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Journal of Plant Pathology (2011), 93 (4, Supplement), S4.25-S4.62 Edizioni ETS Pisa, 2011 S4.25

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Journal of Plant Pathology (2011), 93 (4, Supplement), S4.25-S4.62 Edizioni ETS Pisa, 2011 S4.25 004b_Posters_S25 28-12-2011 14:05 Pagina 25 Journal of Plant Pathology (2011), 93 (4, Supplement), S4.25-S4.62 Edizioni ETS Pisa, 2011 S4.25 CHARACTERIZATION OF A PR-10 PROTEIN FROM VI- tions of specialized cells that coexist within highly structured com- TIS RIPARIA. M. Alessandrini, A. Lovato, L. Bortesi, G. Zoc- munities. Here we report how phenotypic dissociations in B. sub- catelli, S. Zenoni, M. Pezzotti and A. Polverari. Dipartimento di tilis can induce a significant reduction of the secondary metabo- Biotecnologie, Università degli Studi, Strada le Grazie 15, 37134 lites which are associated with antifungal activity. Strain ET-1 ini- Verona, Italy. E-mail: [email protected] tially has a rough, dense, colony morphology phenotype (R-form) with undulated edges, which dissociated into a S-form (smooth, We have started a project aimed at characterizing the expres- soft, whitish) and into a M-form (slimy, translucent, amoeba- sion profile and protein activity of pathogenesis-related proteins of shaped colony). Correlations between colony forms, spore forma- the PR-10 family in grapevine. According to a previous microarray tion and in-vitro antifungal activity of cell-free supernatants were analysis, the expression of several genes, annotated as similar to studied. Germinal inhibition over 95% of Penicillium digitatum PR-10 proteins, is strongly increased in resistant Vitis riparia and, conidia was induced only by S and R forms when cell-free super- to a lesser extent, in V. vinifera, following infection with Plas- natants were diluted 1:16 and 1:128 respectively. Biosynthesis of mopara viticola. PR-10 proteins are induced by pathogen infection antifungal molecules in broth was significantly reduced when an and abiotic stresses in a number of species; because they often old R-form colony was used for inoculation because it was already show ribonucleolytic activity it has been suggested that they may dissociated in M and S phenotypic cells. After 48 h of growth in be involved in antiviral defence; their antimicrobial activity has liquid medium a comparison of the spore-forming colony and an been tested with controversial results. The coding sequence of a examination by phase-contrast microscopy revealed a reduced homologous gene, denoted Vr-PR10, has been cloned from the re- ability of the dissociant M-form to sporulate (104 CFU/ml). A sistant species V. riparia and the corresponding protein expressed complete spore release from the vegetative cells was observed only in E. coli. The purified recombinant protein showed ribonucleolytic for the R-form (109 CFU/ml), whereas the S-form contained a activity in vitro and was used to raise antibodies which recognized mixture of endospores (108 CFU/ml) and few vegetative cells. Dif- the native protein from leaf tissue in Western blot analyses. The ferent levels of Podosphaera fusca control on melon by the three protein was tested in vitro and in vivo against P. viticola, Botrytis form colonies were obtained in in-vivo trials. To reduce secondary cinerea and Grapevine virus A (GVA), without showing any antimi- metabolite production problems, the use, as starter, of a pasteur- crobial or antiviral activity. The possibility that PR-10 proteins have ized spore suspension by young isolated R-form colony of the ET- different or indirect roles in defense should be considered in fur- 1 strain is recommended. ther investigations. AGROBACTERIUM-MEDIATED TRANSFORMATION OF DISTRIBUTION OF BOTRYOSPHAERIACEAE SPECIES FOR INVESTIGATING ASSOCIATED WITH GRAPEVINE DECLINE IN WEST- PYRENOCHAETA LYCOPERSICI MOLECULAR DETERMINANTS OF VIRULENCE. M. ERN SICILY. A. Alfonzo, C. Conigliaro, V. Mondello and S. Bur- Aragona, M.T. Valente and A. Infantino. CRA-PAV, Centro di ruano. Dipartimento DEMETRA, Università degli Studi, Viale delle Ricerca per la Patologia Vegetale, Via C.G. Bertero 22, 00156 Ro- Scienze 4, 90128 Palermo, Italy. E-mail: [email protected] ma, Italy. E-mail: [email protected] Recently, the pathogenicity of different Botryosphaeraceae Pyrenochaeta lycopersici is a soil-borne fungus causing an eco- species on grapevines has been reported from many European nomically important disease known as corky root rot on tomato and extra-European areas. During summer 2008, in a Sicilian and other hosts. A gene coding for an extracellular endo-1,4-b- vineyard located in the Marsala area (TP), we observed a decline glucanase has recently been cloned and, in order to explore its induced by Lasiodiplodia theobromae. The occurrence and quite putative role in virulence, an ATMT (Agrobacterium tumefaciens- rapid spread of the disease seemed to support the