylem and phloem. Canopy reflectance vegetation indices assessed significantly the effects of seasonal drought on rainfed trees and their subsequent recovery. Photochemical reflectance index (PRI) resulted better correlated with photosynthesis (A) (r2= 0.587) than with the other measured parameters, while normalized difference vegetation index (NDVI) showed a stronger relationship with leaf water potential (Ψl) 2 2 (r = 0.668) and stomatal conductance (gs) (r = 0.547). Water index (WI) scaled linearly as gs and 2 2 Ψl increased (r = 0.597 and r = 0.576, respectively) and, even more interestingly, a good correlation was found between WI and whole‐plant transpiration (r2= 0.668) and between WI and A (r2= 0.640). Our findings demonstrate that: a) the correspondence between stem factors (sap flow and diameter variation) and environmental parameters (water availability and evaporative demand) can be determined by a mathematical function b) the seasonal courses of reflectance indices scaled linearly with gas exchange parameters and resulted correlated with whole‐plant transpiration as assessed by stem sap flux density. The proposed analytical approach highlights the importance of combining measured parameters and sensor outputs in order to determine adjustments of specific plant functionality traits, such as the dynamics of water status in trees, for modelling and practical uses.

Keywords: Canopy reflectance, canopy transpiration, drought, leaf gas exchange, water status. OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 84

Irrigating a hedgerow olive orchard to improve production and quality J.E. Fernández1, A. Perez‐Martin1, M.V. Cuevas1, C.M. Rodriguez‐Dominguez1, A. Morales‐Sillero2, J.M. García3, V. Hernandez‐Santana1, A. Diaz‐Espejo1

1Institute of Natural Resources and Agrobiology (IRNAS, CSIC), Seville, Spain 2Department of Agroforestal Sciences, ETSIA, University of Seville, Spain 3Institute of Fats (IG, CSIC), Seville, Spain

Irrigation management in olive orchards with high plant densities is a challenge. A proper balance between water consumption, plant vigour, production and quality must be achieved. In this study main variables related to plant development, plant water stress and quantity and quality of fruit and oil production were measured from 2010 to 2013 in an ‘Arbequina’ olive orchard planted in 2007 near Seville, southwest Spain. Trees were at 4 m × 1.5 m (1667 trees ha‐1), had a single trunk and an average size of 2.10 m wide and 3.2 m high. Irrigation was applied during the dry seasons, from May to October, with a lateral per tree row with a 2 L h‐1 dripper every 0.5 m. Three irrigation treatments were evaluated: full irrigation (FI) to replace daily 100% of the irrigation needs (IN), calculated as IN = ETc‐Pe, where ETc is the maximum potential crop evapotranspiration and Pe is the effective precipitation; and two regulated deficit irrigation (RDI) treatments, which supplied a total of ca. 60% of IN (60RDI) and 30% of IN (30RDI), respectively. The applied RDI strategy was derived from the existing literature for olive when we started the experiments. The average value of IN amounted to ca. 4400 m3 ha‐1. For fresh fruits, the yield/leaf area (kg m‐2) ratio was ca. 1.1 for all treatments. For virgin olive oil this ratio was 0.067 both for FI and 60RDI and 0.084 for 30RDI. During the four years of work we improved the RDI strategy, derived crop coefficient (Kc) values, and found that a water supply of around 45% of IN, i.e. in between the 60RDI and 30RDI treatments, seems to be adequate to achieve the best balance between oil production and quality.

Keywords: Olea europaea, regulated deficit irrigation, water stress, water productivity

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 85

Responses of olive varieties cultivated under different irrigation regimes

M. Aiachi‐Mezghani1, C. Masmoudi ‐Charfi C1

1.Institut de l’olivier, Unité expérimentale de Sousse. BP41‐4061 Ibn Khaldoun, Sousse, Tunisie.

The effects of different irrigation regimes 20%ETc, 50%ETc and 100%ETc.applied on five varieties cultivated in the southern Tunisia, was studied. Trees were six years old and planted with a density of 204 plants.ha‐1.Vegetative growth, yield, water use efficiency WUE and plant water status like leaf water potential Ψleaf, stomatal resistance Rs and conductance gs and relative leaf water contents RWC, were observed throughout the growing season, from March to December. Deficit treatments strongly reduced the vegetative growth. The reduction of water applied seemed to have any effect on the final shoot thickness. It has been observed that further increases in water quantities, did not systematically increased production. Maximum biannual productions were given by Picholine (23.0 kg tree−1) and Coratina cvs., (18.0 kg tree−1) irrigated −1 −1 at 100% ETc, by Manzanille (33.3 kg tree ) and Chetoui (27.1 kg tree ) cultivated at 20% ETc and −1 by Chemlali receiving an amount of 50% ETc (26.4 kg tree ). Measurement of Ψleaf didn’t reveal clearly the response of the tree to water regimes and may be considered as inappropriate to control water potential under different irrigation regimes. Maximum daily Leaf stomatal resistance was observed at midday and showed significant differences between treatments for cultivars Chétoui, Coratina and Chemlali. The most important variations were observed on Chemlali, which seemed to be the most able to regulate stomata aperture. It can be inferred that irrigation amounts varying from 20%ETc and 50%ETc can meet adequately effective water requirements for all varieties cultivated on this experimental field of Olive Institute (Southern Tunisia).

Key words: olive tree, southern Tunisia, intensive orchard, growth, yield, water plant status

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 86

Irrigation Management of Young Olive Trees in Kuwait

A. Al‐Shatti, H. Al‐Menaie, M. Al‐Hadidi, H. Mahgoub, O. Al‐Ragam, and M. Al‐Zalzalah

Kuwait Institute for Scientific Research, P.O. Box 24885, Safat 13109, Kuwait

Olive is a hardy tree that yields reasonably well even under water‐limited conditions, but it is also known from previous research that fruit and oil yields are positively correlated with amount of water applied up to a certain limit. The scarcity and high cost of water in Kuwait demands a careful determination of the optimal economic impact of any amount of water applied. The yield increase in several perennial crops is linearly related to both the level of transpiration and the amount of water supplied. Hence, the effectiveness and economic value of each cubic meter of water used, versus full irrigation systems, must be evaluated. Therefore, this study was carried out to determine the effects of different irrigation levels (50%, 75% or 100%) of estimated ETc on Sourani olive cultivar, and the application of mulching on plant growth under the Kuwait environmental conditions. The optimum irrigation level was determined based on vegetative growth. The results indicated positive effects of both irrigation level and mulching on plant growth. In fact, the best combination in terms of absolute tree growth was 100% ETc + mulch, however relatively high values were also obtained with 75% ETc and 50% ETc + mulch.

Keywords: Olive, irrigation, mulch

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 87

The effect of different irrigation strategies on olive oil (Memecik cv.) antioxidant content and activity

D. Sevim1, O. Koseoglu1, U. Kaya1, E. Akkuzu2, M. Parlak2, N. Pouyafard2

1Olive Research Station Izmir, Turkey 2Ege Universty, Faculty of Agriculture, Izmir, Turkey

In this study, the effects of irrigation levels on the antioxidant content and antioxidant activity of olive oils (cv. Memecik) were investigated in 2012 and 2013 crop seasons. Five different irrigation treatments were applied by drip irrigation. K1: non‐irrigated (rainfed), K2: completion of the diminished water in a depth of 0‐90cm soil to the field capacity for every 5 days, K3: application of the 33% of water given at K2, K4: 3 times application of the 50% of the diminished water in a depth of 0‐90cm soil concerning 3 growing stages; seed hardening, fruit growth and oil accumulation, K5: 3 times application of the 25% of the diminished water in a depth of 0‐90cm soil concerning 3 growing stages; seed hardening, fruit growth and oil accumulation. Olive oil was extracted using the Abencor Method. Total phenolic content, α‐tocopherol content, DPPH• (2,2‐ diphenyl‐1‐picrylhy‐drazyl) and ABTS•+ (2,2‐azino‐bis(3‐ethylbenzothiazoline)‐6‐sulfonic acid) radical scavenging activity (RSA) were determined. In 2012 crop season, the highest total phenolic content was determined at K4 treatments with •+ 115.03 mg CAE/kg oil, the highest α‐tocopherol content and ABTS RSA were identified at K1 and K5 treatments with 625.33 and 600.42 mg/kg and 157.97 and 155.68 µmolTE/100g, respectively. • According to the DPPH RSA results K5 and K4 treatments were the highest with 48.71 and 51.97 µmolTE/100g, respectively. In 2013 crop season, it was determined that K1, K5 and K4 treatmenst were include the highest total phenolic content with 278.35, 315.13 and 357.91 mg CAE/kg oil, • •+ respectively. According to α‐tocopherol content, DPPH and ABTS RSA results, K1 treatment was the highest level with 361.28 mg/kg, 109.17 and 124.67 µmolTE/100g, respectively. And also there was no difference determined at α‐tocopherol content and DPPH• RSA results between the K2, K3, K4 and K5 treatments. In 2012 crop seasons, K1 and K5 treatments showed the highest α‐ tocopherol content and ABTS•+ RSA. But in 2013 crop season, non‐irrigated (rainfed) samples showed the highest α‐tocopherol content, DPPH• and ABTS•+ RSA.

Keywords: Irrigation, deficit irrigation, water stress, DPPH• and ABTS•+ Radical Scavenging Activity, Total phenol, α‐tocopherol

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 88

Effect of deficit irrigation on olive tree performance, yield and oil quality

S. Naoum1, S. Ayoub1, I. Amayreh2, M. Diab1, M. Ammoush2, B. Kawaleet2, L. Daoud2

1National Center for Agricultural Research and Extension, Baqa, Jordan 2National Center for Agricultural Research and Extension, Al‐Mafraq Research Station, Jordan

This research experiment was conducted for two seasons (2012 and 2013) to investigate the effect of deficit irrigation treatments on olive tree performance, yield and oil quality for two local cultivars; Nabali Baladi and Nabali Muhasan. The selected orchard is located at Mugheir Alsarhan village in Al‐Mafraq governorate, north‐east of Jordan. The selected trees were 10 year‐old. Six irrigation treatments were applied; T1:80% ETc full season, T2: 80% ETc, and no irrigation from two months before harvest, T3: 80% ETc until July 1, then 50% ETc until 1 month before harvest stop irrigation, T4: 80% until June for the on year and 80% ETc until July 1, and 50% ETc until 1 month before harvest for the off year, T5: 40% until August 1, and 100% until October 1, stop irrigation for the on year and 80% ETc until July 1, 40% until October 1, stop irrigation for the off year and T6: 100% ETc for the whole season. Sub‐plots of 12 trees (3 rows X 4 trees) were assigned for each treatment, in which two inner trees in the center row were selected and labeled for taking samples and measurements. Crop water requirement, tree yield, crop water productivity, soil and plant analysis, and oil quality parameters were measured. Total applied irrigation water for Nabali Baladi was 2563, 1936, 1809, 2003, 1532, and 3204 m3/ha, for treatments T1, T2, T3 T4, T5 and T6, respectively, and for Nabali Muhasan it was 2838, 2146, 2011, 2225, 1701 and 3559 m3/ha for treatments T1, T2, T3 T4, T5 and T6, respectively. Harvesting was conducted in November. Results showed no significant differences for stem water potential between the treatments. However stem water potential had lower values in July. “Nabali Muhasan” showed less water stress than “Nabali Baladi” for all treatments. No significant effects of irrigation rates on the increase in tree trunk diameter and in shoot growth for two cultivars in both seasons. Results for the two seasons showed no significant differences between irrigation treatments in olive tree yield for Nabali Baladi and Nabali Muhasan cultivars. Average tree yield for Nabali Baladi was 11.97, 12.20, 15.15, 11.73, 7.43, and 9.01 and for Nabali Muhasan was 37.12, 33.50, 18.70, 35.37, 23.16 and 38.36 for treatments T1, T2, T3 T4, T5 and T6, respectively. Irrigation treatments showed no significant effect on fruit oil content and oil quality parameters. It can be concluded that application of deficit irrigation on olive trees had no negative effect on tree vegetative growth, yield and oil quality.

Keywords: deficit irrigation, Nabali Baladi, Nabali Muhasan, stem water potential, tree yield, oil quality.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 89

The timing of water deficit affects oil quality of cv. Frantoio

G. Caruso1, R. Gucci1, C. Gennai1, S. Esposto2, A. Taticchi2, S. Urbani2, M. Servili2

1Department of Agriculture, Food and Enviroment, University of Pisa, Via del Borghetto 80, 56124, Pisa, Italy. 2Department of Agricultural, Enviromental and Food Science, University of Perugia, Via San Costanzo s.n.c., 06126, Perugia, Italy.

Deficit irrigation is a sustainable practice that can be used in orchards for oil production. The timing, duration and intensity of water deficit distinguishes different deficit irrigation strategies. An experiment was carried out in a high‐density olive (Olea europaea L., cv. Frantoio) orchard in Tuscany over two consecutive years to compare the effect of two deficit irrigation regimes on yield components and virgin olive oil (VOO) quality. Subsurface drip irrigation was used to supply 100% (FI), 48‐79% (DI‐1) and 54‐69% (DI‐2) of tree water requirements. Fully irrigated trees (FI) received water during the entire irrigation period (from 23‐28 to 100‐103 days after full bloom, DAFB), whereas DI‐1 and DI‐2 trees were subjected to severe water deficit condition from 23‐28 to 41‐60 DAFB (DI‐1) and from 41‐60 to 71‐85 DAFB (DI‐2). DI‐1 and DI‐2 trees were fully irrigated during the rest of the irrigation period. Fruit yield of DI‐1 and DI‐2 was 70% and 81% of FI, respectively, whereas oil yield of respective deficit treatments resulted 66% and 77% of FI (average of two years). Yield was significantly affected by the year. No differences were found between irrigation treatments when fruit and oil yields were expressed on a trunk cross sectional area basis. In 2012 fruit fresh weight of FI trees was 134% and 123% that of DI‐1 and DI‐2 trees, respectively (110% and 99% in 2013). At the end of the first period of severe water deficit DI‐1 fruits had an higher phenolic concentration than FI fruits. Oils from all treatments were widely within the limits of VOO classification. Differences in UV absorption at 232 and 270 nm were found between FI and DI‐1 treatment in both years, whereas no differences in peroxide values and ∆K emerged between treatments. Over the 2‐year period DI‐1 trees yielded oils with higher concentrations of total phenols (136 % and 121%) and ortho‐diphenols (125 % and 128 %) than those obtained from either FI or DI‐2 trees, respectively. An early deficit (DI‐1) seems sufficient to induce higher phenolic concentrations in VOO.

Keywords: deficit irrigation, oil quality, yield, Olea europaea, polyphenols

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 90

Olive fruit growth indicators for regulated deficit irrigation scheduling

H. Memmi1 , M.C. Gijón1, H.F. Rapoport2, J.F.Couceiro1, D. Pérez‐López3

1 Centro Agrario “El Chaparrillo”, Servicio de Investigación Agraria de la Junta de Comunidades de Castilla‐La Mancha, Ctra de Porzuna km 3.5, 13071, Ciudad Real, Spain. 2 Instituto de Agricultura Sostenible, IAS‐CSIC, 14080 Cordoba, Spain 3 Dpto de Producción Vegetal: Fitotecnia. Technical University of Madrid, 28040, Madrid, Spain.

The correct application of Regulated deficit irrigation (RDI) requires the accurate knowledge of tolerant and critical periods for water stress implementation. For olive trees, RDI has produced good results when water shortage has been restricted to Stage II of fruit growth. The main reference physiological process which occurs during this stage is pit hardening. However measurement of hardening has been subjective, variable and often not even carried out, and opinions vary regarding whether the indicator is the onset, the full process or the end of pit hardening. Recently accurate quantitative measurement of the pit hardening process in olive has been achieved by fitting a machine to measure pit‐breaking pressure. In the current study our aim was to contrast the seasonal pattern of pit hardening with those of fresh and dry weights, longitudinal diameter and oil accumulation, in order to improve the efficiency of RDI application timing. For this purpose, two irrigation regimes were settled in a 16 years ‘Cornicabra’ olive trees orchard; a Control treatment supplied with its full water needs and a stressed treatment irrigated according to midday stem water potential thresholds. Water shortage was implemented during stage II of fruit growth. Each week, olives sample was taken from each treatment to measure the longitudinal diameter of fruits, fresh and dry weights, and pit hardening. For the determination of olives oil content pattern, dry fruit sample was conserved weekly for Nuclear Magnetic Resonance measurements. In contrast to current perceptions, oil accumulation seems to start a few days after the onset of sclerification rather after the end of massive pit hardening. That is, when pit breaking pressure reached its maximum, the olive fruit had accumulated approximately 25 to 40 % of final oil content, suggesting that although mild water deficit may be applied during the pit‐hardening period, a severe water stress could affect oil yield and should be avoided. Longitudinal diameter and pit hardening curves were closely correlated such that the change in the slope of longitudinal diameter growth could be a good indicator to mark the commencement of pit hardening and could be of an important use for RDI scheduling. Additionally results confirmed that olive fruit growth follows a simple sigmoidal pattern and that the double sigmoidal growth pattern mentioned in previous works could be related to water stress conditions.

Keywords: Olea europaea, RDI, growth pattern, pit hardening, oil accumulation

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 91

Effect of foliar amino acid “Pepton” application on growth, flowering, fruit set and oil quality of olive trees (Olea europaea L., cv. Chetoui) grown under intensive planting system.

Ajmi Larbi, Mohamed Ayadi1, Samira Yakoubi1 and Monji Msallem1

1Institut de l’Olivier , BP 286, Cité Mahrajène, Tunis‐ Tunisia

A 3‐year field study was carried out to determine the effects of foliar amino acid (Pepton 85/16, ACP Europe, Spain) application on vegetative growth, flowering, fruit set, fruit drop, yield, fruit oil content and oil quality of olive cv. Chetoui (5‐year‐old, 550 trees/ha). Amino acid foliar treatments (2.5 and 5.0 g l‐1of the commercial product Pepton) were applied at 4 different dates during 2011, 2012 and 2013. The dates correspond to prior to flowering, after fruit set and before fruit maturity. Foliar amino acid application had significant effect on vegetative growth during 3 years of experimentation mainly with the 5 gl‐1 dose. Vegetative growth was 14.3 and 48.1% higher than control trees when using 2.5g l‐1 during 2011 and 2012 while in 2013, no significant effect was observed. For the 5 gl‐1 of amino acid application, a significant increase was observed, being 47, 39 and 13.4% higher than control trees in 2011, 2012 and 2013, respectively. Amino acid foliar application had no significant effect on buds development, except during the third year of experimentation in which the percentage of buds developed into inflorescence increased significantly by 17 and 6% for the 2.5 and 5 g l‐1 doses, respectively when compared to control trees. Both Pepton treatments improved fruit set in 2011 and 2013, with the effect being more pronounced for the higher dose. Furthermore, fruit drop studied only by 2013 decreased by the application of Pepton amino‐acid mainly with the highest dose (5gl‐1). A slight increase of yield was observed after amino‐acid sprays in 2011 followed by an off year (2012) and thereafter the amino‐acid sprays resulted in a significant increase of yield. Therefore, the cumulative yield (2011‐2013) was 22 and 28.2% higher with 2.5 and 5 gl‐1 of amino‐acid spray as compared to control. No effect of amino‐acids foliar spray was observed in oil characteristics such as fatty acid composition and oil content. Thus, the response of olive to foliar Pepton application was significant on vegetative growth and yield but did not decrease alternate bearing characteristic.

Keywords: Amino acid, Olea europaea L, vegetative growth, yield, oil quality

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 92

The effect of foliar fertilization on carbohydrates status of 'Chemlali' olive (Olea europaea L.) leaves cultivated under rain‐fed conditions

I. Zouari1,2,, M. Aïachi‐Mezghani2, B. Mechri3, F. Labidi2, F.Attia4, D. Boujneh2, H. Chehab2, M.Hammami3

1Institut Supérieur Agronomique de Chott Mariem, B.P : 47, 2042 Chott Mariem, Sousse, Tunisia 2Laboratoire d’amélioration de la productivité de l’olivier et de la qualité des ses produits, Institut de l’Olivier, Unité Spécialisée de Sousse, Rue Ibn Khaldoun, B.P.: 14, 4061 Sousse, Tunisia 3Laboratoire de Biochimie, USCR Spectrométrie de Masse, LR‐NAFS /LR12ES05 « Nutrition –Aliments Fonctionnels et Santé Vasculaire », Faculté de Médecine, Université de Monastir, 5019 Monastir, Tunisie. 4Equipe de Recherches Agronomiques, Agronutrition, 3 Avenue de l’orchidée, Parc Activestre, Carbonne 31390, France

Olive tree yield in rain‐fed conditions depends on the adapted orchard management and the growing conditions. A nutrient supply is essential to enhance the product quality and the quantity. Foliar fertilization has the advantage of low application rates, uniform distribution and quick plant responses. The objective of this work is to determine the effect of different foliar treatments on the accumulation of the carbohydrates in olive tree leaves. The studied foliar applications were applied on Chemlali and regrouped into four treatments TC (control, without foliar nutrition), T1 (rich in nitrogen, applied on 18 of February before the vegetative start and 10 days later), T2 (rich in boron, magnesium, sulfur and manganese, applied on 25 of March, before the entry on the flowering and 10 days later) and finally T12 a combined treatment of T1 and T2. Leaves were sampled on two times; the first one was in the end of April (full bloom) and the second one in September (fruit growth). Sugar compounds were determined by gas chromatography. Results showed that fertilization caused changes in the pool of carbohydrates according to the treatments. Mannitol (alcohol sugar), glucose, fructose (monosaccharides) and sucrose (polysaccharides) were the main sugars found in the leaves. The fraction and the amount of these sugars changed between foliar treatments and the period of sampling. For the flowering time, the treatment T1 noted a significantly increasement in the glucose and the fructose concentration which was equal to 17.52 µg/mg dry weight (d.w) and 2.91 µg/mg d.w, respectively. However, the T2 treatment noted the highest concentration of mannitol (32.46 µg/mg d.w). During fruit growth (September), all carbohydrates decreased in all treatments which indicated an intense sugar mobilization on olive fruit.

Keywords: Olive tree, foliar fertilization, sugars, flowering, fruit growth

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 93

Development and validation of an irrigation scheduling tool for olives to improve water use efficiency at farm level

K. Chartzoulakis1, G. Psarras1, I. Kasapakis2, M. Bertaki1

1ELGO, Institute for Olive Tree and Subtropical Plants 73100 Chania, Crete, Greece 2KEPYEL, Department of Chania, 73100 Chania, Crete, Greece

The lack of advisory services for irrigation scheduling leads the farmers to irrigate empirically. They usually tend to “play safe”, increasing irrigation water amount, especially when water price is low. As a result about 20 % of water applied is lost, while at the same time tensions and conflicts with other sectors (urban, tourism) arise. This work presents the development and validation of software for irrigation scheduling of olives to be used by farmers or cooperatives. The software uses the climatic data, the soil characteristics, the crop data the water availability and quality and the method of irrigation to calculate the crop water requirements of the olives. The daily crop evapotranspiration (ETc) is calculated using the formula ETc = Kc x ETo, where ETo is reference Evapotraspiration and Kc is the crop coefficient (Kc) for each growth stage of olive, based on literature and experience in the area. The ETo is calculated using Penman‐Monteith, Hargreaves or class ‘A’ pan method, depending on the available climatic data. Basic elements of the software are the data bases in which all data required are stored. These include a) meteorological data, b) farmers’ data and c) data for each olive orchard of the farmer. After the development of databases and daily input of climatic data, the irrigation requirements are calculated by choosing the orchard and the last day of irrigation. The software is validated at field level for three successive the irrigation seasons (2010‐2012). The plant (physiological and water relations) and soil moisture data showed that the suggested irrigation doses by the software are sufficient for optimal olive growth.

