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© by PSP Volume 24 – No 11c. 2015 Fresenius Environmental Bulletin
CONTENTS
ORIGINAL PAPERS
EFFECTS OF POLY(2-ETHYL-2-OXAZOLINE) EXPOSURE 4072 ON OVARY HISTOLOGY OF ZEBRAFISH (Danio rerio) Nazan Deniz Yön, Cansu Akbulut, Selda Güneş, Müge Alsaran, Uğursoy Olgun, Figen Esin Kayhan and Güllü Kaymak
INVESTIGATION OF MYXOMYCETES DIVERSITY ON 4077 KUSEYR MOUNTAIN; THREE NEW RECORDS IN HATAY/TURKEY Hayri Baba
MONITORING SOIL RESPIRATION DYNAMICS DURING THE BIODEGRADATION OF 4087 CRUDE OIL CONTAMINATED SOIL UNDER DIFFERENT ENVIRONMENTAL CONDITIONS Xinping Zhu, Pujia Yu, Ni Han, Hongtao Jia, Xinjun Wang, and Kobil Gafurovitch Kodirov
THE EFFECT OF PLANTING GEOMETRY AND SEED PRIMING 4095 ON SUNFLOWER YIELD UNDER RAIN-FED CONDITIONS Mehmet Demir Kaya, Suay Bayramin and Gamze Kaya
LIFE TABLE CHARACTERISTICS OF Orius minutus (L.) 4102 (HEMIPTERA: ANTHOCORIDAE) FEEDING ON Bemisia tabaci (GENNADIUS) (HEMIPTERA: ALEYRODIDAE) AT THE LABORATORY CONDITIONS Davut Efe, Mehmet Salih Özgökçe and İsmail Karaca
RELATIONSHIPS BETWEEN CHEMICAL AND PHYSICAL PROPERTIES 4108 OF SOILS AND NUTRIENT STATUS OF PLANTS ON YIELD OF POTATO Kürşat Korkmaz, Özbay Dede, Halil Erdem, Soner Çankaya and Mehmet Akgün
DEVELOPMENT AND TESTING OF A DIATOM-BASED INDEX OF BIOTIC INTEGRITY FOR 4114 RIVER ECOSYSTEMS IMPACTED BY ACID MINE DRAINAGE IN GAOLAN RIVER, CHINA Xiaoyu Dong, Xinghuan Jia, Wanxiang Jiang, Naicheng Wu, Tao Tang and Qinghua Cai
DEVELOPING A CLIMATE-BASED RECREATION 4125 MANAGEMENT SYSTEM FOR A MEDITERRANEAN ISLAND Hossein G.T. Olya and Habib Alipour
QUANTIFICATION OF THE HEALTH EFFECTS OF 4142 EXPOSURE TO AIR POLLUTION (NO2) IN TABRIZ, IRAN Mohammad Ghanbari Ghozikali, Alessandro Borgini, Andrea Tittarelli, Abdeltif Amrane, Kazem Naddafi, Mahmoud Mohammadyan, Gholamreza Goudarzi, Roberto Bono and Behzad Heibati
IMPACT OF COPPER AND ITS Ca MIXTURES ON THE 4149 GLUTATHIONE S-TRANSFERASE ACTIVITY, GLUTATHIONE LEVEL AND METAL ACCUMULATION IN TISSUES OF FRESHWATER FISH Oreochromis niloticus Gülbin Firidin and Ferit Kargın
4070 © by PSP Volume 24 – No 11c. 2015 Fresenius Environmental Bulletin
ANTIMONY DISTRIBUTION AND EFFECT FACTORS 4157 IN THE SURFACE SEDIMENT OF NANSI LAKE, CHINA Tongguang Shi, Lu Zhang, Yuanxin Cao and Shiliang Wang
EVALUATION OF CONTAMINATION DATA 4165 WITH NON-DETECTS USING CENSORED DISTRIBUTIONS Michal Fusek, Jaroslav Michálek and Milada Vávrová
DIRECT APPLICATIONS OF CERIUM AND YTTRIUM ON VEGETABLE PAK CHOI 4173 Yu-Fu Hu, Cheng-Ming Yuan, Shuang-Long Jiang, Ke-Ya Ma, Jia-Jia Peng, Qin Pu and Shu Yuan
A METHOD OF USING ELECTROCOAGULATION-FLOTATION 4179 TECHNOLOGY FOR TREATING ZINC-CONTAINING SIMULATED WASTEWATER Ni Liu, Qiaoping Kong, Hongxiang Zhu, Xuejuan Cao, Jin Wang, Hui He and Shuangfei Wang
PREGNANCY EXPOSURE TO ATMOSPHERIC POLLUTANTS 4184 AND PLACENTAL WEIGHT IN A TUNISIAN POPULATION Abdel Halim Harrath, Saleh Alwasel, Ichrak Khaled, Lamjed Mansour, Alexander V. Sirotkin and Mohammed F. Faramawi
ARE THE PHOTOSYNTHETIC PERFORMANCE INDEXES AND THE 4190 DROUGHT FACTOR INDEX SATISFACTORY SELECTION CRITERION FOR STRESS? Nuran Çiçek, Özlem Arslan, Şeküre Çulha-Erdal, Füsun Eyidoğan and Yasemin Ekmekçi
DOES THE ASSOCIATION OF ARBUSCULAR MYCORRHIZAL FUNGI AND TWO- 4199 SPOTTED SPIDER MITE INCREASE GOSSYPOL SYNTHESIS IN TWO COTTON CULTIVARS? Ozan Demirözer, Hülya Özgönen Özkaya, Tolga Aldemir and Melis Karapire
SYNTHESIS OF SnS2-CNT NANOCOMPOSITES WITH EXIMIOUS 4205 VISIBLE-LIGHT-DRIVEN PHOTOCATALYTIC DEGRADATION OF TBA Ze-Da Meng, Lei Zhu, Kefayat Ullah, Shu Ye and Won-Chun Oh
TRANSFER FACTOR AS INDICATOR OF HEAVY METALS CONTENT IN PLANTS 4212 Nataša Mirecki, Rukie Agič, Ljubomir Šunić, Lidija Milenković and Zoran S. Ilić
TOTAL PHENOLIC CONTENT, PHOTOSYNTHETIC PIGMENT CONCENTRATION AND 4220 ANTIOXIDANT ACTIVITY OF LEAVES AND BULBS OF SELECTED Eucomis L'Hér. TAXA Piotr Salachna, Monika Grzeszczuk and Janusz Wilas
INDEX 4226
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EFFECTS OF POLY(2-ETHYL-2-OXAZOLINE) EXPOSURE ON OVARY HISTOLOGY OF ZEBRAFISH (Danio rerio)
Nazan Deniz Yön1,*, Cansu Akbulut1, Selda Güneş1, Müge Alsaran1, Uğursoy Olgun2, Figen Esin Kayhan3 and Güllü Kaymak1
1Sakarya University Department of Biology, Sakarya, Turkey 2Sakarya University Department of Chemistry, Sakarya, Turkey 3 Marmara Universtiy, Department of Biology, İstanbul, Turkey
ABSTRACT Poly (2-oxazoline)s are one of the class of poly(oxa- zolines) which has been studied since 1960s [4, 5, 6, 7, 8, Poly(2-oxazoline)s have been widely used in biomed- 9, 10]. Poly(oxazolines) are generally used in adhesive and ical applications for the last years. They can mimick natu- coating formulatons [11, 12, 13] as pigment dispersants in ral systems and they generally used as liposomes, drug and inks and drug in delivery applications [14,15]. Polyoxazo- gene deliver and pseudopeptides. In this study histological line-based or polyoxazoline-derived polymers can mimick effects of different doses (10 mg/L, 50mg/L) of poly(2- natural systems so they have significant application poten- ethyl-2oxazoline) on ovary tissue of zebrafish were inves- tial in a large number of technological contexts, whether tigated. After 5 days of administation, ovary tissues were this be the formation of stealth liposomes, or formation of dissected and histomorphological changes investigated un- membrane structures and it’s used as carriers of drugs or as der light microscope. Severe structural deformation in oo- synthetic vectors and antimicrobial materials [16]. Because cytes were monitored. Deterioration in germinal vesicle of their biocompatibilty, poly(2-oxazoline)s have potential and openings at follicular epithelium, zona radiata and vi- to use a variety of applications in biological systems such telline envelope were detected. as drug and gene delivery are the principle application ar- eas for polymers in biological, pharmaceutical and medical contexts [17, 18]. A drug is enclosed in a polymeric matrix and released over time through diffusion [19] and alterna- KEYWORDS: tively, complex multifunctional polymers with covalently poly(2-ethyl-2-oxazoline), zebrafish, ovary, histology attached drug moieties are constructed [20,21]. The com- bination of a polymer with a drug molecule has several main advantages over the use of a pure drug molecule: (1) potentially increased solubility of insoluble drugs or drugs 1. INTRODUCTION of low aqueous solubility, (2) improved pharmacogenetics, (3) protection against deactivation and degradation during Zebrafish, Danio rerio, is a member of Cyprinidae transport and circulation, (4) reduced antigenic activity and family and it lives in India and Pakistan naturally. (5) the possibility to combine a drug with other functional Zebrafish is a widely used model organism because it is components such as contrast agents etc. [21,22]. easy to produce and it has short generation time. In the la- boratory, zebrafish can be stimulated to breed throughout Poly oxazolines have similar behavior similar to poly the year and the development from the fertilised egg to re- (ethylene glycol) so are used in several biomedical appli- cations such as stealth liposomes, membrane structures, producing stage takes only about 3-4 months. Their short 67 generation time makes them an ideal candidate for the ge- carriers of drugs and antimicrobial materials [16]. Ga-la- netic studies and their susceptibility to the mutagens, car- belled liposomes, which were subsequently injected into cinogens, teratogens and toxins makes them ideal environ- the bloodstream of rats in order to monitor their progress mental models for the investigation of ovarian follicle de- and distribution via blood-level and tissue measurements. velopment and maturation because the zebrafish ovary The experiments showed that the behavior of the poly(ox- contains follicles at different stages of development [1,2]. azoline) based liposomes is similar to that of the corre- Zebrafish have been used not only as a general vertebrate sponding PEG-based materials, both in terms of their cir- toxicity model [3], but also as an ecotoxicological test spe- culation in the bloodstream (showing long circulation life- cies to determine the effects of chemicals on fish survival, times) as well as in terms of uptake by liver and spleen [23]. growth, and reproduction. Poly(2-oxazoline)s can be regarded as analogues of polypeptides and also of polypeptoids [21]. So, it is con- sidered as pseudopeptides, thus bioinspired polymers, due * Corresponding author to their structural relationship to polypeptides [24].
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2-methyl, 2-ethyl, 2-isopropyl and 2-phenyl oxazoline are available commercially. In this study, the histological effects of poly(2-ethyl-2-oxazoline) (PEtOx) on ovary tis- sue of zebrafish investigated with light microscopy.
2. MATERIALS AND METHODS
2.1 Animal The zebrafish (Danio rerio) is a small fish about 6 cm in length, characterized by a series of five pigmented stripes running the entire length of each side of its body. The zebrafish’s hardiness makes them excellent stress test subjects, as they can survive fairly severe environmental changes without succumbing, surviving long enough to show developmental defects. Sexually mature zebrafish were maintained under standardized laboratory conditions FIGURE 1 - The ovary of control group, and general apparences of (28oC ± 1 oC and 14h/10h light dark cycle, 61% humidity). different stages of oocytes. Po- Primary oocyte, Ca- Cortical alveoli, The fish were daily fed with Artemia sp. and TetraMin© CoC- Cortical alveolus stage, N- nucleus, Om- Mature oocyte, Ao- Hauptfutter (Tetra Werke,Germany). atretic oocyte, 20x Toluidine blue stain.
2.2 Exposure Following the prelaminary experiment, all determina- tions were repeated three times. In our study we created one control and two experiment groups (n=15) concerning their different PEtOx doses (Group I: 10 mg/L PEtOx, Group II: 50 mg/L PEtOx, Group III: control group). After 48 h adap- tation, different concentrations of PEtOx were added to the experimental aquaria. Mortality was controlled during the test and it was observed that no fish died. After exposure, on the fifth day of the study fishes were anaesthetized with ice water and ovary tissues were dissected immediately.
2.3 Light microscopy
For the histological analysis, ovary tissues were fixed with 10% neutral buffered formalin for 24h. Fixed tissue FIGURE 2 -Ovary tissue of 10 mg/L PEtOx exposed group, Po- Pri- was dehydrated using a series of graded ethanol solutions mary oocyte, CoC- Cortical alveolus stage, Vo- vitellogenic oocyte, (70%-100%) and embedded in the parafin wax and sec- Om- Mature oocyte, Ao- atretic oocyte, 20x, H&E stain. tioned transversely at 5 µm thickness and stained with He- matoxylin-Eosin and toluidine blue staining. 3.2 Exposure Group I In 10 mg/L PEtOx exposed group, the number of fol- licles at vitellogenic oocytes and mature oocytes are 3. RESULTS slightly increased compared with control (Figure 2). Tor- sions at oocyte structures and oocyte shape changings were 3.1 Control group detected. In cortical alveolus stage, openings between zona In control group, normal structure and all development radiata and follicular epithelium were observed. The oo- stages of zebrafish oocytes were observed clearly. Identifi- cyte was also shrinked (Figure 3a, 3b). The zona radiata cation of oocytes were done according to Koç et al. [ 25]. has a corrugated structure. In addition, the zona radiata is In the first growth phase, the multiple nucleoli were ob- clearly visible but follicular epithelium is reserved at corti- served in the nucleus of oocytes. Primary oocytes in the cal oocyte’s surface (Figure 3a). Similar results were also control group were oval-shaped cells with oval nucleus and detected at vitellogenic oocytes. There are no distinct vac- large nucleolus. In cortical alveolar stage, granular struc- uoles at vitellogenic oocyte. (Figure 3b). tures in the ooplasm were increased. It was detected that zona radiata began to form and the follicular epithelium be- 3.3 Exposure Group II come thicker. In vitellogenic phase, oocyte sizes were in- In 50mg/L PEtOx exposed group, different stages of creased and larger granular structures were observed (Fig- maturation at ovarian follicles can be identified. The num- ure 1). ber of follicles at vitellogenic oocytes and mature oocytes
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FIGURE 5 - Ovary tissue of 50 mg/L PEtOx exposed group, openings between zona radiata and vitelline envelope at cortical alveolus stage, CoC- cortical alveolus stage oocyte, Ve- vitelline envelope, ZR- zona radiata, Fe- follicular epithelium, H&E staining, 40x, 100x
FIGURE 3 - Ovary tissues of 10 mg/L PEtOx exposed group, Shrink- ing and openings at oocytes, a) Zona radiata has a corrugated struc- ture, 40x, Toluidine blue, b) There are no distinct vitellogenin vacu- oles, 40x H&E stain. Ca- Cortical alveoli, Vo- vitellogenic oocyte, N- nucleus, No- nucleolus, Fe- follicular epithelium, ZR- zona radiata FIGURE 6 - Ovary tissue of 50 mg/L PEtOx exposed group, deterio- ration at nucleus (germinal vesicle) at cortical alveolus stage, Ca- cor- tical alveoli, N- nucleus, No- nucleolus, H&E staining, 100x
FIGURE 4 - Ovary tissue of 50 mg/L PEtOx exposed group, openings between zona radiata and follicular epithelium at mature and pri- mary oocytes, deterioration at zona radiata (showed with arrow), FIGURE 7 - Ovary tissue of 50 mg/L PEtOx exposed group, cortical CoC-Cortical alveolus stage, Ca- Cortical alveoli, Fe- Follicular epi- alveolus stage oocyte at early phase and deterioration (showed with thelium, ZR- zona radiata, Po- primary oocyte, N- nucleus, No- Nu- star) at vitellogenic oocyte structure, CoC- cortical alveolus stage oo- cleolus, Om- Mature oocyte, Toluidine blue stain, 40x cyte, Po- primary oocyte, Vo- vitellogenic oocyte, H&E staining, 40x
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are increased but primary oocytes are slightly decreased in the number of atretic follicle and decrease in number of compared with control group. This chemical has been acted vitellogenic and mature oocytes. In addition they detected as endocrine disruptors. The ovary of fish exposed to this fragments in vitelline membrane and yolk granules. Dis- chemical presents complete obsence of atretic follicles. A concertingly, our results are different from this study. In follicle at stage of vitellogenic oocyte is atretic with a vi- our study we detected a lot of vitellogenic and mature oo- telline membrane forming characteristic folds. Openings cytes when compared with the control group. Especially in between zona radiata and follicular epithelium at mature exposure groups, increase in the number of vitellogenic and primary oocytes, deterioration at zona radiata were and mature oocytes but decrease in primary oocytes were monitored (Figures 4 and 5). In cortical alveolus stage detected. Our data showed that poly(2-ethyl-2-oxazoline) openings between vitelline envelope and zona radiata were cause early oocyte maturation at zebrafish. On contrary, se- observed (Figure 5). Moreover, deterioration at germinal vere structural deformities have been monitored. This chem- vesicle was detected at cortical alveolus stage. Nucleus ical can be affect endocrine system. Similarly with our study, shape was irregular and corruption at chromatin material Tokumoto et al. [32] studied effects of diethylstilbestrol on was monitored (Figure 6). Even in this group, cortical al- goldfish ovary and they found that diethylstilbestrol induces veolus stage oocyte at early phase was detected. In addi- fish oocyte maturation. In our previous study about acute tion, degeneration at vitellogenic oocyte structure was ob- toxicity of 2,4-diclorophenoxy acetic acid in zebrafish, we served. (Figure 7). found severe damage at follicular epithelium, irregular folding membranes and eccentric nucleus at oocyte struc- ture. On contrary, we detected clumping of karyoplasms 4. DISCUSSION AND CONCLUSION and atretic oocytes [33]. There are some studies about biomedical applications The use area of poly(2-oxazoline)s have been devel- about poly(2-oxazoline)s but toxicological studies are not oped in biomedical applications for the recent years, as a adequate. In this study, toxicological effects of poly(2- result of their biocompability as well as their stealth behav- ethyl-2-oxazoline) were evaluated with histological meth- iour [16, 21]. The biocompability of poly(2-methyl-2-oxa- ods. We chose the zebrafish model at this study because zoline) (PMeOx) was demonstrated in 1989 [26]. 125I-la- they can survive at severe environmental changes and show beled polymers were found to be excreted from mice with- developmental defects. Also they are easy and inexpensive out significant accumulation in organs, although some, pre- to raise, requiring only filtered water, and a minimal invest- sumably high-molecular-weight, polymer was found in ment in fish food, making them an ideal animal model for skin and muscle tissue [26]. In Zalipsky and co-workers’ research laboratories with limited funding. However, more study it was proved that PEtOx liposomes are faster than toxicological studies about this substances should be done PMeOx in rats and mice and revealed enhanced circulation by using other model organisms. times, with similar blood-clearance rates in vitro studies [27]. It was also proved that the liposomes accumulated The authors have declared no conflict of interest. mainly in the liver, kidney, and spleen [27, 28]. The study about poly(2-oxazoline)s was not detected regarding the effects on fish. So, our study is pioneer. 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[19] Kim, S.J., Lee, K.J., Kim, S.I. (2004) Thermo-sensitive swelling behavior of poly(2-ethyl-2-oxazoline)/poly(vinyl alcohol) inter- penetrating polymer network hydrogels. Journal of Macromole- cule Science- Pure and Applied ChemistryA41, 267-274. [20] Kim, S.J., Lee, K.J., Kim, I.Y., Shin, D.I., Kim, S.I. (2006) Temperature and pH-response swelling behavior of poly(2- ethyl-2-oxazoline)/chitosan interpenetrating polymer network hydrogels. Journal of Applied Polymer Science99, 1100-1103. [21] Hoogenboom, R., Schlaad, H. (2011) Bioinspired Poly(2-oxa- zoline)s. Polymers 3, 467-488. [22] David, G., Simionescu, B.C., Albertsson, A.C. (2008) Rapid Received: November 27, 2014 deswelling response of poly(N-isopropylacrylamide)/poly(2- Revised: February 13, 2015 alkyl-2-oxazoline)/poly(2-hydroxyethy-l-methacrylate) hy- Accepted: February 25, 2015 drogels. Biomacromolecules 9, 1678-1683.