regional vari- mediated transformation) system was established. P. lycopersici ability of the species in question, which is retained a tropical and does not produce conidia in culture so the transformation subtropical pathogen to different trees. In the last two years, the method currently available for this species involves protoplast decline syndrome has been observed in the following other transformation in presence of PEG. Despite of the low efficiency, grape-growing areas of western and central Sicily: Alcamo and a GUS-expressing transformant has been obtained and used for Salemi (TP), Montevago (AG) and Milena (CL). From declining investigating the timing of infection and colonization of P. lycop- plants showing typical external and internal symptoms of spurs, ersici inside tomato roots. To increase the transformation efficien- canes and trunks, fungal isolates were obtained in pure and/or cy and to obtain a larger number of transformants for gene re- mixed culture and were identified, by morphological and molec- placement experiments, we tested both mycelium and protoplasts ular analysis, as L. theobromae, Diplodia seriata and Neofusicoc- as recipients for an ATMT-based method. Preliminary results cum luteum. These results, even if preliminary, show the rapid showed that only the protoplasts but not the mycelium were suc- spread of the grapevine decline by Botryosphaeraceae species in cessfully transformed by using the pRF-HU2 vector, harbouring Sicily confirming, at the same time, their different distribution in promoter and terminator genes from Aspergillus nidulans and hy- relation to environmental conditions of the surveyed areas. Path- gromycin coding gene as fungal marker. The average transforma- ogenicity tests are in progress to ascertain the role of these fungi tion frequency obtained with the first experiments was up to 30 in the evoking of the decline syndrome. transformants per 107 protoplasts, all showing the DNA vector insertion. We are going to test the applicability of this technique for targeted gene replacement in P. lycopersici, after cloning the EVALUATION OF THE ANTIFUNGAL ACTIVITY OF endo-1,4-b-glucanase sequences into the pRF-HU2 vector. MORPHOLOGICALLY DISTINCT COLONIES OF BACIL- LUS SUBTILIS STRAIN ET-1. A. Ambrico and M. Trupo. Unità Tecnica Tecnologie Trisaia, Laboratorio Biotecnologie, C.R. ENEA Trisaia, ss106 Jonica km 419.5, 75026 Rotondella (MT), MONITORING FACTORS AFFECTING EPIDEMIOLOGY Italy. E-mail: [email protected] OF ALTERNARIA BROWN SPOT OF CITRUS. P. Bella1, M. Russo2, M. Tomasello2, V. Catara1 and A. Catara2. 1Dipartimento Bacillus subtilis differentiates into distinct adaptive subpopula- di Scienze delle Produzioni Agrarie e Alimentari, Università degli Edizioni ETS Pisa, 2011 004b_Posters_S25 28-12-2011 14:05 Pagina 26 S4.26 Journal of Plant Pathology (2011), 93 (4, Supplement), S4.25-S4.62 Studi, Via Santa Sofia 100, 95123 Catania, Italy. 2Parco Scientifico the farm were conducted and many populations were found, sug- e Tecnologico della Sicilia, Zona Industriale, Via V. Lancia, 95030 gesting that thrips are the main responsible for INSV spreading Catania, Italy. E-mail: [email protected] and widening its natural host range. Alternaria brown spot of citrus, caused by Alternaria alterna- ta, causes considerable damage mainly to hybrids of Dancy tan- gerine. The fungus induces brown to black spots on leaves, fruit FIRST REPORT OF HOSTA VIRUS X INFECTING HOSTA and young twigs resulting in reduced fruit yield and quality. Fol- IN ITALY. M.G. Bellardi1, L. Cavicchi2 and S. Davino3. 1Diparti- lowing the studies carried out primarily in Florida, Brazil, Israel mento di Scienze e Tecnologie Agroalimentari, Patologia Vegetale, and Spain, disease epidemiology observations were performed in Università degli Studi, Viale Fanin 42, 40127 Bologna, Italy. an orchard of cv. Fortune mandarin grafted onto Citrange in 2Plesso Didattico “G. Scarabelli”, Università degli Studi di Bologna, southern Italy (38°09’06.82”N- 16°09’57.40”E). Surveys, carried Viale G. Ascari 17, 40026 Imola (BO), Italy. 3Dipartimento di out over the last eight years (2003-2011), have shown that for Scienze Entomologiche, Fitopatologiche, Microbiologiche agrarie e eight months per year (March to November) temperatures are Zootecniche, Sezione di Patologia Vegetale e Microbiologia Agraria, sufficient for infection, however the annual rainfall distribution Università degli Studi, Viale delle Scienze edificio 5, 90128 Paler- of 600-1000 mm, favours infection in March-April (65-236 mm; mo, Italy. E-mail: [email protected] mean 121 mm) and in September-October (121-462 mm; mean 252 mm). Flower and fruit infections, occurring before fruit set Species of Hosta (Liliaceae) are herbaceous perennials more and fruit growth, directly affect fruit yield and quality, whereas often grown for their foliage (blue, gold, green
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