Keywords: Olea europaea, irrigation, water requirements, water use efficiency, soil water potential

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 94

Arbuscular mycorrhizal fungus (Glomus deserticola) enhance drought tolerance of olive tree (Olea europaea)

W. Khabou1, T. Gargouri2, S. Kammoun2

1Improvement and Protection of olive tree germplasm, Olive tree Institute, 3029 Sfax, Tunisie 2High Horticulture School Chott Mariem, Sousse, Tunisie

In order to study the response of olive (Olea europaea L.) to water stress and to evaluate the effect of inoculation with mycorrhizal strain Glomus deserticola on the performance of the olive variety "Chemlali”, experimental test on 30 plants of this variety of two years old and obtained from semi‐hardwood cuttings, during 4 months. The plant material was grown in plastic containers with a capacity of 8 liters containing sand and irrigated with solutions of polyethylene glycol 4000 according to three water regimes with weekly irrigating (0% PEG, 2% and 4% PEG) applied on two batches of plants (mycorrhizal and non‐mycorrhizal). The study of eco‐physiological, biochemical and anatomical parameters, revealed that the response of the olive to face water stress occurs mainly by membrane damage (resulting in leakage of electrolytes and cell plasmolysis) and that a drop in chlorophyll pigments and phosphorus uptake which affects its growth in percentage of dry matter. However, adaptive strategies have been observed in all plants, such as reducing the relative water content, leaf area and stomatal density and an increase in the synthesis of proline and carotenoids. However, mycorrhizal plants were less affected by water deficit through a development of a far more important than non‐mycorrhizal plants root biomass. The root elongation (especially secondary) was ensured through the mycelial hyphae of Glomus deserticola which increased the surface prospected and therefore water and phosphorus uptake of the host plant. Thus, for all treatments studied, a more or less significant performance has been reported in seedlings inoculated with Glomus deserticola. These results highlight the importance of the production of mycorrhizal olive plants to alleviate water stress.

Keywords: Olea europaea, Glomus deserticola, drought, mineral uptake, proline, relative water content.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 95

Variability of the floral biology of four olive cultivars grown under two watering regimes in an arid region of Tunisia

M. Aiachi‐Mezghani 1, A. Diab1, A. Laroui 2, I. Zouari 2, I. Laaribi2, F. Labidi 1, L. Attia 1, M. Mars 2

1Institut de l’Olivier, station spécialisée de Sousse, BP 14, 4061, Ibn Khaldoun, Tunisia. 2Institut Supérieur Agronomique Chott Mariem Sousse 4042, Tunisia

Floral quality and effective pollination have significant influence on the fruit set, on the fruit number and finally, on the final crop of the olive orchard. These two processes are strongly influenced by environmental conditions and by varietal factor. The aim of the present work was study the effect of two watering regimes (rain‐fed and irrigated conditions) on flowering parameters, pollen viability, inflorescence architecture and fruit set. The trial was carried out in 2012 and 2013 in an olive grove located at the Research Station of Taoues, central Tunisia (34°N, 21°E). Four cultivars, Chemlali and Chétoui (local) and Picholine and Coratina (introduced) were noted. Thirty inflorescences were removed for each variety and watering regimes. Total and perfect flowers were identified in each inflorescence. The number of flowers per node ranged between 6.0 (Coratina) and 10.9 (Chétoui) and between 9.4 (Coratina) and 27.1 (Chemlali), for rain‐fed and irrigated conditions, respectively. The number of flowers varied in the inflorescence for different cultivars and water regimes. The maximum number of branch developed in the inflorescence, was six for rain‐fed conditions and seven for irrigated conditions. Flower position had an effect on gender. Perfect flowers were more frequent for terminal position than for lateral position. Perfect flowers were absent at sixth and seventh branch’s level of the inflorescence for cvs Chétoui and Coratina. However, Picholine and Chemlali showed perfect flowers at these levels. Pollen viability varied between 95 and 97%. Finally, the fruit set percentage was higher in irrigated conditions. The results indicated that floral biology and fruit set of the four cultivars were affected by water availability.

Keywords: Olea europaea L., flowering, inflorescence architecture, pollen viability, fruit set, irrigation

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 96

Efficacy of partial root zone drying application on olive tree in arid climate

L.Trablesi1, S.Maktouf1, M.Ghrab1, M.Khlifi1, N.Soua1, K.Gargouri 1

1 Olive Institute, University of Sfax, PO Box 1087, 3000 Sfax, Tunisia

The current situation in Tunisia is constrained by an increase in water demand from the agricultural sector. Scarcity of water available for irrigation is one of the most serious problems for agriculture in arid regions. Deficit irrigation system seems to be an adequate technique to improve water efficiency.Therefore, an irrigation experiment started in Sfax region (central Tunisia) to evaluate the long term effect of partial root zone drying (PRD) as a deficit irrigation management on olive trees. The experiment was carried out in an olive orchard of the Olive Tree Institute in Sfax in Tunisia during 2012‐2014. The region is characterized by an arid climate of Mediterranean type with mean annual precipitation and reference evapotranspiration ETo of 220 and 1390 mm respectively.Trees were planted on deep sandy soil (89.9% sand, 5% clay, 5.1% silt) with field capacity and wilting point of 12% and 6% respectively and irrigated with saline water (EC 6.73 mS cm−1) from ground water source. Water was delivered using two dripper with a dripper of 2.3 l h−1m−1. Four irrigaon treatments were applied. Control: Fully irrigated that applied daily the necessary water to match crop evapotranspiration ETc. PRD1: Alternate irrigation switched every 15 days, that supplied 50% of the irrigation water applied in the control. PRD2: Alternate irrigation switched every 30 days, that supplied 50% of the irrigation water applied in the control and Dry (rain‐fed): Olive trees were grown under rain‐fed conditions without irrigation supply.Ten trees were investigated per treatment. Yield components of the adult olive trees under the different treatments especially changes in endogenous sugars, relative water content (RWC) and mineral content (Na, P, K, Ca, Cl) were evaluated. Simultaneously, physicochemical characteristics of the different soils samples were analysed. Our results showed a significant difference in leaves sugars concentration varying from 29 to 106 (mg/g) respectively for FI and rainfed treatment. Moreover, high calcium concentration (3.4 %) for the PRD2 treatment was detected. Leaves phosphorus contents varied from 0.093% to 0.113% and potassium content was almost 1.03 % for all treatments. The RWC is among the important parameters of water status of the plant and it ranged between of 79% ‐ 81%. For soil analyses, we noticed a value of 1, 17% of organic matter for PRD1 and full irrigated systems versus 0,9% for the control. Similarly the control system showed the lowest electric conductivity (2,23 mS/cm) and the lowest humidity (2%) values.

Keywords: Olea europaea, PRD, soil, mineral nutrients, irrigation, soluble sugars

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 97

Physiological responses of young olive (Olea europaea L. cv Ayvalık) trees to water stress N. Pouyafard1, E. Akkuzu1,, U. Kaya2

1Ege University, Faculty of Agriculture, 35100, İzmir, Turkey 2Olive Research Station, Universite cad. 43,35100, Izmir, Turkey

Olive is a drought tolerant species that can survive under prolonged periods of drought by developing resistance mechanisms to stress. In this study, some phsiological response of young olive tree to drought stress under different irrigation levels and the relationships of some stress indicators with each other and with some atmospheric parameters were investigated. This research was carried out on two years old olive (cv Ayvalık) trees grown in pots (33x29 cm) at field condition in Bornova Olive Research Station, in İzmir in 2011. For this reason, the experiment consisted of 4 treatments: 1) Irrigated at the level of 100% of the ETa (I100), 2) Irrigated at the level of 66% of the ETa (I66), 3) Irrigated at the level of 33% of the ETa (I33), 4) Non‐irrigated (I0). All pots were manually irrigated once every four days during the irrigation season. The pots were weighed just before irrigation in the morning to calculate evapotranspiration.Some physiologic responses of plants such as stomatal conductance (gs), leaf temperature and air temperature difference (Tl‐Ta), leaf water potential (LWP) and leaf relative water content (RWC) were investigated. In this study, irrigation water requirement was ranged from 0 to 82.45 liters/pot and actual evapotranspiration was ranged from 86.11 to 10.78 liters/plant. The leaf water potential and leaf relative water content values statistically significant differences were found among the treatments. On the other hand belong to treatment I100, Tl‐Ta with conductivity of the stoma with a polynomial relationship between them as a result of regression analysis (R2= 0.71) was obtained. The result of regression analysis between VPD with Tl‐Ta was a linear relationship (R2 = 0.74), and the result of regression analysis between VPD and stomatal conductivity was founded as a polynomial relationship with R2 = 0.54. Ayvalik olive, is a kind of drought tolerant plant, recommended for growing in arid or inadequate water supplied areas. However, as a proposal determining the level of water stress in olive plants leaf water potential, leaf relative water content can be used.

Keywords: Olive, water stress, Leaf Water Potential, Leaf Relative Water Content, Stomatal Conductance, Vapour Pressure Deficit, Ayvalık

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 98

Effect of deficit irrigation strategies on canopy temperature and leaf water potential in olive (Memecik Cv.)

U.Kaya1, M.Parlak2 , N.Pouyafard 2, T.Cakır 2, M. Gurbuz1, G.P.Mengu2, E.Akkuzu2

1Olive Research Station, Universite cad. 43 Bornova 35100, Izmir, Turkey 2Ege University, Faculty of Agriculture, Bornova, 35100 Izmir, Turkey

Plants respond to drought by regulating the exchange of CO2 and H2O between their leaves and the air using their stomata. Decrease in turgor status of the leaves, concomitantly occurring with the reduced leaf water potential and transpiration, causes an increase in the canopy temperature. In this study, the response of Memecik cv., which has been commonly grown cultivar in Turkey, to water stress and the air vapor pressure deficit was determined by monitoring the difference between canopy and air temperature (Tc‐ Ta) during the irrigation season. For this purpose, a trial was designed in the olive orchard of Olive Research Station with the Memecik cv. which was 25 years of age. The trial was conducted in randomized block design with 5 treatments and 3 replicates and was repeated for 2 years (2012‐2013). Water treatments; K1: non‐irrigated (rainfed), K2: completion of the diminished water in a depth of 0‐90cm soil to the field capacity for every 5 days, K3: application of the 33% of water given at K2, K4: 3 times application of the 50% of the diminished water in a depth of 0‐90cm soil concerning 3 growing stages; seed hardening, fruit growth and oil accumulation, K5: 3 times application of the 25% of the diminished water in a depth of 0‐90cm soil concerning 3 growing stages; seed hardening, fruit growth and oil accumulation. In the study, leaf water potential measurements were carried out at two‐week intervals in predawn using pressure chamber device. Following that, canopy temperatures were measured between 11.30‐13.00 in the same day in order to monitor the Tc‐ Ta difference using infrared thermometer. In order to determine the relationship between Tc‐Ta and vapor pressure deficit, wet and dry bulp temperature were measured and discussed in the study.

Keywords: Olive, deficit irrigation, leaf water potential, canopy temperature, Memecik cv.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 99

Water relations and gas exchanges of olive trees (cv. chetoui) in response to different deficit Irrigation regimes

S. Dbara1, K. Lahmar2, M. Ben Mimoun3

1Centre Régional des Recherches en Horticulture et Agriculture Biologique à Chott Mariem ; BP57, Sousse 4042, Tunisie 2Université de Monastir, Rue Salem Bachir, BP56, Monastir 5000, Tunisie 3Laboratoire d’arboriculture fruitière, Institut National Agronomique de Tunisie, 43 Avenue Charles Nicolle, 1082, Tunis Mahrajène, Tunisie

The increase of water demand for irrigation in olive orchards is leading to adopt deficit irrigation approaches. The present study aims to compare two different regimes: regulated deficit irrigation (RDI) and partial root drying (PRD) applied on olive orchard (Olea europaea. L, cv Chetoui). Field experiment was carried out on trees grown under semi‐arid climate of Tunisia. Five treatments were applied: Rainfed (without irrigation), Control (irrigated with 100% of crop evapotranspiration on the two sides of trees), RDI (irrigated with 50% of the control on both sides of trees), PRD100 and PRD50 (irrigated respectively with 100% and 50% of the control on one side, alternating the wetted and dried sides of the root system every ten days). Results showed significant difference between treatments. In fact, the relative water content of leaves and fruits was slightly decreased by the decrease of water supply however the net carbon assimilation was less affected. So the improvement of water use efficiency was noted seen that water relations were less affected than gas exchanges. The PRD treatments affected significantly olive water relations, starting with a decrease in stomatal conductance, and subsequently an increase of stem water potential (ψ), with a small non‐significant effect on leaf relative water content. Also, the implication of non hydraulic limitation was highlighted. The leaf proline content increased in the most stressed treatments (Rainfed, PRD50 and RDI).

Key words: Olive, water relation, gas exchange, efficiency

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 100

Ecophysiological response of young olive plants under three irrigation regimes.

M. Gouiaa, F. Zaouay, D. Boujnah

1IRESA‐Université de Sousse, Institut Supérieur Agronomique de Chott‐Mariem, U.R. Agrobiodiversité, Chott Mariem, 4042, Sousse, Tunisie. 2 Institut de l’Olivier. Route de l’aéroport, km 1,5. BP 1087. TN‐3000, Sfax, Tunisie.

In order to study the behavior of olive cv. Chemlali under drought stress, irrigation intervals of 6 days was studied on potted young olive trees. Three irrigation treatments (T1: full irrigation, three liters per week; T0.5 50% irrigation, 1.5 liters per week and T0: zero irrigation, as control treatment) were applied. Stomatal conductance, net photosynthesis rate and leaf water potential were measured over hours on consecutive days. Anatomical observations were also made on half, one and two‐years‐old stems to determine the hydraulic diameter, the theoretical hydraulic conductivity level and the lumen area of vessels. The results showed that the highest stomatal ‐2 ‐1 conductance (680 mmol m s ) was observed with T1 treatment. In the unirrigated treatment, the drought decreased stomatal conductance from 54.26 to 28.08 mmol.m‐2 s‐1 and leaf water potential from −15.6 to −34.8 bars over hours (from the morning to the aernoon). Stressed young plants showed also the lowest net photosynthesis rate (3.23 μmol m‐2 s‐1) compared to other treatments. Similar response to unirrigated treatment (T0) was observed in young plant shoots for the theoretical hydraulic conductivity level and the size of vessels. The vessel lumen area decreased as the day temperature increased mainly for the stressed young trees. It was also revealed that for all treatments the hydraulic conductivity is high while the vessel diameter is large. The increase in the theoretical hydraulic conductivity depends on the substrate water content. Although the olive tree is characterized by its hardiness, young plants could not resist to water stress and they eventually wilt. This information about ecophysiological approach contributes to the knowledge about the most efficient irrigation strategy of olive orchard at their young age under drought‐related hydraulic impairments induced by the environment.

Keywords: Olea europaea, water stress, stomatal conductance, net photosynthesis, lumen area, water absorption.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 101

Rainfed olive production as influenced by low quantity of applied water during the first stage of fruit growth

R. Razouk1, A. Kajji1, M. Karrou2

1National Institute for Agricultural Research, Meknes, Morocco. 2Integrated Water and Land Management Program, International Center for Agricultural Research in the Dry Areas, Allepo, Syria.

Irrigation of rainfed olive trees using a low amount of water may have an important production gain if it is applied during critical stages of olive growth. However, adoption of this technique requires hydro‐agricultural investments that must be economically efficient. For this reason, it is essential to determine the optimal periods of irrigation water application. Within this context, an experiment was carried out over two years to evaluate the effects of low quantity of supplemental irrigation (0.5 m3/tree, equivalent to 2/3 of soil water reserve on 70 cm of depth) applied during one day at three stages of fruit growth: beginning of stage I (June 15); beginning of stage II (July 25); beginning of stage III (September 5) and no irrigation used as control. Treatments were applied to thirty years old olive trees (cv. Picholine Marocaine) grown under rainfed conditions, planted at a distance of 8x6 m in the experimental field of Regional Agricultural Research Center of Meknes in Northern Morocco. Results showed that fruit weight and shoot length were more affected by irrigation applied during the first period of fruit growth (stage I) as compared to rainfed regime, and hence fruit and oil yields were increased, in average, respectively by 43% and 37%. Water use efficiency for this irrigation was 23 kg/m3/tree. The two irrigations applied during stages II and III of fruit growth improved only oil content and shoot length without significant effect on yield. According to the obtained results, we recommend to undertake at least one major irrigation of rainfed olive orchards during the early phase of fruit growth (June for Morocco). The obtained production gain under this irrigation is promising to promote hydro‐agricultural investments in several rainfed olive‐growing areas.

Keywords: Morocco, rainfed olive, supplemental irrigation, vegetative growth, fruit yield, oil yield

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 102

Contribution of the study of the behavior of the olive tree (Olea europaea L.) under different water regimes

H. Benjeddou1, B. Ben Rouina2

1 Laboratory of Biology and Plant Ecophysiology of Arid Area, Faculty of Sciences of Sfax, Tunisia 2Laboratory of Improvement of Oleo Culture productivity and Fruit Trees, Olive tree Institue of Sfax, Tunisia

Mediterranean type climate is characterized by high temperatures, low precipitation, high irradiance levels and vapor pressure deficits during the growing season. As a consequence, crops are often subjected to severe water stress. In this context, three‐year‐old olive trees (Olea europaea L. cv. Chemlali) were transplanted into 15‐L pots filled with sand and peat and subjected to three irrigation treatments during four months, under ambient environmental conditions: control plants (T1) were irrigated with 100% of their needs calculated from the amount of water lost by transpiration, the two other treatments were subjected to gradual water depletion that received only 66% and 33% of their needs, respectively. The main objective of this work was to investigate the adaptive mechanisms used by the olive trees in response to drought stress. To this aim we carried out measurements of photosynthetic performance (net photosynthesis (Pn), stomatal conductance (Gs) and transpiration rates (E), relative water content (RWC), soluble sugars and proline. During drought periods, plants undergo many physiological and biochemical changes. It is obvious that, with the increase of stress intensity leaves exhibit large reduction in relative water content with significant decrease in photosynthetic activity. In fact, T3 plants exhibit decreases of 81%, 72% and 55% (in Pn, Gs and E respectively) as compared to decreases of 67%, 60% and 44% observed in T2. These decreases mainly due to stomatal closure which is one of the very early responses to water deficit. Furthermore, the biochemical analysis revealed a significant increase in proline and soluble sugars contents. High concentrations of these compatibles solutes contribute to the lowering of osmotic potential and allow water to move into cells, thereby maintaining turgor and increasing tissue tolerance to low soil water potential.

Keywords: Olive tree, Chemlali, water stress, adaptive mechanisms

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 103

Application of exogenous organic matter promote soil fertility and carbon sequestration

Rayda Chaker1 , Noha Sallemi1 , Sameh Maktouf1 , Nabil Soua1 , Mouna Khilifi1 , kamel Gargouri1

1Olive institue, University of Sfax , Sfax , Tunisia

The effects of application of five organic amendments compost of manure and olive pomace (C), a palm leaf based compost (CP), a green manure derived from olive pruning waste (GW), an olive mill wastewater (OMW) and ovine manure (OM) on the rate of mineralization, sequestration of Carbon in soil as well as on the microbial biomass content and level of selected enzymatic activities were studied. A single dose of 39 mg of organic carbon/ 100 g of soil was tested. These soils in addition with the control soil were incubated at 28 C° and a 12% of humidity for 56 days. Soil respiration, organic C and microbial biomass were measured. Soil respiration increased in the amended soils on the first week of incubation the highest values of all studied parameters were found when OM and GM were applied. Increases in CO2 emission resulting from OM and GW amendments were strongly associated with microbial biomass content. The highest number of fungi was observed in the OM amendment while the number of bacteria was most important when GM was applied. Organic carbon and microbial biomass (fungi and bacteria) were also increased in amended soils compared to the control. Most of the parameters studied were found to be highly correlated, indicating a balanced enhancement of soil biological activity after the application of organic residues. In conclusion, organic amendment can be a technique that sequesters carbon in soils that enhanced carbon storage in soils.

Keywords: organic amendments, enzymatic activities, microbial biomass, Soil respiration

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 104

Effects of soil management practices on Particulate Organic Matter (POM) fraction involved in soil fertility of rainfed olive orchards

A.M. Palese, A. Arous, C. Xiloyannis, G. Celano

Dipartimento delle Culture Europee e del Mediterraneo: Architettura, Ambiente, Patrimoni Culturali (DiCEM) – Università degli Studi della Basilicata, via San Rocco, 3 – 75100 Matera (Italy)

Soil Organic Matter (SOM) plays a key role in maintaining soil fertility and, consequently, in assuring plant productivity. Soil management techniques can strongly affect SOM content. Particularly, tillage has detrimental effects being a non‐conservative practice of SOM. Particulate Organic Matter (POM) is the youngest SOM fraction and a sensitive indicator to changes induced by soil management practices. For its characteristics, it is a microbially active fraction, readily decomposable and thus it can contribute to soil fertility through nutrient cycling. The aim of this research was to evaluate the medium‐term effect on POM content of soil sampled within olive orchards managed according to different soil management systems (spontaneous vegetation cover + pruning material provision versus tillage). The trial was carried out in a rainfed olive orchard located in Southern Italy whose soil was covered since 2000 by spontaneous vegetation and by pruning material cut up in loco (Sustainable System – SS). Soil sampling was carried out in November 2013 at 0‐5 and 5‐10 cm depths from the inter‐row positions. Soil was also sampled from a tilled olive orchard which was taken as control (Conventional System – CS). POM fraction (> 53 μm), was physically separated from SOM on the basis of particle size. Total organic carbon (TOC) and the mineral‐associated organic carbon (MOC) were analyzed. POM carbon (POM‐C) was calculated as the difference between TOC and MOC. TOC was significantly higher in the SS than in the CS especially in the shallow layer (0‐5 cm) (p < 0.05). Under our experimental condition, TOC was effective in showing changes in soil after 14 years since the beginning of the sustainable soil management. Similarly, differences in POM‐C content between soil management systems were significant (p < 0.05) being more than the double in the SS than in the CS (13.3 g kg‐1 versus 6.3 g kg‐1 in the 0‐10 cm layer). The ratio of POM‐C to TOC was 61.3 and 45.0% (mean 0‐10 cm) in the SS and CS, respectively. This indicated a more rapid decomposition in the CS probably due to the soil disturbance and other soil conditions such as temperature, moisture, substrate type and availability, all affecting decomposition rates. According to our results, the sustainable soil management improved SOM and POM concentration of rainfed olive orchards. The use of such techniques, which also lead to others agronomical and environmental advantages, should be strongly encouraged with opportune strategic policies.

Key words: Olea europaea, residue handling, SOM fractions

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 105

Effect of nitrogen and phosphorus fertilizer on the growth of young olive trees in Kuwait

H. Al‐Menaie, O. Al‐Ragam, H. Mahgoub, M. Al‐Hadidi, A. Al‐Shatti and M. Al‐Zalzalah

Kuwait Institute for Scientific Research, PO Box 24885, Safat 13109, Kuwait

An appropriate fertilization is important during the first years of olive growth when the tree has to take in nutrients and produce assimilates for the development of its root system and canopy perennial structures and to prepare itself for future fruit production. Nitrogen and phosphorus which are essential nutrients in olive orchards play an important role in plant growth, and are usually required in greater amounts than other minerals. Knowledge regarding the optimum nutritional requirements is important to ensure a balanced development of the plant and to avoid surplus administration of fertilizers, which could also damage the environment. Hence, this study was initiated by Kuwait Institute for Scientific Research to determine the growth response of olive to different nitrogen and phosphorus concentrations in order to establish suitable fertilization program for olive young trees under Kuwaiti growing conditions. One‐year old acclimatized plants of the Sourani variety were subjected to three levels of nitrogen (100, 150, 200 g N/ tree), and phosphorus (0, 80,160 g P2O5/ tree). Results showed significant effects of the treatments, the combination N2P2, corresponding to 150g/tree of N, and 80 g/tree of P was found to be the best for all studied parameters.

Keywords: Olive, Nitrogen fertilizer, Sourani

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 106

Nutritional status of olive orchards in rainfed and irrigated area of Tensift‐Haouz region in Morocco

H. Boulal1, L. Sikaoui2, A. Bouizgaren2, M. El Gharous1

1International Plant Nutrition Institute (IPNI) North Africa, INRA BP. 589 Settat, Morocco 2INRA Marrakech, BP. 533 Menara Marrakech, Morocco

Olive (Olea europaea L.) is one of the most important tree crops in Morocco. It represents 60% of the total cultivated area with tree crops. Despite the importance of the olive and the significant government support, olive yields are low with an average fruit production of 1.7 tons per hectare. One of major constraints of olive orchards is nutrient deficiencies and imbalance in some nutrients. Therefore, the nutritional status of olive trees is among the most important factors which affecting the yield. Hence, a study was conducted in two sites in Tensift‐Haouz region of Morocco in olive orchards under irrigated and rainfed areas respectively. The nutritional diagnosis was made on 15 olive orchards in rainfed area and on 19 olive orchards in irrigated area in Essaouira and Marrakech provinces respectively. In summer 2013 leaves were sampled from all sites for foliar analysis (N, P, K, Ca, Na, Mg, Fe, Cu, Mn, Zn, Ni). Leaves were picked from the current year’s new growth shoots which are not bearing fruit, lower and apical leaves were discarded. Results of foliar analysis showed that leaf nutrient concentrations were less than nutrient critical values of olive orchards in both regions in the case of nitrogen, phosphorus and potassium. Concerning micronutrient concentration of copper represents the most deficient micronutrients. Those results are an indicator that nutrient management still one of the main constraint to manage by farmers for improving olive production in Morocco.