[23] Lasic, D.D., Martin, F.J., Gabizon, A., Huang, S.K., Papa- hadjopoulos, D. (1991) Sterically stabilized liposomes: a hy- CORRESPONDING AUTHOR pothesis on the molecular origin of extended circulation times. Biochimica et Biophysica Acta: Biomembranes 1070, 187- 192. Nazan Deniz Yön Sakarya University [24] Aoi, K., Suzuki, H., Okada, M. (1992) Architectural control of Department of Biology sugar-containing polymers by living polymerization: Ring- opening polymerization of 2-oxazolines initiated with carbo- Sakarya hydrate derivatives. Macromolecules 25, 7073-7075. TURKEY [25] Koç (Yön), N.D., Aytekin, Y., Yüce, R. (2008) Ovary Matu- ration Stages and Histological Investigation of Ovary of the E-mail: [email protected] Zebrafish (Danio rerio). Brazilian Archieves of Biology and Technology 51(3), 513-522. FEB/ Vol 24/ No 11c/ 2015 – pages 4072 - 4076
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INVESTIGATION OF MYXOMYCETES DIVERSITY ON KUSEYR MOUNTAIN; THREE NEW RECORDS IN HATAY/TURKEY
Hayri Baba
Department of Biology, Faculty of Arts and Science, Mustafa Kemal University, Alahan-31000 Antakya-Hatay, Turkey
ABSTRACT 2. MATERIALS AND METHODS
In this study, myxomycetes collected from Kuseyr 2.1 Description of research area Mountain of Hatay province between 2010-2013 have been Kuseyr Mountain is (36' 09" 46N, 36' 10" 19E) located investigated. As a result of field and laboratory studies, at the Hatay province of Turkey (Figure 1). 43 species belonging to 11 family and 19 genera were re- The area is situated at the Mediterranean phytogeo- ported. Three species, including Didymium clavus (Alb. & graphical region in C6 square grid system of Davis (1965- Schwein.) Rabenh., Perichaena pedata (Lister & G. Lister) 1985) [19]. The vegetation of reseach area is partially ruined Lister ex E. Jahn and Trichia scabra Rostaf.) were rec- Pinus brutia Ten. forest (350-520 m) and different types of orded for the first time from Hatay, Turkey. Distribution, shrubs are dominate for example Arbutus andrachne L., habitat characteristics, relative abundance of fruit bodies, Laurus nobilis L., and Quercus coccifera L. species are com- taxonomic diversity indices and diversity of species were mon. In settlement areas, Ficus carica L. and Olea europea discussed in details. In addition, photographs belonging to L. species are common. Platanus orientalis L., Salix baby- these new records were taken with SEM. lonica L., Myrtus communis L. and Nerium oleander L. are
common near the stream while in empty areas, Sarcopo- terium spinosum (L.) Spach, Cistus creticus L. and Thymbra spicata L. are common [20]. KEYWORDS: Biodiversity, Distribution, Myxomycota, New records, Turkey. 2.2 Sample collection and moist chamber culture
The specimens were collected from natural substrata, bark and debris, bark of living trees, as well as on decaying fruits, bark, wood, leaves, litter and animal dung. Natural 1. INTRODUCTION mature fructifications were gently collected from substra- tums, and placed in cardboard fungarium boxes. In addi- Myxomycetes, which are also known as myxogastrids tion, fructifications of myxomycetes were obtained from or plasmodial slime molds, are common and relatively cos- the moist chamber culture in laboratory. All moist chamber mopolitan in their distribution. They are characterized by cultures were prepared within the week after returning an amorphous, multinucleated, and protoplasmic mass from field survey. Substratum samples were placed in petri called plasmodium and fruiting bodies. They occur in as- dishes lined with filter paper. Distilled water was added to sociation with decaying or living plant materials in terres- each petri dishes and the samples allowed to soak over- trial forest ecosystems. Humidity and temperature are the night. After 24 hours, excess water was poured off. Cul- two main factors in their diversity and abundance [1]. The tures were maintained under diffuse light at room temper- majority of species are mainly cosmopolitan but a few ap- ature (22-25°C) for a period of approximately three month. pear to be confined to the tropics or subtropics, and some All cultures were checked weekly for the presence of myx- others have been collected only in temperate regions [2]. omycete plasmodia or fruiting bodies. Although Myxomycetes have been extensively studied worldwide [3-9], a few studies have been conducted on 2.3 Morphology and identification Myxobiota in Turkey. In recent years, some Myxomycetes Microscopic and macroscopic features of samples have been added to the Turkish myxobiota by some re- were determined in laboratory. Fruiting bodies were iden- searchers [10-16]. The number of known Myxomycetes tified to species using light microscopy in consideration of species in world is 964 [17] but there are only about 236 morphological characters such as fruiting bodies; shape, species known from Turkey [18]. There has been no previ- size and colour, peridium; colour, presence and calyculus ous studies in Kuseyr Mountains. In this study, three taxa situation, spore; size, colour and ornamentation, capillit- were added to the Turkish Myxomycota as new records. ium and columella; colour and branching, lime crystal size
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FIGURE 1- Map of Hatay and Kuseyr Mountain
and morphology, stalk; length, colour and proportion to In this study, 3 samples (1 species) are members of fruiting bodies, hypothallus; presence or absence, colour, ordo Ceratiomyxales (3.5%); 2 (1 species) are members of shape, width. Myxomycetes specimen was identified ac- Echinosteliales (2.4%); 10 (9 species) are Liceales cording to references, from both the published literatures (11.8%); 24 (11 species) are Trichales (28.2%); 17 (8 spe- and web-based identification keys [17, 21-27]. The sam- cies) are Physarales (20%); and 29 (13 species) are ples were prepared as fungarium material and stored in the Stemonitales (34.1%). Among six ordo of myxomycetes, laboratory of Department of Biology, Faculty of Arts and Trichiales and Stemonitales are the most specious, whereas Science, Mustafa Kemal University. The relative abun- Ceratiomyxales and Echinosteliales are poorest in species. dance of fruit bodies of a particular species was determined The most common families are Stemonitidaceae, Trichia- by placing it in categories following a modified method ceae and Arcyriaceae. These percentages are similar to this proposed by Stephenson et al. (1993) [28]. For this, species study of Yağız & Afyon (2007) and Baba (2008) [11, 12]. represented by more than 3.0 % of the total number of col- The number and percentage of families are at Figure 2. The lections were considered as abundant (A), those falling be- most common genera are Arcyria, Comatricha and tween 1.5 % and 3.0 % as common (C), between 1.5 % and Perichaena with 5 species (each of genera), Didymium and 0.5 % as occasional (O), and those less than 0.5 % as rare Licea with 4 species, Cribraria and Stemonitis with 3 spe- (R). Taxonomic diversity indices (species per genera, S/G cies, Badhamia and Physarum with 2 species, Cerati- ratio) for the study sites as well as for the seasons were cal- omyxa, Echinostelium, Enteridium, Dictydiaethalium, Col- culated based on the methodology described by Stephen- laria, Enerthenema, Macbrideola, Stemonitopsis, Symphy- son et al. (1993) [28]. tocarpus and Trichia with 1 species. Myxomycetes commonly occur with decaying plant materials. In our study, species were collected from forest on 3. RESULTS AND DISCUSSION decomposed or dead wood (36 species), bark (20 species), debris (13 species), trunks (2 species), twigs (4 species), logs In this study 43 species belonging to three subclasis, (9 species) and on living Pinus sp. (1 species) (Figure 3). In six ordo, eleven familia and nineteen genera were identi- general, bark and woods were characterized by the highest fied using both field and moist chamber culture (Table 1). diversity and species richness [7, 9, 22, 26, 30]. Among sub- Totally, 85 samples (43 species) were collected. These strates, dead twigs, trunks and living substrates was charac- include; 9 Myxomycetes (3 species) were collected in field, terized by the lowest diversity of myxobiota. 76 myxomycete (34 species) were obtained in laboratory Forty myxomycetes were collected in March; 22 col- with moist chamber technique, and 6 species were obtained lected in October; 22 collected in December and 1 collected from both field and laboratory. in January. The best months for finding Plasmodial slime
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TABLE 1 - Myxomycetes sample number, substrates, habitat and seasons.
No Species S.N Substrates Habitat Season 1 Ceratiomyxa fruticulosa 2 a,b N, MCT Spring, Autumn 2 Echinostelium minutum 3 a,b,c,h,ı MCT Spring,Winter, Autumn 3 Cribraria cancellata 3 a N Spring, Winter, Autumn 4 C. violaceae 2 a MCT Spring, Winter 5 C. purpurea 1 a MCT Spring 6 Licea biforis 2 a MCT Spring, Autumn 7 L. castanea 3 a MCT Spring, Winter, Autumn 8 L. kleistobolus 2 a MCT Spring, Winter 9 L. tenera 1 a MCT Spring 10 Reticularia lycoperdon 2 f N Spring, Winter 11 Dictydiaethalium plumbeum 1 a MCT Spring 12 *Didymium clavus 2 a,b MCT Winter, Autumn 13 D. eximium 2 a MCT Spring, Autumn 14 D. difforme 2 k,l MCT Spring, Winter 15 D. squamulosum 2 a,h, MCT Spring, Winter 16 Badhamia macrocarpa 2 g MCT Spring, Autumn 17 B. utricularis 1 b MCT Autumn 18 Physarum leucopheum 2 e MCT Spring, Autumn 19 P. notabile 2 h MCT Spring, Autumn 20 Collaria lurida, 2 a,c MCT Spring, Autumn 21 Comatricha ellae 2 a,c,d,j N, MCT Spring, Winter 22 C. laxa 2 a,e MCT Spring, Autumn 23 C. nigra 3 a,d N,MCT Spring, Autumn, Winter 24 C. pulchella 2 e MCT Spring, Winter 25 C. tenerrima 1 h MCT Winter 26 Enerthenema papillatum 2 h MCT Spring, Winter 27 Macbrideola cornea 2 b MCT Spring, Autumn 28 Stemonitis flavogenita 2 a,b N,MCT Spring, Autumn 29 S. smithii 1 b MCT Spring 30 S. splendens 1 a MCT Spring 31 Stemonitopsis amoena 3 a,h MCT Spring, Autumn,Winter 32 Symphytocarpus impexus 2 a MCT Spring, Autumn 33 Arcyria cinerea 3 a,b,c N,MCT Spring, Autumn, Winter 34 A.pomiformis 3 e,h MCT Spring, Autumn, Winter 35 A. incarnata 3 a MCT Spring, Winter, Autumn 36 A. obvelata 3 b,h N,MCT Spring, Autumn, Winter 37 A. minuta 1 a MCT Spring 38 Perichaena corticalis 2 b MCT Spring, Winter 39 P. chrysosperma 1 a MCT Spring 40 P. depressa 2 a MCT Spring, Winter 41 P. vermicularis 2 c,i MCT Spring, Winter 42 *P. pedata 2 a MCT Spring, Autumn 43 *Trichia scabra 1 a N Spring Abbreviations: S.N: sample number; Substrates: Dead wood: a, Dead bark: b, Fallen twigs: c, Fallen bark: d, Dead log: e, Living substrat: f, Dead trunk: g, Dead debris: h, Fallen leaves:ı, Fallen branch: i, Dead twigs:j, Petri dishes: k, Filterpaper:l ; Habitat: MCT: Moist Chamber Technique, N: Natural; *: New records
35 30 The Number of the Families
25 The percentage of the Families 20 15 10 5 0
FIGURE 2 - The number and percentage of families
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45 40 Substrate of Myxomycetes 35 Percent of Myxomycetes 30 25 20 15 10 5 0 Dead Bark Dead Trunk Twigs Logs Living wood debris Pinus sp.
FIGURE 3- The distribution of myxomycetes as to where the development
FIGURE 4- Seasonal distribition of Myxomycetes
molds in our research are October, December and March are the two main factors in diversity and abundance of this (Figure 4). Because there is rain, relative humidity is ap- group [27]. parently optimum and temperature is mild. Primary char- In our study, 10 species are abundant (A), 9 species are acteristics of these months in our research area are the al- common (C), and 24 species are occasional (O) (Table 2). ternation of rainy and sunny periods. These provide favour- The mean number of species per genus (S/G) was calcu- able conditions of adequate levels of moisture and suitable lated from the data sets of study area. The research area temperatures to allow Plasmodial slime molds to complete species/genus ratio (S/G) is 2.3. As Simberloff (1970) and their life cycle. others have pointed out, a biota in which species are di- Arcyria cinerea, A. pomiformis, Comatricha ellae, C. vided among many genera is intuitively more diverse than nigra, Didymium difforme, and D. squamulosum are the one, in which most species belong to only a few genera. most common species in our investigation. Most Myxomy- Consequently, a low value for S/G implies a higher overall cota species are spread globally and in most studies these diversity than a high value [28]. This rate is calculated from species were observed to be most widely spread in many Stephenson et al. (1993) and from the data set of southern different substrates [26, 29-31]. Humidity and temperature India had 2.24 and North America 4.13. Alexopoulos [28]
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TABLE 2 - Myxomycetes name and occurrence
No Species Occurrence No Species Occurrence 1 Ceratiomyxa fruticulosa A 23 C. nigra A 2 Echinostelium minutum C 24 C. pulchella C 3 Cribraria cancellata O 25 C. tenerrima O 4 C. purpurea O 26 Enerthenema papillatum O 5 C. violaceae C 27 Macbrideola cornea O 6 Licea biforis O 28 Stemonitis axifera O 7 L. castanea O 29 S. flavogenita A 8 L. kleistobolus O 30 S. splendens O 9 L. tenera O 31 Stemonitopsis amoena C 10 Reticularia lycoperdon C 32 Symphytocarpus impexus O 11 Dictydiaethalium plumbeum O 33 Arcyria cinerea A 12 *Didymium clavus C 34 A. incarnata C 13 D. difforme A 35 A. minuta O 14 D. eximium O 36 A. obvelata C 15 D. squamulosum A 37 A. pomiformis A 16 Badhamia macrocarpa O 38 Perichaena chrysosperma O 17 B. utricularis O 39 P. corticalis O 18 Physarum leucopheum O 40 P. depressa O 19 P. notabile O 41 *P. pedata O 20 Collaria lurida A 42 P. vermicularis C 21 Comatricha ellae A 43 *Trichia scabra O 22 C. laxa A
reported that species diversity of myxomycetes is lower in smooth small spores, 6-8 μ in diameter [22]. The columella tropical forests than in temperate forests. is inconspicuous or represented merely by the thickened base of the peridium [26]. In the field it could be mistaken Three species are new records for Turkey; only for Diderma hemisphaericum (Bull.) Hornem. which, Didymium clavus (Alb. & Schwein.) Rabenh., however, has double peridium, pale stipes, smaller spores, Deutschl. Krypt.-Fl. 1: 280, (1844). and granular lime [32] (Table 3). Fructifications sporangiate, stipitate, total height up to Habitat: Especially broad-leaved trees, very rarely as a 1 mm, sporangia loosely gregarious or scattered, erect de- casual on bark of living trees in moist chamber culture [4], pressed-globose to discoid, deeply umblicate below, white dead leaves, twigs, wood [32] or greyish white or dark (when limeless) 0.5-1 mm in di- Distribution: Argentina, Bolivia, Brazil, Costa Rica, ameter. Stalk erect or sometimes bent, thick at the base, ta- Dominica, Germany, Haiti, India, Jamaica, Mexico, New pering upwards, longitudinally rugose, dark brown, lighter Zealand, Puerto Rico, USA, Venezuela. above, 0.2- 1 mm long, sometimes so short as to be con- tained within the base, the sporangium then appearing ses- Perichaena pedata (Lister & G. Lister) Lister ex E. sile. Hypothallus circular to venulose, membranous, dark Jahn, Ber. Deutsch. Bot. Ges. 36: 667, (1919). brown to almost black rotate. Peridium single, thin, mem- Fructifications sporangiate, scattered, solitary, stalked branous, dark grey, white crystals of lime except at the sporangia, 0.3-0.7 mm tall, the sporangium 0.3-0.5 mm thickened, umblicate base, dehiscence irregular. Columella diam., globose to subglobse, pale yellowish to orange inconspicuous, represented by the thickened, brown, brown. Hypothallus inconspicuous. Stalk slender longer dome-like base of the sporotheca and merely by the light than sporongial diam., erect, cylindrical, stout, roughened, brown, raised basal portion of the sporangium, sometimes dark brown to black, 0.4-0.6 mm high. Peridium single absent. Capillitium abundant delicate, flexuose composed membranous, dehisce into lobes. Capillitium abundant, of of purple brown, slender, sparsely branching and sparsely branching threads 3-4 μm diam. with scattered small spines anastomosing, hyaline. Spores sometimes with scattered or and small pits, often completly smooth, elastic, consisting distinctly clustered verrucose, black in mass, violet brown of yellow threads, the threads scarcely branched, with oc- in transmitted ligth, globose, inconspicuously verrucose to casional bulbous expansions, prominently marked with almost smooth, (5) 6-7 (8) μ in diameter. Plasmodium gray spines or minute scattered warts, 3-5 μm in diam. Spores or colourless (Figure 5). in mass bright yellow, in transmitted light yellow, 9-10 μm Locality: Hatay: Altınözü way, on Cupressus sp. bark, diam, covered in small spines. Plasmidium pale reddish- 304 m, 21.01.2011, Baba 5. Hatay: Kale hill, on dead Pinus brown (Figure 6). sp. wood, 509 m, 03.10.2011, Baba 32. Locality: Hatay: Altınözü way, on dead wood, 391 m, Didymium clavus is easily recognized by white, erect, 30.10.2011, Baba 4. discoid or depressed-globose sporangia, strongly umblicate The typical feature of Perichaena pedata is of the re- below, dark stipe and inconspicuously verrucose to almost lationship with P. chrysosperma. They have common fea-
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tures but its major resemblance is Hemitrichia minor G. threads of our specimens also resemble those of H. minor Lister, which has been classified in Perichaena by many in diameter and surface markings except for the spiral authors. Two species are certainly similar but P. pedata bands. The capillitial threads of H. minor bear faint spiral never has spiral markings on the capillitium and has a more bands [34] (Table 4). slender stalk, where as H. minor has a shorter, fatter stalk and faint spiral bands on the capillitium [26, 34]. The spo- Habitat: on straw and garden litter [26,34]. rangia of this species and H. minor are similar in shape and Distribution: England, Japan, Taiwan, USA. colour. They both have stalked sporangia. The capillitial
FIGURE 5- SEM photographs of Didymium clavus. A: sporangia, B: spore, C, D: capillitial threads.