Key words: Olea europaea, nutritional diagnosis, foliar analysis, Morocco

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 107

Olive tree nutritional status survey following a non‐typical dry growing season in the area of Chania, Crete, Greece

G. Psarras, A. Digalaki, M. Darioti, M. Bertaki

ELGO Demeter, Institute for Olive Tree and Subtropical Plants of Chania, Agrokipio, 73100, Chania, Greece

Soil water availability is a crucial factor affecting nutrient uptake and is more important in rainfed orchards, since it cannot be under human control. Recently, due to the economic crisis, several Cretan farmers do not irrigate their olive orchards in non‐cropping years, a practice that may negatively affect the nutritional status of the trees. A nutritional survey (leaf and soil analyses) was carried out in 61 commercial olive orchards located in the Municipality of Platanias at Chania Prefecture in Crete, with sampling between November and December 2013. The preceeding growing season was characterized by a very low crop, as well as by a long dry period (56 mm of rainfall between February 15th and October 31st). About half of the orchards in the survey were rainfed, while both ‘Koroneiki’ and ‘Mastoides’ orchards (the dominant cvs in the area) were included. The results highlighted the fact that soil analysis is not the best indicator for olive tree nutritional status, as compared to leaf analysis, especially in the case of micronutrients. Potassium and B were the nutrients that were more frequently deficient in the leaves, at considerably higher percentages, as compared to previous surveys in the area (deficient orchards: 51% for K and 87% for B). The extended dry period resulted in higher frequencies of N and K deficiency in rainfed orchards (27 and 58% for N and 41 and 65% for K in irrigated and rainfed orchards respectively). Under rainfed conditions, the majority (74%) of the orchards with deficient K levels in the leaves, corresponded to adequate or high levels of the element in the soil, a fact that could be attributed to the unfavorable soil moisture conditions. Phosphorus deficiency was not a significant problem in the area, with low levels appearing in only 6% of the orchards. In contrast, excess fertilizing with P was common, with 46% of the orchards showing high or excess levels of P in the leaves. This resulted in cases of unbalanced nutrition, with Zn being the most evident example. Low levels of Zn were observed in 26% of the orchards, but in 81% of these cases the deficient Zn levels were linked with excess P levels in the leaves.

Keywords: Olea europaea, mineral nutrition, leaf analysis, soil analysis

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 108

Role of indigenous weed flora in carbon and main nutrient elements storage in olive orchards in Southern Greece

E. Markakis1, N. Kourgialas1, C. Sergentani1, V. Borraccia1,2, L. Mikalef1, N. Digalaki1, S. Reppas3, A. Aggelaki4, C. Manolaraki5, S. Malliaraki4, G. Giakoumaki5, K. Giannopolitis6, N. Kavroulakis1, G. Psarras1, G. Koubouris1

1ELGO Demeter, Inst. for Olive Tree & Subtropical Plants, 73100, Chania, Greece 2University of Basilicata, Potenza, Italy 3FG “Nileas”, Chora Messinias, Greece 4UAC Meramvello, Neapoli, Crete, Greece 5UAC Peza, Peza, Crete, Greece 6AgroTypos SA, Maroussi, Greece

The carbon footprint is a recent term that has been introduced as a way to characterize if a product has been produced in an environmental friendly way, regarding the global warming issue. The first step in managing the carbon footprint of olive oil, as an agricultural product, at farm level is to have an estimate of how the applied agricultural practices contribute in the carbon balance of the orchard. Within the framework of the LIFE+ project oLIVECLIMA, several alternative orchard management practices are applied to commercial olive orchards, as a way to cope with the upcoming climate changes and improve the carbon balance within the orchard ecosystem. Among others, applied practices include the conservation of the local weed population during winter (as compared to farmer practice of soil cultivation or chemical weed control) and intervention for alteration of the indigenous flora in order to achieve a more efficient carbon and nutrient fixation. In the present work, preliminary data on the carbon and nutrient fixation in the indigenous orchard flora are reported. Three representative sampling areas per orchard were established in 70 commercial olive orchards in the areas of Nileas in Peloponnese, and Meramvello and Peza in Crete. The percentage of surface coverage by weeds and the dominant weed species within each sampling area were recorded, while carbon and mineral nutrient content in the above‐ground weed mass was determined. The results indicate that carbon fixation was significantly variable among the different areas and averaged 0.92, 0.55 and 0.80 tn of C per ha for Nileas, Meramvello and Peza respectively. This variation was in accordance to variation in annual precipitation among the three areas. A significant amount of readily available mineral nutrients was also removed from the soil into the weed above ground tissues. This was estimated as 38, 16 and 28 kg.ha‐1 for N, 8, 3 and 5 kg.ha‐1 for P and 56, 22 and 39 kg.ha‐ 1 for K respectively for Nileas, Meramvello and Peza. The following aim of the 5‐year project is to introduce an appropriate mix of plant species as winter cover crop in each area, as a way to achieve a more efficient carbon and nutrient fixation within each orchard.

Keywords: Olea europaea, mineral nutrients, carbon, weeds

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 109

Particulates fallout in the southern Sector of the city of Sfax: monitoring and impact on the soil and plants. The olive tree as an example

C. Mbadra1, M. Bahloul2, K.Gargouri3, A. Chafai2, F. Ben Abdallah1

1Biology Department, Faculty of Sciences, Sfax (Tunisia) 2Geology Department, Faculty of Sciences, Sfax (Tunisa) 3Olive Tree Institute, Sfax (Tunisia)

The spatio‐temporal variation of particulate fallout in the southern sector of the city of Sfax (Tunisia) and its impact on neighboring soils and plants was studied using the technique of platelets DIEMs. The study of the temporal evolution of particulate deposits above 5 selected in the southern sector of the city of Sfax site showed high levels that can acheived 25 g / m² / 15 day. The simulation of particulate fallout on the ground under the axis industrial plumes in two extremely different conditions of stability (very stable and unstable) shows that our study area is threatened mainly by emissions of soap (SIOSZETEX) and smelting lead.. In addition, the maximum particulate fallout occurred mainly in very unstable atmospheric conditions due to the effect of the winding and smoke phenomenon. Chemical analysis of the collected particles showed a progressive enrichment in Ca 2+, K + and Cl‐ . Consequently, a neutral to alkaline pH and an increasing electrical conductivity were observed while away from the polluting area. The enrichment of metalliferous dust was also marked. The results also showed that the Cd contents of the studied soils exceed the standards set at 0.68 mg / kg MS, whereas Pb remained below the standards set at 30.2 mg / kg. The use of numerical approaches (Rate enrichment, enrichment factor) showed the intervention of SIAPE and lead smelter in the metalliferous contamination of surrounding soils. Especially, the impact of metals was pronounced in various vegetative species (Nerium oleander, Prunus amygdalus, Olea europea mainly). The leaves of the olive tree exposed to industrial plumes are covered with a film of whitish dust. It has been noted among species in the most polluted area (500 meters from the source), apical and marginal leaf necrosis that generally do not exceed the upper third of the leaf. The border between the healthy part of the necrotic part is fine. The exposed sheets are covered with a film of dust whitish. In addition to their persistence, the leaves of the olive tree are characterized by a low leaf area for their resistance to environmental stress.

Keywords: particulate, southern sector, soil, plants, pollution

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 110

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 111

PEST AND DISEASE CONTROL

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 112

Integrated Pest Management of olive tree in Tunisia

M. Ksantini

Olive Tree Institute‐ BP 1087‐ 3000 Sfax – Tunisia

In Tunisia, the olive and oil sector plays an important socioeconomic role since its vast distribution (1670 000 ha) and the high proportion of oil production reserved for export (65%). However, because of the severe climatic conditions, the productivity of olive orchards is low and must be improved especially at the levels of production costs and the oil quality in order to increase the competitivity of Tunisian olive oil. In this way, among the factors influencing the oil quality, emphasis must be focused on the control of olive pests. Several insect pests infest olives in Tunisia. Olive fruit fly (Bactrocera oleae), the olive moth (Prays oleae) and the olive psylla (Euphyllura olivina) are the most important pest in cost area. Other insects: olive bark borer (Phleotribus scarabeoides and Hylesinus oleiperda), olive black scale (Saissetia oleae) and diaspine scales (Aspidiotis nerii and Parlatoria oleae) … cause serious economical damage some years. The research developed since more than twenty years permitted to establish the basis of IPM strategy which was applied at large scale in the beginning of the years 1990 through national campaign of control. On the basis of data collected for several years, the present note try to synthesize the main results obtained particularly those related to the status of harmful insect pests, their geographic distribution and sources, population dynamics and the control strategy adopted. In addition, it points out the economical and ecological impact of the strategy applied and brings out the large possibilities to promote organic farming on olive growing.

Keywords: olive pests, integrated pests control, economical and ecological impact, organic farming.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 113

Effective, environmentally‐sensitive pest management in olive production

V. Sergeeva

Centre for Plants and the Environment, UWS, Penrith, NSW 1797, Australia

In recent years increasing global interest in health and the environment has stimulated studies of alternatives to the usual procedures for controlling pests and diseases. As a result, management decisions now take into account the impact of control methods on native flora and fauna. Control of olive pests continues to attract interest and requires a high level of attention. Pest control is fundamental to sustainable olive crop production, in terms of both quality and quantity, however the protection of the environment is now factored into the equation. In particular, diseases and pests associated with environmental stress and unfavourable climatic conditions impact greatly on sustainable olive production and olive oil quality. They must be controlled, while also ensuring the exclusion of chemical residues in table olives and olive oil, a central aspect of product quality and food safety for the industry. Integrated Pest Management (IPM) is an effective and environmentally sensitive approach to pest management that relies on a combination of common‐sense practices, and has proven a viable answer to both of these needs. The regime includes cultural agronomic practices for maintaining tree health through proper cultivation techniques, irrigation, pruning and soil health. It also utilizes natural products as an excellent alternative to synthetic pesticides, reducing negative impacts on both human health and the environment. These increasingly‐common biological products are in general far less expensive than their synthetic counterparts and provide safe, eco‐friendly yet highly effective pest control solutions. Combined with successful control and management using beneficial insects, ecological insect traps, pheromone traps, sterile insect introduction, soil solarization, these ‘green’ pesticides contribute to reduce pest populations and increase olive food production. Timing is also an important consideration in IPM. Outbreaks often occur during the abundant olive tree flowering season, with significant damage from high pest populations coinciding with the development of olive inflorescence. Timely action in spring can therefore prevent a pest outbreak from getting out of control and stop the cycle of infestation in its early stages. IPM proves that by using our knowledge of the life cycles of pests and their interaction with the environment, in combination with available pest control methods, we can manage pest damage by the most economical means and with the least possible risk to people, property and the environment. The result is a sustainable olive farming system which not only protects the environment it operates in but also produces high quality, safe table olives and oil at a viable cost.

Keywords: Olea europaea, environment, pest management.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 114

Evaluation of olive fly Bactrocera oleae oviposition preference on Cobrançosa, Madural and Verdeal Transmontana cultivars in Portugal

R. Malheiro1,2, S. Casal2, L. Pinheiro1, P. Baptista1, A. Bento1, J. A. Pereira1

1Mountain Research Centre (CIMO), School of Agriculture, Polytechnic Institute of Bragança, Campus Stª Apolónia, Apartado 1172, 5301‐855, Bragança, Portugal 2REQUIMTE/Laboratory of Bromatology and Hydrology, Faculty of Pharmacy, Porto University, Rua Aníbal Cunha 164, 4050‐047 Porto, Portugal

The objective of this work was to assess the oviposition preference of B. oleae towards cvs. Cobrançosa, Madural, and Verdeal Transmontana. Infestation levels were monitored from olives of the three varieties collected during crop ripening. Bioassays were run in vitro to assess oviposition preference. Two different approaches were tested: i) three‐choice assays (20 fruits of each variety exposed simultaneously to 5 independent populations of B. oleae composed by 10 males and 10 females with 15 days old, during 10 consecutive days; each exposure of 24 hours); ii) one‐choice assays (60 fruits of each variety exposed according to the conditions described in three‐choice assay). Olives maturation index, number of stings per fruit, and pupae emergence percentage were recorded. Three choice‐assays clearly showed that the number of fruits attacked was similar in the three varieties, however the number of stings were significantly higher in cv. Verdeal Transmontana (1319) comparatively to the other two varieties (991 and 1096 for Madural and Cobrançosa varieties respectively). Verdeal Transmontana cultivar is attacked sooner than other varieties. Stings per fruit were higher in cv. Verdeal Transmontana (2.25) than cv. Cobrançosa (1.98) and cv. Madural (1.82). Pupae emergence was higher in cv. Madural, maybe due to the lower rate of stings per fruit, being easier the development of larvae in these fruits. Results obtained in three‐choice assays were corroborated by those obtained in one‐choice assay. Verdeal Transmontana reported 2.30 stings per fruit while cv. Madural reported 1.78 and cv. Cobrançosa 1.54. Bioassays showed preference of B. oleae towards cv. Verdeal Transmontana, also in accordance to the data obtained in the field. Therefore the study of the interaction of B. oleae with the different olive varieties is crucial to improve and develop efficient control strategies.

Keywords: Olea europaea, Bactrocera oleae, olive variety, oviposition preference; bioassays

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 115

New insect pest species recently established on olives groves in Marrakech ‐ Tensift – Al Haouz region, Morocco

Y. Ouguas1, M . Karama 1, I. Zafzafi 2 , T. Bamhamed 2

1Institut National de la Recherche Agronomique, Marrakech, Morocco 2Faculté des Sciences et Techniques Marrakech, Morocco

Climate change has a major impact on insects, diseases and host plants. Until now, most studies have focused on the effects of these changes on forest and agricultural pests. Indeed, a rise in temperature may influence the winter survival, the number of generations, the early arrival of pests, or the appearance of pests known as being of secondary importance. This was the case of pests encountered in the region of Marrakech‐Tensift ‐ Al Haouz. In spring 2013, we witnessed the establishment of insect pests on young olive groves. Xylomedes coronata ‐ branch and twig borer ‐ (Coleoptera, Bostrychidae) attacks young twigs causing breakage of these and their subsequent fall. In Morocco, this pest has been observed for the first time in 1930 on the vine and plum. This species also attacks olive trees; it causes serious damage in Kabylia (Algeria) since 1995. In Morocco, this pest has been reported on citrus trees in 1964. Moreover, a very large amount of caterpillars of Acherontia atropos (Lepidoptera, Sphingidae) commonly called Death's Head Hawk‐moth was picked up by growers in the same region during October and November of the same year, 2013. This insect feeds on leaves of young olive trees, 3 to 6 years old, and causes drying of olive trees. Also, in an orchard of 1400 trees belonging to the worldwide collection of olives in Tassaout region, we detected Hylesinus oleiperda (Coleoptera, Curculionidae) whose appearance took place in April 2014. Attacked varieties belong to different geographic origins. Some parameters of the insect attack will be discussed.

Keywords: Xylomedes coronata, Hylesinus oleiperda, Acherontia atropos, olive trees

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 116

Fruit Fly Net Project: E‐monitoring and pest management design issues of a location–aware system for olive fly and other fruit fly pests

M. AlQasem1, A. Sciarretta2, M. R. Tabilio3, M. Miranda‐Chueca4, D. Nestel5, N. Papadopulos6, T. Tsiligiridis7

1National Center for Agricultural research and Extension (NCARE), Jordan 2Agricultural Research Council – Fruitculture Research Center (CRA‐FRU), Italy 3University of Balearic Island (UIB), Balearic, Spain 4Agricultural Research Organization (ARO), Israel 5University of Thessaly (UTH), Hellenic Republic 7Agricultural University of Athens (AUA), Hellenic Republic

Fruit flies of the family Tephritidae consist one of the most economically important groups of insect pests, threatening a multibillion fruit producing industry of the Mediterranean countries. There are major invasive species such as the peach fly, B. invadens and others that have dispersed over the last few years either in some countries of the area or in neighbouring countries, which expand the list of pestiferous Tephritids beyond the “native” Med fruit fly, European cherry fruit fly, and the olive fly. Conventional pest monitoring methods, including special designed traps are been utilized; however, these methods are time and labour consuming, mainly because they are not able to provide an automated insect‐specific detection/identification method. In this paper we present the basic architectural issues of a Location–Aware System (LAS) for fruit fly e‐ monitoring and spraying control. LAS is based on a real‐time Wireless Multimedia Sensor Network (WMSN) able to acquire and transmit scalar data and images from the field to a host station. In addition, a semi‐automatic trapping and insect counting warning system, based on existing traps, with specific characteristics will be developed and integrated with LAS for the detection of invasive species. A decision support system (DSS) will perform the final optimization of the control treatments. Efforts are concentrated on the design issues of a WMSN platform, able to collect and integrate multimedia data from the field. Finally, a flexible, architecture is adopted for integration of a WMSN to the cloud for multimedia sensor data collection and sharing using Web services. The project is running in five Mediterranean countries. The LAS is already under development and it will be deployed and tested for Bactrocera oleae (in Jordan and Spain), Ceratitis capitata (in Italy), Rhagoletis cerasi (in Hellenic Republic), Dacus ciliatus and Bactrocera zonata (in Israel). To facilitate the experiments a test site is also under development (in Hellenic Republic).

Keywords: Bactrocera oleae, Fruit flies, Invasive species, Traps, Sensors, Spraying

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 117

Evaluation of susceptibility of the most cultivated olive trees cultivars in Tunisia to Verticillium wilt disease (Verticillium dahlia)

M. A. Triki1, H. Hassairi1, F. Ben Amar 1, Y. Gharbi,2, I. Hammami1, S. Krid1, W. Khabou1, A. Rhouma1 and R. Gdoura2

1Laboratoire d'Amélioration et Protection des Ressources Génétiques de l’Olivier, Institut de l’Olivier, Université de Sfax, Tunisia 2Unité de Recherche Toxicologie ‐ Microbiologie Environnementale et Santé (UR11ES70), Faculté des Sciences de Sfax, Université de Sfax, Tunisia.

Verticillium wilt caused by Verticillium dahliae is one of the most important diseases occurring in Tunisia and all olive growing Mediterranean countries. To control the disease, the varietal resistance of olive tree could be an advantage to associate with other cultural measurements. For this purpose, two years old of sixteen olive tree cultivars (Olea europaea) potentially interesting in conventional production obtained from herbaceous cuttings, were root‐dip inoculated by the non defoliant isolate of V. dahliae V30. Sensitivity to wilt disease was evaluated by assessing symptom severity using a 0‐4 rating scale and estimating the area under disease progress curves. Results indicate that Verticillium wilt affected the most tested varieties. The olive cultivars Chemlali, Chetoui, Rakhmi and Arbequina were highly sensitive to Verticillium disease. The Interaction between V30 isolate of V. dahliae with different olive cultivars show higher disease severity index and stem colonization. However, Touffehi Tataouine and Coratina varieties have proved tolerant to Verticillium wilt with an average severity index not exceeding 1. This study demonstrates also the high performance of Zarrazi variety by showing the ability to reduce the potential for severe attack of V. dahliae. through the recovery phenomenon.

Keywords: Olea europaea, non defoliating isolate of V. dahliae, Verticillium wilt, sensitivity, tolerance.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 118

Genetic structure of Verticillium dahliae isolates infecting olive tree using AFLP, Pathogenicity and PCR Markers

Y. Gharbi1,2, M.A. Triki3, F. Daayf1, R. Gdoura2

1Department of Plant Science, Faculty of Agriculture and Food Sciences University of Manitoba, Manitoba R3T 2N2, Winnipeg, Canada; 2Unité de Recherche Toxicologie ‐ Microbiologie Environnementale et Santé (UR11ES70), Faculté des Sciences de Sfax, Université de Sfax, Tunisia. 3Laboratoire d'Amélioration et Protection des Ressources Génétiques de l’Olivier, institut de l’Olivier, Université de Sfax, Tunisia

Since 2006, verticillium wilt of olive induced by Verticillium dahliae has caused considerable economic losses in olive orchards in Tunisia. Using virulence tests, Vegetative Compatibility Grouping (VCG) and Amplified Fragment Length Polymorphism (AFLP) analyses, we investigated the genetic structure of V. dahliae isolates collected from different olive growing regions. In all, 42 isolates of V. dahliae from diseased olive trees were tested. Cluster analysis of AFLP data revealed that geographic origin was the main factor determining the genetic structure of V. dahliae populations. Isolates were divided into two major groups: the AFLP‐I group included all isolates from Sidi Bouzid, Kairouan, Kasserine and Sfax (central region of the country) and the AFLP‐II group included isolates from Monastir, Sousse, Mahdia (Coastal region), and two isolates from Sfax. Analyses of both the defoliating (D) and non‐defoliating (ND) pathotypes and VCG markers indicated that most of the isolates belong to VCG 2A and 4B/ND pathotype. These techniques were unable to differentiate V. dahliae isolates based on their geographical origin. The virulence patterns of V. dahliae isolates were investigated by assays on “Chemlali”, the most popular olive cultivar in Tunisia.

Keywords: Olea europaea; VCG ; Pathotype ; genetic diversity ; phylogeny.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 119

PESTOLIVE: a Mediterranean research project for understanding and managing soil‐borne parasites on olive using historical and ecological approaches

T. Mateille1, M. Achouri2, M. Ater3, A. Belaj4, G. Besnard5, P. Castillo6, E. Chapuis1, R. De La Rosa4, F. De Luca7, A.M. D’Onghia8, H. El Maraghi3, C. El Modafar9, A. El Mousadik10, A. El Oualkadi11, Z. Ferji2, N. Horrigue‐Raouani12, R.M. Jimenez‐Diaz6, M. Kadiri3, S. Kallel13, B. Khadari14, B.B. Landa6, L. Leon4, M. Montes‐Borrego6, A. Moukhli11, J.A. Navas‐Cortes6, A. Öcal15, N. Sasanelli7, J. Tavoillot1, M.A. Triki16, A. Troccoli7, E. Tzortzakakis17, M. Ulas18, N. Vovlas7, T. Yaseen9

1IRD, UMR CBGP, Campus Baillarguet, CS30016, 34988 Montferrier‐sur‐Lez Cedex, France. 2IAV Hassan II, Complexe d’Agadir, BP 18/S, 80000 Agadir, Morocco. 3UAE‐FST, BP 2062, 93030 Tétouan, Morocco. 4IFAPA, Avenida M. Pidal, Apdo. 3092, 14071 Córdoba, Spain. 5CNRS‐UPS, UMR EDB, Bât. T4R3, 118 Route de Narbonne, 31062 Toulouse, France. 6IAS‐CSIC, Alameda del Obispo, Apdo. 4084, 14005 Córdoba, Spain. 7IPP‐CNR, Via Amendola 122/D, 70126 Bari, Italy. 8IAMB‐CIHEAM, Via Ceglie 9, Valenzano, 70010 Bari, Italy. 9UCAM‐FSTG, Bd. A. Khattabi, BP549, 40000 Marrakech, Morocco. 10UIZ‐FSA, LBVRN, BP 8106, 80000 Agadir, Morocco. 11INRA, CNRA, BP513, Menara Marrakech, Morocco. 12ISA, Chot Meriem, BP47, 4042 Sousse, Tunisia. 13INAT, Cité El Mahrajene, 43 Av. C. Nicole, 1082 Tunis, Tunisia. 14SUPAGRO, UMR AGAP, TA A‐108/03, Av. Agropolis, 34398 Montpellier Cedex, France. 15BATEM, Pasakavaklari Cad. Demircikara Mah. Muratpasa, Antalya, Turkey. 16IO, Rte. de Soukra, Km1.5, BP1087, Sfax, Tunisia. 17NAGREF, PPI, POBox 2228, 71003 Heraklion, Crete, Greece. 18ZAIM, University Street No43, 35100 Bornova‐Izmir, Turkey.

PESTOLIVE (Contribution of olive history for the management of soil‐borne parasites in the Mediterranean basin) is a project funded by ARIMNet, an ERANET action supported by the 7th European Framework Programme and by non‐European Mediterranean countries (www1.montpellier.inra.fr/CBGP/pestolive). PESTOLIVE aims at producing knowledge and tools for a new and efficient management of plant‐parasitic nematodes (PPN) and plant‐pathogenic fungi (PPF) in olive (Olea europaea L.) cropping systems and nurseries, while reducing the use of pesticides. Because of the anthropic continuum from Olea post‐glacial refuges to oleasters (domestication) and then to olive‐trees (breeding and cropping), the fragmentation of the PPN and PPF communities and of their natural enemies could explain the scattered diversity of the control techniques (especially resistance rootstocks, biocontrol, cropping strategies) developed and applied all around the Mediterranean basin. The novelty of PESTOLIVE is based on i) the analysis and the management of the parasite diversity (ecology of communities) instead of controlling emblematic species (population approach) and ii) the involvement of knowledge about the historical co‐adaptation of soil‐borne parasite and natural enemies communities to olive‐tree domestication (origins and past assemblages) and breeding that follows the history of O. europaea around the Mediterranean basin.

Keywords: breeding, domestication, management, olive, root‐parasites. OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 120

Biopesticide production by solid‐state fermentation using olive industry by‐ products

S. Roussos1, M.S. Tranier1, Q. Carboué1, M. Mehjoub1‐2, C. Périssol1, I. Perraud‐Gaime1, R. De la cruz‐Quiroz3, C.N. Aguilar3, M. Ait Hamza4, A. Cherif2

1Aix Marseille University, CNRS, IRD, Avignon University, IMBE UMR‐237, Ecotechnology & Bioremediation Team, Faculty of Sciences of St Jérôme ; 13397 Marseille cedex 20, France 2Laboratoire de Biotechnologie et Valorisation des Bio‐Géo Ressources; ISBST; Université de la Manouba ; BioTechPole Sidi Thabet, 2020, Sidi Thabet, Ariana, Tunisie 3Food Research Department (DIA‐UAdeC). School of Chemistry. Autonomous University of Coahuila. Saltillo, 25280, Mexico 4Laboratory of Biotechnologies & Valorization of Natural Ressources ; Department of Biology, Faculty of Sciences, Ibn Zohr University, BP 8106, Agadir, Morocco

The four main causes of diseases affecting olive trees are: insects, bacteria, fungi and nematodes. These phytopathogenic pests cause important agricultural losses each year. In order to control them, chemicals are widely used. Because of their highly ravaging effects on biodiversity, environment and human health, numerous products have been removed from the market, and more and more will be withdrawn in the future. A friendly alternative to replace chemical pesticides consists in the use of natural antagonists to control pests. These antagonists could be insects, bacteria, filamentous fungi, but also pheromones, which are biodegradable natural specific toxins against insects. In our laboratory, a great number of nematophagous fungi have been isolated from phytoparasitic nematodes affecting olive‐tree roots in Morocco. These native antagonists have been isolated in pure culture and a study of their physiology of growth and sporulation has been done in order to produce new biopesticides. To produce them in a high range, the solid‐state fermentation technique was used for the cultivation of some filamentous fungi like Paecilomyces lilacinus, Trichoderma harzianum, and Verticillium lecanii. These fungi were cultivated on solid substrates, in particular on olive pomace, olive oil, mixed with other agro‐industrial substrates (wheat bran, sugarcane bagasse). For the scaling up and the production of high amounts of biomass and metabolites, a new disposable Bioreactor “FMS‐Unique” had been set up. It permits the growth of fungi in optimal conditions, ensuring aseptia and application during the process of a hydric stress stimulating sporulation and production of secondary metabolites to finally obtain a virulent and viable dry product that could be stored for more than one year. Analyses were done and have permitted to measure the quantity of viable and virulent conidiospores over time and set up conservation conditions for biopesticides obtained by SSF in that new bioreactor. In this article, the strategy of isolation, culture and sporulation of antagonistic fungi, and also Solid‐State Fermentation techniques in the FMS‐Unique bioreactor‐ mainly the interest of the use of by‐products of the olive production industry (olive pomace) to maintain the virulence of the biopesticides‐ will be discussed.

Keywords: olive cake, olive oil, Solid State Fermentation, filamentous fungi, nematode, Trichoderma harzianum

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 121

Physical parameters of olive fruits and olive leaves from Portuguese olive varieties and their role in oviposition preference of Bactrocera oleae

R. Malheiro, S. Casal, N. Rodrigues, P. Baptista, A. Bento, J. A. Pereira

1Mountain Research Centre (CIMO), School of Agriculture, Polytechnic Institute of Bragança, Campus Stª Apolónia, Apartado 1172, 5301‐855, Bragança, Portugal 2REQUIMTE/Laboratory of Bromatology and Hydrology, Faculty of Pharmacy, Porto University, Rua Aníbal Cunha 164, 4050‐047 Porto, Portugal

In this work was intended to evaluate physical parameters of olive fruits (weight, maximum and minimum diameters, length, and color parameters (a*, b* and L*)) and olive leaves (weight, width, length, and color parameters) from three Portuguese olive varieties (cvs. Cobrançosa, Madural and Verdeal Transmontana) during olives maturation (6 sampling dates) and try to establish correlations with cultivars susceptibility towards olive fly. Infestation levels were monitored in the three varieties during the crop maturation. Results clearly showed that cv. Verdeal Transmontana is more susceptible (24.5% infested fruits) than cv. Madural (16.0%), being cv. Cobrançosa the most resistant olive cultivar (8.5%). Maturation of the different olive varieties showed to be an important aspect in olive fly preference, reporting cv. Verdeal Transmontana a slower maturation process comparatively to cvs. Madural and Cobrançosa. Such fact influenced color parameters of olive fruits. An interesting observation was that olive leaves from cv. Verdeal Transmontana reported higher L* values (luminosity), while cv. Cobrançosa reported lower values. This result could suggest that olive leaves luminosity may attract olive fly at a long‐range. The same observation was checked in olive fruits. Results obtained appear to show two kinds of attractant/repellency mechanism. Our hypothesis is that olive leaves may function as a long‐range guide for olive fly, while olive fruits may act at close‐range. Color parameters may exert an important function in attractant/repellency mechanisms, being directly related to crop maturation in the case of olive fruits. Such results included in a broader project, considering chemical and molecular factors, may contribute for the adoption of effective control means to control olive fly.

Keywords: Bactrocera oleae, olive variety, oviposition preference; physical parameters; maturation process

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 122

Distribution of olive fruit fly in California based on fruit infestations since the 1998 invasion

V. Yokoyama

USDA‐ARS, San Joaquin Valley Agricultural Sciences Center, Parlier, California, 93648, U.S.A.

Olive fruit fly, Bactrocera oleae (Rossi), was first discovered in Los Angeles, California in 1998. Eradication and containment programs were immediately initiated, but within four years the olive pest was detected throughout the state. Olive fruit fly is not tolerated in canned fruit and the insect has potential to destroy the annual table olive crop which is solely produced in California for domestic consumption. Pest abundance and distribution has been traditionally determined with yellow sticky traps with adult pheromone lures and female baits, but trap captures are not good indicators of fruit damage. To determine the extent of fruit infestations, olives were collected throughout the state and evaluated for olive fruit fly larval emergence. Olives were maintained in the laboratory in plastic containers and mature larvae emerging from the fruit were collected as pupae. Larval age was evaluated by emergence date and insect density was determined by numbers per fruit or fruit weight. Olive growing regions were delimitated by climate and county and classified as inland valley including the Sacramento and San Joaquin Valleys, or coastal including the northern, central, and southern coast. Infested fruit was collected primarily during the fall when olive fruit fly populations were prevalent. Collection sites were reported by coordinates and compared among the olive growing regions from 1999 to 2013. A large amount of infestation data was obtained during an intensive biological control program in which a parasitoid imported from Guatemala was released throughout the state from 2002‐2011. Olive fruit fly adults were shown to be heat sensitive and populations in table olives were limited in distribution by high summer temperatures. Fruit infestations were highest and as many as three adults were reared per fruit in olives grown along the humid Pacific coast where temperatures are mild throughout the year. Lowest infestations or < 1 per fruit were collected from olives grown in the hot and dry inland valleys. Fruit grown in the foothills at or above 125 m where temperatures were mild, also had infestations more typical of coastal populations. Olive fruit fly has not yet become an economic pest of new high density plantings of oil olives grown in the Central Valley of California. Although mature olive fruit grown for oil is smaller than fruit grown for canning, unripe table fruit 1 cm in length can produce one olive fruit fly adult, so the pest has potential to migrate to oil olives.

Keywords: Bactrocera oleae, Olea europaea, larval infestations, pest populations, canned olives

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 123

Study of the duration of attractant action of various attractant solutions used in bait sprays against Bactrocera oleae (Diptera: Tephritidae)

Α. Kalaitzaki1, E. Malandraki2, E. Kapogia3, A. Ioannou3

1Nagref‐Institute of Olive Tree and Subtropical Plants of Chania, Agrokipio, 73100 Chania, Greece 2Regional Unit of Chania, Directorate of Agriculture and Veterinary Medicine, Αgia, 73103 Chania, Greece 3Hellenic Ministry of Rural Development and Food, Department of Plant Protection, Sygrou Avenue 150, 17671, Athens, Greece

Bait sprays constitute the current standard control method against Bactrocera oleae and when properly and timely applied, are very effective and more friendly to olive agro‐ecosystems compared to cover sprays. One of the most important requirements for high efficacy of this method is the prolonged residual activity of the bait since the currently used lasts only 5‐6 days, which is not enough to cover the practically continuous emergence of adults. The objective of this study was to evaluate the duration of attractant action of commercial attractant solutions used in bait sprays, tested at different doses. The study was conducted in an olive grove of Institute of Olive tree and Subtropical Plants in Aspreas area, in the Chania, Greece of about 1,000 olive trees (cvs “Koroneiki”, Konservolia, Kalamon and “Tsounati”). To test the attractant solutions an offhand paper trap (55cm x 45cm x 0.4 cm) was developed to better approach the real conditions of application of attractive solutions with bait spray. The trap was coated with entomological glue and five to six olive shoots of about 30‐50 cm in length were placed in the trap’s surface. The following treatments were used: Success 0.24 CB (Elanco Hellas) (at rates 3.3%, 6.7%), BAS31063 (BASF) (at rate 50%), Dacus Bait 100 SL (ΕVIP) (at three rates 2, 5, 10%), Dacus Bait 100 (at three rates 2, 5, 10%) combined with the registered insecticides a‐cypermethrin (Fastac), lambda cyhalothrin (Κarate), dimethoate (Perfection 40 EC) και thiacloprid (Biscaya 240 OD) and the water. The shoots were sprayed with the tested attractants until runoff. For the evaluation of attractant solutions, three trials were applied from August to October 2013. The experimental design was a randomized complete block with three replications. The evaluation was based on measurements of captured olive fruit fly adults on traps, every 2‐3 days, for about 20 days. According to the results the combination of insecticides Karate and Fastac with Dacus Bait at denser doses (5% and 10%) than the recommended (2%) attracted higher number of insects compared with other treatments up to 13 and 11 days after application during August and September respectively. Significantly lower number of insects was attracted by the Dimethoate + Dacus Bait at the three tested doses almost throughout the studying period.

Keywords: Bactrocera oleae, attractant solutions, bait spray, olive tree

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 124

Combined effectiveness of tebuconazole‐trifloxystrobin against infections of Fusicladium oleagineum in comparison to conventional products

V. Vizzarri, N. Iannotta , I.Muzzalupo, T. Belfiore

Olive Growing and Oil Industry Research Centre CRA‐OLI, 87036, Rende (CS), Italy

The fungus Fusicladium oleagineum (Castagne) Ritschel causes the “scab” or “peacock spot” disease in olive, and is responsible for severe yield losses in many olive‐growing regions in the world. The disease is manifested by severe leaf drop that alters the metabolism of the plant, with serious nutritional and hormonal imbalances that interferes with flower induction, with lower production. Infections develop mainly during spring and autumn, with serious vegetative and productive damages mainly for susceptible plants placed in moist conditions favorable for disease development (olive groves into the bottom of valleys, in plains, or adjacent to water courses etc). Currently, the control is carried out with cupric or translaminar (Dodine) products having high toxicological and environmental risks. This study tested a new systemic fungicide (Flint®Max) against the pathogen, that combines the effect of tebuconazole and trifloxystrobin. Efficacy is compared to traditional fungicides (Dodine and Copper oxychloride). Tests are carried out in Calabria in the experimental fields of the Research Center for Olive and Oil Industry Rende (Cosenza) on young plants of cv. Carolea, highly susceptible to fungal infection. Four experimental theses with three replicates were used in the experimental design, of wich three treated with copper oxychloride, tebuconazole‐trifloxystrobin and dodine, and one used as control. The aim of the research was to evaluate the effectiveness of the new fungicide, safer for the environment and with lesser toxic residues within the product than traditional products. The results demonstrate that the new fungicide contains the infection of the pathogen within limits comparable to dodine and copper oxychloride under the conditions in which the test was carried out.

Keywords: Olea europea, Fusicladium oleagineum, systemic fungicide, copper, Calabria.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 125

Genetic diversity of Verticillium dahliae from olive trees in Tunisia based on RAMS and IGS‐RELP

Y. Gharbi1,2, M.A. Triki3, F. Daayf1, R. Gdoura2

1Department of Plant Science, Faculty of Agriculture and Food Sciences University of Manitoba, Manitoba R3T 2N2, Winnipeg, Canada; 2Unité de Recherche Toxicologie ‐ Microbiologie Environnementale et Santé (UR11ES70), Faculté des Sciences de Sfax, Université de Sfax, Tunisia. 3Labortoire d'Amélioration et Protection des Ressources Génétiques de l’Olivier, institut de l’Olivier, Université de Sfax, Tunisia

Forty‐two Verticillium dahliae isolates were recovered from stem and root samples of olive trees showing typical symptoms of verticillium wilt in various olive‐growing regions in Tunisia. Each isolate was identified based on microscopic observations of morphological and cultural characteristics, as well as PCR amplification using Vd1/Vd2 primers. Genetic diversity among the isolates was investigated using random amplified microsatellites (RAMS) and PCR‐RFLP of intergenic spacer region (IGS) of ribosomal DNA (rDNA). A single fragment of approximately 1.7‐ 2.1 kb was amplified from all isolates by PCR using primers CNL12 and CNS1. Digestion of the amplified IGS region with restriction enzyme RsaI produced similar banding patterns (1200 and 800 bp) for 40 isolates. Interestingly, individual and distinctive banding patterns (1100, 850 and 150 bp) were observed for 2 isolates. Using RAMS primers, nine, eight, and four bands were produced when using primers CGA (2200, 1400, 1200, 1100, 1000, 650, 550, 500 and 350 bp), CCA (2000, 1200, 950, 850, 800, 550, 500 and 400 bp), and GT (2500, 2400, 700 and 500), respectively. When data from the RAMS experiments were combined, a total of 21 polymorphic markers were scored. The sources and extent of this difference are unknown. Phylogenetic analysis of the RAMS profiles indicated that V. dahliae isolates are randomly spread across Tunisia.

Keywords: Olea europaea,; IGS‐RFLP,; RAMS,; genetic diversity,; phylogeny.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 126

Biological control against olive Psyllid Euphyllura olivina by releases of indigenous bug Anthocoris nemoralis

N. Gharbi1 , A. Dibo2

1Institut de l’Olivier, Rue Hédi Karray, 2080 Ariana, Université de Sfax, Tunisia, 2Institut de l’Olivier, P.O. Box. 1087, 3000 Sfax, Université de Sfax, Tunisia.

This study investigated the impact of field releases of Anthocoris nemoralis (Heteroptera: Anthocoridae) nymphs against the olive psyllid, Euphyllura olivina (Homoptera: Psyllidae), a serious problem pest in Tunisia. Field trials were conducted in two successive years (2009 and 2010) in two organic olive orchards located in the regions of Taous and Chaal (Sfax : Southern East of Tunisia). For this purpose, two dose treatments were applied: a low dose (A. nemoralis 10 nymphs/tree) and a high dose (A. nemoralis 30 nymphs/tree). The first and the second releases of indigenous bugs were made during the second and the last week of March, respectively. The obtained results showed that augmentative releases of A. nemoralis nymphs were an efficient method for controlling the build‐up of E. olivina spring population. Compared to the control, the use of the high dose reduced significantly the densities of pest, whereas no significant difference was recorded between results from the low dose use and the control. After 8 weeks from the first release, the anthocorid nymphs were able to bring at about 9.34 % – 85.27 % in olive psyllid linear densities.

Keywords: Anthocoris nemoralis, biological control, Euphyllura olivina, olive tree

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 127

Learning from olive evolution and cultivation to understand the diversity of associated plant‐parasitic nematodes communities in Morocco

N. Ali1, E. Chapuis1,2, J. Tavoillot1, M. Aït Hamza3, Z. Ferji4, A. El Mousadik3, A. El Oualkadi5, G. Besnard6, A. El Bakkali7, A. Moukhli5, B. Khadari8, C. El Modafar9, M. Ater10, T. Mateille1

1IRD, UMR CBGP, Campus de Baillarguet, CS30016, 34988 Montferrier sur Lez Cedex, France. 2IRD, UMR RPB, BP 64501, 34394 Montpellier Cedex 5, France. 3UIZ‐FSA, LBVRN, BP 8106, 80000 Agadir, Morocco. 4 IAV Hassan II, Complexe d’Agadir, BP 18/S, 80000 Agadir, Morocco. 5INRA, CNRA, BP513, Menara Marrakech, Morocco. 6CNRS‐UPS, UMR EDB, Bât. T4R3, 118 Route de Narbonne, 31062 Toulouse, France. 7INRA, BP 578, Km 13 Route Haj Kaddour, Meknes, Morocco. 8SUPAGRO, UMR AGAP, TA A‐108/03, Av. Agropolis, 34398 Montpellier Cedex, France. 9UCAM‐FSTG, Bd. A. Khattabi, BP549, 40000 Marrakech, Morocco. 10UAE‐FST, BP 2062, 93030 Tétouan, Morocco.

Plant‐parasitic nematodes (PPN) significantly contribute to economic losses in the top‐ten olive producing countries, especially in the Mediterranean basin. Instead of controlling the main pathogenic nematode species as usual, one innovative strategy to control PPN would be to manage diversity in communities in order to lead them to be less pathogenic. Then, knowing assemblage mechanisms in communities due to evolution and environmental forces is a prerequisite.This study was conducted in Morocco, because (i) information about PPN diversity is lacking, (ii) different forms of olive occur (relict O. europaea subsp. maroccana, wild and cultivated olive), and (iii) typical cropping systems are present (traditional, high density, irrigated or not). Morphobiometric observations revealed a very diverse parasite nematofauna (99 species), six new taxa being recorded for the first time on olive. Hoplolaimidae and Telotylenchidae nematodes were dominant (80% of the samples), whereas root‐knot nematodes (Meloidogyne spp.) were detected in 40% of the samples. Multivariate analyzes showed that the development of Heterodoridae and Longidoridae nematodes was favored in PPN communities on wild olive, while lesion (Pratylenchidae) and root‐knot nematodes multiply in orchards. Three Meloidogyne species were identified: M. javanica on feral and cultivated olive in southern and center Morocco; M. arenaria and M. hapla on wild olive in the north. COI, 63R and ITS2 genes were good markers for species differenciation, but they were not able to distinguish M. javanica and M. arenaria populations and were not adapted for intraspecific differenciation. However, a significant morphological variability was observed between the Meloidogyne species, and within and between M. javanica populations. The response of the diversity of PPN communities as well as of Meloidogyne populations to olive genotype, geo‐climatic and soil physico‐chemical characteristics, and diversity of plants associated with olive trees is discussed.

Keywords: diversity, communities, Morocco, olive, plant‐parasitic nematodes.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 128

Application of antagonistic bio‐products (Bio‐Zeid and Bio‐Arc) to control Verticillium wilt disease of olive in nursery conditions

T. Yaseen, R. Abou Kubaa , A. M. D’Onghia

Centre International de Hautes Etudes Agronomiques Méditerranéennes (CIHEAM)/Mediterranean Agronomic Institute, Via Ceglie 9, 70010 Valenzano (BA), Italy.

Two antagonistic commercial bio‐products namely Bio‐Arc® 6% (Bacillus megaterium) and Bio‐ Zeid® 2.5% (Trichoderma album) were used in this study to evaluate their effect on vegetative parameters of olive plantlets and on the inoculum density of Verticillium dahliae microsclerotia in nursery produced olive planting stocks, under controlled conditions. Trials were carried out on both 8‐month old self‐rooted and 18‐month old grafted olive plantlets of cv Leccino. The bio‐ products were added to the growing media (20% sand, 45% peat moss, 20% pomice, 15% perlite, macro and microelements) seven days before transplanting. Plantlets were inoculated by dipping the roots in a conidial suspension (4×106 CFU/ml‐1) of the pathogen. Real‐time PCR assessed inoculum density of V. dahliae and the incidence of infected olive plantlets, after 15 months. Plant height, fresh weight, and root weight were also determined. All grafted olive plantlets and inoculated with V. dahliea treated or not with bio‐product showed wilting symptoms up to death. Bio‐products efficacy was more evident on self‐rooted plantlets. The bio‐control agents also increased the plant growth parameters. Nevertheless, Real‐time PCR results indicated that all the tested bio‐products reduced, significantly, inoculum density of V. dahliae in the growing media and the incidence of infected roots.

Keywords: Olea europaea, Biological control, V. dahliea, Real‐time PCR, Bacillus megaterium¸ Trichoderma album.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 129

Endophytic fungi in olive tree cultivars with different susceptibilities to anthracnose

F. Martins, J.A. Pereira, A. Bento, P. Baptista

Mountain Research Centre (CIMO), School of Agriculture, Polytechnic Institute of Bragança, Campus Stª Apolónia, Apartado 1172, 5301‐855, Bragança, Portugal

Olive anthracnose (OA), caused by Colletotrichum acutatum and C. gloeosporioides, is one of the most damaging fungal diseases of olive fruits worldwide. The available methods to control OA are mostly preventive, based on the selection of resistant cultivars and on early harvesting, since no effective fungicides are commercially available. During the last decade, endophytic fungi have received considerable attention due to their ability to protect their host against pathogens. Thus, in this work we intend to assess the diversity of fungal endophytes in olive tree cultivars with different susceptibilities to OA: cv Cobrançosa ‐ moderately resistant; and cv Galega – susceptible. It is expected to reveal if the differences on the fungal endophyte community between olive cultivars are somehow correlated with OA disease susceptibility. The fungal endophytes were isolated from roots, leaves and twigs of healthy trees and identified by rDNA sequencing. The fungal endophyte frequency and diversity were higher on the OA‐susceptible cv Galega than on the moderately resistant cv Cobrançosa, in all of the plant tissues analyzed (roots, leaves and twigs). Among the species identified in the cv Cobrançosa, Paecilomyces lilacinus, Fusarium oxysporum and Trichoderma gamsii were the most frequently isolated endophytes; whereas in the cv Galega were Phomopsis columnaris, F. oxysporum and Macrophomina phaseolina. A clear distinction on fungal endophyte community was observed between olive cultivars and plant organs. Further, in all of the organs analyzed, some endophytic fungi were exclusively found in OA‐susceptible cv Galega as well as on moderately resistant cv Cobrançosa. Although preliminary, the results emphasized the potential importance of endophytic fungi in the OA disease resistance in olive tree. Further studies are needed to confirm these findings.

Keywords: Olive tree,; cv Galega,; cv Cobrançosa,; endophytic fungi,; plant resistance, Colletotrichum.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 130

Phytophthora palmivora: Geographic distribution in different olive tree regions in Morocco and pathogenicity

M. Chliyeh1, Y. Rhimni1, K. Semaoui1, A. Ouazzani Touhami1, A. Filali‐Maltouf2, C. El Modafar3, A. Moukhli4, A. Oukabli5, R. Benkirane1, A. Douira1

1 Laboratoire de Botanique et de Protection des Plantes, Département de Biologie, Faculté des Sciences, Université Ibn Tofail, Kénitra; 2Laboratoire de Microbiologie et Biologie Moléculaire, Faculté des Sciences, Université Mohammed V Agdal, Av. Ibn Batouta; 3Laboratoire de Biotechnologie, Valorisation et Protection des Agroressources, Faculté des Sciences et Techniques, Guéliz, Marrakech; 4 UR, Amélioration génétique des plantes, INRA Marrakech; 5INRA, Amélioration des Plantes et Conservation des Ressources Phytogénétiques, CRRA, Meknès, Maroc.

Surveys were carried out, Spring 2012 ‐ Summer 2013, in different regions of the olive sector in Morocco: Meknes Souk Larbaa, Kenitra (nurseries in Sidi Taibi), Sefrou, Ouazzane, Marrakech (Azouzia, Aataouia, Ait Aourir, Sraghna Tamnsourt, Jaidate and Tassaout). The isolation of fungi was carried out from the roots and bark of trees that show symptoms of decay and drying. Phytophthora palmivora was detected in six regions with different isolation percentages. Souk Larbaa was the region where P. palmivora was the most (85%) followed by Sidi Taibi nurseries (73.6%), Aattaouia (64%) and Jaidate (47%). P. palmivora existed in the region Tassaout with the lowest isolation percentage (32%). Two isolates of P. palmivora were selected to study the pathogenicity of P. palmivora on plants of two olive varieties: Haouzia and Dahbia. Both isolates of P. palmivora have demonstrated their ability to induce root rot, dieback of olive plants twigs for both varieties (Haouzia and Dahbia). The percentages of the affected branches (Pab%) of the olive plants inoculated with isolate 1 were higher (81.8% for Dahbia and 68% for Haouzia) than those inoculated with the isolate 2 (43% Dahbia and 32% for Haouzia). Koch's postulates have been verified by isolating P. palmivora from the inoculated olive plants.