TABLE 3 - Comparison of Didymium clavus according to different researchers
Thind (1977) [34] Stephenson (2003) [1] This study (2015)
Sporocarps Depressed- globose to discoit, deeply Scattered to gregarious, discoit, often Loosely gregarious, or scattered, erect depressed- umbilicate below white or greyish white umbilicate globose to discoid, white or greyish Peridium Single, thin, membranous, dark grey Membranous, covered lime chrystals Single, thin, membranous, dark grey, dehiscence dehiscence irregular, persisting below, above, greyish white or dark, thick- irregular translucent ened Stalk Erect or sometimes bent, thick at the base Tapering upward, longitudinally Erect, thick at the base, tapering upwards, 0.2-1 mm long striate, dark brown or black Capillitium Dense, delicate, composed of purple brown Abundant, delicate, threads nearly Abundant delicate, flexuose hyaline purple brown branching and anastomosing threads hya- colourless, sparsely branched line, branching and anastomosing Columella Inconspicuous, raised basal portion of Thickened discoid to dome-like base Inconspicuous raised basal portion, sometimes the sporangium of the sporotheca absent Spores Globose, inconspicuously verrucose to Pale violaceous brown nearly smooth Globose, violet brown almost smooth, (5) 6-7 (8) µm almost smooth 6-8 µm 6-7 µm Hypothallus Dark brown to almost black, rotate Membranous, discoid to venulose, Membranous, dark brown to almost black black
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FIGURE 6- SEM photographs of Perichaena pedata. A: sporangia, B: spores, C, D: spores and capillitial threads
TABLE 4 - Comparison of Perichaena pedata according to different researchers
Ing (1999) [26] Liu et al. (2007) [33] This study (2015) Sporocarps Yellow or pale brown 0.3-0.4 mm Globose to subglobse pale yellowish to Scattered, solitary, 0.3-0.7 mm globose to diam. extent, bright orange yellow orange brown, 0.3-0.6 mm subglobse, pale yellow or golden brown
Peridium Single membranous, dehiscing into Single, membranous, with deposition of Single, membranous, dehisce into lobes lobes granular matter
Stalk Slender black Erect, cylindrical, stout, roughened, dark Slender, erect, cylindrical, stout, dark brown to black roughened, brown to black Capillitium Dense, branching threads 3-4 µm, Consisting of yellow threads, threads Abundant, branching threads 3-4 μm Scattered small spines or smooth scarcely branched, with occasional bulb- diam, often completly smooth ous expansions, prominently marked with spines or minute scattered warts, 3-5 μm in diameter
Columella Absent Absent Absent Spores Globose, covered in small spines 9- Globose to subglobse minutely warted, Yellow, 9-10 μm diam in small spines 10 µm 9.5-10 μm in diam Hypothallus Inconspicuous Inconspicuous Inconspicuous
Trichia scabra Rostaf., Sluzowce monogr. 258, tightly clustered in large developments several centimetres (1875). in extent, bright or dull orange–yellow or golden brown. Sporocarps crowded in large groups, sessile sporangia, glo- Hypothallus membranous dark brown continuous below bose, cylindrical or turbinate (0.4) 0.5-0.7 (0.8) mm diam, the whole development and surrounding the bases of the
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sporangia so that when they have disintegrated it is left like Trichia scabra is usually characteristic deeply colored, a honeycomb of cups, thin. Stalk absent. Peridium single, strongly iridescent, densely crowded sporangia with dark thin membranous delicate smooth, shining partially eva- yellow or orange contents [22, 34]. Yellow or olivaceous nescent remaining as deep funnel-shaped calyculus. Colu- collections are easily distinguishable from Trichia favo- mella absent. Capillitium tubular, elastic deep yellow to ginea (Batsch) Pers., Trichia lutescens (Lister) Lister and rusty orange of simple elaters, 5-6 μm diam, marked with 3- Trichia varia (Pers. ex J.F. Gmel.) Pers. by the characters 4 tight, regular, spiral bands which are densely spinulose, the of spores and capillitium. The bright orange-yellow, ends short and pointed. Spores in mass deep orange-yellow, densely packed small, sessile sporangia on brown honey- in transmitted light yellow, (9) 10-12 μm diam, delicately comb hypothallus, the rough elaters and the delicately re- and completely reticulate. In optical section showing as a ticulate spores all help to identify these species. Habitats border of 0.2-0.5 μm thickness. Plasmodium white or oc- and colours of these species are much like those of the T. casionally pale yellow (Figure 7). favoginea but spores reticulation, fine meshed and faint [1]. Locality: Hatay: Altınözü way, on dead Pinus sp. (Table 5). wood, natural, 479 m, 21.03.2011, Baba 4.
FIGURE 7- SEM photographs of Trichia scabra A: sessile sporangia, B: capillitial threads and spore, C: spore and peridium.
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TABLE 5 - Comparison of Trichia scabra according to different researchers
Ing (1999) [26] Stephenson (2003) [1] This study (2015) Globose, tightly clustered in large de- Globose or turbinate, dull orange or Crowded in large groups globose, cylindri- Sporocarps velopments several centimetres in ex- golden brown cal or turbinate yellow- golden brown tent, bright orange yellow Peridium Smooth, shining Delicate, smooth, shining Single, thin, delicate smooth Stalk Sessile Sessile Sessile Deep orange yellow, 5-6 µm, marked Deep yellow to rusty orange 5-6 µm, Tubular, elastic deep yellow 5-6 µm, ends Capillitium with 3-4 tight, ends short and pointed bearing 3 or 4 closely wound short
Columella Absent Absent Absent Globose, 10-12 µm reticulate Globose, fine-meshed reticulum yel- Yellow, (9) 10-12 μm diam, completely re- Spores low, 10-12 µm ticulate Dark brown continuous below whole Continuous for a grouop of sporangia, Membranous dark brown Hypothallus of sporangia membranous, brown
Habitat: on rotten logs, wood and barks [22, 26] espe- [7] Heherson, N., Dagamac, A., Stephenson, S. and Cruz, cially elm and beech, characteristic of the ancient wood- T.E.E.D. (2014) The occurrence of litter Myxomycetes at dif- ferent elevations in Mt. Arayat National Park, Pampanga, Phil- land [34], decaying wood or bark [1]. ippines, Nova Hedwigia 98: 187-196 Distribution: Brazil, Costa Rica, Germany, Jamaica, [8] Shchepın, O., Novozhilov, Y. and Schnıttler, M. (2014) Nivi- Scotland, Venezuela. colous myxomycetes in agar culture: some results and open problems. Protistology, 8 (2): 53-61. [9] Tee, D.C., Mad, R., Tran, H.T.M., Ko, K.T.W. and Stephen- 4. CONCLUSIONS son, S.L. (2014) A comparative species listing of myxomy- cetes from tropical (Philippines) and temperate (United States) forests. Mycosphere, 5 (2): 299-311. In the study area, climatic conditions and vegetation are suitable for Myxomycetes, that we have observed very [10] Ocak, I. and Hasenekoğlu, I. (2003) Myxomycetes from Erzu- rum, Bayburt and Gümüşhane Provinces, Turk J Bot, 27: 223- rich Myxobiota. Three new species of Myxomycete rec- 226. orded for the first time from Kuseyr Mountains in Hatay- [11] Yağız, D. and Afyon, A. (2007) Three new records for Myxo- Turkey. mycetes of Turkey, Turk J Bot, 31: 470 - 476.
[12] Baba, H. (2008) A new Myxomycetes genus and three species record for Turkey, International Journal of Botany, 4: 336-339. ACKNOWLEDGEMENT [13] Ergül, C.C. and Akgül, H. (2011) Myxomycete diversity of Uludağ national park. Turkey, Mycotaxon 116:479. This study was supported by the Mustafa Kemal Uni- [14] Baba, H. (2012) Myxomycetes of Mustafa Kemal University versity Scientific Research Projects (BAP) (Project No: campus and environs. Turk J of Bot 36: 769-777. 1102 M 0102). [15] Demirel, G. and Kaşık, G. (2012) Four new records for Physar- ales from Turkey. Turk J of Bot, 36: 95-100 The authors have declared no conflict of interest. [16] Baba, H., Gelen, M. and Zümre, M. (2013) Three new Trichiales (Myxomycetes) record for Turkey. Fresenius Envi- ronmental Bulletin, 22(11): 3338-3342. REFERENCES [17] Lado, C. (2015) An on-line nomenclatural information system of Eumycetozoa. (Last updated May 29, 2015) http://www.no- [1] Stephenson, S.L. (2003) Myxomycetes of New Zealand. Hong men.eumycetozoa.com. Real Jardín Botánico, Madrid, CSIC. Kong: Fungal diversity Press. [18] Sesli, E. and Denchev, C.M. (2014) Checklists of the myxo- [2] Martin, G.W., Alexopoulos, C.J. and Farr, M.L. (1983) The mycetes, larger ascomycetes, and larger basidiomycetes in Genera of Myxomycetes, Univ. Iowa Pres. Iowa City, p. 438. Turkey. 6th edn. Mycotaxon Checklists Online [Updated and uploaded in February 2014] (http://www.mycotaxon.com/re- [3] Schnittler, M., Lado, C. and Stephenson, S.L. (2002) Rapid sources/checklists/sesli-106-checklist.pdf):1-136 biodiversity assessment of a tropical myxomycete assemblage - Maquipucuna Cloud Forest Reserve, Ecuador, Fungal Diver- [19] Davis, P.H. (1965-1985) Flora of Turkey and the East Aegean sity, 9: 135-167. Islands. Vols. 1-9, Edinburgh: Edinburgh Univ. Press. [4] Ko, K.T.W., Stephenson, S.L., Hyde, K.D. and Lumyong, S. [20] Duman, H., Vural, M. (1991) C6 karesinden yeni floristik (2011) Influence of Seasonality on the occurrence of Myxo- kayıtlar. Turk J of Bot 15(2): 201-203. mycetes, Chiang Mai Journal Science, 38 (1): 71-84. [21] Martin, G.W. and Alexopoulos, C.J. (1969) The Myxomycetes [5] Joshaghani, A.A., Falahian, F., Nejadsattari, T., Khavarinejad, Univ. Iowa Pres. Iowa City, p. 560. R.A. and Saadatmand, S. (2013) Two new myxomycetes rec- [22] Farr, M.L. (1976) Flora Neotropica, Monograph No:16. N.Y. ords for Iran mycoflora, Pak. J. Bot, 45(5): 1813-1816. Bot. garden. pp.298. [6] Liu, C.H., Chang, J.H. and Yeh, F.Y. (2013) Myxomycetes of [23] Neubert, H., Nowotny, W. and Baumann, K. (1993) Die Myx- Taiwan XXIV. The genus Physarum. Taiwania, 58(3): 176-188. omyceten (Band I), Gomaringen: Karlheinz Baumann Verlag.
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[24] Neubert, H., Nowotny, W. and Baumann, K. (1995) Die Myx- omyceten (Band II), Gomaringen: Karlheinz Baumann Verlag. [25] Neubert, H., Nowotny, W., Baumann, K. and Marx, H. (2000) Die Myxomyceten (Band III). Karlheinz Baumann Verlag Gomaringen.
[26] Ing, B. (1999) The Myxomycetes of Britain and Ireland. Slough, England: Richmond Publish Comp. [27] Stephenson, S.L. and Stempen, H. (2000) A Handbook of Slime Moulds. Timber Press, Portland, Oregon p. 183. [28] Stephenson, S.L., Kalyanasundaram, I. and Lakhanpal, T.N. (1993) A comparative biogeographical study of Myxomycetes in the Mid-Appalachians of Eastern North America and two regions of India, Journal of Biogeography, 20(6): 645-657.
[29] Farr, M.L. (1981) True Slime Molds, In Wm. C. Brown Comp., Dubuque, Iowa pp. 132.
[30] Stephenson, S.L. and Stempen, H. (1994) Myxomycetes: A handbook of Slime Molds, Timber Press, Portland, Oregon.
[31] Novozhilov, Y.K., Schnittler, M., Rollins, A.W. and Stephen- son, S.L. (2000) Myxomycetes from different forest types in Puerto Rico, Mycotaxon, 77: 285–299.
[32] Lado, C. and Pando, F. (1997) Flora Mycologica Iberica Vol. 2. Madrid, Spain, pp. 323. [33] Liu, C.H., Chang, J.H. and Yang, F.H. (2007) Myxomycetous Genera Perichaena and Trichia in Taiwan, Botanical Studies 48: 91-96.
[34] Thind, K.S. (1977) The Myxomycetes of India. I.C.A.R. New Delhi p.702.
Received: November 28, 2014 Revised: June 05, 2015 Accepted: June 26, 2015
CORRESPONDING AUTHOR
Hayri Baba Department of Biology Faculty of Arts and Science Mustafa Kemal University Alahan-31000 Antakya-Hatay TURKEY
E-mail: [email protected]
FEB/ Vol 24/ No 11c/ 2015 – pages 4077 - 4086
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MONITORING SOIL RESPIRATION DYNAMICS DURING THE BIODEGRADATION OF CRUDE OIL CONTAMINATED SOIL UNDER DIFFERENT ENVIRONMENTAL CONDITIONS
Xinping Zhu1, Pujia Yu2,3, Ni Han1, Hongtao Jia1,*, Xinjun Wang1, and Kobil Gafurovitch Kodirov4
1 College of Grassland and Environment Sciences, Xinjiang Agricultural University, Urumqi, 830052, China; 2 Northeast Institute of Geography and Agroecology, Chinese Academy of Sciences, Changchun 130102, China; 3 University of Chinese Academy of Sciences, Beijing 100049, China; 4 Agricultural College of Tajikistan Agrarian University, Dushanbe, 734017, Tajikistan
ABSTRACT soil nutrient availability, thereby affecting the soil respira- tion [2, 3]. Therefore, as the sensitive indicators to deter- Soil respiration was usually used to be the sensitivity mine microbial activity in contaminated soils, soil respira- indicators of contamination due to their importance in cy- tion was usually used as the valuable technique to test the cling of organic matter in soils. In order to understand the bioremediation treat-ability [4, 5]. effect of environmental factors on the bioremediation of Hydrocarbons, such as crude oil or petroleum, are the crude oil contamination at arid and semi-arid regions, the most common occurring environmental contaminants dynamics of soil respiration and degradation rate of total found in soil as a result of industrial activity. Jia et al. [6] petroleum hydrocarbons (TPH) were determined using a found that the total petroleum hydrocarbons in soil organic 59 days laboratory incubation experiment under a range of matter concentration accounted for about 60% in some oil crude oil contaminated levels, soil water contents and C:N fields of China, which indicated that hydrocarbons had be- ratio in soils. Results showed that crude oil contaminated come an important part of soil organic matter in contami- soil of lower C:N ratio (6:1) had significant greater soil res- nated soils. Contaminated soils are a system of great com- piration and TPH degradation than that with the higher C:N plexity and the behavior of many enzymatic activities in ratio (25:1). Although the TPH degradation rate decreased such soils may be very variable [7]. So, bioremediation of with the increase of contamination levels, the soil respira- hydrocarbons contaminated soils requires an understanding tion and TPH degradation of contaminated soil is higher of the processes that limit degradation in order to choose and than the uncontaminated soils, especially when the con- develop the most suitable methodology, especially in arid -1 tamination level was more than 20 g kg . Unlike the fac- regions where site conditions are very different to those tors of C:N ratio and contamination levels, the soil water found in temperate and humid regions [8]. The level of en- content has no significant effect on soil respiration and vironmental contamination depends on the chemical com- TPH degradation. These results indicated that low nitrogen position and concentration of the contaminants and the concentration rather than soil water content was the main properties of the soil [9]. Researchers have found that high limiting factors for soil respiration and TPH degradation in molecular weight compounds by very low solubility in wa- contamination soils at Northwest China when the soil wa- ter and high level contamination were not easily degraded ter content reach 20%. during the course of the degradation experiment [10].
The contamination of soils by hydrocarbons could in- KEYWORDS: soil respiration, total petroleum hydrocarbons, C:N duce changes of biological activity in soils. So, many bio- ratio, contaminated soils, Northwest China remediation techniques, such as optimizing conditions with the addition of nutrients, organic matter, microbial inocula, surfactants and also by aeration, control of temperature and 1. INTRODUCTION water content, was used to accelerate the naturally occur- ring biodegradation [11, 12]. The importance of nitrogen Soil respiration is the main output of soil carbon path- in amino acids to microbial processes has long been ways and an important source of atmospheric CO2, and it known. Contaminated soils that have intrinsically high C:N is also an important part of the world’s terrestrial ecosys- ratio will require nitrogen additions to allow a sufficient tem carbon cycle [1]. Different forms of C source can in- increase in biomass for environmentally significant hydro- fluence the activity of soil microorganisms by changing the carbon degradation to occur [8]. Many studies indicated positive effects from supplemental N, such as increase cell * Corresponding author growth rate [13], decrease the microbial lag phase [14], and
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increase the microbial activity and the rate of hydrocarbon petroleum hydrocarbons in contaminated soils at the arid degradation [10, 15]; however, large number paper report or semi-arid regions of Northwest China. no effect or even inhibitory effects when much higher lev- els of N was supplied in incubation experimental [8, 16- 18]. Soil water content is one of the important factors to 2. MATERIALS AND METHODS limit the soil respiration rate especially in the arid or semi- arid regions. Yu et al. [19] found that the mean value and 2.1. Soil sampling and experimental design the extreme deviation of soil respiration increased after the Soil Sampling was performed at the experimental plot increasing of soil water content in Gurbantunggute desert. of Xinjiang Agriculture University, Urmqi, China in Au- In addition, the relationship between nutrient and water gust 2012. The study area is characterized by an arid and content in soils is also having great effect on the soil respi- semi-arid desert climate. The annual average air tempera- ration and soil microbial activity, because of the toxic of ture is about 7.7℃, and annual average precipitation is about nutrients to microbial populations at high osmotic poten- 310 mm. The pan evaporation approximates 2570 mm [22]. tials [20, 21]. Walworth et al. [21] suggested that reporting The soil type was gray desert soil with soil texture of 36.62% N concentrations as a function of soil water rather than the sand (2-0.02 mm), 30.50% silt (0.02-0.002 mm), and more commonly cited concentration relative to dry soil 32.88% clay (<0.002 mm), which was classified as Aridisols weight. So it is important to estimate how much nitrogen in Chinese soil taxonomic system or as a Calcisols in World and water supplement is suitable for the biodegradation of Reference Base for Soil Resources (WRB). Hand shovel was contaminated soils. used to sample soil. Samples were collected at 0-20 cm soil layer. The soil samples were put in bags and transported to As part of a comprehensive investigation into factors the laboratory immediately after sampling. The basic prop- that affect the degradation of crude oil hydrocarbons at erties of the soil samples were shown in Table 1. Northwest China, this study focused on nitrogen, soil wa- ter, crude oil contaminated levels as a priority. In this All soil samples were air-dried, separated from the vis- study, the dynamics of soil respiration and degradation rate ible plant materials, and ground to pass through a 5 mm of total petroleum hydrocarbons were determined using a sieve to prepare for making the different crude oil level laboratory incubation experiment under a range of crude contaminated soil. The crude oil used in this experiment oil contaminated levels, soil water contents and C:N ratio was bought from the oil transportation department of Kara- in soils. The objectives of this study were to: (1) evaluate may oilfield in Xinjiang. Different weights of crude oil the effect of environmental factors on the dynamics of soil were added into 2000 mL petroleum ether (Boiling point respiration at a crude oil contaminated soils; (2) assess the 30-60℃) to make the crude oil solution. Then, we mixed the best soil environmental factors for the degradation of total crude oil solution with the sieved soil to homogenize the
TABLE 1 - The basic properties of the soil samples
Organic carbon Total salt Total nitrogen Total phosphorus Total potassium pH (g kg-1 ) (g kg-1 ) (g kg-1 ) (g kg-1 ) (g kg-1 )
46.51 0.58 0.73 1.75 17.91 7.87
TABLE 2 - The experimental scheme Crude oil concentration Treatment No. Moisture content (%) C:N ratio Combination (g kg-1 soil) 1 Ck 50 6:1 A1B1C1 2 Ck 20 25:1 A1B2C2 3 0 20 6:1 A2B2C1 4 0 50 25:1 A2B1C2 5 5 50 6:1 A3B1C1 6 5 20 25:1 A3B2C2 7 10 20 6:1 A4B2C1 8 10 50 25:1 A4B1C2 9 15 50 6:1 A5B1C1 10 15 20 25:1 A5B2C2 11 20 20 6:1 A6B2C1 12 20 50 25:1 A6B1C2 13 40 50 6:1 A7B1C1 14 40 20 25:1 A7B2C2 15 80 20 6:1 A8B2C1 16 80 50 25:1 A8B1C2
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samples. These soil samples with different levels of crude flasks, mixed and the Erlenmeyer flasks were put on a 55 ℃ oil contamination were placed in a fume hood for 15 days water bath shaker again for 0.5 h. After collecting the extract to make sure that the petroleum ether completely volatized solution to the beaker, the above procedure was repeated from the soil sample. In addition to a control sample (with- twice. The collected extract solution was concentrated to out petroleum ether and crude oil), soil samples containing dryness in a 58 ℃ water bath evaporator. The TPH content -1 crude oil at concentrations of 0, 5, 10, 15, 20, 40, 80 g kg was determined by gravimetric analysis, and the increase soil were also used in this experiment. gravimetric was the extracts of trichloromethane. The re- The experiments were carried out according to the de- covery rate of the result was over 99%. The TPH degrada- 3 -1 sign of orthogonal table L16(8×2 ) (Table 2). Except the tion content (g kg soil) and degradation rate (%) was cal- different crude oil level contaminated soil, the other two culated as follows: factors were designed to test the effect of environment con- CTPH=Ct-Cm ditions on the biodegradation of crude oil contaminated soil. The three factors were labeled as A (crude oil contam- DRTPH=(CTPH/Ct)100 inated soil), B (soil moisture content) and C (C:N ratio). where CTPH is the TPH degradation content in each A1, A2, A3, A4, A5, A6, A7, and A8 represent the crude sample, Ct is the total TPH content in each sample before oil contamination level of Ck, 0, 5, 10, 15, 20, 40, and 80 incubation, and Cm is the measured TPH content in each g kg-1 soil. The Ck treatment means only 2000 mL petro- sample after incubation; DRTPH is the TPH degradation rate leum ether was added in the soil sample. The 0 g kg-1 soil in each sample. (matrix black) means the air-dried soil, which is no crude oil and petroleum ether was added in it. Deionized water 2.4. Statistical data analyses (DI water) was added to the soil to 50% (B1) and 20% (B2) All statistical analyses were carried out with the of maximum water holding capacity, which was main- SPSS16.0 software package (SPSS 16.0 for windows, USA). tained by adding water periodically (2 days) throughout the The means of three replicates and standard error for each var- incubation duration. C:N ratio of 6:1 (C1) and 25:1 (C2) in iable measured (i.e. soil respiration rate, total soil respiration, these soil samples was produced in this experiment using and biodegradation rate) are provided. One-way ANOVA carbamide (CO(NH2)2). In order to calculate the adding was conducted to test the treatment effects (i.e., soil water weights of carbamide for each soil sample, the soil organic content, C:N ratio) for total soil respiration and biodegra- carbon concentration was determined using the K2Cr2O7- dation rate. Treatment significance was determined at sig- H2SO4 oxidation method [23] and total nitrogen concentra- nificant level of 0.05. tion was measured using the Kjeldah method [24].
2.2. Soil respiration rate measurement 3. RESULTS AND DISCUSSION Three replicates per treatment soil (each 1 kg dry weight) were incubated in plastic pans (Top width×Bottom Soil respiration is defined as the release of CO2 from width×Height: 17cm×13cm×10cm) at the room tempera- soil to atmosphere, and includes soil microbial respiration, ture. The top of plastic pans covered with plastic wrap to soil fauna respiration, and plant root respiration [26]. So, prevent the loss of soil water, and five small ports in each soil respiration has been widely used to understand the mi- plastic wrap was left for gas exchanging. In order to main- crobial response to contaminated soil and to monitor pol- tain the soil moisture content, DI water was added period- lutants degradation [10, 12]. Many factors, such as the car- ically (2 days) throughout the incubation duration accord- bon composition, the microbial activity, and environmental ing to the weight difference. Soil respiration rate were de- factors can be effect the soil respiration rate. Incubations termined at the 10:00-11:00 am of incubation days 1, 2, 3, are usually limited by the fact that the truly resistant C with 4, 5, 6, 7, 8, 9, 10, 11, 12, 14, 16, 21, 25, 30, 47, and 59 mean residence times of millennia is not accessed by the using CIRAS-2 photosynthetic apparatus connected to the microbes, so respiration rate was usually leveled off by the SRC-1 soil chamber (PP System’ analyzer, England). At end of incubation [27, 28]. Similar with the previous study, the end of the 59 day experiment, pans were sacrificed for respiration rate in this study almost leveled off for all treat- analysis of TPH. ment soils after 6 days incubation, but differences in the respiration rates among the treatments were still apparent 2.3. Total petroleum hydrocarbons (TPH) at the end of incubation (Fig. 1). Like the change trend of Using the gravimetric analysis method [25] to deter- soil respiration rate during the incubations days in other mine the TPH content. 15 g air-dried soil samples and studies [12, 29], soil respiration rate of most treatments 30 ml trichloromethane (CHCl3) were put into 100 ml Er- soils in our study increased in the beginning incubation lenmeyer flasks. Then the Erlenmeyer flasks were shaken days, and almost peaked at the incubation day 4. The in- for 2 mins to make sure that the solution fully mixed and crease and slight fluctuation of soil respiration rate may be then were stilled for 24 hours. The stilled Erlenmeyer ascribed to the physiological answer of microbial commu- flasks were put on a 55 ℃ water bath shaker for 1 hour and nity to the environmental factors change; the more suitable the extract solution was collected in a 100ml beaker. Then environmental conditions increase the microbial activity 15 ml trichloromethane was added into the Erlenmeyer and microbial biomass [29, 30]. The behavior of leveled off
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confirmed that a lack of optimum microbial activity in the higher C:N ratio (25:1, lower supplement of nitrogen) (Fig. 1). final phase when nutrients were reduced. Throughout 59 The highest soil respiration rate of lower C:N ratio was about days of incubation, average soil respiration rate of contami- 8 g m-2 h-1 and it peaked in the early incubation days, while nated samples was 38.8% higher than the uncontaminated the highest soil respiration rate of higher C:N ratio was -2 -1 soils, which indicated that the crude oil stimulated CO2 evo- only about 2.6 g m h and there were almost no big lution. The lowest value of average soil respiration rate was changes in all treatments. Ferguson et al. [8] found similar observed at the A3B2C2, while the highest value was found result that natural biomineralisation of petroleum hydrocar- at the A8B2C1. These results reflected that the three factors bons at 10 ℃ in the soil is limited by lower nitrogen concen- (crude oil contaminated soil, soil water content, and C:N ra- trations, and the mineralization of unamended microcosm tio) have great effect on the soil respiration rate. Results of treatments with 85 mg N kg-1 soil was almost 10 times less the univariate analysis of variance showed that C:N ratio (F= than the highest mineralization in nitrogen–amended treat- 46.47, P < 0.001) and crude oil contamination level (F= 4.557, ments. However, much more excessive levels of N supple- P < 0.05) have a significant effect on the average soil respira- ment may be have no benefit, or even deleterious effects tion rate , while soil water content has no significant effect on [18, 20]. Walworth et al. [20] found that when the C:N ratio the average soil respiration rate (F=2.519, P = 0.16). of soil was much lower, hydrocarbon mineralization was Nitrogen is the most often limiting nutrients for soil severely inhibited, because the sandy soil used in their microbial community to support microbial activity and bi- study was more sensitive to over-fertilization due to its lim- ological hydrocarbon degradation in soils [21]. Many stud- ited water holding capacity and the over- fertilization was ies reported that supplemental N has positive effects on the more toxic to microorganisms than the chemically inhib- hydrocarbon degradation of crude oil contaminated soil, ited. In the current study, higher soil respiration rate was because N could decrease the microbial lag phase and help observed at the crude oil contaminated soil with lower C:N to maintain microbial community at high activity levels [8, ratio, suggesting that the soil used has not reach the over- 15]. In the present study, C:N ratio has significant effect on fertilization level and microbial has adapted to the soil en- the soil respiration rate. Crude oil contaminated soil of vironments. lower C:N ratio (6:1, higher supplement of nitrogen) had Previous studies showed that soil organic matter content significant greater soil respiration rate than that with the and soil respiration rate was positively correlated [12, 31].
9 A 9 B 8 8 A2B2C1 ) ) -1 7 -1 h 7 A4B2C1 h -2 A1B1C1 -2 A6B2C1 6 6 A3B1C1 A8B2C1 5 5 A5B1C1 4 A7B1C1 4 3 3 2
Soil respiration rate (g m 2 Soil respiration rate (g m 1 1 0 0 1 2 3 4 5 6 7 8 9 10 11 12 14 16 21 25 30 47 59 12345678910111214162125304759 Days Days
3.0 3.0 C D 2.7 2.7 A2B1C2
A1B2C2 )
) A4B1C2 -1 2.4 -1 2.4 A3B2C2 h h -2 A6B1C2 -2 2.1 2.1 A5B2C2 A8B1C2 1.8 A7B2C2 1.8 1.5 1.5 1.2 1.2 0.9 0.9
Soil respiration rate (g m 0.6
Soil respiration rate(g m 0.6 0.3 0.3 0.0 0.0 1 2 3 4 5 6 7 8 9 10 11 12 14 16 21 25 30 47 59 1 2 3 4 5 6 7 8 9 10 11 12 14 16 21 25 30 47 59 Days Days
FIGURE 1 - Daily soil respiration during the biodegradation of crude oil contaminated soil. Values are the means of three replicates. (A) soil water content 50%, C:N ratio 6:1; (B) soil water content 20%, C:N ratio 6:1; (C) soil water content 20%, C:N ratio 25:1; (D) soil water content 50%, C:N ratio 25:1. The mean of combination treatment (A, B, and C) is listed in Table 2.
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The more the soil organic matter has, the more sub- current study also verified the Ferguson et al. [8] conclu- strate can be used by the microbial community, and then sions that low nutrient concentrations rather than water the soil respiration rate was higher. The different concen- content was the main limiting factor for biodegradation of tration of crude oil contaminated soil had different effect hydrocarbon in the soil when incubated at 10 ℃. The rea- on soil respiration rate (Fig. 1). When the crude oil concen- sons for the results of no effect on the respiration may be tration was less than 20 g kg-1, there was no significant dif- due to that all the water content of 20% and 50% can meet ference for the soil respiration rate between the contami- the requirement for microbial population to maintain high nated soil and uncontaminated soil (P>0.05), which indi- activity levels [35]. cated that microorganism had good resistance to the low Throughout 59 days of incubation, the total soil respi- oil-contaminated soil. When the crude oil concentration -1 ration can be used to assess microbial activity and biodeg- was greater than 20 g kg , the soil respiration rate was sig- radation. It has been shown that respiration correlates with nificant higher than the uncontaminated soil (P<0.05). The contamination level, pointing towards a relationship be- results in this study were as same as Saviozzo et al. [12] tween TPH degradation and CO2 production [36]. Increased findings that the increase of crude oil concentration stimu- total soil respiration was observed for contaminated soil lated CO2 production, increased C mineralization and total samples as compared to uncontaminated soils in our study microbial activity. During the 59 days incubation, higher (Fig. 2), especially for contaminated soils with 40 and 80 g variability of soil respiration rate was observed in contam- kg-1 soil. Liner correlation and regression analysis showed inated soil samples regardless the soil water content and significant correction between 6 days and 59 days total res- soil C:N ratio (Fig. 1), this indicated the utilization of car- piration (R2=0.625, P< 0.05), which indicated that the bon source with higher conversion efficiencies in contam- higher variability of soil respiration rate of soil samples inated soils. with lower C:N ratio (6:1) didn’t change the overall trend As one of the most important environmental factors, for all 59 days incubation. Regardless the soil water content soil water content has a positive effect on soil respiration, and incubation days (6 and 59 days), the total soil respira- especially when the soil is dry and the soil temperature is tion of C:N ration for 6:1 is significant higher than it for -1 suitable for microbial activity [32]. However, soil water 25:1 except the soil samples of 0 and 15 g crude oil kg content may has a negative effect or irrelevant with the soil soil at the 59 days incubation. The result also indicated the respiration [33]. In this study, the soil respiration rate of great effect of the C:N ratio on soil respiration and biodeg- soil samples at 50% soil water content have no clearly radation. change tendency compare with it at 20% soil water content Like the change trend of soil respiration rate and total (Fig. 1). Even the changes of soil water content from 10% soil respiration, the total TPH degradation increased with to 60%, Jin et al. [34] also found that biodegradation activ- the increase of crude oil contamination level after 59 days ities and CO2 production were not distinguishable. Results incubation (Fig. 3). The highest degradation value was of the univariate analysis of variance also indicated that 33.38 g TPH kg-1 soil, which found at the 80 g TPH kg-1 soil soil water content has no significant effect on the soil res- with lower C:N ratio and lower soil water content, while piration rate (P > 0.05) in this study. These results in the the lowest value was 3.93 g TPH kg-1 soil, which found at
2400 )
-2 2000 6 days total respiration 59 days total respiration 1600
1200
800
400 Total soil respiration (g m (g respiration soil Total
0
2 C1 1C2 C2 2C1 C C1 1B1 1B2C2 2B 3B1C1 3B2C2 4B2C1 4B1 5B1C1 6B 6B1 7B1C1 7B2C2 8B2 8B1C2 A A A2B2C1 A A A A A A A5B2C2 A A A A A A Treatment group
FIGURE 2 - Total soil respiration at incubation days 6 and 59 of crude oil contaminated soil. Values are the means of three replicates (±SE). The mean of combination treatment (A, B, and C) is listed in Table 2.
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90 40
80 35
70 TPH biodegradation rate 30
60 TPH biodegradation soil) 25 -1 50 20 40 15 30
TPH degradation rate (%) rate TPH degradation 20 10 TPH degradation (g kg TPH degradation 10 5 0 0 A3B1C1 A3B2C2 A4B2C1 A4B1C2 A5B1C1 A5B2C2 A6B2C1 A6B1C2 A7B1C1 A7B2C2 A8B2C1 A8B1C2 Treatment groups
FIGURE 3 - TPH degradation and TPH degradation rate after 59 days incubation of crude oil contaminated soil. Values are the means of three replicates (±SE). The mean of combination treatment (A, B, and C) ) is listed in Table 2.
the 5 g TPH kg-1 soil with higher C:N ratio and higher soil piration rate than that with the higher C:N ratio (25:1). The water content. Compared with the high crude oil contami- highest soil respiration rate of lower C:N ratio was about nation level (20 g TPF kg-1 soil), the low crude oil con- 8 g m-2 h-1, which is more than 3 times than the highest soil taminated soils have the higher degradation rates (Fig. 3). respiration rate of higher C:N ratio (2.6 g m-2 h-1). Although Saviozzi et al. [12] study the effect of compost on diesel the soil respiration of low contaminated level soils (less contaminated soil degradation through 120 days incuba- than 20 g kg-1) was higher than the uncontaminated soil, tion, and they found that the decrease of TPH was about there was no significant difference between them. When 60-66%. Except the contaminated soil of 5 and 10 g TPH the crude oil concentration was greater than 20 g kg-1, the kg-1 soil, the degradation rate of other contamination level soil respiration rate was significant higher than the uncon- in our study was lower than 60%. This mainly due to the taminated soil (P<0.05). The soil water content has no sig- high contamination level used in our study, which the die- nificant effect on the soil respiration, which indicated that sel oil contaminated level used in Saviozzi et al. [12] study the water content of 20% and 50% all can meet the require- was only 10 g kg-1 soil. The lowest degradation rate of ment for microbial population to maintain high activity at 10.47% was found at A7B2C2, which was much lower the Northwest China. Clearly, getting the right environ- (P<0.05) than the degradation rate of other contamination mental factors, such as C:N ratio and soil water level is levels. The main reasons may be due to high concentration very important when considering the biodegradation of of crude oil enhanced toxicity to microorganisms, and the crude oil contaminated soils. The main conclusions found two limitation of lower soil water content and higher C:N from this research are that TPH degradation rate was al- ratio make the toxicity more than other samples. Many most more than 30%, and lower C:N ratio has a significant studies showed that volatilization has a great effect on the effect on the degradation of TPH, while soil water content decrease of TPH in incubation experiment. Saviozzi et al. has no significant effect. So, when the soil water content [12] found that the decrease value of volatilization can be was more than 20%, the best way to increase the degrada- reach to 40%, but they think the frequent mixing of soil tion of TPH for contaminated soils at the Northwest China samples was the main reason for the high volatilization. was to supply enough N to the soils. Our result of the lowest degradation rate of 10.47% indi- cated that the volatilization decrease of TPH in our study is less than 10%, which is close to the result of Namkoong et al. [37]. The lowest volatilization decrease of TPH and ACKNOWLEDGEMENT high degradation rate in our study also indicated that the microbial community plays an important role in the degra- This work was supported by the Strategic Priority Re- dation of crude oil contaminated soils. search Program – Climate Change: Carbon Budget and Re- lated Issues of Chinese Academy of Sciences (XDA05050504), the International Science and Technol- 4. CONCLUSIONS ogy Cooperation Program (2010DFA92720-13), and Xin- jiang Key Disciplines of Soil Science Foundation. The current study showed that crude oil contaminated soil of lower C:N ratio (6:1) had significant greater soil res- The authors have declared no conflict of interest.