Keywords: Morocco, Olea europaea, Phytophthora palmivora, geographic distribution, pathogenicity.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 131

Effect of olive varieties and rearing substrates on plant‐parasitic nematode communities in southern Morocco olive nurseries

M. Ait Hamza1,2, Z. Ferji 2, N. Ali 3, H. Tazi1, J. Tavoillot 3, A. Moukhli 4, H. Lakhtar 1, S. Roussos 5, H. Boubaker 1, A. El Mousadik1 and T. Mateille 3

1UIZ‐FSA, LBVRN, Agadir, Morocco ; 2IAV Hassan II, CHA, Agadir, Morocco ; 3IRD, UMR CBGP, Montferrier‐ sur‐Lez Cedex, France ; 4INRA, CRRA, Marrakech, Morocco ; 5IRD, UMR IMBE, Marseille, France.

Plant‐parasitic nematodes (PPN) affect significantly the production of susceptible plants, including olive tree (Olea europaea) in the ten‐top olive producing countries in the world, especially in the Mediterranean basin (Spain, Italy, Greece, Tunisia, and Morocco). Moreover, these parasites strongly affect young olive trees in nurseries and contribute to economic losses. PPN are everywhere detected in communities (i.e. species population mixtures). Knowing that plant production partly depends on parasite diversity, more information about PPN communities and about interactions with their habitats is a prerequisite for the development of any control strategy. In this context, PPN communities were determined in 210 soil samples collected from several plant varieties in 20 olive nurseries located in the Souss and the Haouz regions (southern Morocco), during March and April 2013. Morphobiometric observations revealed a significant diversity of PPN, belonging to 12 families and 22 genera. Results showed dominance of the genera Helicotylenchus, Tylenchorhynchus, Rotylenchus, Trichotylenchus and of Tylenchidae that are known for their damages on Mediterranean olive. The most frequent PPN were Helicotylenchus spp. (86.45 %), Meloidogyne spp. (56.28%), Tylenchoryhnchus spp. (50.37%), Trichotylenchus spp. (42.31%), and Rotylenchus spp. (37.17%). However, Helicotylenchus spp., Meloidogyne spp., Tylenchorhusyhnchus spp., Trichotylenchus spp. and Rotylenchus spp. population levels were significant (up to 56,640 individuals /dm3 of soil) in most of the olive nurseries surveyed. The contribution of olive varieties and soil substrates on biodiversity indices and on PPN community patterns is discussed.

Keywords: Morocco; Olea europaea; nurseries; plant‐parasitic nematode communities; soil substrates; varieties.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 132

Comparative effect between biopesticides and biofertilizers on bioagressors of Olea europaea

L. Brahimi, H. Halladj, Z. Djazouli

University of Blida, Faculty of Agro‐Veterinary Sciences and Biological Department of Agricultural Sciences, BP270, road Soumaa, Blida, Algeria.

Integrated production in sustainable agriculture seeks to improve the efficiency of organic inputs across formulations thus prolonging persistence at field by incorporating synergists products themselves being non‐toxic at the doses used in order to increase protective action and maintain the culture viability. In this context the present study considers the evaluation of the effectiveness of biofertilizer (vermicompost) and a biopesticide (formulated essential oil of oregano) compared to a synthetic pesticide Methomyl, on pests of the olive tree (Olea europaea). From elementary plots used for our investigations, weekly monitoring of the abundance of pests was carried before the application of molecules and has spread out to 10 days after treatment. The overall analysis of the results shows a disparity in the degree of toxicity of various treatments, the three molecules which have a repressive effect diverges depending on the physiology of each pest, Unlike the olive bud mite (Aceria Olea) which has a similar sensitivity for the three molecules, olive psyllid (Euphyllura olivina) has a sensitivity for essential oil of oregano has made a. The results show a moderate biocenotic resumption in species having received biological molecules, however as a result of Methomyl, there is a pronounced disturbance of the abundance of different groups of pests. In the end, we can conclude that biological control is a good alternative to chemical control, including the use of biopesticides and biofertilizers that provide major results in protecting the environment

Keywords: biofertilizer, biopesticide, synthetic pesticide, olive tree, biocenotic resumption.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 133

First report of Botryosphaeria dothidea and Diplodia seriata associated with olive tree wood lesions in Algeria

F. Ammad1, S. Benarous 2

1Departement of Biotechnology, University Blida1, BP 270 Blida 09000 Algéria. 2Departement of plant protection , SRPV Chelef

In Algeria, dieback of olive tree is becoming important and is very common in old and young trees. This study was conducted during 2014 to assess the state of dieback and to determine the fungal pathogens associated with olive tree decline. Some fields were chosen in Chelef province revealed that an average disease incidence was 7% based on herbaceous symptoms. A cross section of an infected wood trunk collected from two localities showed two kinds of necrosis: a dead brown of tissue of wedge shape and central light‐colored necrosis of soft consistency. Botryosphaeria dothidea and Diplodia seriata were isolated from infected wood. Our results represent the first report of a canker disease of olive tree associated with species of the Botryosphaeriaceae in Algeria.

Keywords: Olea europaea, trunk diseases, epidemiology, Botryosphaeria sp.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 134

Biological control of Colletotrichum species associated with olive anthracnose in Tunisia

M. Chattaoui, M. Msallem, A. Rhouma

Lab. Improvement and protection of olive genetic resources, BP 208 Cite Mahrajene – 1082 Tunis TUNISIA

Species of Colletotrichum collectively cause anthracnose on olive (olea europaea L.), which is considered the most dangerous infection of olive fruit. In fact, it can inflect considerable yield loss under the appropriate environmental conditions. In Tunisia, disease symptoms were first reported to be caused by the Colletotrichum gloeosporioides species. More recently, in 2014, the Colletotrichum acutatum complex has been reported for the first time as the causal agent of olive anthracnose. The objective of this research was to evaluate the use of rhizobacteria for the biological control of this disease. For that, a total of forty isolates were obtained from the rhizosphere of three olive tree cultivars (cv. Chetoui, cv. Chemlali and cv. Meski). After “In vitro” screening, three isolates (IOT.MC39, IOT.MC82, and IOT.MC85) were selected and used as antagonists. Detached healthy olive fruits were first inoculated with C. glaeosporioides (IOT.COL02) and C. acutatum (IOT. COL 14.2) isolates and treated with bacterial suspensions. Results showed that among the three isolates, IOT.MC85 was the most effective with an inhibition of 65%. The antagonistic activity of the three isolates was also tested on the conidial germination and results showed a significant difference. IOT.MC82 and IOT.MC85 had the most important inhibition, with respectively 75% and 80% of non‐germinated conidia.

Keywords: Olea europaea, soapy olive, mycelium

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 135

Current status of Olive Weevil Rhynchites cribripennis Desbrochers (Coleoptera: Attelabidae) in Montenegro

T. Perovic, S. Hrncic, M. Cizmovic

University of Montenegro, Biotechnical Faculty, Center for Subtropical Cultures, Bar, Montenegro University of Montenegro, Biotechnical Faculty, Department for Plant Protection, Podgorica, Montenegro

It seems that olive weevil Rhynchites cribripennis becomes a serious olive pest in Montenegro. It was detected for a first time in the area of the peninsula Luštica in 2004th, when his presence was observed in the abandoned olive groves, on rocky terrain. In recent years, the damage caused by weevil was registered in intensive plantations of autochthon and introduced olive varieties. The aim of this study was to determine distribution and level of infestation of the R. cribripennis in Montenegro. One‐year study was conducted at seven localities along the Montenegrin seaside. In order to determine the level of infestation, at weekly intervals over the period from the end of June to the end of September 2013, at all localities, a sample of 50 randomly selected fruits was taken and examined in the laboratory under a stereomicroscope. The results show that R. cribripennis is widespread pest at Montenegrin seaside. The level of infestation was high on most localities, reaching over 80% except one location in the area of Bar (Belveder), where 35% of damaged fruit was recorded. The most widespread autochthon and introduced varieties, Žutica and Leccino, are susceptible to attack of this pest.

Keywords: Rhynchites cribripennis, olive weevil, distribution, infestation

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 136

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 137

OLIVE OIL QUALITY HEALTH TABLE OLIVE

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 138

Olive Oil Quality and Health

A. Kiritsakis1, D. Gerasopoulos 2, E. L. Iorio3 and K. Kiritsakis4

1Prof. AOCS Fellow, Greek Observatory of Oxidative Stress, 2Prof. Aristotle University of Thessaloniki, Greece 3International Observatory of Oxidative Stress, IOOS, 84127 Salerno (SA), Italy. 4Food Scientist, Ph.D Candidate Aristotle University of Thessaloniki.

The formation of the oil quality starts in the orchard and is affected by cultivar, climatic and environmental factors. These factors, in turn, affect the oil aroma, polyphenol and sterol constituents, among others. The aroma and the flavour compounds of olive oil, as well as the chlorophyll and pheophytin pigments, increase the stomach secretion and facilitate the absorption of the polyphenol antioxidants, which furthermore protect the body tissues from oxidation. The processing systems (Pressure or Centrifugation) and the conditions applied during processing also affect the quality of olive oil and its composition. E.g. the addition of high amount of water (with temperature higher than 27 oC) in the olive oil mill results in a considerable decrease of the hydro soluble (polyphenols) and in a partial decrease in the lipid soluble (chlorophylls and tocopherols) compounds. Deterioration of olive oil quality during storage is favored by factors such as temperature, presence of metals, light, moisture and others. A strong linear correlation between the polyphenol content and the induction period of the oil has been observed. The presence of phenols, tocopherols and other natural antioxidants prevent lipid oxidation and protect the oil during storage. Oxidation has been noticed since antiquity as a serious deterioration of fatty substances. The reactive oxygen species (ROS) – free radicals needed for the oxidation mechanism are generated not only from auto‐oxidation of lipids during storage but also from many cellular oxidative pathways in the body. An imbalance between generation and removal of ROS can cause an oxidative stress in which excess ROS attacks and damages virtually all biomolecules (including DNA) in the cells, leading to serious chronic diseases and cell death. This presentation will focus in the total olive oil quality, the factors affecting it from the fruit to the oil consumption and to the relationship existing between olive oil quality and health. Emphasis will be given in constituents such as oleuropein, oleocanthal, oleacin and others, which greatly contribute to human health. Oleocanthal for example which is existing in extra virgin olive oil shows potential as a therapeutic agent in the treatment of inflammatory degenerative joint diseases. Generally, phenolic compounds which characterize a good quality of olive oil not only protect olive oil from oxidation but also modulate ROS and other oxidising agents, thus controlling oxidative stress. It becomes obvious that for a good olive oil quality (low acidity, negligible oxidation and best phenols and sensory characteristics) many factors must be considered and that olive oil of good quality has a great contribution to human health.

Key words: Olive oil, quality, ROS, oleocanthal, human health

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 139

Which of the factors: cultivar, extraction systems or storage conditions is determinant on the variation of olive oil physicochemical and the sensory profiles?

K. Ben‐Hassine, D. Malouche, K. Talmoudi, T. Ameni, H. Sebei, M. Hamami, N. Grati‐Kamoun

1Biochemistry Laboratory ‐ UR03/ES08 ’Human Nutrition and Metabolic Disorders USCR ’Mass Spectrometry Lab of Monastir Medicine University’, Tunisia 2 National Institute for Applied Sciences and Technologies, Tunisia 3 Institut de l’Olivier, Unité Technologie et Qualité, BP 1087, 3018 Sfax, Tunisia 4Higher School of Agriculture, Mograne 5Engineering School of Statistics and Information Analysis, University of Carthage, Tunisia 6National School of Engineers of Tunis, Unit Systems and Signals – (ENIT‐U2S), Tunisia 7Center of Biotechnology of Borj Cedria, Tunisia

This research was carried out to evaluate the effects of variety, extraction system and packaging material on the virgin olive oil’s physico‐chemical and sensory profile. The oils were analyzed before and after being stored for 9 months. Principal compound analysis and Graphical Modeling were applied to fully explore the influence of the studied factors. Graphical Modeling is considered as a way to determine which of these factors is more determinant in the variation of the different analytical descriptors. We mainly assume that the variables are sampled from a Gaussian distribution and where the interactions between the variables can be represented by a directed graph where the vertices are the variables. Once these graphs are estimated we are able to determine direct factors that influence the olive oil aromatic and sensory profile. Thus, two olive varieties Chemlali (Tunisia) and Coratina (Italy) were selected. The olive oils were extracted by different industrial processes (super press, dual and triple phase decanter) then stored in the established conditions (ambient and refrigerator temperature) in the following packaging materials: clear and dark glass bottles and metal bottles. The application of PCA to the samples on the basis of the extraction system employed does not show a good separation among VOOs especially for the 2‐phase and 3‐phase centrifugation system. According to the packaging factor, no important separation was observed especially between clear and dark glass samples. The separation between the Coratina and Chemlali VOOs according to the different studied factors shows that Coratina oils are characterized by higher amounts of 2 methyl butanal, cis 3 hexen 1 ol, cis 3 hexenyl acetate, hexyl acetate, cis‐2‐pentenol, 2‐pentanone, and 3‐pentanone and it is noteworthy for its higher content of phenolic compounds, while Chemlali samples were richer in pentanol, trans‐3‐hexenol, octanal, hexanal and penten‐3‐ol. The results of graphical models (fig. 1) on the phenolic, volatile compounds and the sensory descriptors shows that the volatile profiles of oils were influenced mainly by the storage date, that was influenced by the packaging material, where a considerable decrease was observed after storage which was reflected by the change of sensory characteristics of stored oils: loss of positive attributes fruitiness, bitterness and pungency and onset of defects which were mainly influenced by the storage date (fruity and bitter attributes), packaging material (pungent, rancid and fusty attributes) and extraction system.

Keywords: Virgin olive oil, volatile compounds, storage, sensory analysis, extraction system, graphical modeling. OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 140

Potential usefulness of a colorimetric test to measure olive oil peroxidation

E. L. Iorio1, A. Kiritsakis2, M. Carratelli1

1International Observatory of Oxidative Stress, IOOS, 84127 Salerno (SA), Italy. 2Greek Observatory of Oxidative Stress, GOOS, Thessaloniki, Greece.

The nutritional properties and the safety of olive oil can be impaired by lipid peroxidation, which rate has been related to early aging and many chronic, degenerative and inflammatory diseases. Therefore we developed and validated a ready‐to‐use colorimetric test kit (svelarancido) for consumers able to detect olive oil lipoperoxides level by exploiting the chromogenic properties of an aromatic amine; the intensity of the developed colour (from pink to red) is proportional to the initial amount of lipoperoxides in the olive oil sample (10 uL) and can be read after 2 minutes on a visual‐chromo‐analogical scale. The results obtained on 25 samples of extra virgin olive oil of the Italian olive germplasm from different brands were comparable with those done by the conventional test for lipoperoxides (potassium iodide method) by photometric way (Carpe Diem, Diacron International, Grosseto, Italy). Noticeably the test allowed to identify 2 samples over the maximum allowed range (>20 mEq/kg O2), which presumable cause was a transparent bottle. Because the close relationships between dietary lipid peroxides and oxidative stress‐related disorders (e. g. early aging, atherosclerosis and so on) this novel test appear as an easy, fast and suitable method to evaluate together other chemical parameters the quality and therefore the safety of olive oil even at home or in a restaurant.

Keywords: Olive oil, lipoperoxides, oxidative stress, svelarancido, safety, consumers.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 141

Fatty Acid Alkyl Esters in Extra Virgin Olive Oil: an Evolving Parameter

R. B. Gómez‐Coca1, G. D. Fernandes1,2, M. C. Pérez‐Camino1, W. Moreda1

1Department of Characterization and Quality of Lipids, INSTITUTO DE LA GRASA –CSIC‐, Avda. Padre García Tejero 4, E‐41012‐Sevilla, Spain. 2Fats and Oils Laboratory, Department of Food Technology, Faculty of Food Engineering, University of Campinas, Campinas‐São Paulo, 13083‐970, Brazil.

According to the Food and Agriculture Organization olive crops occupy the highest expanse of land in the world. Clearly the importance of olive oil does not only lie in its nutritional properties, but also in its repercussions in the Mediterranean countries’ economy. The European Union ‐the most important manufacturer and consumer of olive oil worldwide‐ and the International Olive Council have given a series of quality and purity parameters to distinguish among oil categories. Oil quality is directly related with both agricultural aspects and manufacturing practices. Olive fruit quality, grade of ripeness, and handling during and after harvesting, together with oil processing and storage conditions will determine the quality of the final product. Low quality olive oils with weak organoleptic defects are the objects of illegal practices channeled into concealing their negative flavor. One of the strategies commonly used consist of a neutralization and/or soft deodorization at low temperature, and then blending with extra virgin olive oil. This practice is difficult to detect and the determination of the fatty acid alkyl esters (FAAE) concentration could give unmistakable reasons to evidence it. FAAE are formed by esterification of free fatty acids with short chain alcohols, mainly methanol and ethanol, yielding methyl and ethyl esters, respectively. A relationship between the FAAE concentration in olive oils and their sensory classification is established. We have also proven the connection between the presence of large quantities of FAAE and the oil’s fermentative organoleptic defects. The strict demands regarding the FAAE presence in extra virgin olive oil –limits are thought to be lower by the years‐ were made under the assumption that the concentrations of such analytes were something fixed, and that they did not evolve over time. Our results demonstrate otherwise. After a two‐year study on the FAAE behavior under controlled olive‐oil storage conditions (taking into account light and temperature exposition together with the initial FAAE content), we will present our data and discuss the fact that the FAAE concentration does not only change in accord with fruit ripeness, but also over time once the oil is bottled. In this line will also present our research and conclusions on the study of the FAAE kinetics.

Keywords: olive oil, fatty acid alkyl esters, fermentative defects, organoleptic assessment, panel of experts.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 142

Hepatoprotective activity of oleocanthal extracted from olive oil amurca in the rat

S. Janakat, A. Al Amour

Department of Nutrition and Food Technology Faculty of Agriculture, Jordan University of Science and Technology, Irbid PO Box 3030, Jordan

Oleocanthal is a phenolic compound responsible for the pungent sensation of olive oil in the throat. The aim of this study was to evaluate oleocanthal potential hepatoprotective activity in the rat. Hepatoprotective activity of crude oleocanthal extracted from olive oil amurca was assessed by giving oleocanthal to Wistar albino rats via gavage for 5 days. On the penaltemate day, a single dose of 3 g/kg body weight of paracetamol was given to the rats via gavage, then liver function was evaluated by measuring total bilirubin and the activities of alkaline phosphatase (ALP), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in addition to ratio of liver/body weight. Crude oleocanthal was found to possess hepatoprotective activity at 1 and 5 mg/kg body weight. These concentrations reduced liver weight/body weight ratio significantly (P < 0.05) in comparison with the positive control (Paracetamol only group) (from 4.7 ± 0.26 % to 3.9 ± 0.13% and from 4.7 ± 0.26% to 4.09 ± 0.05 % respectively). One and 5 mg/kg body weight of crude oleocanthal were able to reduce plasma bilirubin level significantly (P < 0.05) in comparison with the positive control group (from 0.30 ± 0.03 mg/dL to 0.20 ± 0.009 mg/dl and from 0.30 ± 0.03 mg/dL to 0.17 ± 0.01 mg/dL respectively). A significant reduction in ALP activity (P < 0.05) was noticed using 1 mg/kg body weight of crude oleocanthal in comparison with paracetamol group (from 193.50 ± 24.5 U/L to 125.12 ± 8.02 U/L). A non‐significant reduction (P > 0.05) was noticed when using 5 mg/kg of crude oleocanthal in comparison with paracetamol group. Plasma AST activity was reduced significantly (P < 0.05) in comparison with the positive control group (from 99.33 ± 20.9 U/L to 53.35 ± 10.94 U/L and from 99.33 ± 20.9 U/L to 42.68 ± 3.36 U/L respectively). A significant reduction (P < 0.05) in ALT activity was noticed using 5 mg/kg body weight of crude oleocanthal in comparison with paracetamol (from 54.51 ± 12.31 U/L to 32.59 ± 2.35 U/L). A non‐significant reduction (P > 0.05) was noticed when using 1 mg/kg of crude oleocanthal in comparison with paracetamol. In conclusion oleocanthal could act as a hepatoprotective agent in rats against paracetamol induced liver damage. Nevertheless, further studies are needed to confirm these results in the humans.

Keywords: Olea europaea, amurca, oleocanthal, liver function tests, hepatoprotection

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 143

Effect of cultivar and climatic conditions on olive (Olea europaea L.) oil fatty acid composition

A.A. Zeinanloo

Horticulture Dept. Seed and plant improvement institute Karaj, Iran

Olive oil effects on health have rendered it as a beneficial product for consumers. Many factors, particularly the cultivar used in oil production and climatic condition, can affect olive oil quality. This research was conducted with 12 olive cultivars in 5 olive research stations with different climatic conditions (Tarom, Roodbar, Gorgan, Kazeron and Sarpolezehab). Fruits with 4‐4.5 ripening indices were harvested in each region. Fatty acids composition data were analyzed in a factorial experiment on the base of complete randomized block design. The results indicated that palmitic acid was significantly higher in warm climates, Kazeron and Sarpolezehab. The cultivar and climate had significant effects on oleic acid content and this fatty acid was higher in Tarom, Roodbar, and Gorgan regions than Kazeron and Sarpolezehab. ‘Amphissis’ cultivar had high Oleic acid content in all conditions. Mean linoleic acid content was 19.09% in warm condition of Sarpolezehab which was higher than mean value in Gorgan, 14.04%. The result indicated that ‘Koroneiki’ oil exhibited the best fatty acid composition in Tarom, Roodbar and Gorgan regions and ‘Amphissis’ was the only cultivar suitable for warm climate. ‘Shengeh’ cultivar produced the lowest quality oil with regard to fatty acid composition especially in warm region.

Keywords: Olive Cultivars, Oil quality, Adaptability Temperature, Iran

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 144

Role of chemometric techniques in the typicality determination of virgin olive oil from Moulay Driss Zarhoun (Volubilis)/ Morocco

S. Rouas1, M. Rahmani1, A. Antari², E. Baamal3, N. Maata4

1Department of Food Science & Nutrition, Section of the Agricultural and food Industries, IAV Hassan II, Madinat Al Irfane, B.P. 6202 Rabat‐Institutes Morocco, 2Laboratory of Oil Technology, Program of the Olive Tree INRA‐Marrakech, BP533 Marrakech, Morocco, 3Department of Statistics and Applied computer Science, IAV Hassan II, Madinat Al Irfane, Rabat – Institutes, Morocco, 4Official Laboratory of Analyses and Chemical Researches (LOARC), Casablanca, Morocco

The exact authentification of virgin olive oil origin becomes one of the priorities of the international community, both at the legislative and commercial levels. It also concerns the ultimate destination of this product; at the consumers’ level. The uses of geographical indications are considered a reliable way to guarantee the traceability of this product. The objective of the present work is to study the typicality of virgin olive oil from Moulay Driss Zarhoun's (Volubilis) region, with the ultimate objective to attribute a protected designation of origin (PDO) to the oil. Previous recommendations of such PDO were made in the past by the International Olive Oil Council (IOOC) and the Moroccan Ministry of Agriculture, based on a set of characteristics (typicality, originality, knowledge of production, fame, and a strong historic anchoring). The present research deals with the chemical characterization (composition in fatty acids (AG) and triacylglycérols (TAG)) and sensory evaluation of the olive oil. Based on chemometric techniques, the interpretation of the results will allow drawing the fingerprints of olive oil originating from “Volubilis”. A protocol of olive sampling (40 samples) was made in the zone of study. Samples, taken in double, were either submitted to oil extraction by the farmer (1st lot), or submitted to oil extraction in the laboratory (2nd lot), using hammer crusher. The analysis revealed a big wealth in polyphenols which exceeds on average 430ppm, an increase of oleic acid according to the altitude, a normal rate of triglycerides, and an average organoleptic note of the fruity attribute greater than 6.5. The oil is very aromatic, given the calcareous soil prevailing in the zone. Besides the chemical and sensory characterization of the oil, this study aims to establish the link between quality, origin of production and the system of oil extraction. A raising awareness of the participating farmers on the importance of good manufacturing practices is necessary for the improvement of olive oil quality.

Keywords: olive oil, chemical characterization, sensory evaluation, quality, geographical indication, Moulay Driss Zarhoun.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 145

Deodorized olive oils: from fraud to nutrition

R. Aparicio‐Ruiz1, D.L. García González2, I. Romero2, C. Oliver2, R. Aparicio2, M.T. Morales1

1Department of Analytical Chemistry, US, 41012 Seville, Spain 2Instituto de la Grasa, CSIC, 41012 Seville, Spain

Virgin olive oils are classified into different categories according to current regulations. The classification is based on a series of physical and chemical parameters, and organoleptic characteristics, allowing assign the oil to a specific category. In some instances, the oils can be clearly assigned to the lampante category , which means that they are intended for refining due to exceed certain levels of these parameters, related to hydrolytic, fermentative or oxidative alteration, and is unfit for human consumption. Other oils meet all physical and chemical parameters, but have a slight presence of sensory defects, and they do not fit current regulations, due to the presence of volatile compounds responsible for sensory defects. These oils may undergo a slight deodorization at low temperature, under vacuum, using nitrogen as carrier‐gas, to produce deodorized oils which maintain the lipid matrix virtually unchanged and have been used by fraudsters to be mixed with virgin olive oils of good quality, which constitute a kind of fraud difficult to be detected. These oils have not volatile compounds, but the content of other minor components, such as phenols, may remain unchanged. In this work the quality parameters, sensory properties, and phenols content of 50 virgin olive oils with different sensory profiles, from extra virgin to high intensities of sensory defects, and deodorized olive oil samples, have been evaluated. Phenolic compounds were determined by SPE‐HPLC‐DAD. Data were acquired at 235, 280, and 335 nm. Quantification of individual phenols was carried out using two internal standards. The results show significant differences in the phenol profiles between high and low quality virgin olive oils depending on the sensory defect, and also differences between the phenolic profiles of the standard samples of particular sensory defects. Virgin olive oils, characterized with slight intensity of certain sensory defects, maintain the profile of phenols virtually intact after deodorization under mild conditions. It is a deceived practice whether the resulting oil is labelled as virgin olive oil, but is not when labelled as olive oil but containing its natural phenols, what allows suggest the possibilities of this kind of oils as a source of bioactive components.

Keywords: virgin olive oils, deodorized, phenols, sensory defects, bioactive components.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 146

Olive Oil: An Exceptional Functional Lipid Food Protecting Our Body from Oxidative Stress

A. Kiritsakis1, E. Iorio2, F. Shahidi3, D. Gerasopoulos4, K. Kiritsakis5

1Prof. AOCS Fellow, Greek Observatory of Oxidative Stress. 2International Observatory of Oxidative Stress. 3Memorial University of Newfoundland, Canada. 4Associate Prof. Aristotle University of Thessaloniki, Greece. 5Food Scientist, Ph.D Candidate Aristotle University of Thessaloniki.

Certain populations have traditionally enjoyed overall good health. While a number of wide ranging factors certainly contributed to this good health, available information would suggest that contributions of specific components of the diet played a very important role. Olive oil, a natural juice obtained from the fruit of the tree Olea europaea, is not only a flavorful lipid food but also a product with a unique composition, playing an important role in our diet. It is a naturally monounsaturated oil containing polyphenols, flavonols and other compounds that have strong antioxidant properties and is believed to be protective against cardiovascular heart diseases (CHD). It also contains other micro‐ components, such as aroma and taste compounds, pigments (chlorophyll, pheophytin), both of which, contribute to the good aroma and taste of this oil and facilitate the absorption of all the useful components (e.g antioxidants) existing in the food.Composition and quality of olive oil are affected by several factors such as cultivar, fruit maturity, micro‐environment, processing technology (two or three‐phase decanter) and conditions of preservation. Metabolic studies of blood lipids and evidence regarding oxidation of LDL cholesterol suggest that consumption of olive oil, mainly virgin, as a principal fat is eliminating the oxidative stress and therefore the presence of free radicals in our body and contributes in part for traditionally low rates of CHD, breast cancer, and human longevity in general. Good quality virgin olive oil, containing phenol antioxidants , aroma and other important compounds helps indeed the functions of our body and protects it from different diseases related to oxidative stress. The later is a result of the unbalance between the antioxidant constituents of the diet and the presence of free radicals in our body. The purpose of this paper is to discuss how the quality and the main constituents of olive oil are affected by several factors and to provide an overview of evidence that olive oil, is a very important and original functional lipid food, eliminating the oxidative stress and protecting our body from different diseases.

Keywords: olive oil composition, quality, antioxidants, functional lipid food, oxidative stress, body protection

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 147

Reduction of acrylamide in potato chips using olive oil as frying mean. Immunoassay vs LC‐MS/MS techniques for acrylamide determination.

G. Siragakis1, G.Miliadis2, A.Vrettakou3

1Food Allergens Lab, Violi Haraki, 74100 Rethimno, Crete, Greece 2FA Food Allergens Lab Ltd, Kalopsidas 38, 7060, Livadia, Larnaca, Cyprus 3Athens Branch Food Allergens Lab, K.Varnali 40, 14231 N.Ionia, Athens, Greece

Acrylamide is formed in food containing starch such as French fries, potato crisps, crunch muesli, and crisp bread through the processes of turning brown like baking, roasting and frying. High acrylamide concentrations in food were first detected in 2002. This finding caused distinct concern in the scientific community, since animal experiments had shown that acrylamide is a genotoxic and carcinogenic agent. The top 5 food products by average acrylamide intake are French fries, French fries oven baked, potato chips, breakfast cereals and cookies. Although there was a myth that olive oil is not the best edible oil for frying potatoes, on 2010 Goody’s the greatest fast food chain in Greece and Balcans replaced all other edible oils with olive oil for frying potatoes. This proved to be better in smoke point, anisidine value and polar compounds. In this paper the relationship between olive oil (OO) vs other edible oils for the formation of acrylamide in potato chips was investigated. The aclylamide level of 20 packed potato chips samples delivered from Super Markets were analyzed by Elisa and LC‐MS/MS after Dispersive Solid Phase Extraction. An Elisa kit from Morinaga ( Japan) was tested versus LC‐MS/MS method. ELISA is cheaper but its range of quantification is limited and most samples analyzed were found out of the quantification range. Additionally the total time for ELISA analysis is 7 hours while the LC‐MS/MS technique demands only 4 hours if we use the QuEChERS ( Quick, Easy, Cheap, Effective, Rugged and Safe) sample extraction and clean‐up technique. Furthermore, different edible oils selected for frying experiments (corn oil, sunflower oil and olive oil) were tested. Slices of potatoes were fried at 180oC, for 15 min, and acrylamide content was determined by LC‐ MS/MS. Results demonstrated that olive oil yielded lower acrylamide concentration. Sunflower gave the higher concentration and baked potato chips (without frying) yielded also low acrylamide levels.

Keywords: acrylamide, potato chips, olive oil, LC‐MS‐MS, QuEChERS, ELISA,

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 148

The effect of chlorophyll pigments and squalene addition on refined olive oil oxidative stability

G. Rigane1, R. Ben Salem2

1Organic Chemistry‐Physics Laboratory UR11ES74, Sciences Faculty of Sfax, B.P « 1171 » 3000, Sfax, Sfax University, Tunisia. 2Chemistry‐Physics Department, Sciences and Technics Faculty of Sidi Bouzid, B.P « 380 » 9100, Sidi Bouzid, Kairouan University, Tunisia.

Refined olive oil has been analyzed in order to evaluate the influence of storage time on their quality after enrichment with chlorophyll pigments and squalene. At the end of the storage, 90% of chlorophyll pigments decomposed into their derivatives while the amount of carotene pigments diminished around 25% in all the oil samples stored at 20°C and 50°C. The reduction of total phenolic compounds exhibited similar degradation profiles, reducing by 81% for enriched refined olive oil stored at 20°C and 50°C after six months. For all studied samples, squalene content decreased significantly (p< 0.05) only after four months of storage. Oils with added pure ‐1 squalene had the lower peroxide values (52 meq O2.kg ) at the end of storage at 20 °C. After six months of storage the oxidative stability increased to 1h 28 min and 2 h 06 min for refined olive oil stored at 20 °C enriched with chlorophyll + squalene and only by squalene, respectively. Therefore, squalene supplemented with 800 mg.kg‐1 exhibited favorable antioxidant effects and is preferable for effectively avoiding oxidation. This study could provide useful information for industry to produce high‐stability vegetable oil.

Keywords: Refined olive oil, squalene, chlorophyll pigments, Chemlali, oxidative stability

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 149

Healthy and sensory properties in virgin olive oil: new approach to extraction process

A. Taticchi, S. Esposto, R. Selvaggini, S. Urbani, B. Sordini, M. Servili

Department of Agricultural, Food and Environmental Sciences, UNIPG, 06126 Perugia, Italy

The healthy values and the sensory properties of Virgin Olive Oil (VOO) are ascribed to minor components, in particular phenolic and volatile compounds, which are more affected than other substances by technological variables applied during mechanical extraction process. In fact, crushing and malaxation are directly involved in the releasing of phenols and in the forming of the aroma in VOO. The impact of a technological innovation on the structural modification of crushed paste and on the quality of the relative VOO has been extensively studied. The new approach consists in flash thermal conditioning (FTC) of the crushed paste, by its transit on a heat exchanger tube bundle followed by a very short malaxation. The assessment of olive paste structure modification by cryo‐scanning electron microscopy (cryo‐SEM) showed that the introduction of FTC after the crushing determines significant differences on the breaking of the parenchyma cells and the aggregation of oil droplets with respect to the paste obtained from a traditional crushing. In terms of VOO quality, the adoption of this new technology allowed a significant increase of the content of volatile compounds responsible for the olfactory positive attributes, whereas it had the effect of reducing significantly the percentage of the oil recovered and of the polyphenols content. However, a further short period of malaxation on the paste heat‐ treated can greatly reduce these negative aspects. This experiment, carried out on an industrial scale and on various cultivars, allowed to draw important considerations about the possibility of drastically reducing the malaxation time, improving the efficiency of industrial mill. So far, the optimal operative conditions (in terms of time and temperature) adopted during malaxation after the FTC treatment, can be properly chosen for improving the related VOO quality, according to the characteristics of olives such as the olives’ cultivar, sanitary state, or state of ripeness or the growing climatic conditions of the olive fruit.

Keywords: virgin olive oil, phenols, volatile compounds, flash thermal conditioning, healthy value and sensory properties

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 150

Comparison of olive oils from autochthonous and foreign cultivars of the region of Qom, Iran

M. Sharifani1, M. Asheri1, A.Yamchi1 and S. M. Hosseini Mazinani2

1Faculty of Plant Production, Gorgan University of Agricultural Sciences and Natural Resources, Gorgan, Iran2National Institute of Genetic Engineering and Biotechnology, Tehran, Iran

This study was aimed to compare olive oil quality and purity in ten olive cultivars grown in arid region of Qom, Iran. The cultivars were included; Mari, Roghany and Shangoh as autochthonous samples and the others were Arbequina, Mission, Beladi, Oblonga, Frantoi, Nociara and Koroneiki. The physicochemical variables were; peroxide value, fatty acid composition, chlorophyll, and carotenoid contents, K232 and K270 values were determined to identify the cultivar/s with high quality. Olive fruits were handpicked at the equal maturity stage in autumn 2012.The fruits were crushed, the paste was malaxed at 30˚C, and then centrifuged at 4000 rpm for 20 minutes. The extracted oils from fruits' mesocarps were collected and kept in dark glass jars in refrigerator. Peroxide values were measured according to the modified method of Garcia et al., (1996). Chlorophyll and carotenoid concentrations were measured by Minguez‐Mosquera method (1991). Fatty acid composition of the oils was analyzed by gas chromatography. Significant differences were observed in oleic acid percentage among the analyzed cultivars. The highest mean value of oleic acids were determined in oils from Mari and Koroneiki cultivars with 75.6 and 74.9%, respectively, while Beladi showed the lowest mean value of 54.6%. Among the samples, Beladi had the highest peroxide value (PV) (7.73), while Shangoh extracted oil had the lowest PV (6.4). Chlorophyll and carotenoid content ranged from 1.98 to 3.20 mg/kg and from 1.44 to 2.44 mg/kg oil, respectively. Oblonga and Roghany extracted oils owned the highest amount of chlorophyll pigments (4.5mg/kg and 4.3mg/kg oil, respectively) and carotenoids (3.75 mg/kg and 3mg/kg respectively). However, Shangoh extracted oil had the lowest amount of the pigments. The data presented indicate that Beladi as a table olive is the one cultivar with the lowest oil purity having the lowest oleic acid content. However, it contained the highest PV and pigments content, with the latter interfering with oxidation of the oil.

Keywords: fatty acid content, peroxide value, pigment contents, olive oil

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 151

The Evaluation of Turkish Virgin Olive Oil Fatty Acid Profiles By Chemometrics During Different Ripening Index and Storage Conditions

O. Koseoglu1, M. K. Unal2, D. Sevim1, D.ozdemir3

1Olive Research Station Izmir,Turkey 2Ege Universty, Faculty of Food Engineering, Izmir, Turkey 3Izmir Institute of Technology, Faculty of Science Department of Chemistry, Izmir,Turkey

In this study, the effect of growing area and ripening index of olives and also storage conditions of olive oils were examined. Extra virgin olive oils of Ayvalık and Memecik olive varieties harvested from different orchard in Ayvalık and Aydın region which have a significant potential for olive oil production in Turkey. Olives were harvested by hand at 2 different maturation index during two harvest years and processed by an Abencor system. The olive oils were divided in two parts in order to observe storage effect in daylight and in dark at room temperature for a period of 15 months. Analyses of fatty acid composition were carried out every 5 months (0, 5, 10 and 15 months). Multivariate classification and clustering were done by the application of unsupervised chemometric methods such as principal component analysis (PCA) and hierarchical cluster analysis (HCA) based on the fatty acid profiles of the olive oil samples. According to the results of PCA, the score plot of first (PC1) and second (PC2) principal component showed that samples were classified into two main group with respect to the harvest years. In addition, the samples were also successfully clustered into two sub‐groups according to cultivars (Ayvalık and Memecik). In terms of storage effect, there was no significant change on the fatty acid compositions among the samples during 15 months storage and also there was no significant change observed between the samples stored in dark and in daylight conditions. PCA results indicated that the variables C18:0, C20:0, C22:0, C24:0 and C18:1, C20:1, MUFA played an important role in the characterization of the olive oil samples obtained in 2009 harvest years, Ayvalık and Memecik, respectively. In addition, olive oils were essentially characterized by fatty acid such as variables C16:0, C16:1, C17:0, C17:1, C18:3T, C18:2T+C18:3T, SFA and C14:0, C18:2, C18:2T, C18:3, PUFA obtained in 2010 harvest years, Ayvalık and Memecik, respectively. In summary, it was showed that the applications of PCA and HCA to fatty acid profiles are successfull for the classification clustering virgin olive oil samples based on varieties and harvest years.

Keywords: Turkey, virgin olive oil, fatty acid profiles, chemometrics, PCA, HCA

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 152

Chemical composition and antibacterial activity of two different oils of Olea europea L.

A. Chaker1, H. Boukhebti1, M. Ramdhani1, S. Dahamna.2

1Laboratoire de Valorisation des ressources biologiques naturelles. Département d'Ecologie et Biologie Végétale, Faculté des Sciences de la Nature et de la vie, Université Sétif1, Alegria 2Laboratoire de Phytothérapie Appliquées aux Maladies Chroniques

The essential oil obtained by hydro distillation of the aerial parts “stems and leaves” of olive tree (Olea europea L.), was analyzed by gas chromatography coupled with mass spectrometry (GC/MS). The olive oil of fruits which was obtained by traditional way of pressing, were also analyzed by CGMS after the methylation process. Thirty eight compounds, representing 84.91% of the total essential oil of Olea europea L. were identified, Hexadecanoic and Linoleic acid are the major compounds; while sixteen compounds representing 99.98% of the total olive oil were identified, where the major compounds are Oleic, Palmitic and Linoleic acids. The antibacterial activity was tested by using the agar diffusion test. The essential oil of Olea europea L. showed modest effect against Escherichia coli ATCC 25922, Staphylococcus aureus ATCC25923and Pseudomonas aeruginosa ATCC27853, while olive oil has no effect against these bacteria strains.

Key words: Olea europea L., antibacterial activity, chemical analysis, olive oils, essential oils

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 153

Steryl glucosides composition in monovarietal extra virgin olive oils

R. Cruz‐Hidalgo1, R. B. Gómez‐Coca1, G. D. Fernandes1,2, M. C. Pérez‐Camino1, W. Moreda1

1Department of Characterization and Quality of Lipids, INSTITUTO DE LA GRASA –CSIC‐, Avda. Padre García Tejero 4, E‐41012‐Sevilla, Spain. 2Fats and Oils Laboratory, Department of Food Technology, Faculty of Food Engineering, University of Campinas, Campinas‐São Paulo, 13083‐970, Brazil.

Plant glycolipids consist mainly of steryl glucosides, sphingoglycolipids, and glyceroglycolipids. Those steryl glucosides can be found either as free molecules or as esterified compounds. Free steryl glucosides (SG) are phytosterol conjugates where one glucose moiety binds at the C3 position of the sterol residue via an acetyl bond. When this kind of sterol is esterified to a fatty acid at the C6 position of the sugar moiety, it gives rise to esterified (also named as acylated) steryl glucosides (ESG). The richest natural sources of dietary phytosterols are edible vegetable oils, like those from corn, palm, rapeseed or sunflower, relatively little information is available regarding their glycosylated sterol content. Published data on the content and sterol composition of glycosylated sterols in more than forty plant matrices shows the potential contribution of different foods as sources of steryl glucosides when included in the diet. Although, there are data on the content of SG in many vegetable oils, there is scarce information about these compounds in olive oils. Mainly due to the inexistence of a method, Gómez‐Coca et al.1 developed a procedure to specifically analyse SG and ESG in olive oil. The next step was to determine the concentration range, together with a thorough study of the profile, since up to now just ‐ sitosterol derivatives have been identified in olive oils. Besides the influence of the cultivar in the profile and content has also been studied. In order to have control of the traceability of each variety, olive fruits were harvested from a variety garden and processed using an Abencor® system. The oils were analysed using the method already published. Additionally, determinations of other minor compounds were performed in order to seek for a relationship between them and the content of sterol derivatives.

Keywords: olive oils, sterol derivatives, steryl glucosides

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 154

Changes in Virgin Olive Oil Quality Characteristics Due to Heating in Comparison with Vegetable Oils

A. Yorulmaz

Department of Food Engineering, Faculty of Engineering, Adnan Menderes University, Aydın, Turkey

The aim of the study was to determine effect of various heating techniques on olive oil quality and fatty acid composition in comparison with other vegetable oils. For this purpose; extra virgin olive oil, hazelnut, canola and corn oil samples were heated in microwave, conventional and hybrid ovens for six different periods. Heated oil samples were evaluated for free fatty acid content, peroxide value, photometric color index, K232 ‐ K270 values and fatty acid distribution. Results have shown that hybrid ovens which have combined conventional and halogen effect, enabled faster heating till 130°C but have slower heating effect at higher temperatures, when compared to conventional heating. Microwave, hybrid and conventional heating did not influence the free fatty acid content of olive oil, while the other vegetable oils’ either rose or fell during heating. Peroxide value of olive oil was constant till 129 ̊C and then decreased as the result of hydroperoxide formation and degredation whereas corn, canola and hazelnut oils’ peroxide values increased till 173 ̊C and decreased afterwards. K232 values of olive oil increased comparative with the ascending oven temperatures, while K270 values increased in low amounts. The similar trend were also observed for other vegetable oils. Photometric color index of olive oil decreased till 70 °C and increased then during heating. Oleic and palmitic acids were the main fatty acids of extra virgin olive oil. Significant independent correlations between heating temperature and fatty acid composition, oleic/palmitic acid ratio, linoleic/palmitic acid ratio were not observed for virgin olive oil and remaining vegetable oils.

Key words: Fatty acid, hybrid oven, quality, convection, microwave, olive oil

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 155

Preliminary results of olive zoning study in the province of Sassari: the peculiarities of cv Bosana oils produced in different growing areas

L. Morrone1, A. Rotondi1,N. Di Virgilio1,B. Alfei2, C. Cantini3,E. Vagnoni4,P. Duce4

1Institute of Biometeorology, IBIMET‐CNR, Bologna, Italy 2Agenzia per i Servizi nel Settore Agroalimentare della regione Marche, ASSAM, Ancona, Italy 3Institute of Trees and Timber, IVALSA‐CNR, Firenze, Italy 4Institute of Biometeorology, IBIMET‐CNR, Sassari, Italy

In Sardinia olive growing is widespread to almost all municipalities and it covers primary importance to the agricultural economy of the area for both landscape and land conservation. The aim of this work is to study the territory of the province of Sassari analyzing the correlations between pedo‐climatic conditions and chemical and sensory characteristic of cv Bosana monovarietal olive oil, one of the most widespread cultivar of Sardinia. As a first step of the experimental plan a morpho‐pedological land characterization was performed using a Territorial Information System (GIS‐Geographical Information System. From the maps found in the Geo‐ portal of the Region of Sardinia has been developed a Digital Elevation Model (DEM) from which morphometric maps of altitude, slope and orientation of territory studied were processed. A geological and soil characterization was made using the Sardinia Region’s maps. As a second step chemical characterization, in terms of quality indices contained in the Regulation EC 2568/91, 61/2011, 299/2013 of the Commission of the European Union, fatty acids profile, pigment profile, total phenol content, phenolic profile and sensory characterization of cv. Bosana olive oil, performed by the “ASSAM – Marche panel” a fully‐trained analytical taste panel, were carried out during two consecutive crop seasons. The province of Sassari has been divided in three macro areas (Alghero, Sassari and Ittiri) characterized by different environmental conditions and in each of the macro areas three olive groves has been selected in which to perform the harvest. Olive fruits were manually harvested when 50% achieved the black stage, in order to have a similar maturity index in each olive grove. The olive oil production was carried out using a low scale mill in order to standardize all the technological parameters during the extraction. The data obtained were analyzed using a full factorial design to investigate the effect of the geographical origin and of the seasonality on the olive oil characteristics The different content in some fatty acids observed in the oil samples is attributable exclusively to the production area without any influence of seasonality, while total phenol content, pigment, phenolic and organoleptic profiles are not able to discriminate the production area.

Keywords: olive zoning, fatty acids, GIS, olive oil, seasonality.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 156

Virgin olive oil carotenoid fraction improvement by addition of microalgae extracts of Scenedesmus almeriensis

P. Limon1, R. Malheiro2,3, G. Acién‐Fernández4, J. M. Fernández‐Sevilla4, S. Casal3, N. Rodrigues2, R. Cruz3, R. Bermejo1, J. A. Pereira2

1 Department of Physical and Analytical Chemistry, Jaén University, Linares High Polytechnic School (EPSL), 23700 Linares, Spain 2 Mountain Research Centre (CIMO), School of Agriculture, Polytechnic Institute of Bragança, Campus Stª Apolónia, Apartado 1172, 5301‐855, Bragança, Portugal 3. REQUIMTE/Laboratory of Bromatology and Hydrology, Faculty of Pharmacy, Porto University, Rua Aníbal Cunha 164, 4050‐047 Porto, Portugal 4 Departement of Chemical Engineering, Almeria University, E‐04071, Almeria, Spain.

Carotenoids intake in daily diet is insufficient to reach the recommendable dairy dose, since fruits and vegetables consumption is far below the recommended as well. The remaining necessary carotenoids fraction is compensated by supplements. In a contradictory tendency to fruits and vegetables consumption is olive oil. The consumption of this premium vegetable oil is increasing steadily through the years. Virgin olive oils (VOO) possess carotenoids in good amount, but even though not enough for daily recommendations. In this context, the present work intended to add different concentrations (0, 0.1 and 0.21 mg mL‐1) of an extract rich in carotenoids from the microalgae Scenedesmus almeriensis. Five commercial EVOO’s (extra‐virgin olive oils) were selected for the experiment, being added the different concentrations of S. almeriensis extract. Color evaluation (a*, b* L*, YI and ΔE), quality parameters (free acidity, peroxide value, UV analysis, ρ‐anisidine value), chemical composition (fatty acids, tocopherols, and pigments) and oxidative stability were studied. Olive oils color was affected, becoming olive oils more reddish‐ yellow according to the concentration of extract. Quality was improved since peroxidation was inhibited in EVOO with extracts. Levels of free fatty acids wasn’t affected by S. almeriensis extracts addition, as well as EVOO’s chemical composition (fatty acids profile and tocopherols). Pigments concentration (lutein and β‐carotene) was considerably improved, mainly β‐carotene content, being responsible for EVOO’s color changes. Oxidative stability in some cases was improved about 40%, having repercussions in oil stability and shelf‐life. Overall, the behavior observed was similar in the five EVOO’s studied. The results obtained also allowed to consider that the concentration of 0.1 mg mL‐1 of S. almeriensis extract give similar results to those obtained with 0.21 mg mL‐1. The addition of S. almeriensis extracts to EVOO is a good strategy to counteract peroxidation, increasing the nutritional value and bioactive constituents of olive oil, being also an excellent alternative to carotenoids supplements in human diet.