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Received: December 06, 2014 Revised: June 08, 2015 Accepted: June 26, 2015
CORRESPONDING AUTHOR
Hongtao Jia 311 Nongda East Road Urumqi P.R. CHINA
Phone: +86- 13609915585 E-mail: [email protected] [email protected]
FEB/ Vol 24/ No 11c/ 2015 – pages 4087 - 4094
4094 © by PSP Volume 24 – No 11c. 2015 Fresenius Environmental Bulletin
THE EFFECT OF PLANTING GEOMETRY AND SEED PRIMING ON SUNFLOWER YIELD UNDER RAIN-FED CONDITIONS
Mehmet Demir Kaya1,*, Suay Bayramin2 and Gamze Kaya3
Department of Field Crops, Faculty of Agriculture, Eskişehir Osmangazi University, 26160 Turkey 2 The Central Research Institute of Soil, Fertilizer and Water Resources, Yenimahalle, Ankara, Turkey 3 Provincial Directorate of Turkish Republic, The Ministry of Food, Agriculture and Livestock, 26160 Eskisehir, Turkey.
ABSTRACT 1. INTRODUCTION
Sunflower is an indispensable crop for rain-fed condi- Sunflower (Helianthus annuus L.) is a major oil seed tions in Turkey, where productivity of sunflower has been crop in arid and semi-arid regions due to its high edible oil dependant on rainfall during growing season. This research content in the seeds, high adaptability, low labor needs and was carried out to investigate the potential of row direction, suitability for mechanization [1, 2]. In Turkey, sunflower plant arrangement and seed priming in increasing yield and is cultivated on an area of 550.000 hectares with an annual oil ratio of sunflower under rain-fed conditions in Turkey. seed production of 1.480.000 tons, giving an average seed The experiment was established during the summer sea- yield of 2690 kg ha-1 [3]. Approximately, 70% of total sun- sons of 2010 and 2011 using four row directions (North- flower cultivation has been done under rain-fed conditions. East, South-West, Northeast-Southwest and Northwest- Its tolerance to drought makes sunflower more important un- Southeast), three plant arrangements (70 × 30, 60 × 35 and der dry conditions in Turkey, where the local climatic condi- 50 × 42 cm) and seed priming treatments (control, hydro- tions are characterized as semi-arid due to irregular and insuf- priming and KNO3) with the cultivar Sanbro MR. In the ficient rainfall and high temperatures during the sunflower study, days to flowering, plant height, head diameter, thou- growing season [4]. Average productivity of sunflower un- sand seed weight, seed yield, oil ratio and photosynthetic der these conditions is relatively low because it is subjected parameters were investigated. The results showed that seed to high temperatures and water limitation during blossom- yield, yield components, oil ratio, and photosynthetic pa- ing, which is the most drought-sensitive growth stage [5, rameters were higher in 2011 due to high precipitation and 6]. However, some agronomic practices can mitigate low temperature. Seed yield obtained from all row direc- drought effects. Early sowing, which allows the plants to tions ranged from 2106-2367 kg ha-1 in 2010 and 4115- use the late winter and early spring rainfall [7], and using 4263 kg ha-1 in 2011, with no significant differences be- drought tolerant cultivars, as reported by several research- tween row directions. None of the row directions tested en- ers [8, 9], reduces the effects of drought on plant growth hanced seed yield or oil content. Hydropriming and the and seed yield. plant arrangement of 60 × 35 cm were found to be effective Alternative agricultural practices to improve seed yield for increasing seed yield. The rate of photosynthesis also of sunflower under dry conditions should be developed. was higher in 2011, along with the investigated agronomic Row direction, plant arrangement and seed priming are characteristics. Seed priming increased the rate of photo- promising approaches to increasing seed yield in sunflower synthesis and the increased photosynthetic activity was without additional expenses. Shafiullah et al. [10] reported evaluated as an indicator of improved seed yield. It was that northwest to southeast (NW-SE) row orientation in- concluded that a plant arrangement of 60 × 35 cm and hy- creased seed yield of sunflower, although results were not dropriming should be the preferred techniques for increas- statistically significant. Diepenbrock et al. [11] found higher ing seed yield of sunflower regardless of row direction. seed yields in plants oriented in east-west rows, while Rob-
inson [12] determined that there were no significant differ-
ences in seed yield between north-south and east-west orien-
KEYWORDS: Helianthus annuus L.; planting geometry; priming; tations. However, Kaul et al. [13] found higher fruit pro- row orientation; photosynthesis duction in okra plants when they were planted in a north- south direction. Also, wheat and barley crops oriented east-
west had lower weed biomass and higher grain yield than
crops oriented north-south [14]. The studies on row direc-
tion have given inconsistent results, and this subject needs
to be studied in more depth. Optimum sunflower plant den- * Corresponding author sity varies with climate, soil properties and plant growth
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habit. Higher plant density has been preferred in fertile and after, the seeds were rinsed with tap water three times. The irrigated fields, while lower densities have been used in treated seeds were surface-dried back to their original arid areas. Vahedi et al. [15] reported that equidistant inter- moisture content at room temperature (about 22 °C, 45% and intra-row spacing with a constant density gave higher relative humidity) as determined by changes in seed seed yield relative to wider row spacing and narrow intra- weight. Moisture content of untreated seeds (control), hy- row distances. On the other hand, plant arrangement can droprimed and KNO3 treated seeds was equilibrated at optimize land use for plant productivity and provides room temperature for 2 days. shaded areas on the soil surface, preventing evaporation of Photosynthetic parameters, including net photosyn- moisture from soil [16, 17]. Seed priming increased the thetic rate (A), stomatal conductance (gs) and transpiration germination performance and decreased germination time rate (E), were determined using an LCi (leaf chamber anal- in sunflower under drought and saline conditions [18] and ysis) portable photosynthesis system (ADC, Analytical De- shortened the time to emergence [19-21]. The superiority velopment Company, Hoddesdon, UK) with a leaf cham- of primed seeds is exhibited in seed yield and oil content, ber area of 11.25 cm2. All measurements were performed as reported by Hussain et al. [22], who demonstrated a 10% on the upper third fully expanded leaves. Three measure- increase in the yield of sunflower. ments were done at two day intervals during flowering, and The objective of the present study was to determine the the data were pooled for statistical analysis. Each measure- appropriate combination of row direction, plant arrange- ment was recorded from 10:00 to 11:30 h. ment and seed priming to increase seed yield of sunflower At maturity, ten plants were randomly selected from under dry conditions in Central Anatolia, Turkey. each plot and were used to determine yield and yield com- ponents. The oil percentage was determined by Soxhlet ex- traction (Gerhardt Soxtherm 414, Germany) method using 2. MATERIALS AND METHODS n-Hexane as a solvent. The combined analysis of variance of the data and the comparison of the means on the basis of This study was conducted at the experimental field of Duncan’s Multiple Range Test were carried out using the Central Research Institute for Field Crops in Haymana- MSTAT-C (Michigan State University) software. Ankara, Turkey during the summer seasons of 2010 and 2011. The soil type in the experimental field was clay loam, and the soil was slightly alkaline (pH 7.6-8.1), rich in po- 3. RESULTS AND DISCUSSION tassium (385-404 kg ha-1), intermediate in phosphorus (107- -1 149 kg ha ) and low in organic matter (1.33%). The climate Mean squares with significance levels of the main ef- was characterized by hot, dry summers and cold, snowy win- fects, two and three way interactions for all of the investi- ters. Rainfall from sowing to harvest was 131 mm in 2010 gated yield components and oil ratio of sunflower as influ- and 172 mm in 2011, with a long-term average of 166 mm. enced by row direction, plant arrangement and seed prim- As shown in Figure 1, precipitation in May and June in ing are shown in Table 1. 2011 occurred considerably higher than in 2010. Mean temperatures during the growing period were 17.6 °C and The average values of the investigated parameters in 16.4 °C in successive years. 2011 were higher than in 2010 (Table 1). Higher precipita- tion and lower temperature in 2011 relative to 2010 caused The sunflower hybrid cv. Sanbro MR, extensively cul- significant differences between two years. As shown in tivated in dry conditions, was sown on 22 April 2010 and Figure 1, the high rainfall in April, May and July in 2011 27 April 2011. In both years, 150 kg ha-1 N and 375 kg ha- resulted in promoting plant growth and increasing seed 1 P2O5 (DAP, Diammonium phosphate) were applied be- yield of sunflower. Also, the factors examined did not sig- fore sowing according to the results of soil analysis. Pre- nificantly influence the number of days to flowering, plant emergence herbicide (Linuron 450 g L-1) was applied to height or thousand seed weight. No significant main effects control weeds after sowing and the field was manually of row direction were found on days to flowering, plant hoed once at 30 days after emergence. Plants were grown height, head diameter, thousand seed weight, seed yield or under rain-fed conditions and no irrigation was performed. oil ratio in either year. The NW-SE direction produced Four row directions [(north-south (N-S), east-west (E- slightly higher seed yield than the other directions. How- W), northeast-southwest (NE-SW) and northwest-southeast ever, plant arrangement significantly affected plant height (NW-SE)], were assigned to the main plots. Plants were ar- and seed yield in 2010 and head diameter, seed yield and ranged in three levels (70 × 30, 60 × 35 and 50 × 42 cm) oil ratio in 2011. In contrast to oil ratio, greater plant with a constant density (4.8 plants m-2) in sub-plots, and height, head diameter and seed yield were obtained in plants from the 60 × 35 cm plant arrangement. Seed treat- seed primings (Control, KNO3 and hydropriming) were performed as described by Kaya et al. [18] in sub-sub plots. ments significantly affected head diameter, seed yield and For hydropriming, sunflower seeds were immersed in dis- oil ratio in the 2010 growing season. tilled water at 25 °C for 18 h under dark conditions. For A significant two-way interaction of row direction × plant KNO3 treatment, the seeds were immersed in 500 ppm arrangement for plant height, head diameter, thousand seed KNO3 solution at 25 °C for 2 h in the dark condition. There- weight and seed yield was pronounced in 2010, but was not
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present in 2011 (Table 2). The tallest plant was located in between 42.3 g and 49.9 g, and plant arrangements of 70 × the NW-SE and 50 × 42 cm treatment. Similarly, the max- 30 cm in NE-SW and NW-SE, 60 × 35 cm in N-S and 50 imum head diameter (15.9 cm) was recorded in the NW- × 42 cm in E-W gave the highest thousand seed weights. A SE and 50 × 42 cm treatment. Thousand seed weight varied similar trend was observed in seed yield, which ranged be-
TABLE 1 - Main effects of row direction, plant arrangement and seed priming on the investigated traits of sunflower in 2010 and 2011
Days to flower- Plant height (cm) Head diameter Thousand seed Seed yield Oil ratio (%) Traits ing (day) (cm) weight (g) (kg ha-1) Year 2010 2011 2010 2011 2010 2011 2010 2011 2010 2011 2010 2011 Row direction (A) N-S 78.2 83.3 127 165 14.8 24.3 43.9 65.3 2106 4115 36.6 48.5 E-W 78.5 83.8 127 165 14.9 24.1 47.2 66.2 2201 4178 38.5 48.7 NE-SW 78.2 83.5 127 157 14.9 24.6 47.2 67.3 2208 4263 38.7 47.6 NW-SE 78.3 83.5 129 167 15.5 24.4 48.1 67.4 2367 4205 39.0 47.9 Plant arrangement (B) 70×30 cm 78.0 83.6 128ab 166 15.0 24.1b2 46.7 66.5 2222ab 4084b2 38.1 48.5a1† 60×35 cm 78.0 83.6 129a 161 15.1 24.8a1 47.0 66.8 2289a 4397a1 38.5 47.5b2 50×42 cm 79.0 83.6 126b 163 15.0 24.2b2 46.0 66.3 2150b 4089b2 37.9 48.5a1 Seed priming (C) Control 78.3 83.6 127 166 14.8b 24.5 46.1 66.1 2156b2 4199 38.7a 48.4 Hydropriming 78.2 83.4 128 161 15.3a 24.4 47.1 66.8 2274a1 4229 38.4a 48.1 b a12 b KNO3 78.4 83.7 127 163 14.8 24.2 46.6 66.8 2232 4142 37.5 47.9 F test A ns ns ns ns ns ns ns ns ns ns ns ns B ns ns * ns ns ** ns ns * ** ns * A×B ns ns ** ns ** ns ** ns ** ns ns ns C ns ns ns ns ** ns ns ns ** ns * ns A×C * ns ns ns * ns ** ns ns * ns ns B×C ns ns ns ns * ns ns ns ns ns * ns A×B×C ns ns ns ns ** ns ns ns ns ns * ns *, ** significant at p < 0.05 and p < 0.01 respectively; ns: not significant. †: Means followed by the same letter(s) in each column are not significantly different at p < 0.05.
100 2010 2011 90 80 70 60 50 40 30
Precipitation (mm) 20 10 0 April May June July August September Months
FIGURE 1 - Monthly precipitation at the experimental location during sunflower vegetation period
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TABLE 2 - Interaction of row direction × plant arrangement on yield and yield components of sunflower in 2010
Row direction Plant arrangement N-S E-W NE-SW NW-SE Plant height (cm) 70×30 cm 126cd 127a-d 130ab 128a-d* 60×35 cm 130ab 128a-d 128a-d 129abc 50×42 cm 125de 127bcd 121e 131a Head diameter (cm) 70×30 cm 14.3c 14.9bc 15.2abc 15.5ab 60×35 cm 15.3abc 14.7bc 15.0bc 15.1abc 50×42 cm 14.7bc 15.0bc 14.4c 15.9a Thousand seed weight (g) 70×30 cm 42.3de 45.5bcd 49.3ab 49.9a 60×35 cm 46.0a-d 46.3a-d 48.5ab 47.2abc 50×42 cm 43.5cd 49.7ab 44.0cd 47.1abc Seed yield (kg ha-1) 70×30 cm 1932c 2168ab 2396a 2393a 60×35 cm 2322a 2183ab 2306a 2344a 50×42 cm 2064bc 2252ab 1921c 2365a Means followed by the same letter(s) are not significantly different at p < 0.05.
TABLE 3 - Interaction of row direction × seed priming on yield and yield components of sunflower in 2010 and 2011
Row direction Seed priming N-S E-W NE-SW NW-SE Days to flowering (day) Control 78.4ab 78.6a 78.0bc 78.3abc* Hydropriming 77.9c 78.2abc 78.4ab 78.3abc abc a abc abc KNO3 78.2 78.6 78.3 78.3 Head diameter (cm) Control 14.8cd 14.8cd 14.8cd 15.0bcd Hydropriming 15.2bc 15.2bc 14.8cd 16.2a d cd bc b KNO3 14.4 14.6 15.1 15.5 Thousand seed weight (g) Control 41.8c 45.6ab 49.1a 47.7ab Hydropriming 44.9bc 47.7ab 46.4ab 49.5a bc ab ab ab KNO3 45.0 48.2 46.2 47.0 Seed yield (kg ha-1) in 2011 Control 4275ab 3988c 4258ab 4275ab Hydropriming 4026bc 4340a 4303a 4248ab bc abc abc abc KNO3 4043 4205 4227 4091 *: Means followed by the same letter(s) are not significantly different at p < 0.05.
tween 1920 and 2400 kg ha-1. The highest seed yield was the experimental fields. In our study, row direction influ- obtained from the NE-SW and 70 × 30 cm plant arrange- enced seed yield and other traits more dramatically in 2010 ment. Our results demonstrated that row direction slightly than in 2011. improved seed yield in the NW-SE and NE-SW rows; how- ever, significant differences were not observed among the Plant arrangement is specified as the dispersion of row directions. The results of this study are in agreement plants on inter- and intra-row spacing with constant den- with the observations of Shafiullah et al. [10], who reported sity. It determines the potential of the canopy to intercept that plants in NE-SW rows had higher average seed yields more light and permits more shadow on the soil surface to than plants in N-S and E-W rows. Diepenbrock et al. [11] prevent moisture loss [16]. Our results indicated that plant observed positive yield and oil responses to row direction in arrangement affected the seed yield of sunflower remarka- sunflower, and E-W rows gave higher values than N-S rows. bly in both study years, and the highest seed yield was rec- However, Robinson [12] found no significant differences orded in the 60 × 35 cm arrangement, in spite of reduced oil between N-S and E-W row directions with respect to seed content. Riahinia and Dehdahti [23] reported that the maxi- yield of sunflower. The findings of previous studies are con- mum seed yield was observed in a 35 × 33 cm arrangement tradictory due to different soil and weather conditions in (4.64 t ha-1) relative to other plant arrangements of 35 × 33 cm,
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50 × 23.2 cm, 65 × 17.9 cm, 80 × 14.5 cm and 100 × 11.6 cm. priming enhanced head diameter and seed yield in 2010. Similar findings were reported by Vahedi et al. [15], who The effectiveness of hydropriming may primarily be due to found that a plant arrangement of 45 × 27.5 gave greater more rapid germination and root growth than in the control head diameters and higher seed and biological yields than seeds. Hussain et al. [22] reported that hydropriming and 75 × 16.5, 60 × 20.5 and 45 × 27.5 cm arrangements. osmopriming with NaCl reduced mean emergence time Although differences between treatments in days to and increased achene yield in sunflower. Also, the findings flowering were statistically significant, it occurred at ap- of Singh and Rao [20] and Wahid et al. [21] indicate that proximately 78 days (Table 3). These differences were too primed seeds produced longer roots compared to control small to evaluate for practical purposes. Head diameter was seeds. significantly affected by the interaction of row direction × Gas exchange characteristics are shown in Figure 2. seed priming. The highest values were detected after hydro- There was a significant difference between 2010 and 2011. priming in the N-S, E-W and NW-SE rows. Potassium ni- Higher values were obtained from the plants grown in 2010 trate (KNO ) enhanced head diameter in the NE-SW direc- 3 relative to 2011. Transpiration rate (E) was significantly tion. KNO treatment in the N-S and E-W row directions in- 3 influenced by the interaction of plant arrangement × seed creased thousand seed weight, while hydropriming in- priming in both years. The lowest value in 2010 was meas- creased it in NW-SE rows. Hydropriming improved seed ured in control seeds at 50 × 42 cm. In addition, a signifi- yield of sunflower during 2011 season in the E-W and NE- cant effect of the interaction of row direction × plant ar- SW directions. The maximum seed yield (4340 kg ha-1) was rangement on stomatal conductance (g ) was detected in achieved in the NE-SW direction with hydroprimed seeds. s both years. The maximum stomatal conductance in 2010 Head diameter and oil ratio were significantly influ- was recorded in the NE-SW row sown at 70 × 30 cm, while enced by a three-way interaction in 2010. Hydropriming it was the highest in the N-S direction when it was sown at gave higher head diameters in all row directions except for 70 × 30 cm in 2011. All of the plant arrangements in 2011 NE-SW (Table 4). The 50 × 42 cm plant arrangement pro- gave the highest stomatal conductance in the N-S direction. duced wider heads in the N-S and NW-SE directions, while Our results showed that photosynthetic rate (A) was signif- plants in the 60 × 35 cm and E-W, 70 × 30 cm and NE-SW icantly affected by seed treatments, and primed seeds gave treatments performed better. The maximum oil ratio was higher rates of photosynthesis relative to control seeds in observed in plants sown in the NW-SE direction with a both years. Gimenez et al. [24] and Noreen and Ashraf [25] plant density of 70 × 30 cm and hydrated seeds. Hydro- reported that the rate of photosynthesis was greatest under priming proved advantageous for increasing oil ratio in the conditions of no stress and was a good indicator for deter- E-W, NW-SE and NE-SW row directions. mining plant productivity in sunflower. Sunflower is a pho- Seed priming permits better and quicker germination totropic plant from emergence to onset of flowering. Its of seeds and improves emergence and seedling growth un- heads and leaves face east in the morning and west in the der unfavorable conditions [18]. In our experiment, hydro- evening [26]. Due to this characteristic, its light intercep-
TABLE 4 - Interaction of row direction × plant arrangement × seed priming on yield and yield components of sunflower in 2010
Plant arrangement Row direction Seed priming N-S E-W NE-SW NW-SE Head diameter (cm) Control 14.3bcd 14.7bcd 15.6bc 15.1bcd* 70×30 cm Hydropriming 14.6bcd 14.9bcd 15.1bcd 15.6bc cd bcd bcd b KNO3 14.1 15.0 14.9 15.9 Control 15.4bcd 14.6bcd 14.8bcd 14.8bcd 60×35 cm Hydropriming 15.3bcd 15.5bcd 14.9bcd 15.2bcd bcd cd bcd bcd KNO3 15.2 14.2 15.4 15.3 Control 14.6bcd 15.2bcd 13.9d 14.9bcd 50×42 cm Hydropriming 15.6bc 15.2bcd 14.3bcd 17.7a d bcd bcd bcd KNO3 14.0 14.6 14.9 15.2 Oil ratio (%) Control 38.4a-g 39.3a-g 38.3a-g 38.8a-f 70×30 cm Hydropriming 36.8d-g 39.0a-f 39.2a-e 41.1a h a-g b-g a-g KNO3 33.3 37.8 37.6 38.3 Control 36.6b-g 39.3a-e 39.7a-d 38.6a-g 60×35 cm Hydropriming 35.9e-h 39.8a-d 40.4a-c 36.8d-g a-g a-g a-e a-d KNO3 38.0 38.5 39.2 39.4 Control 37.6b-g 38.4a-g 39.8a-d 39.5a-d 50×42 cm Hydropriming 37.6b-g 38.7fgh 37.4c-g 40.9ab g-h a-f d-g c-g KNO3 35.4 38.8 36.7 37.1 *: Means followed by the same letter(s) are not significantly different at p < 0.05.
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2010 2011 8,6 9 Control Hydropriming KNO3 a a 8,8 a 8,4 a abc a a abc abc abc ) ) 8,6 -1 -1 a abc s s 8,2 a a a b -2 -2 c 8,4
8 8,2 mmol m mmol ( 7,8 8 7,8 7,6 b iration rate p 7,6 7,4 Trans Transpiration (m rate m ol m 7,4 7,2 7,2 7 7 70x30 60x35 50x42 70x30 60x35 50x42 0,48 0,6 a a 70x30 60x35 50x42 ab 0,46 ab ab a 0,55 ) )
ab -1 -1 0,44 ab s
s ab ab -2 -2 ab a ab 0,5 0,42 bc ab ab 0,4 ab bc 0,45 bc c c bc c 0,38 c 0,36 0,4
0,34 m (mol conductance Stomatal Stomatal (mol conductance m 0,35 0,32
0,3 0,3 N-S E-W NE-SW NW-SE N-S E-W NE-SW NW-SE 27,5 24,5 a 27 ab 26,5 b )
-1 ) s -1 a -2 s 24 26 -2
25,5 µmol m µmol ( ab 25 23,5 24,5 nthetic rate rate nthetic
y b 24 23 Photos Photosynthetic rate (µmol m rate Photosynthetic 23,5 23 22,5 22,5 Control Hydropriming KNO3 Control Hydropriming KNO3
FIGURE 2 - Transpiration rate (mmol m-2 s-1), stomatal conductance (mol m-2 s-1) and photosynthetic rate (µmol m-2 s-1) of sunflower plants subjected to varying row direction, plant arrangement and seed priming. Letters on each bar show significance level of means at p < 0.05.
tion is almost equivalent to that of C4 plants. Among the plant density and increasing seed yield of sunflower under investigated parameters, seed priming has the potential to rain-fed conditions. The optimum plant density was deter- increase the rate of photosynthesis and ultimately seed mined to be 60 × 35 cm, while the most promising seed yield under rain-fed conditions. treatment method for increasing seed yield was hydroprim- ing. In the study, no row direction showed superiority for increasing seed yield and oil content; however, orientation 4. CONCLUSIONS in the NW-SE direction should be beneficial for sunflower cultivation. Uniform adjustment of the crop spacing in the field is one of the most important factors determining yield and quality in sunflower [27, 28]. Among a variety of factors ACKNOWLEDGEMENTS responsible for lower sunflower yields in our country, non- uniform plant distribution exhibits a remarkable effect on This research was funded by the Scientific and Tech- the productivity of the crop. It was concluded that seed nological Research Council of Turkey (TÜBİTAK), with priming was a promising method for providing uniform project number TOVAG 109O641.
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The authors have declared no conflict of interest. [19] Kathiresan, K., Kalyani, V. and Gnanarethinam, J.L. (1984) Effect of seed treatments on field emergence, early growth and some physiological processes of sunflower (Helianthus annuus L.). Field Crops Research, 9, 215-217. REFERENCES [20] Singh, B.G. and Rao, G. (1993) Effect of chemical soaking of sun- flower (Helianthus annuus L.) seed on vigour index. Indian Jour- [1] Ozer, H., Polat, T. and Ozturk, E. (2004) Response of irrigated nal of Agricultural Sciences, 63, 232-233. sunflower (Helianthus annuus L.) hybrids to nitrogen fertilization: growth, yield and yield components. Plant Soil and Environment, [21] Wahid, A., Noreen, A., Basra, S.M.A., Gelani, S. and Farooq, M. 5, 205-211. (2008) Priming-induced metabolic changes in sunflower (Helian- thus annuus) achenes improve germination and seedling growth. [2] Kazemeini, S.A., Edalat, M. and Avat, S. (2009) Interaction effects Botanical Studies, 49, 343-350. of deficit irrigation and row spacing on sunflower (Helianthus an- nuus L.) growth, seed yield and oil yield. African Journal of Agri- [22] Hussain, M., Farooq, M., Basra, S.M.A. and Ahmad, N. (2006) In- cultural Research, 4, 1165-1170. fluence of seed priming techniques on the seedling establishment, yield and quality of hybrid sunflower. International Journal of Ag- [3] Anonymous (2015) Web page of Turkish Statistical Institute. riculture and Biology, 8, 14-18. http://tuikapp.tuik.gov.tr/bitkiselapp/bitkisel.zul (accessed on: 10.09.2015) [23] Riahinia, S. and Dehdahti, S.M. (2008) Row spacing effects on light extinction coefficients, leaf area index, leaf area index affect- [4] Kaya, M.D. and Kolsarıcı, Ö. (2011) Water use features of sun- ing in photosynthesis and grain yield of corn (Zea mays L.) and flower (Helianthus annuus L.) hybrids irrigated at different growth sunflower (Helianthus annuus L.). Journal of Biological Science, stages. Tarla Bitkileri Merkez Araştırma Enstitüsü Dergisi, 20, 1- 8, 954-957. 5. [24] Gimenez, C., Mitchell, V.J. and Lawlor, D.W. (1992) Regulation [5] Kadayıfçı, A. and Yıldırım, O. (2000) The response of sunflower of photosynthetic rate of two sunflower hybrids under water stress. grain yield to water. Turkish Journal of Agriculture and Forestry, Plant Physiology, 98, 516-524. 24, 137-145. [25] Noreen, S. and Ashraf, M. (2008) Alleviation of adverse effects of [6] Göksoy, A.T., Demir, A.O., Turan, Z.M. and Dagustu, N. (2004) salt stress on sunflower (Helianthus annuus L.) by exogenous ap- Responses of sunflower to full and limited irrigation at different plication of salicylic acid: growth and photosynthesis. Pakistan growth stages. Field Crops Research, 87, 167-178. Journal of Botany, 40, 1657-1663. [7] Flagella, Z., Rotunno, T., Tarantino, E., Caterina, R., Caro, A., Di [26] Robinson, R.G. (1978) Production and Culture. Sunflower Science Caterina, R., Di Caterina, A. and De-Caro, A. (2002) Changes in and Technology, pp 89-132. Carter, J.F. (ed.) Wisconsin, USA. seed yield and oil fatty acid composition of high oleic sunflower (Helianthus annuus L.) hybrids in relation to the sowing date and [27] Barros, J.F.C., Carcalho, M. and Basch, G. (2004) Response of the water regime. European Journal of Agronomy, 17, 221-230. sunflower (Helianthus annuus L.) to sowing date and plant density under Mediterranean conditions. European Journal of Agronomy, [8] Angadi, S.V. and Entz, M.H. (2002) Water relations of standard 21, 347-356. height and dwarf sunflower cultivars. Crop Science, 42, 152-159. [28] Long, M., Feil, B. and Diepenbrock, W. (2001) Effects of plant [9] Bakht, J., Shafi, M., Yousaf, M., Raziuddin and Khan, M.A. density, row spacing and row orientation on yield and achene qual- (2010) Effect of irrigation on physiology and yield of sunflower ity in rainfed sunflower. Acta Agronomica Hungarica, 49, 397- hybrids. Pakistan Journal of Botany, 42, 1317-1326. 417. [10] Shafiullah, S., Baitullah, Rana, M.A. and Khan, A.S. (2001) Effect of row directions on yield and yield components of sunflower (He- lianthus annuus L.). Helia, 24, 137-144. [11] Diepenbrock, W., Long, M. and Feil, B. (2001) Yield and quality of sunflower as affected by row orientation, row spacing and plant density. Die Bodenkultur, 52, 29-36. [12] Robinson, R.J. (1975) Effect of row direction on sunflowers. Agronomy Journal, 67, 93-94. Received: December 10, 2014 Revised: July 13, 2015; September 11, 2015; [13] Kaul, K., Greer, E.C., Kasperbauer, M.J. and Mahl, C. (2001) Row September 17, 2015 orientation affects fruit yield in field-grown okra. Journal of Sus- tainable Agriculture, 17, 169-174. Accepted: September 17, 2015
[14] Catherine, P.D.B., Hashem, A. and Pathan, S. (2010) Manipulating crop row orientation to suppress weeds and increase crop yield. Weed Science, 58, 174-178. CORRESPONDING AUTHOR [15] Vahedi, B., Gholipouri, A. and Sedghi, M. (2010) Effect of plant- ing pattern on radiation use efficiency, yield and yield components Mehmet Demir Kaya of sunflower. Recent Research in Science and Technology, 2, 38- Department of Field Crops 41. Faculty of Agriculture [16] Tetio, F.K. and Gardnar, F.P. (1988) Response of maize to plant Eskişehir Osmangazi University population density. 1. Canopy development, light relationship and vegetative growth. Agronomy Journal, 80, 430-435. 26160 Eskişehir TURKEY [17] Zarea, M.J., Ghalavand, A. and Daneshian, J. (2005) Effect of planting patterns of sunflower on yield and extinction coefficient. Agronomy for Sustainable Development, 25, 513-518. Phone: +90 222 3242790 Fax: +90 222 3242791 [18] Kaya, M.D., Okçu, G., Atak, M., Çıkılı, Y. and Kolsarıcı, Ö. (2006) Seed treatments to overcome salt and drought stress during E-mail: [email protected], germination in sunflower (Helianthus annuus L.). European Jour- nal of Agronomy, 24, 291-295. FEB/ Vol 24/ No 11c/ 2015 – pages 4095 - 4101
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LIFE TABLE CHARACTERISTICS OF Orius minutus (L.) (HEMIPTERA: ANTHOCORIDAE) FEEDING ON Bemisia tabaci (GENNADIUS) (HEMIPTERA: ALEYRODIDAE) AT THE LABORATORY CONDITIONS
Davut Efe1, Mehmet Salih Özgökçe2,* and İsmail Karaca3
1 University of Bingöl, Agricultural Faculty, Plant Protection Department, Bingöl, Turkey 2 University of Yüzüncü Yıl, Agricultural Faculty, Plant Protection Department, Van, Turkey 3 University of Süleyman Demirel, Agricultural Faculty, Plant Protection Department, Isparta, Turkey
ABSTRACT 1. INTRODUCTION
In the study, life table of Orius minutus (L.) (Hemip- Orius spp. are important biological control agents that tera: Anthocoridae) feeding on Bemisia tabaci (Gennadius) can be abundantly observed in many agro ecosystems. (Hemiptera: Aleyrodidae) was constructed at the labora- Apart from the fact that they feed on important pests such tory conditions adjusted to 25±1 oC, 65±10 % relative hu- as thrips, mites, whiteflies, aphids, their omnivorous char- midity and 16 hours of artificial light. B. tabaci used as acteristics bring them advantage in surviving in the nature prey was reared on cotton plant at climatic chamber. As a and sustain their activity. Since they are reared relatively result of the study, it was determined that total develop- easy at the artificial conditions, it is quite common in com- mental period and mortality rate of O. minutus female and mercial insect production markets in Europe and North male were 19.80, 20.54 days and 23%, 24% respectively; America [1]. It is known that various types of Orius spp. longevity of O. minutus were 57.40 days for female and are used successfully for long years in greenhouse and in 29.98 days for male. Pre-oviposition, oviposition and agricultural fields [1]. postoviposition periods were 3.84, 34.20 and 19.00 days, re- The reason why species included within biological spectively, daily egg numbers were 3.69 and total egg num- control program depends on knowing various biological bers were 180.97. Reproductive value reached the highest characteristics such as survival, reproduction of the species th value at 5 days of oviposition period. Intrinsic rate of in- apart from various external factors such as adaptation they crease (rm) was 0.121 female/female/day, net reproductive show against climate factors, nutrition habits, being easily rates (Ro) was 60.446 females/female, mean generation produced. Although biology of Orius species feeding on time (T0) was 33.892 days, gross reproductive rate (GRR) different preys has been studied well, there has been no de- was 78.684, doubling time (T2) was 5.727 days and finite tailed study about biology of Orius minutus feeding on Be- rate of increase (λ) was 1.129. The most appropriate repro- misia tabaci [1-5]. With this study it was aimed to reveal duction curve was calculated by Enkegaard equation and specific biological characteristics of reproduction and sur- parameters were found as follows; a = 0.711±0.049; b = vival of Orius minutus on Bemisia tabaci that is quite an 2 0.091±0.004; R = 0.926. Weibull distribution was used to important pest on various crop and greenhouses. determine the best curve showing survival rate for females and males of Orius minutus and parameters were found as 2 follows; for females, b = 57.4±0.019, c = 2.26±0.007, R = 2. MATERIALS AND METHODS 0.818; for male, b = 31.67±0.014, c = 1.77±0.0.06, R2 = 0.918. Orius minutus (Hemiptera: Anthocoridae) was collected from cotton fields near Adana in the east Mediterranean re- gion of Turkey and colonized on 4-6 week-old cotton plants KEYWORDS: Orius minutus, age-specific life table, Bemisia (Gossypium hirsutum cv. Çukurova 1518) infested with Be- tabaci, Enkegaard, Weibull distribution misia tabaci as prey at 25±1oC, 65±10 % RH and 16 h of artificial light. B.tabaci were obtained from laboratory cul- tures, and they were reared on cotton under the same con- ditions. 2nd instar nymph of B. tabaci was used as prey. * Corresponding author To obtain this stage, females of B. tabaci was confined onto
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leaves of young cotton plants for oviposition period of Mathematical model, Enkegaard equation (most ap- 24 h. Experiments on the life history parameters of O. propriate reproduction curve equation, mx), was used to fit minutus fed on B. tabaci were employed on excised cotton the data on fecundity [16-18]. leaves placed upside down on wetted foam rubber in petri Enkegaard equation: dishes (Ø 9 cm). For 24 h, the predators were enclosed to- gether with its prey onto the leaf by a small leaf cage (4 F(x) = a.x.e(-b.x) [16, 17] cm), the upper side closed with fine mesh. 10 replications Where; F(x) i sdaily fecundity rate related to age were set up for each immature stage and adults of the pred- (eggs/female/day), x is female age (in days), a and b are o ator. All experiments were conducted at 25±2 C, 65±10 % constants. Day 1 is the first day of oviposition period. De- RH and 16 h of artificial light in a climate room. velopment times of male and females were tested by using The data obtained from daily observations were used t-student test. Analayses were done by using JMP (ver. 5), to construct the age specific life table according to Euler- MS Excel (ver. 2003) ve SPSS, (ver. 17) softwares. Lotka equation [6-8]. The Weibull frequency distribution was used to de- (rm.x) scribe the age-specific survival of all individuals [19]: e .l x .mx 1 [6] t c Where e is the base of natural logarithm; x, the age of b S (t) e t,b,c 0 [20] individual in days; lx , the proportion of individuals alive p at day x, or age-specific survivorship; and m , the number x where Sp(t) represents the probability of surviving to of offsprings/female/day x, or age-specific fecundity. The a given age, b is the parameter that describes the scale, c net reproduction rate, R0, which was calculated as the sum is shape of the curve and t is time. The shape parameters of the lx.mx, column in life table. cc>1, =1 and c <1 correspond to type I, II and III survi- The mean generation time, T0, was then calculated vorship curves, respectively [20; 19]. Statistical analyses from the formula; were done by using following softwares, CurveExpert pro (ver. 1.6.7), SPSS (ver. 17), MS Excel (ver. 2003). ln.R0 T0 [6-10], rm 3. RESULTS Gross reproduction rate, GRR mx [6; 11], 3.1 Development period In adition; r Development periods of egg, immature stages and as Finite rate of increase, e m [6], population in- well as total development periods of Orius minutus feeding creases per unit time, on Bemicia tabaci were not statistically different between ln 2 female and male (Table 1). Total development periods and longevity for female and male were 19.80, 20.54 days and Doubling time, T2 [10] the time it takes a rm 57.40 and 29.98 days, respectively. population to double in size, 3.2 Fecundity Reproductive value of the females When fecundity periods of Orius minutus females rm .y (e .ly .my ) were analyzed, it was observed that pre-oviposition, ovipo- yx V [12], age-specific expectation of sition and postoviposition period were 3.84, 34.20 and x rm .x lx.e 19.00 days respectively (Table 2). Generation time is 24.80 future offspring, days, egg number laid daily is 3.69 and total offspring num- ber of predator was 180.97. Reproductive rate of the pred- Expected remaining life time of the females ator increased quickly at beginning of the oviposition pe- th ly ly1 riod and reached maximum value at the 10 day (Figure 1) 2 Later, the number of the eggs gradually decreased with lit- y x tle fluctuations (2-3 individuals). At the 26th day of ovipo- Ex [13, 14], lx sition period, reproductive rate reached its maximum level and oviposition finished at 69th day (Figure 1). This rela- The stable age distribution of population tion related to age was described by Enkegaard equation rm .x lx.e and calculated parameters and the curve was illustrated in Cx r .x [6] were calculated. Figure 1. The optimum curve equation for the model was ob- (l .e m ) x tained via non-linear regression. Parameters were found to be: x0 a = 0.711±0.049; b = 0.091±0.004; correlation coefficient All parameters were obtained by using RmStat-3 [15]. (R2) = 0.926. The females laid eggs every day during the
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TABLE 1 - Development periods of males and females of Orius minutus feeding on Bemisia tabaci (Mean±S.E.).