Keywords: Virgin olive oil; Scenedesmus almeriensis; quality; composition; oxidative stability; oxidation.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 157

Effect of olive oil on lipid profile in female climacteric ‐ Research project in progress

F. A. conte1, L. B. Franz2, O. B. Oliveira3

1Subproejct of the Research: Multidimensional Research on Women’s Aging Process and TCC (Course Conclusion Essay) / Nutrition / Unijuí. 2Fellow PIBIC / CNPq; 3. Professor at Unijuí/Advisor/ PhD in Public Health (FSP / USP). 3Nephrologist at Charity Hospital, Ijuí.

The female aging is characterized by many changes, being the hypoestrogenism one of the most important, which among other causes interfere negatively on metabolism and lipid profile. The improvement of the lipid profile happens through the modification of feeding habits as the reduction of animal fat and trans fat (margarine) intake, increased consumption of polyunsaturated fats (soy, corn, sunflower) and mainly monounsaturated extra virgin olive oil and canola oil, which have positive effects on HDL cholesterol. The extra virgin olive oil acts preventing the oxidation of LDL cholesterol, since it presents antioxidant compounds such as vitamin E and its phenolic compounds; also prevents the formation of thrombi, has a positive effect on the prevention of breast, colon, rectum cancers, and assist in reducing LDL cholesterol and increasing HDL‐cholesterol. This study aims to determine the effect of olive oil on the lipid profile in dyslipidemic climacterical women in the city of Ijuí /RS/Brazil. Intervention research conducted with 20 climacterical women (35‐65 years old), linked to the Family Health Strategies VII and VIII of the city. Criteria to participate in the survey: have some degree of dyslipidemia, no use of lipid‐lowering medications and sign the Instrument of Consent. These women are assigned to two groups: case and control, both with 10 women, all attending to monthly workshops of Food and Nutrition Education. During three months the case group is consuming 2 tablespoons of extra virgin olive oil daily, and after this period the participants will be switching group, and those who were not consuming will use olive oil also during three months, and who were consuming will only participate in the workshops. The lipid profile is being accompanied by laboratory tests: total cholesterol; LDL cholesterol and HDL cholesterol, in three different moments. The first will be before the intervention, the second will be at the end of the first 3 months, and the third at the end of the survey, after six months. This project was approved by the UNIJUÍ’s Ethics Committee, embodied in the document number 294 456.

Keywords: olive oil; climacteric; dyslipidemia; hypoestrogenism.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 158

Effect of Olea Europaea L. folium ethanolic extract in rats submitted to LPS

M. Ramos¹, O. B. Oliveira²,C. Rasvickas³, A. Pereira4 , N. Schor5 .

1Department of Nephrology, UNIFESP, SP, Brazil 2Department of Nephrology, HCI, Ijui, Brazil 3Department of Nephrology, UNIFESP, SP, Brazil 4Department of Agronomy, UNAERP, SP, Brazil 5Department of Nephrology, UNIFESP, SP, Brazil

Sepsis and septic shock are characterized by a systemic inflammatory response accompanied by arterial hypotension, arterial vasodilatation, decreased vasopressor sensitivity and may progress to multiple organ failure, acute renal failure with high mortality rates. The systemic inflammatory response of sepsis develops a significant production of reactive oxygen and nitrogen species, leading to oxidative stress with an imbalance of antioxidants and pro‐antioxidant components forming a system that consists of many substances as the glutathione, which is considered as the major intracellular antioxidant compound. It is believed that there in an intake of antioxidants in cases of severe sepsis with multiple organ failure. There are reports in the literature that olive leaves has a considerable antioxidant effect and has a potential protective effect in individuals in sepsis. In the present study we evaluate the effect of ethanolic extract of olive leaves in a sepsis in rats submitted to LPS, which resulted in a potential protective effect to the systemic inflammatory response on sepsis.

Keywords: sepsis, olive leaves extract, septic shock

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 159

Nutritional value and health benefits of table olives

F. Gungor, A. Yildırım

Olive Research Station, Izmir, Turkey

Table olives are the main fermented foods in Mediterranean countries and constitute an important part of the Mediterranean diet and also the diet of many non‐olive‐producing countries. Due to consumers’ increasingly demand to incorporate foods into their diets that can help boost or maintain health, as well as help prevent some diseases, interest in table are increased resently. Table olive is a healthy food with high nutritional value consist primarily of water, fat and carbohydrates. Olives are an excellent source of oleic acid, also known as omega 9 or monounsaturated fatty acids (MUFA). And they are a good source of fiber, amino acids, the potent antioxidant vitamin E, iron, copper and other minerals. Moreover, olives are a rich dietary source of phenolic compounds that are linked to powerful antiinflammatory, antibiotic, antimicrobial, antiviral and antinociceptive effects. The disease‐preventing effect of table olives is mainly attributed to its unique fatty acid profile and to the presence of some minor components such as tocopherols, carotenoids, phospholipids, triterpenic acids and biophenols. As part of a well‐balanced, nutritious diet olives and olive products also provide added protection against many chronic diseases. Olive benefits have been demonstrated for the cardiovascular system, respiratory system, nervous system, musculoskeletal system, immune system, inflammatory system, and digestive system. In this review, it was focused on the active compounds present in table olives and also the contribution of these compounds to the human health was examined in detail.

Keywords: Table olive, health, nutritional value, active compounds

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 160

Comparison of single strain starter culture and a selected inoculum enrichment in the processing of natural table olives

M. Campus1, P. Sedda1, E. Cauli1, R. Comunian2, A. Paba2, E. Daga2, S. Schirru2, R. Zurru1, G. Bandino1

1DIRARB ‐ Department of wood and fruit tree, Agricultural Research Agency of Sardinia (AGRIS Sardegna), S.S. 291, Loc. Bonassai, Sassari, Italy 2DIRPA – Animal Production Research Department, Agricultural Research Agency of Sardinia (AGRIS Sardegna), S.S. 291, Loc. Bonassai, Sassari, Italy

The processing of natural table olives is the result of complex biochemical interactions between the indigenous microflora and the olives. This empirical process has drawbacks and is far to be controlled. The control of the fermentation processes is carried out through chemical, chemico‐ physical and microbiological approaches. The use of selected microorganisms to drive the fermentation, able to supplant the indigenous microflora and, in particular, the adventitious microflora responsible for spoilage, has been thoroughly investigated. Single or dual strain starter cultures have been developed and used, while more complex mixes of lactic acid bacteria (LAB) cultures have not yet been studied. In the present study, taking natural fermentation (NF) as control, the performances of a single strain LAB starter culture (SSL) and a selected inoculum enrichment (SIE), coming from previous successful fermentations, were compared. All trials were performed in triplicate. The results showed that the SIE was more efficient in controlling spoiling microflora (Enterobacteriaceae spp.) than the single strain starter. Indeed, Enterobacteriaceae were not detected in 10‐days SIE samples, while they were still detected in 16‐days SSL and NFS samples (1.08x104 and 1.80x103 CFU/ml, respectively). Both starters rapidly acidified the brine, lowering the pH to safety levels (<4.3), reaching pH 3,8 after 12 days, while NF samples reached pH 4,3 after 45 days and this value was kept constant up to 153 days. Moreover, both showed debittering activity, as stated through sensory analyses. Texture profile analysis (TPA) showed that olives processed with SIE inoculum were firmer and more elastic, compared to SSL inoculated olives, thus resulting more similar to NF samples. One out of the three batches inoculated with a single strain culture failed to rapidly acidify the brine, probably due to phage infection, and showed no differences with NF profiles. Indeed, this is a major obstacle in the use of single starter cultures as inoculum, while complex starter cultures, such as SIE, made up of an undefined number of strains are less sensitive to phages. In fact, different strains can sustain an attack by a phage which, having a strain‐specific action, selects the resistant strains. Furthermore, indigenous complex microflora, as a result of the adaptation to local environments, constitutes the basic genetic resource to make a product typical and not reproducible in other areas than in that of production, assuring the preservation of the local microbial biodiversity.

Keywords: Olea europaea, selected starter cultures, undefined starter cultures, LAB

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 161

Natural fermented table olives from Trás‐os‐Montes region as a source of bioactive compounds with antioxidant and antimicrobial properties

R. Malheiro1,2, P. Mendes1, A. Bento1, J. A. Pereira1

1Mountain Research Centre (CIMO), School of Agriculture, Polytechnic Institute of Bragança, Campus Stª Apolónia, Apartado 1172, 5301‐855, Bragança, Portugal 2REQUIMTE/Laboratory of Bromatology and Hydrology, Faculty of Pharmacy, Porto University, Rua Aníbal Cunha 164, 4050‐047 Porto, Portugal

Table olives are a food fermented product recognized for their rich content in bioactive compounds, namely phenolic compounds. Among the most representative types of tables olives worldwide, natural fermented table olives present higher phenolic content, related with their technological process, conferring to these products high added value due to their bioactive potential. In the present work, our objective was to assess the phenolic composition and bioactivity of natural fermented table olives produced in Trás‐os‐Montes region, one of the most important olives producing regions in Portugal. For that, 24 samples of natural fermented table olives were collected from local producers and their phenolic composition was assessed (HPLC/DAD) as well as their antioxidant activity (DPPH and reducing power methods) and antimicrobial activity (against Bacillus subtilis, B. cereus, Escherichia coli, Staphylococcus epidermis, Staphylococcus aureus, and Pseudomonas aeruginosa). Ten phenolic compounds were identified and quantified in the 24 table olives samples, being hydroxytyrosol, a derivate from verbascoside and hydroxytyrosol glycol the most abundant phenols. The extracts contained from 2.4 to 64.2 µg of phenolic compounds per mg of extract. Samples reported strong antioxidant activity in both methods tested and significant correlations were established between phenolic composition and antioxidant activity. Table olives extracts reported antimicrobial activity mainly against Gram‐negative pathogenic bacteria. Extracts rich in phenols reported higher inhibition in bacterial growth. Chemometrics, by a principal component analysis, showed that samples can be grouped according to their phenolic composition and related bioactivity. Natural fermented table olives from Trás‐os‐Montes region revealed to be rich in phenolic compounds with antioxidant and antimicrobial potential.

Keywords: natural fermented table olives; phenolic compounds; antioxidant activity; antimicrobial activity OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 162

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 163

OLIVE MILL WASTE MANAGEMENT

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 164

Alternative technologies for olive mill wastewater management with emphasis on soil application

K. Chartzoulakis1, G. Psarras1, N. Kalogerakis2, F. Santori3

1ELGO‐Institute for Olive Tree and Subtropical Plants, 73100 Chania, Crete, Greece 2Dept of Environmental Engineering, Technical University of Crete, 73100 Chania Greece 3ISRIM SC ar.l., 05100 Terni, Italy

The disposal of olive mill wastewater (OMW) produced during oil extraction in Mediterranean countries creates a significant environmental problem due to great volumes of effluent produced (10‐12 Mm3/year) and heavy pollution load (40–80 g/l BOD, 50–150 g/l COD) and phytotoxic properties (due to phenolic compounds). During the last 30 years many OMW treatment methods have been proposed and tested, but their application at olive mill level is limited due to high investment and/or running cost and technical expertise required. Within the LIFE+ OLEICO+ project have been identified and evaluated existing technologies ffollowing certain criteria that are currently being used in Portugal, Spain, Italy and Greece. The results show that several viable technologies for OMW treatment do exist (composting, electro‐coagulation, hydrolysis‐ oxidation, phyto‐remediation, co‐digestion, energy production); however, they require capital investment and maintenance costs which cannot be afforded by a small or medium olive mill plants. On the other hand, application of OMW to olive orchards can be a low cost alternative method for OMW treatment, in regions with small size olive oil mill enterprises. Annual rates up to 1,500 l/tree were applied in five equal doses, at 20‐days intervals between November and February. Soil analyses were performed before the onset and after the end of OMW application period. The response of olive trees to OMW application was monitored by measuring plant nutritional status, photosynthesis and yield. Furthermore, the possibility of groundwater pollution by the application of fresh OMW was investigated in lysimeters. OMW‐treated soil had higher K content throughout the experiment. Phenols were decomposed rapidly, and therefore the phenolic content before the onset of a new OMW application period was negligible. The nutritional status, the physiology and yield of olive trees were not affected by the application of OMW. The total olive orchard area required for the annual OMW production is 3.6 ha (280 trees per ha), which is easily available around an olive mill. The cost of application is 0.007 €/lt of OMW, seems reasonable compared with sophisticated methods. Detailed study for each case to determine the application dose according to soil and climatic conditions of the area is required.

Keywords: Composting; aerobic‐anaerobic; hydrolysis; oxidation; phyto‐remediation; energy production; land application.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 165

Agronomic and environmental valorization of olive mill wastewaters

A. Abichou1, L. Mohamed2

1Institut de l’Olivier Zarzis 4170, Tunisia 2Institut des Régions Arides 4119 Médenine, Tunisia

The quantities of the OMWW that accumulate from one year to the next constitute a real environmental problem because, in the more part of case, the physicals and chemicals treatments are limited by some technical and economic requirements. On the other hand, in the Tunisian arid zones, where the soils are sandy and very poor in organic and mineral matters, wind erosion process is very active and triggered the departure of the thinnest elements of the soil and the apparition of dune building. Facing to these situations, many simple and efficient practices for combating desertification had been tested since many years in the Tunisian arid zones. The mulching of the olive mill wastewaters is one of these techniques. Otherwise, an olive mill wastewaters spray experimental device, with these respective doses of 50 m3/ha, 100 m3/ha and 200 m3/ha next to a witness parcel without OMW, has been putted in place since 1995 in the Chammakh –Zarzis, Tunisia ( the size of different parcel is one hectare). The mulching of the OMW in the southern Tunisian regions constitutes an interesting alternative for the controlled and rational evacuation of this polluting sewage and for the soil cohesion improvement by : (i) the increase of the organic matter rate which raised, in proportion to the dose, from 0.06% to 1.67% after 10 years of OMW mulching, and (ii) the improvement of the soil structural stability or “the mean weight diameter” where the aggregation rate superior to 2 mm is increased until 34 % with the dose of 200 m3/ha. Therefore, that treatment will bind the sand particles and thereof control of sandstorms is gained. Indeed, the results in wind tunnel tests on the soil treated with these different doses of OMW showed that the threshold friction velocity u*t was raised from 8.5 m/s to 12 m/s for 50 m3/ha and 200 m3/ha, respectively. The obtained result revealed that OMW is an interesting soil fertilizer that can be used in sandy soils to improve not only its physical and chemical characteristics but also its biological activity.

Key words: olive mill wastewater, organic matter, wind erosion, south Tunisia

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 166

Impact of olive mill wastewater treated by different techniques on seed germination of Barley

M. Rusan, A. Balasmeh, S. Zuraiqi, M. Bashabsheh

Department of Natural Resources and Environment; Faculty of Agriculture; Jordan University of Science and Technology (JUST); PO Box 3030.

Olive‐mill wastewater (OMW), an effluent of olive oil extraction process, is annually produced in huge amounts in olive growing areas. Olive mill wastewater (OMW) is a by‐product effluent of olive oil extraction process, that is produced in large amount in the Mediterranean region. OMW is believed to induce phytotoxic effect on organisms including seed germination and plant growth. The objective of this study was to evaluate the impact of untreated and treated with different technologies OMW on seed germination of barley (Hordeum vulgare). The following treatments were investigated: 1) Potable water (Control); 2) Untreated OMW (UOMW); 3) OMW treated by aerobic biological technology JR; 4) OMW treated by solar fenton oxidation (SFO); 5) OMW treated by an integrated microfiltration/reverse osmosis (MF+RO) process; 6) OMW treated by microfiltration followed by nanofiltration (MF+NF); 7) Untreated OMW; 8) Diluted OMW by 75%; 9) Diluted OMW by 50%; and 10) Diluted OMW by 25%. A germination test was conducted in an incubator at temperature of 23 C. In each petri dish a filter paper was mounted and ten seeds of barley were placed on the filter paper. Five mls of water were added to each dish. With time, the seed germination was determined by counting the number of germinated seeds to calculate the percentage of germination. Then the fresh and dry weights of the shoots and the roots of the germinated seeds were recorded. The results show that untreated OMW and the 25% and 50% diluted OMW were very phytotoxic to barley seeds and completely prohibited seed germination. However, phytotoxicity decreased significantly following treatments of OMW with all technologies investigated and by 75% dilution, as results of removing the phenols and other phytotoxic organic compounds from the OMW. It was concluded that if OMW will be used for irrigating crops, it has to be first treated or diluted by 75% at least.

Keywords: Olive mill wastewater, seed germination phytotoxicity, barley.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 167

The different points of view of olive mill wastewater in Turkey: A result of New Ecological Paradigm

R. Tunalioglu1, R.Yildirim2

1Dr. Adnan Menderes University, Faculty of Agriculture, Department of Agricultural Economics, Aydın, Turkey 2Adnan Menderes University, Faculty of Agriculture, Department of Agricultural Economics, Student of Master Degree, Aydın, Turkey

Turkey takes part among the important olive and olive oil producer countries in the world. Olive and also olive oil production has increased in Turkey during the recent years, partially, because the government have support to producers substantially for certified plant from 2000. This resulted in increasing the olive production both quantitatively and qualitatively. On the other hand, increasing the quantity of the Olive Mill Wastewater (OMWW) as a byproduct of olive oil extraction, resulted in negative environmental impact on the environment. Currently, usually OMWW which discharges directly to the nature (lake, river, soil…etc.), pose a threat to the environmental especially in dry years. As a matter of fact, the Ministry of Environmental and Urban Planning offer several solutions and suggestions to olive oil firms to prevent discharging the OMWW and enforce penalties to those who do not comply with those solutions. Lack of evaluation in economical terms and the high cost of clarification of olive mill wastewater in Turkey unfortunately cause management difficulties. Aydın is one of the most significant province of Turkey, because of its olive and olive oil production potential. Thus, this problem also concerns Aydın province. In this study, interviews have been conducted with almost 50% of the olive oil business managers of Aydın. These olive oil business management’s ecological sensitivities, how they comprehend olive mill wastewater and their general ecological attitudes has been analyzed with using the New Ecological Paradigm Methodology. In addition, as a result of this research, it has been identified business manager’s level of general ecological sensitivity is mediocre (3,52).

Keywords: Olive mill wastwater, new ecological paradigm, Turkey.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 168

Valorization of olive husk into valuable organic amendment: does initial C/N ratio affect the quality of the produced compost?

K. Azim1 , S. Roussos2 , C. Périssol2, I. Thami Alami3 , B. Soudi4

1Research Unit of Integrated Crop Production, INRA‐Agadir CP. 80350, Morocco 2Aix Marseille Université, CNRS, IRD, Avignon Université, IMBE UMR 7263, 13397 Marseille, France 3Laboratory of Microbiology, INRA‐Rabat, 10101, Morocco 4Department of Environment and Natural Resources. IAV Hassan II‐Rabat. 10101, Morocco

Huge amounts of by‐products are generated per year both by the olive oil industry and the olive tree cultivation. Thirty millions m3 of olive mill wastewater and twenty million tons of solid waste called olive husk are generated per year by the olive oil mills. Current environmental requirements require treatment of the olive mill wastes. This action will help to minimize the spread of olive mill wastes to the surrounding environment. Different experiments have studied the effect of composting of the olive husk and the quality of the produced compost on soil fertility and crop performance. The objective of this study was to evaluate the effect of initial Carbon to Nitrogen (C/N) ratio on the quality of the produced compost from olive mill husk and other organic waste (tomato leaves and stems, melon end cycle plants and sheep manure). The four studied initial C/N ratios (treatments) were: C/N=25; 30; 35 and 40, which contain respectively 5; 20; 20 and 15% (w/w) of olive mill husks. Results showed a difference of behavior at the level of the monitoring parameters such as temperatures that were higher and more stable for the mixture of C/N=35. Similarly, the relative humidity recorded was too high and stable compared to other mixtures of other C/N ratios. The Cresson test showed a high percentage of germination (86%) for the produced compost of initial C/N=35 that contains the highest percentage of olive mill husk (20% w/w). This result was confirmed by the phytotoxicity test, which also showed a decrease in the percentage of germination by increasing the concentration of the compost water extract. The results of physicochemical analysis and phytotoxicity showed that the compost of initial C/N=35 ratio is the most stable and the less phytotoxic compared to other composts.

Keywords: olive mill husk, horticultural wastes, composting, monitoring, compost quality.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 169

Coagulation/flocculation followed by solar Fenton oxidation at a pilot scale for the purification of olive mill wastewater

I. Michael1, A. Panagi1, L. A. Ioannou1, Z. Frontistis1, D. Fatta‐Kassinos1

1Nireas‐International Water Research Centre, School of Engineering, University of Cyprus, P.O. Box 20537, 1678 Nicosia, Cyprus

This study investigated the application of a solar‐driven advanced oxidation process (solar Fenton) combined with previous coagulation/flocculation, for the treatment of olive mill wastewater (OMW) at a pilot scale. The pre‐treatment of OMW was performed using FeSO4.7H2O (6.67 g L‐1) and an anionic polyelectrolyte (FLOCAN 23, 0.287 g L‐1). The aim of the pre‐treatment was to remove the solid fraction of OMW, in order to improve the efficiency of the subsequent treatment by solar Fenton that was later on applied. The solar Fenton experiments were carried out in a compound parabolic collector (CPC) pilot plant, in the presence of varying doses of H2O2 and Fe2+. The optimization of the oxidation process using reagents at low concentrations 2+ ‐1 ‐1 ([Fe ]=0.08 g L ; [H2O2]=1 g L ), resulted in high COD removal (87%), while the polyphenolic fraction, which is responsible for the biorecalcitrant and/or toxic properties of OMW, was eliminated. This study demonstrated also the efficacy of solar Fenton to operate at the inherent pH of the OMW, yielding only a slightly lower organic removal (80%), compared to that obtained under acidic conditions. Under both operating pH conditions studied, mineralization in terms of DOC removal, was found to be relatively low compared to the COD removal, a fact that clearly indicates that a considerable organic load remained in the treated OMW solution, which could be attributed to the presence of stable oxidation products. A kinetic study using a modified pseudo first‐order kinetic model was performed, in order to determine the reaction rate constants. The modified kinetic equation indicated the occurrence of two sequential oxidation steps during the solar Fenton process where, on a first stage, the COD is converted quickly to intermediates, which undergo further slower degradation during a second stage into final oxidation products. Moreover, solar Fenton was found to be efficient in reducing the initial OMW toxicity against the examined plant species (Sorghum saccharatum, Lepidium sativum, Sinapis alba), and the water flea Daphnia magna. The OMW treated samples displayed a varying toxicity profile for each type of organism and plant examined in this study, a fact that can potentially be attributed to the varying oxidation products formed during the process applied. Finally, the overall cost of solar Fenton oxidation for the treatment of 50 m3 of OMW per day was estimated to be approx. 2.1 € m‐3.

Keywords: coagulation, flocculation, olive mill wastewater, pilot scale, solar Fenton, toxicity

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 170

Production of extracellular lignocellulolytic enzyme by Streptomyces Sp grown on olive pomace

L. Medouni‐Haroune1,3, M. Kecha1, F. Zaidi 2, S. Roussos3, V. Desseau4

1Applied Microbiology Laboratory University of Bejaia, Algeria 2 Food Biochemistry Laboratory University of Bejaia, Algeria 3Aix Marseille University, CNRS, IRD, Avignon University, IMBE UMR‐237, Ecotechnology & Bioremediation Team, Faculty of Sciences of St Jérôme ; 13397 Marseille cedex 20, France 4Aix Marseille University, CNRS, ISM2 UMR, Faculty of Sciences of St Jérôme ; 13397 Marseille cedex 20, France

Actinomycetes constitute an economically important class of bacteria for their ability to produce degradative enzymes and secondary metabolites. Ninety actinomycetes strains were isolated from diverse environmental sources of Algeria. They were screened for their cellulolytic and xylanolytic activities on carboxymethyl cellulose (CMC) and xylan‐ containing agar plates. Streptomyces sp. S1MIII showed the highest carboxymethyl cellulase (CMCase) and xylanase activity. The production of extracellular lignocellulose‐degrading enzymes, xylanase and CMC‐ase during the growth of Streptomyces sp S1MIII in olive pomace medium was studied. A comparison of enzyme activities revealed that the activity of xylanase (33Uml‐1) was greater than those of CMC‐ase activity (4.13 Uml‐1).