Development period n Female n Male Egg 13 5.60±0.213a* 17 6.02±0.122a Stage 1 13 2.71±0.090a 17 2.89±0.077a Stage 2 11 2.03±0.031a 15 1.94±0.102a Stage 3 10 1.87±0.070a 13 1.97±0.143a Stage 4 10 2.77±0.124a 13 2.88±0.106a Stage 5 10 4.48±0.177a 13 4.24±0.219a Total development time 10 19.80±0.416a 13 20.54±0.526a Life span 10 57.40±3.830a 13 29.98±3.140b * Within rows, means followed by the different letter between sexes differ significantly (t-test, P< 0.05).
TABLE 2 - Fecundity of Orius minutus feeding on Bemisia tabaci. (T0) was 33.892, gross reproductive rate (GRR) was 78.684, doubling time (T2) was 5.727 days and finite rate of in- Fecundity n Mean±St.E. crease (λ) was 1.129 (Table 3). Preoviposition 10 3.84±0.145 Oviposition 10 34.20±2.426 TABLE 3 - Life table parameters of Orius minutus on Bemisia tabaci. Postoviposition 10 19.00±3.246 Generation time 10 24.80±0.467 Life Table Parameters
Daily offsprings 10 3.69±0.360 Intrinsic rate of increase, rm 0.121
Totally offsprings 10 180.97±16.281 Net reproductive rate, Ro 60.446
Mean generation time, To 33.892 Gross reproduction rate, GRR 78.684
Doubling time, T2 5.727 Finite rate of increase, λ 1.129 n 13
3.4 Survival Immature mortality rates of female and male were cal- culated as 23% and 24% respectively. Only second and third stage individuals of the both sex dead. While survival rate of females were stable in during of oviposition period, it regularly reduced from beginning of postoviposition to end of 91th day (Figure 2). Survival rate of males regularly and quickly reduced and last individual dead at 69th day.
The stable age distribution of population (Cx) during their entire life displayed stable slope similar to survival FIGURE 1 - Age-specific fecundity (female/female/day) of Orius rate but this changed rapidly due to the high mortality rates minutus as a function of female adult age (days) (Parameters: a = 0.711±0.049 ; b = 0.091±0.004; R2 = 0.926). at the end of life (Figure 2). First period of life of male in- dividuals a fairly stable slope is shown like female popula- oviposition period. Mean generation time (T0) was theoret- tion but due to dead rates observed second period of life ically calculated as 33.892 days (Table 2). According to the showed fluctuations, due to high rates of dead at end of life graph of life table, T0 value, which covered the first 13 days have changed rapidly. of the oviposition period lasting 49 days, constituted the Expected remaining lifetime (Ex) of both sexes due to 36% of the total eggs. Mean egg number per day during 23-24% mortality rates observed showed a rapid decline at overall oviposition period was 3.69, egg number per day first 9-10 days. Expected life time of females which was for the first 13 day of the period was 4.95, and that for the calculated as 61.22 days decreased 53.27 days at 9th day; last 36 days was 3.24. Age specific reproductive value of expected remaining life time was calculated as 63.50 days th the females (Vx) reached maximum level at the 5 day of at 14th day since remainder population showed high sur- oviposition (Figure 1). vival rate for long time. After this day, depending on age till end of life it shown a regularly decreased slope and total 3.3 Life table parameters life time took more 30 days than expected life time. Ex- Life table parameters were calculated by RmStat-3 pected lifetime of males was calculated as 40.91 days. [15] and the results were presented in Table 3. As a result, These estimated days were 31.91 days at 10th day and 37.04 th intrinsic rate of increase (rm) was 0.121, net reproductive at 14 day. Total lifetime took more 28 days than expected rate (Ro) was 60.446 females/female, mean generation time lifetime.
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FIGURE 2 - Survival rate (lx), fecundity (mx) and reproductive value (Vx) of Orius minutus.
FIGURE 3 - Stable age distribution (Cx) and expected life time (Ex) of Orius minutus males and females.
Weibull distribution was used to determine the best O. minutus and optimal slope and fitted parameters were curve showing survival rate for females and males of Orius given at Figure 2. The slope for both sexes was found as minutus. [19] reported that Weibull distribution has been convex. Parameters were found as follows, for female; b = using for natural populations of plants and animals. In this 57.4±0.019, c = 2.26±0.007, R2 = 0.818; for male; b = study, Weibull distribution was applied for both sexes of 31.67±0.014, c = 1.77±0.0.06, R2 = 0.918.
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FIGURE 4 - Survival probability of males and females of Orius minutus and lines of best fit by Weibull function (Parameters: Female, b = 57.4±0.019,c = 2.26±0.007, R2 = 0.818; Male, b = 31.67±0.014,c = 1.77±0.0.06, R2 = 0.918).
4. DISCUSSION ities were 47.6% and 67.4%, rm were 0.097, 0.045, R0 were 13.78, 3.109, GRR were 16.23, 3.97, λ were 1.102, 1.047, To our knowledge, there isn’t any study on the biology To were 26.98, 24.83 and T2 were 7.129, 15.17 days, re- of O. minutus on Bemisia tabaci. However, our findings can spectively, for both preys. Compared with this study, de- be compared with the results of the studies performed on dif- spite low development period and mean generation time, it ferent preys under similar laboratory conditions. [1] reported is seen that all other parameters have lower values. Accord- that development time of O. minutus fed on Ephestia kuehni- ing to these values, it shown that B. tabaci is a better prey ella was 11.70 and 11.53 days for females and males, respec- than T. urticae and T. tabaci. tively, while Toyoshima [4], reported the same values, Parameters calculated by Weibull distribution summa- which were closer to our results, as 21.0 and 19.6 days on rizing survival rate of an organism were found as follows, Tetranychus urticae. There was no statistical difference be- while this parameter (c) was close to 1 for male population, tween the development times of females and males, lon- it was bigger than 1 for both sexes. Weibull distribution pa- gevity of female and male adults were found to be signifi- rameter and survival slope were similar type 1. The ad- cant contrary to findings of [1]. When fecundity rate was vantages of the Weibull distribution as survival slope in an analyzed, it was observed that fecundity rate slope of O. ecological environment are flexibility of equality belonging minutus was different from many other insects. While fe- to slope, availability as statistical curve and be able to make cundity rates among many species reached maximum level important explanations from model parameters [21]. Second with a rapid increase at the beginning of oviposition period important topic is related to the model parameters determin- and gradually decreased depending on the age, in this study ing three basic survival slopes that were explained by [22]. it has a slope similar to the one in study of [1]. The similar slope draws attention of O. insidiosus and O. niger as well It was shown that c parameter calculated from slope [2]. Fecundity rate rapidly increased and reached maxi- equal is type 1 survival curve increasingly of mortality rates mum at the beginning of oviposition period, but without a as a function of age of populations in both sexes. decrease depending on the age, stayed constant for a long The c parameter is calculated from the curve equation time and decreased at the end of oviposition period. Egg and identifies curve form; If c > 1, then the life curve of the number per day and total number of eggs found in this study population is defined as type 1 life curve, in which where were 3.69 and 180.97 and rm= 0.121, while Honda et al. [1] death ratio increases as a function of age. In a type 2 life reported the same values as 6.02 and 105.36 and rm= 0.137. curve c = 1; death ratio is constant rather than age-depend- [3] reported rm= 0.107 for O. minutus on E. kuehniella at 26 ent. In a type 3 life curve, c < 1 and death ratio decrease as ºC. It is difficult to explain the difference between those a function of age [19; 21]. studies, since the humidity conditions of the laboratories As can be seen from the model parameters, survival were not mentioned in both studies, if it is because of the rate was high during of adult survival except 23-24% mor- conditions or the different preys used. tality rates at immature period in both sexes but towards to The results of life table parameters of study that [5] end of life decreased regularly. Both sexes of O. minutus used a little bit lower values (24±1 ºC and 50±5% RH) and showed typically survival slope as increased populations. made with O. minutus fed on T. urticae and Thrips tabaci have become lower than the present study. [5] reported that The authors have declared no conflict of interest. development times were 15.7, 16.4 days, nymphal mortal-
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REFERENCES [18] Enkegaard, A. and Brødsgaard, H.F., (2001) Biological con- trol of pests in glasshouse ornamentals / biologyand interac- tions between pests and their natural enemies] (Danish, Eng- [1] Honda, J. Y., Nakashima, Y. and Hirose, Y. (1998) Develop- lish summary). DJF-Rapport 49,59_/63. ment, reproduction and longevity of Orius minutus and Orius sauteri (Heteroptera: Anthocoridae) when reared on Ephesti- [19] Pinder, J.E. III, Wiener, J.G. and Smith, M.H., (1978) The akuehniella eggs. Appl. Entomol. Zool. (Jpn.) 33: 449-453. Weibull distribution: a new method of sumarizing survivor- ship data. Ecology, 59, 175–179. [2] Meiracker, R. A. F. v. d., (1999) Biocontrol of western flower thrips by hetereopteran bugs. (Thesis). Universiteit van Am- [20] Deevey, E.S., (1947) Life tables for natural populations. sterdam. Retrieved from http://dare.uva.nl/record/79156 Q.Rev. Biol. 22: 283‹314. [3] Kakimoto, K., Urano, S., Noda, T., Matuo, K., Sakamaki, Y., [21] Hogg, D.B. and Nordheim, E.V., (1983) Age-specific survi- Tsuda, K. and Kusigemati, K., (2005) Comparison of the re- vorship analysis of Heliothis spp. populations on cotton. Res. productive potential of three Orius species, O. strigicollis, O. Popul. Ecol. 25, 280-297. sauteri, and O. minutus (Heteroptera : Anthocoridae), using eggs of the Mediterranean flour moth as a food source. Appl. [22] Pearl, R. and Miner, J. R., (1935) Experimental Studies on the Entomol. Zool. 40: 247–255. Duration of Life. XIV. The Comparative Mortality of Certain Lower Organisms. The Quarterly Review of Biology, Vol. 10, [4] Toyoshima, S., (2006) Development, Prey Consumption, and No. 1 (Mar., 1935), pp. 60-79. Fecundity of Orius minutus (Heteroptera: Anthocoridae) when fed on Tetranychus urticae (Acari: Tetranychidae). Journal of the Acarological Society of Japan; 15(2):151-156. [5] Fathi, S.A.A., (2009) The abundance of Orius niger (Wolf.) and O. minutus (L.) in potato Welds and their life table param- eters when fed on two prey species. J Pest Sci., 82:267–272.
[6] Birch, L.C., (1948) The intrinsic rate of natural increase of an insect population. Journal ofAnimal Ecology,17: 15–26.
[7] Howe, R.W., (1953) The rapid determination of the intrinsic of increas of an insect population. Annals of Applied Biology, 40, 134-151.
[8] Watson, T.F., (1964) Influence of host plant condition on pop- ulation increase of Tetranychus telarius (L.) (Acarina: Tetranychidae). Hilgardia, 35, 273-322. [9] Chazeau, J., Bouye, E. and Larbogne, L.B., (1991) Cycle de developpement et table de vie d’ Olla v-nigrum (Coleop- tera:Coccinellidae) ennemi naturel d’ Heteropsylla cubana (Hom.:Psyllidae) introduit en nouvelle Cale´donie. Ento- mophaga,31(2): 275–285. [10] Kairo, M.T.K. and Murphy, S.T., (1995) The life history of Rodolia iceryae Janson (Coleoptera: Coccinellidae) and the potential for use in innoculative releases against Icerya patter- soni Newstead (Homoptera: Margarodidae) on coffee. Journal of AppliedEntomology, 119: 487–491. [11] Leslie, P.H. and Park, T., (1949) The Intrinsic Rate of Natural Increase of Tribolium castaneum Herbst. Ecology, 30:469- 477. [12] Imura O. (1987) Demographic attributes of Tribolium freeman Hinton (Coleoptera: Tenebrionidae). Applied Entomology and Zoology,22(4): 449–455. [13] Southwood, T.R.E., (1978) Ecological Methods, with Particu- Received: August 31, 2014 lar Reference to the Study of Insect Populations. Chapman and Accepted: October 23, 2014 Hall, London, 524 p. [14] Carey, J.R., (1993) Applied Demography for Biologists with Special Emphasis on Insects. Oxford University Press, 211 p. CORRESPONDING AUTHOR [15] Özgökçe, M.S. and Karaca, İ., (2010) Yaşam Çizelgesi: Temel Prensipler ve Uygulamalar. Türkiye Entomoloji Derneği 1. Mehmet Salih Özgökçe Çalıştayı, Ekoloji Çalışma Grubu, 2010, Isparta. University of Yüzüncü Yıl [16] Enkegaard, A., (1993) The poinsettia strain of the cotton Agricultural Faculty whitefly,Bemisia tabaci (Homoptera; Aleyrodidae), biological Plant Protection Department anddemographic parameters on poinsettia (Euphorbia pulcher- rima) in relation to temperature. Bull. Entomol. Res. 83: 535– 65080 Van 546. TURKEY
[17] Hansen, D.L., Brødsgaard, H.F., Enkegaard, A., (1999) Life table characteristics of Macrolophus caliginosus preying upon E-mail: [email protected] Tetranychus urticae. Entomologia Experimentalis et Appli- cata 93, 269–275. FEB/ Vol 24/ No 11c/ 2015 – pages 4102 - 4107
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RELATIONSHIPS BETWEEN CHEMICAL AND PHYSICAL PROPERTIES OF SOILS AND NUTRIENT STATUS OF PLANTS ON YIELD OF POTATO
Kürşat Korkmaz1,*, Özbay Dede2, Halil Erdem3, Soner Çankaya4, and Mehmet Akgün1
1University of Ordu, Faculty of Agriculture, Department of Soil Science and Plant Nutrition, Turkey 2University of Ordu, Faculty of Agriculture, Field Crop Department, Turkey 3University of Gaziosmanpaşa, Faculty of Agriculture, Department of Soil Science and Plant Nutrition, Turkey 4University of Ordu, Faculty of Medicine, Biostatistics Department, Turkey
ABSTRACT commercial importance. Nutritionally, potato is an excel- lent source of carbohydrate, vitamins and essential miner- The relationships among nutrient levels in the soil, als; its protein notably contains a higher proportion of the plant tissue, and tuber with yield of potato were investi- essential amino acid lysine than most cereals and it is used gated by multiple regression analysis. The concentration to fortify cereal products such as rice and pasta [3]. Growth levels of selected macronutrients (N, P, and K) and trace and quality of potato is influenced by environmental fac- metals (Fe, Mn, Cu, and Zn), of 140 samples were analyzed tors such as temperature, moisture, light, soil type and nu- in this study. Mineral nutrient concentrations were deter- trients [4]. Several factors are closely related to yield pa- mined in leaves, tubers of potato and soil samples col- rameters and the plant nutrition is one of them. Potato’s lected. The nutrient concentrations in the samples were yield relies largely on the use of fertilizer. During the past found at different levels. In leaves, the nutrient concentra- decade, commercially available soluble chemical fertiliz- tion levels ranged from 4.25 to 6.28 % for N, 0.10 to 0.42% ers are used extensively in potato cropping systems, partic- for P, 1.00 to 5.58% for K, 103.0 to 329.4 mg kg-1 for Fe, ularly nitrogen, phosphorus, and potassium has increased 92.3 to 637.8 mg kg-1 for Mn, 7.7 to 70.8 mg kg-1 for Cu potato yields and quality [5].To get optimum yield, nutrient and 19.4 to 69.9 mg kg-1 for Zn. In tuber, the nutrient con- concentrations should be at sufficient level in potato. How- centration levels ranged from 1.31 to 2.27 % for N, 0.03 to ever, soil and plant analysis are the best method to deter- 0.28 % for P, 1.14 to 3.54% for K, 14.5 to 49.1 mg kg-1 for mine plant nutritional status affecting yield as well as other Fe, 0.7 to 8.3 mg kg-1 for Mn, 2.6 to 9.1 mg kg-1 for Cu and important agronomic attributes of this crop [6]. Soil and 12.0 to 22.7 mg kg-1 for Zn. Results indicated that three plant analysis and nutritional diagnosis may be adopted to parameters of this regression model were statistically sig- reach the efficient use of fertilizer. By monitoring the nu- nificant (P<0.05) and the R2 was 0.522. Plant tissue con- tritional status of a crop, potential deficiencies can some- centrations of N and Mn were significantly correlated with times be detected early enough for treatments of nutrient variations on yield of potato. These results prove that espe- disorders during the growing season. Such studies are im- cially N and Mn are important in the nutrition of potatoes. portant because this information can be employed to opti- mize the fertilizer use in potato crops. Several reports of concentrations of various nutrients in the plant tissue of po-
KEYWORDS: tato can be found in the literature [6-12]. However, nutrient Multiple regression, stepwise, potato yield, soil and plant analysis concentrations in leaves and soils have not been related to yield of the potato. The objective of this study was to de- termine if there are relationships between nutrient concen- trations of the plant tissue and tuber with nutrient status of 1. INTRODUCTION the soil which may be related to differences in productivity of potato, among genotypes. Potato (Solanum tuberosum L.), a major food in many countries is one of the most important agricultural crops in the world, which occupy fourth place in total annual produc- 2. MATERIALS AND METHODS tion at global level after wheat, maize and rice [1] . It is also an important crop in Turkey, cultivated on 174 000 ha and Soil and plant samples were collected in 2011 from produced nearly 5 million tons of tubers [2] with increasing Ordu in Black Sea region of Turkey, Three sampling loca- tions were selected in Ordu (Kabatas, Korgan and Aybastı), * Corresponding author the most representative potato production area. Five samples
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were taken and analyzed from 28 different potato varieties performing a normal multiple regression. If all variables in this study. Figure 1 shows the approximate location of are shown as significant (P-values < 0.05), then stop - the the sampling area. Soil samples from the rooting zone (0– complete model is good. But if significance F is low, but 30 cm) were taken from each location. The soil samples one or more of the p-values for the t-tests are high, stepwise were air dried and screened through a 2 mm stainless steel regression can be used to develop the best model that con- sieve. Each samples were analyzed for nutrient status. De- tains some of the variables. Stepwise regression can be tails of the soil pH, salt (EC), organic matter (OM) content, achieved either by trying out one independent variable at a texture, total N, available P, K, Fe, Cu, Zn and Mn are in- time and including it in the regression model if it is statis- dicated in Table 1. tically significant, or by including all potential independent variables in the model and eliminating those that are not statistically significant, or by a combination of both meth- ods. Thus, stepwise multiple regression analysis were used to predict the model in this study. All the computational work was performed by means of SPSS [15]
3. RESULTS AND DISCUSSION
Descriptive statistics (means, standard deviations, coef- ficients of variation) and significant values of normality test for the soil physical and chemical properties at the three lo- cations are given in Table 1. The results of particle-size anal- ysis showed that the texture was sandy loam with clay loam in Aybastı, loam in Kabataş and Korgan at 0-30 cm depth in soils. Also, these soils have adequate the contents of soil or- ganic C, 4.5%, and calcareous with salt was free. The pH of the soil samples ranged from 4.5‐6.4 with a mean of 5.5. A total of 7 elements as a macro and micro elements (N, P, K, Zn, Cu, Fe and Mn) were determined in the soil samples (Table1). The soils varied widely in their chemical proper- ties. The nitrogen (N), potassium (K), manganese (Mn), copper (Cu), zinc (Zn), and iron (Fe) in soils were present in adequate quantities, but phosphorus (P) was considera- bly lower for normal potato production in soils.