Keywords: Actinomycete, Streptomyces, Olive pomace, enzymes, cellulase, xylanase, enzymatic essay.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 171

Olive mill wastewater agronomic application on olive tree (Olea europaea L.) in the South of Tunisia: Some biochemical and nutritional traits

S. Magdich 1, 2, 3, C. Ben Ahmed 1, 3, B. Ben Rouina 1 and E. Ammar 2

1Laboratoire d’Amélioration de la Productivité Oléicole et des Arbres Fruitiers, Institut de l’Olivier de Sfax, B.P. 1087 ‐ 3000 Sfax, Tunisia 2UR Etude et Gestion des Environnements Urbain et Côtier, Université de Sfax, Ecole Nationale d’Ingénieurs de Sfax, B.P. 1173 ‐ 3038 Sfax, Tunisia 3Laboratoire d’Ecologie Végétale, Faculté des Sciences de Sfax, B.P. 1082 ‐ 3018 Sfax, Tunisia

The purpose of this study was to investigate olive mill wastewater (OMW) spreading effects on some biochemical (pigments, soluble sugars and total phenols contents) and nutritional status characteristics in leaves of adult olive tree (Olea europaea L. cv. Chemlali). In this study, four olive orchards were subjected over eight successive years (2004‐2012) to OMW treatments (0, 50, 100 and 200 m3/ha/year). The experimental orchard was constituted by a sandy soil (86.63% sand, 13.26% silt and 0.20% clay). Total chlorophll and carotenoid concentrations did not present significant differences between the three OMW‐levels if compared to the control. Total phenols and soluble sugars contents showed a decrease with OMW‐applied doses. Indeed, the lowest content of total phenols leaves was observed in soil treated with 200 m3 OMW/ha, with a decrease percent equal to 6%. In contrast, application of OMW was associated with the increase in potassium, sodium and chloride contents in comparison to the control. For the different nutrients elements, the contents increment is in function of the OMW applied level. The three doses (50, 100 and 200 m3/ha/year) at which OMW was assayed were noted to induce negatively effects on biochemical and nutritional status characteristics of olive tree.

Keywords: Olive mill wastewater (OMW), spreading, olive tree, biochemical parameters, nutritional status.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 172

Influence of olive mill wastewater applied at different quantities on an alkaline soil fertility

R. Razouk1, K. Oubella2

1National Institute for Agricultural Research, P.O. Box 578, Meknès, Morocco 2Department of Biology, UMI, P.O. Box 4010, Meknès, Morocco

The trials regarding use of olive mill wastewaters (OMW) as organic fertilizer, gross or composted, carried out in several Mediterranean countries portend use them as fertilizer with condition to not exceed a certain amount depending on crops and soil types. However, the decision‐making of this technique still confronted to some adverse effects reported by some authors, especially related to the acidity, salinity and polyphenol content of OMW. For this reason a trial was carried in 10 liters pots to identify the seasonal evolution of these three parameters on an alkaline soil (pH 7.98), silty clay and moderately rich in organic matter (1.40%), fertilized by four amounts of OMW (50, 75, 100 and 150 m3/ha) in comparison to water treatment, using fenugreek as an indicator plant, planted one day after OMW application. The indicators of stress measured on plants were leaf chlorophyll content, stomatal conductance and leaf temperature. Results showed that application of the four amounts of OMW induced a decrease of soil pH and an increase of electrical conductivity (EC) whereas soil polyphenol content (PC) following significant regression models. Practically, changes observed on soil pH and electrical conductivity values under all treatments, were unable to adversely affect plants growth. The higher dose of OMW (150 m3/ha) decreased soil pH by 0.3 points and EC by 50 µS/cm during the first week of its application. However, soil polyphenol content was amply increased under effect of OMW. Indeed, after one week of OMW application, the initial soil PC (0.175 mg/g) doubled under the treatment 100 m3/ha to reach three fold value under 150 m3/ha. The moderate treatments 50 and 75 m3/ha increased soil PC, respectively by 17 and 40%. However, the physiological parameters of the fenugreek showed a certain stress under 150 m3/ha of OMW, marked by a considerable degradation of chlorophyll and decrease of stomatal conductance. In conclusion, OMW may be used as organic fertilizer up to 100 m3/ha on alkaline soil without adversely affecting its quality in the arable layer. Beyond this amount, a particular attention should be paid, to soil polyphenols as far as their phytotoxicity is concerned, since a high concentration of polyphenols may exert an allelochemical effect.

Keywords: olive mill wastewater, alkaline soil, soil pH, salinity, soil polyphenol content, Trigonella foenum‐ graecum

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 173

Performance of Jet‐Loop reactors with ultrafiltration membrane system (JACTO.MBR) for olive mill wastewaters biotreatment

B. Ribeiro1, I. Torrado1, L. Baeta‐Hall1, A. Amer2, M. Rusan2, A. Eusébio1

1Bioenergy Unit, LNEG, Estrada do Paço do Lumiar, 22, 1649‐038 Lisboa, PORTUGAL 2Department of Natural Resources and Environment, JUST, PO Box 3030, Irbid, JORDAN

Olive mill wastewaters (OMW) disposal is a large‐scale environmental problem particularly in the Mediterranean area. Jordan is facing limited water resources and the reuse of wastewater is considered an additional water source and an important solution from an economical and environmental point of view. The present work, carried out within the EU project "Mediterranean Cooperation in the Treatment and Valorisation of Olive Mill Wastewater (MEDOLICO)", aims to improve the use of jet‐loop reactors technology for the aerobic biological treatment of OMW. Two pilot jet‐loop reactors coupled with an ultrafiltration membrane system (JACTO.MBR) were tested for the influence of hydraulic parameters. In Portugal, one JACTO.MBR with a volume of 100 dm3 was tested under batch and hydraulic retention times (HRT) of 40, 9 and 6 days, with the respective organic loading rates (OLR) of 1, 6 and 14 kgCOD.m‐3.d‐1. In Jordan, JACTO.MBR was scaled‐up to 1000 dm3 and was tested under batch conditions. Chemical parameters, such as chemical oxygen demand (COD), decreased markedly (up to 88%) by the biological treatment, and the efficiency of the process was significantly affected by aeration and inlet flow rates. The total polyphenols content of inlet OMW was reduced around 80% after the treatment. Resident microbial sub‐populations of the microbiota were detected and isolated during OMW treatment in 100 dm3 ‐ JACTO.MBR. The data obtained by the experimentation at pilot scale provided results that support the suitability of the technology for the treatment of wastewaters with the purpose of its reutilization. The treated OMW (UF permeate) was tested for irrigation purposes. Preliminary results of agronomic tests on the corn crop cultures have evaluated the fertilizing effect of the final effluent resulting from the biological treatment.

Keywords: Jet‐loop reactor, olive mil wastewater, aerobic treatment, microbiota, irrigation

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 174

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 175

ECONOMICS MARKETING

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 176

Olive Oil Promotion: A must for the future.

M. Rappou

Agronomist, former Head of the IOC Promotion Department, Hellenic Ministry of Rural Development and Food.

Olive growing plays an important socio‐economic and environmental role in the development of rural populations of many countries in the world, especially those located in the Mediterranean Basin. However, gradually olive growing has gained significant ground in many other countries where the climate and soil conditions made it possible to cultivate the olive tree, thus creating a significant number of new producers. Given the importance of olive growing and in order to safeguard the future of olive producers, it is necessary to strike a balance between supply and demand for olive products. If we take a look at the current situation and also the prospects of olive oil production, we can clearly see that it is following an upward trend both in countries which are traditional producers like Spain, Portugal, Morocco, Algeria, Turkey etc and new emerging ones like China, Chile, S. Africa, etc. Given the upward production‐trend it is necessary to safeguard a parallel trend for consumption. Stimulating demand enough to align consumption with production will permit the smooth function and development of the global olive‐oil market. Obviously the demand for olive oil in the traditional producing countries has reached or is near a saturation point. Therefore the solution to additional production will come only through the expansion of demand for olive oil, not only in the traditional consuming countries, but primarily through increased consumption in the nontraditional new consuming countries. In order to achieve this and to avoid the creation of structural surpluses, we need a well designed plan. A key component of this plan would incorporate a well structured and ongoing promotional campaign to educate and inform consumers in the previously mentioned countries of the health and nutritional benefits of olive products. By definition, the most appropriate organization to carry out this task is the International Olive Council.

Keyword: olive oil market, production and consumption trend, consumer, promotional campaign.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 177

Performance of the olive trees and economic profitability of olive production in the Haouz region in Morocco: effect of drip irrigation with full and deficit water regimes compared to flooding method

A. Ait Hmida1 , L. Sikaoui1 , M. Karrou2 , V. Nangia3

1INRA Marrakech, BP 533, Marrakech, Morocco 2ICARDA Rabat, Morocco 3ICARDA Amman, Jordan

In Morocco, Olive trees cover more than 950,000 ha. The irrigated olive areas are 230,000 ha but only around 24,000 ha are conducted under drip irrigation. The Government policy encourages and supports the use of localized irrigation technology. The main objective of this study is to assess the productivity of olive trees, the production cost and the economic profitability of the orchards in intensive system under the drip irrigation with full (100% ETc) and deficit (70% ETc) water treatments compared to traditional irrigated system with flooding method in the Haouz region. The experiment was conducted at Sâada experiment station located at 10 Km from Marrakesh city. This region is characterized by a semi‐arid climate with a rainfall of about 200 mm/year. The experimental plot was sub divided into two main plots, one for drip irrigation system on 1.15 ha and the other for traditional irrigation with flooding on 0.52 ha. The density of orchards studied is 156 trees/ha (8m x 8m) of the “Picholine Marocaine” variety. The results of this study are the average of 2011‐12 and 2012‐13 agricultural campaigns. The amount of irrigation water applied is 600 mm, 200 mm and 136 mm in 2011‐12 and 673 mm, 192 mm and 132 mm in 2012‐13, respectively for flood irrigation, full drip irrigation and deficit drip irrigation. The yield was 6.22, 8.31 and 8.06 tons/ha, the production cost of a kilogram of olives is 0.45, 0.30 and $ 0.28 and gross margin of production of olive oil is 1,539 ; 3,190 and 3,445 $/ha respectively for the three irrigation treatments. Irrigation cost represents 41% of global production cost in the case of flood irrigation. This part is about 27% and 21% in the cases of full drip irrigation and deficit drip irrigation respectively. In semi‐arid regions, the deficit drip irrigation does not have any negative effect on yield of olive trees. It is more profitable than the other two irrigation treatments studied.

Keywords: Olea europaea, Flood irrigation, Drip irrigation, Cost of Production, Gross margin, Morocco.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 178

Estimating price and income elasticity of olive oil demand in Libya for the period 1980‐2010

K. Elbeydi1 , A. Hamuda2

1Department of Agricultural Economics, Tripoli University 2Biotechnology Research Center, Tripoli – Libya

This study empirically estimate the short and long run price and income elsticities of olive oil demand in Libya in order to provide evidence about their sensitivity to changes in prices and income by using time series data for the period 1980‐2010. Auto Regressive Distributed Lag Model (ARDL) proposed by Pesaran et al. (2001) has been used to analyze the long run relationship amongst the variables. We also implement CUSUM and CUSUMSQ stability tests on the oil olive demand function. The empirical results indicate that olive oil consumption is highly price elastic both in short and long run. The estimated long and short‐run elasticities suggest that per capita income are more significant determinants of olive oil consumption than price of olive oil, indicating that consumers are more sensitive to income changes.

Key words: Demand, Price Elasticity, Income Elasticity, ARDL, olive oil, Libya

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 179

Preference mapping of Extra Virgin olive oil with variety, extraction systems and analytical parameters: Multiple Factor Analysis and Principal Component Analysis

K. Talmoudi1,2, K. Ben‐Hassine4,5,6,7, A. Taamalli8, M. Hamami4, N. Grati‐Kamoun6, D. Malouche1,2,3

1Engineering School of Statistics and Information Analysis, University of Carthage, Tunisia 2National School of Engineers of Tunis, Unit Systems and Signals – (ENIT‐U2S), Tunisia 3Yale MacMillan Center, USA 4Biochemistry Laboratory ‐ UR03/ES08 ’Human Nutrition and MetabolicDisorders USCR ’ Mass Spectrometry Lab of Monastir Medicine University’,Tunisia 5National Institute for Applied Sciences and Technologies, Tunisia 6Institut de l’Olivier, Unité Technologie et Qualité, BP 1087, 3018 Sfax, Tunisia 7Higher School of Agriculture, Mograne 8Laboratoire de Biotechnologie de l’Olivier. Centre de Biotechnologie de Borj Cedria. Tunisia

Preference mapping approach is used to explore and understand the structure and tendencies of consumer preferences and to link consumer preference information to the physico‐chemical data. The objective of this study is to evaluate the physico‐chemical profile and acceptability of 14 extra virgin olive oils (EVOOs) obtained using discontinuous and continuous processing systems. Thus, three studies were then conducted between 2012 and 2013: the first is a survey on 274 consumers, randomly selected, who evaluated 14 EVOOs according to their preferences in different Tunisian regions: Beja, Kairouan, Mahdia and Sfax. In the second study, descriptive sensory profiles of EVOOs were determined using a trained panel (n=8) recognized by the International Olive Oil Council. A third descriptive analysis of these EVOOs was conducted in parallel in Tunisian Olive Tree Institute to characterize them by 70 physico‐chemical parameters. Two different statistical methods were used in order to determine the factors that discriminate between the different olive oils’ preference areas. We have also performed two external preference mapping in order to make a link between consumer preferences and the physicochemical statistical analysis, the first was based on principal component analysis (PCA) map of products and the second based on multiple factor analysis (MFA) map of products. In order to realize the MFA, we have divided the variables of the descriptive dataset into 7 groups predefined by EVOOs tastes, while the PCA was carried out on the 70 physico‐chemical and sensory variables. The external preference mapping performed on the basis of PCA did not show a significant segmentation in the consumers’ ratings (45% and 55%). While the external preference mapping applied in the basis of MFA allowed more detailed descriptions and more classification of the EVOO samples in terms of less versus most prefered EVOO samples (45% and 60%). High quality indices in the case of Coratina olive oil obtained by both processing systems and fruity taste in the case of Chetoui olive oil obtained by continuous processing system were positive drivers of choice for the consumers, whereas richeness in polyphenols was a negative driver in the case of Zalmati olive oil obtained by both processing systems.

Keywords: MFA, External preference mapping, Extra Virgin Olive Oil, Tunisian consumer preferences, Graphical representations.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 180

Olive production in Algeria: situation and perspective

B. Lotmani1, A. Berkani, D. Labdaoui

Laboratoire Protection des Végétaux, Faculté SNV, Université A. Ibn Badis Mostaganem, Algeria

In recent years Algeria gives great attention to the development of olive growing in its agricultural development programs. Olive production is expanded throughout the steppe of the Algerian south regions. The objectives in the plan were to achieve one million hectares as a total olive area in 2014. The main objective of this study was to evaluate the impact of the national Algerian agricultural programs for olive production. To investigate the level of agricultural development, a literature review and consultation of archives and agricultural statistics have been used. In 2000 the olive trees cultivation in Algeria occupied an area of 168,080 hectares, representing only 2% of agricultural land. In 2012, the olive area increased to 400,000 ha, representing increasing of 140% in twelve years. This increase was achieved through the Agricultural Development National Fund that began in 2001. The aim of the agricultural program project is to achieve one million Ha of olive area in 2014. We notice a slight progress in the olive production, olive production fluctuates from year to year. In 2012, 190,000 ha of olive trees were in production excluding isolated trees. 43% of total olive trees were over 50 years old. The total production of olive in 2012 was 0.39 million tonnes. The olive harvest for production of olive oil in 2010‐2011 showed a record harvest of 66,980 tonnes. While, in 2011‐2012, it was only of 39,558 tonnes. In 2012‐ 2013, it was 45,000 tonnes. The cultural conditions, agricultural practices and fluctuated rainfall partly explain the fluctuations in production. The fluctuation from year to another in production is certainly linked to old age of the trees and to the phenomenon of alterne bearing that affect olive growing in Algeria. Table olive production is concentrated in irrigated areas. Table olive production has increased 5 to 6% each year. Algerian exports in terms of olive oil were very small compared to that of neighboring countries; it didn’t exceed 2500 tonnes/year.

Keywords: Algeria, olive production, table olive, olive oil.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 181

The diversity of the Italian monovarietal extra virgin olive oils: from trained panel to consumer experience

B. Alfei1, M. Magli2, A. Rotondi2, L. Morrone2

1Agency for Agrofood Sector Services of Marche, ASSAM, Ancona, Italy 2Institute of Biometeorology, IBIMET‐CNR, Bologna, Italy

Italian olive oil production is based on autochthonous cultivars and these contribute to maintain a large part of its ancient genetic diversity. Since the chemical and sensory characters of extra virgin olive oil are strongly affected by the genotype, the safeguard and the characterization of cultivars and clones play a key role to ensure the marketing of high quality standard olive oils. In the last nine years (2006‐2014) of the Italian National Review of Monovarietal Extra Virgin Olive Oils more than 2000 olive oil samples originating from 154 varieties and 18 different Italian regions were sensory characterized by the Agency for Agrofood Sector Services of Marche (ASSAM) trained panel. This large amount of data was analysed using techniques based on cluster analyses in order to identify similarity among the monovarietal oils and six EVOOs typologies with different sensory properties were identified. The method used to define sensory properties of EVOOs involves evaluation of oils on their own and do not take into account the conditions that consumers often experience when consuming olive oil (i.e. mixed with other food). The present study investigated the impact of this consumer perception to the different monovarietal EVOOs typologies on liking and product discrimination. A total of 540 different consumers, all describing themselves as regular EVOO users, were recruited during four editions of the Italian National Review of Monovarietal Extra Virgin Olive Oils. During the consumer tests, information about hedonic ratings and sensory profiles by the Check‐All‐That‐Apply (CATA) technique were collected. Subjects were asked to taste and evaluate different culinary preparations each one paired with two different monovarietal EVOO samples (belonging to two different typologies). After the tasting, consumers gave hedonic ratings for the two culinary preparations and assessed the presence of specific sensory properties of the EVOO by CATA. Consumers' hedonic responses in terms of liking showed that the use of different monovarietal EVOO typologies influenced the sensory properties of the different culinary preparations. In some pairing, consumers did not discriminate the two EVOOs in terms of acceptability, but the CATA technique discriminated between the two EVOOs used. These results indicate that the six typologies of monovarietal olive oils obtained from the analysis of a trained panel are discriminated by consumers, so this type of information may prove useful in orienting products, i.e. monovarietal olive oil, towards the real expectations and preferences of consumers.

Keywords: monovarietal olive oil, consumer, pairing effect, liking, Check‐All‐That‐Apply; consumer test

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 182

Analysis of efficiency in organic and conventional Italian olive farms using F.A.D.N. dataset N. Galluzzo

Association of Economic and Geographical Studies of Rural Areas, ASGEAR, 02100, Rieti (RI), Italy

Since the 1990s there has been a growth of organic farms in all European states and specifically in some countries such as Spain and Italy, where is concentrated, according to the recent data published in 2013 by the National Institute of Agricultural Economics (INEA), an organic cultivated surface equal to 1.46 and 1.10 million of hectares. The level of income for organic farmers is lower than conventional ones because of specific techniques aimed at improving the quality of production with significant consequences on the economic and technical farms’ efficiency. The purpose of this research is to investigate, over five years (2008‐2012), the level of technical, allocative and economic efficiency in Italian olive farms part of the Farm Accountancy Data Network (FADN) dataset with two different systems of farming such as organic farming system versus conventional one. To estimate the efficiency it has used the dataset FADN, which is a standardized dataset published by the European Union to assess the impact of some actions of Common Agricultural Policy towards European farmers. The Italian FADN sample is set up by 50 organic farms and 500 conventional olive farms analyzing net income, land capital, agrarian capital, labor capital, general costs and financial supports allocated by the Common Agricultural Policy. In order to evaluate the efficiency we used a non parametric quantitative methodology such as the Data Envelope Analysis (D.E.A.). The results have pointed out as conventional olive farms (90% of the sample) are more efficiently than organic farms; over the time there has been an increase of technical efficiency in the organic olive farms. The allocative efficiency has been lower in two‐thirds of analyzed farms. Some pivotal variables such as land capital, agrarian capital and labor capital should be improved in order to ameliorate both the level of economic and also allocative efficiency.

Keywords: economic efficiency, Data Envelope Analysis, Farm Accountancy Data Network, Italy.

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 183

Olive growing in Morocco: Competitive agribusiness

M. Ismaili Alaoui

Department of Food Sciences IAV Hassan II, Rabat Institutes, BP 6202, Morocco

The olive is the main tree crop cultivated in Morocco, spread across the whole of the country due to its ability to grow in a variety of conditions and its adaptability to highly adverse soil and climatic conditions. According to ‘Green Morocco Plan (GMP), Morocco is set to nearly double the area under olive cultivation by 2020, from 720,000 Ha to 1.3 million Ha in 2020. Fruit production for oil is expected to triple, from 550,000 tons to 1.65 million tons in 2020; while olive oil production is expected to nearly quadruple, from 90,000 tons to 330,000 tons in 2020. The olive sector has a vital socio‐economic role and plays an active part in stemming rural depopulation by providing more than 20 million workdays, permanent employment for more than 60 000 people, and livelihood for two million people, covering 16% of Morocco’s consumption requirements in edible vegetable oils. Furthermore, it helps to strike equilibrium in the balance of trade by generating the equivalent of more than one thousand million dirhams in foreign currency earnings on exports of table olives and olive oils. This places Morocco in second place in the world ranking of table olive exporters, after Spain. In addition, it contributes to develop marginal land and protect the environment. The new strategy of the Green Morocco Plan (GMP) is to develop a new label using Geographical Indication (IG) and/or Protected Origin Appellation (AOP) indications giving to olive sector a significant role in the competitiveness of small agribusinesses. The present study investigates the main determinants of Moroccan olive sector according to ‘Green Morocco Plan (GMP) strategy.

Keywords: Olive sector, GMP, label, IG, AOP, Competitive agribusinesses

OLIVEBIOTEQ 2014 3‐6 November, 2014 Amman – Jordan 184

Factors affecting consumers' knowledge of olive oil Protected Designation of Origin label

Y. Erraach, S. Sayadi, C. Parra‐López

Department of Agricultural Economics and Sociology. Institute of Agricultural Research and Training (IFAPA). P.O. Box 2027 – 18080 Granada, Spain.

Food labelling aims to remove information asymmetry or subsidization of search costs that benefits the consumer, by providing information about the attributes of the product. This is only possible if consumers are aware of the different food labels existing in the market and they have a high level of knowledge of the peculiarity and advantages of these labels. One particular strategy of labelling which has been developed in EU countries is based on the use of geographical indications such as Protected Designation of Origin (PDO). Despite the time lapse since their inception, the debate continues concerning knowledge of these labels and their effect on consumers' attitudes, preferences and quality perception. Spain is the first world producer and exporter of olive oil and table olives. Near of 38% of Spanish olive‐oil‐orchard surface area was labelled with a PDO, having 27 PDOs; among them 12 are located in Andalusia (southern Spain) which is the major olive‐oil production area worldwide. The potential benefits of olive oil PDOs are related not only to economic profit gained through the differentiation of the product but to a wide array of social and environmental externalities (employment, high biodiversity, landscape, etc.) that contribute to the sustainable development of rural areas and territorial governance. In this framework, the objective of this study is twofold: 1) examining the Andalusian consumers' knowledge of PDO label on olive oil, and 2) to explore the socio‐demographic and lifestyle variables determining this knowledge. With this purpose a binomial Probit model was fitted based on data gathered through 439 face‐to‐face interviews to Andalusian olive oil consumers in 2010. The survey was stratified by random sampling proportionally to age, gender, and place of residence. The results indicate that consumers' knowledge of PDO olive oil is low. Furthermore, this knowledge is determined by some key socio‐economic and lifestyle variables, such as gender, education level, household income and if consumer comes from or lives in an olive‐oil zone. Finally, based on these results, marketing strategies to improve PDO labelled olive oil knowledge and consumption are drawn.

Keywords: Olive oil, Protected Designation of Origin (PDO), Consumers, Knowledge, Andalusia.