The macro and micro elements (N, P, K, Zn, Cu, Fe FIGURE 1 - The approximate location of the sampling area. and Mn) were determined in the leaves and tuber of potato Potato leaf samples were taken at flowering using the by plant analysis. The macro and microelements composi- most recently matured compound leaf. Plant samples were tion of potato is presented in Table 1. The nitrogen (N), po- thoroughly washed with running tap water and rinsed with tassium (K) manganese (Mn), copper (Cu), zinc (Zn), and deionized water to remove any soil/sediment particles at- iron (Fe) in leaf were present in adequate quantities although tached to the plant surfaces. The aboveground and under- P concentration was considerably lower for potato. In leaves, ground tissues were dried at 70°C for 48 hour to constant the nutrient concentration levels ranged from 4.25 to 6.28 % for N, 0.10 to 0.42% for P, 1.00 to 5.58% for K, 103.0 to weight. The dried tissues were weighed and ground into -1 -1 powder for analysis. 329.4 mg kg for Fe, 92.3 to 637.8 mg kg for Mn, 7.7 to 70.8 mg kg-1 for Cu and 19.4 to 69.9 mg kg-1 for Zn The dried and ground analytical samples were analyzed (Table 2). In tuber, the nutrient concentration levels ranged for total nitrogen by the Kjeldahl method [13]. Plant samples from 1.31 to 2.27 % for N, 0.03 to 0.28 % for P, 1.14 to were analyzed for P colorimetrically by the method of mo- 3.54% for K, 14.5 to 49.1 mg kg-1 for Fe, 0.7 to 8.3 mg kg-1 lybidate [14] by spectrophotometry, K by flame emission, for Mn, 2.6 to 9.1 mg kg-1 for Cu and 12.0 to 22.7 mg kg-1 Fe, Cu, Zn and Mn by atomic absorption spectrometry. Tu- for Zn (Table 1). The yield varied from 0.36 to 2.33 kg per ber yields of potato are calculated on a per-hill basis. hill with a mean of 0.97 kg per hill and there were also dif- Generally, multiple regression analysis is used to pre- ferences in the yield of the potato tubers between the culti- dict the model explaining the relationships among nutrient vars considered (Table 1). The results above indicated that levels in the soil, plant tissue, and tuber with yield of po- the tubers contain large amounts of nutrients such as Fe, tato. The aim of the multiple regression was to estimate Mn, Cu, Zn, N, and K but have a low concentration for P (0,1,..,p) from the data (Xi1, Xi1,.., Xip; Y). In multiple in this study. Tuber Fe, and Zn concentrations have been linear regression models, several independent variables shown to vary significantly between cultivars considered. were used to model a single dependent variable. Begin by There is considerable genetic variation in tuber mineral con-
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TABLE 1 - Descriptive Statistics for Examined Parameters
Mean* Std. Deviation CV % Mean* Std. Deviation CV % Leaf Soil N (%) 5.35 0.52 10 N (%) 0.13 0.05 39 P (%) 0.21 0.07 35 P (mg kg-1) 7.38 5.05 68 K (%) 3.27 1.04 32 K (mg kg-1) 284.93 126.59 44 Cu (mg kg-1) 16.87 13.86 64 Cu (mg kg-1) 1.41 0.75 53 Mn (mg kg-1) 255.30 119.01 47 Zn (mg kg-1) 1.00 0.56 56 Fe (mg kg-1) 178.51 56.18 31 Mn (mg kg-1) 25.00 16.10 64 Zn (mg kg-1) 35.75 13.58 38 Fe (mg kg-1) 43.32 13.94 32 pH (1:2.5) 5.48 0.45 8 Tuber OM (%) 4.47 1.78 40 N (%) 1.73 0.26 15 P (%) 0.12 0.08 65 K (%) 2.04 0.58 29 Yield Cu (mg kg-1) 5.28 1.96 37 Yield (per hill) 0.97 0.49 51 Mn (mg kg-1) 3.61 1.59 44 Fe (mg kg-1) 28.53 9.07 32 Zn (mg kg-1) 17.13 3.40 20 * Mean of 140 samples
TABLE 2 - The relationships between nutrient concentration of leaves and tubers
Tuber N Tuber P Tuber K Tuber Cu Tuber Mn Tuber Fe Tuber Zn Leaf N 0,489** -0,223 -0,224 0,279 0,289 0,387* 0,098 Leaf P -0,102 0,663** 0,133 -0,330 0,081 0,124 -0,284 Leaf K 0,133 -0,308 0,410* 0,288 0,299 0,104 -0,032 Leaf Cu 0,217 -0,176 -0,215 0,072 0,009 0,325 0,112 Leaf Mn -0,317 0,169 0,029 -0,264 -0,064 -0,125 0,058 Leaf Fe 0,564** 0,119 -0,120 0,285 0,522** 0,247 0,026 Leaf Zn -0,319 0,158 0,125 -0,210 -0,193 -0,076 0,058 **. Correlation is significant at the 0.01 level (2-tailed), *. Correlation is significant at the 0.05 level (2-tailed).
centrations within potato genotypes. White et al. [16] re- environmental and genetic factors. One of the most signif- ported that higher-yielding potato genotypes have lower icant environmental factor is the availability of mineral el- concentrations of mineral elements in their tubers than ements in the soil. The nutrient levels obtained in plant are lower yielding genotypes when grown in the same condi- in the same range of those reported by other authors for tions. Moreover, they found no significant relationships be- potatoes from several countries, our data agree with those tween tuber yield and tuber N, K, S, Ca, Mg, Fe, Zn, or Mn published in other regions, such as Rivero et al. [6] War- concentrations, but tuber P and Cu concentrations in- mann et al. [7], and Tekalign and Hammes [9], Christelle creased significantly with increasing tuber yield. et al. [10], Galdon et al. [11], Lombardo et al. [12]. The behavior of trace elements was very interesting, Correlation analysis was used to estimate the relation- with the highest mean concentrations of Fe, Mn, Cu and Zn ships between nutrient concentrations in leaf and tuber of in the leaves samples. Among vegetables crops potatoes potato. There were significant influence of N and Fe con- are relatively tolerant of high Mn and level 700 mg kg-1 has centration in leaves on uptake N of potato at the 1% prob- been proposed as a critical concentration for toxicity, but ability level (Table2). Statistical analysis of these data also reported levels of up to 3000 mg kg-1 Mn in healthy showed that N concentration of tubers was linearly corre- potato leaves [17]. Some mineral contents in the potato lated with N and Fe concentration of leaf (Table2). There samples must be influenced by the region of production, was also significantly positive relationship between leaf Fe which is mainly influenced by the mineral contents of the and Mn in tuber. Manganese and iron (Fe) has an interac- soils and climatic factors as well as fertilization. tion in plants, iron uptake by plants affects with high Correlation analysis used to estimate the relationships amounts of manganese in the soil [18]. Also, manganese between nutrient concentrations in the plant tissue, nutrient promotes the activity of various enzymes that helps in the status of the soil with yield of potato. The results showed photosynthetic light reactions, respiration and protein syn- that nutrient levels of plant weren’t significantly associated thetic processes leading to better utilization of Fe in leaves. with variations in yield of potato. It is interesting to note The results indicated that N in the leaves was nega- that there was no significant correlation of yield with the tively related to the Zn and Fe in the soil and K in leaves soil properties measured. The concentrations of mineral el- was negatively related to Cu and Zn in soil (Table 3). The ements in leaf and tubers of potato are influenced by both significantly negative relationships may be an explanation
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TABLE 3 - The relationships between nutrient concentration of leaves and soils
Soil N Soil P Soil K Soil Cu Soil Zn Soil Mn Soil Fe Leaf N 0,078 0,113 -0,270 -0,235 -0,517** -0,321 -0,445* Leaf P -0,172 0,236 -0,047 -0,111 0,159 -0,035 -0,580** Leaf K 0,333 -0,103 0,267 -0,394* -0,490** -0,310 -0,062 Leaf Cu 0,174 0,083 0,130 -0,211 -0,328 -0,113 -0,290 Leaf Mn 0,223 0,156 0,031 -0,179 -0,032 0,163 -0,077 Leaf Fe -0,264 -0,239 -0,458* -0,030 -0,331 -0,185 -0,334 Leaf Zn 0,279 0,261 0,343 -0,074 0,047 0,079 -0,150 **. Correlation is significant at the 0.01 level (2-tailed), *. Correlation is significant at the 0.05 level (2-tailed).
to dilution effect from the increase in plant biomass. In- responded in N2-fixing bacterial inoculation with N com- creasing plant biomass increased Fe and Zn uptake from binations than N fertilizer applications and microorgan- soil by plant. There was significantly negative relationship isms significantly promoted growth and increase yield in between Fe concentrations in leaves and K concentration potato. There was significantly negative relationship be- in soil (Table 3). Çelik et al. [19] suggested that increasing tween leaf P and Fe in soil. At acidic pH values, phosphate applications of K can inhibit the Fe uptake and transloca- ions react with aluminum (Al) and iron (Fe) to again form tion of Fe in plants. less soluble compounds and in fact are optimally available at a slightly acidic pH, e.g. 6.5 to 6.8. Leaf P concentration Relationships between soil chemical and physical was also positively correlated with soil pH, indicating that properties on nutrient status of leaves were found (Table soil P availability is directly affected by soil pH (Table 4). 4). Significantly negative correlation between N concen- Significant correlations between leaf P and K with soil tex- trations in leaves and Fe in soil was found. Potassium in ture were found (Table 4). Leaf P was negatively corre- the leaf was negatively associated with Cu and Fe levels in lated with loam whereas leaf P concentration was posi- the soil. This adverse effect was related to increase plant tively correlated with sand in soil. There was significantly biomass. Iron and Cu uptake of plant increases with in- negative relationship between leaf K and soil pH. Besides, creasing K uptake. Increasing nitrogen by plants also in- leaf K was positively correlated with loam. The propor- creases iron uptake from soil. Biological nitrogen fixation tions of sand and loam in soil might be related to internal is an essential process in the nitrogen cycle, providing water-holding capacity or drainage in these soils. These available nitrogen for plants. Especially, a symbiotic N2- complex interactions between minerals in the soil, plant tis- fixing bacteria lived in the rhizosphere is important for sue and tuber with the yield of potato may be due to inter- plant nutrition by increasing N uptake by the plants [20]. actions between uptake of mineral nutrients and distribu- The enzyme responsible for this process, nitrogenase, con- tion within the plant. Besides, there is significant genetic sists of two proteins, dinitrogenase (Mo-Fe protein) and ni- variation in tuber mineral concentrations both between and trogenase reductase (Fe protein) [21]. Iron has a key func- within Solanum species [16]. However, not much is known tion in nitrogenase enzyme and contributes nitrogen uptake about the genetic variation and nutrient uptake mechanisms of potato in particularly high soil organic matter condi- that contribute to the high accumulation of mineral ele- tions. Ekin et al. [22] reported that Potato could be more ments in the tuber of different potato species.
TABLE 4 - The relationships between nutrient concentration of leaves and soil properties
Soil Texture Soil pH Soil O M Clay Loam Sand Leaf N -0,259 0,065 0,211 0,114 -0,169 Leaf P 0,441* -0,168 -0,369 -0,627** 0,561** Leaf K -0,460* 0,325 0,189 0,395* -0,333 Leaf Cu -0,136 0,169 0,131 0,131 -0,142 Leaf Mn 0,027 0,210 -0,169 -0,061 0,116 Leaf Fe -0,148 -0,276 0,064 -0,143 0,059 Leaf Zn 0,190 0,269 -0,201 -0,063 0,133 **. Correlation is significant at the 0.01 level (2-tailed). *. Correlation is significant at the 0.05 level (2-tailed).
TABLE 5 - The ANOVA result of Stepwise Regression Analysisa (Adj R2=0.447)
Source df SS MS F P‐values Regression 2 14.498 7.249 55.364 <0.001b Residual 137 17.938 0.131 Total 139 32.436 a. Dependent Variable: yield, b. Predictors: (Constant), Mn, N
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TABLE 6 - The Regression Coefficients estimated by Stepwise Regression Analysisa
Coefficients Standard Error t‐values P‐values Lower 95% Upper 95% Constant 2.055 0.342 6.005 <0.001 1.378 2.732 Mn 0.002 <0.001 8.276 <0.001 0.002 0.003 N ‐0.308 0,061 ‐5.091 <0.001 ‐0.428 ‐0.188 a: Dependent Variable: yield
Multiple regression analysis was used to predict the REFERENCES model explaining the relationships among nutrient levels in the soil, plant tissue, and tuber with yield of potato. The [1] Rusinovci, I., S. Aliu, S.H. Fetahu, S. Kaçiu, S. Salihu, D. independent variables were measurements of soil chemical Zeka and D. Berisha (2012). Contents of mineral substances in the potato (Solanum Tuberosum L.) tubers depending on cul- and physical properties and nutrient concentrations in the tivar and locality in the agro-ecological conditions of Kosovo. plant tissue and tuber at three regions. Stepwise methods ActaHort. 960, 289-292 were used to determine effective variable on yield of pota- [2] FAO, (2010). Statistical Database. From: http://www.fao- toes in regression analysis (Table 5 and Table 6). The re- stat.org/. gression coefficients estimated for equation of relationship [3] Abebe, T., S., Wongchaochant, T. Taychasinpitak, and O. between the leaf parameters and yield are given in Table 5. Leelapon (2012). Variation of Mineral Concentrations among The model used in this study has three parameters: an Different Potato Varieties Grown at Two Distinct Locations in Ethiopia. Kasetsart J. (Nat. Sci.) 46, 837-850 intercept, b0, and two regression coefficients, b1, and b2, as [4] Khan, A.A., M.S. Jilani, M.Q. Khan and M. Zubair (2011). , where = dependent variable, Effect of Seasonal Variation on Tuber Bulking Rate Of Potato. yield (kg per hill); Xi= independent variables: Mn and N The Journal of Animal and Plant Sciences, 21(1), 31-37 concentrations, respectively. Thus, by using the equation, stepwise regression model’s parameters were predicted [5] Davenport, J. R., P.H., Milburn, C.J. Rosenand and R.E. Thornton (2005). Environmental Impacts of Potato Nutrient and given as following model, Management. Amer J of Potato Res. 82, 321-328 2.055 0.002 0.308 . [6] Reis, R.A. and P.H. Monnerat (2000). Nutrient Concentrations Results indicated that three parameters of this regres- in Potato Stem, Petiole and Leaflet in Response to Potassium Fertilizer. Scientia Agricola, 57(2), 251-255 sion model were statistically significant (P<0.05) and the R2 (multiple correlation coefficient) was 0.447 (Table 5). [7] Warman, P.R. and K.A. Havard (1998). Yield, vitamin and mineral contents of organically and conventionally grown po- Plant tissue concentrations of N and Mn were significantly tatoes and sweet corn. Agriculture, Ecosystems and Environ- correlated with variations in yield of potato. Important var- ment 68: 207–216 iables positively associated with yield included Mn con- [8] Rivero, R.C., P.S. Hernandez, E.M.R. Rodriguez, J.D. Martin centration in the plant tissue. Variables negatively associ- and C.D. Romero (2003). Mineral concentrations in cultivars ated with yield included N concentrations in the plant tis- of potatoes. Food Chemistry 83, 247–253 sue. The trend of lower N concentrations in higher yielding [9] Tekalign, T. and P.S. Hammes (2005). Growth and productiv- plants was probably an indication of a greater dilution of N ity of potato as influenced by cultivar and reproductive growth in the plant tissue due to higher yield of potato. It was pos- II. Growth analysis, tuber yield and quality. Scientia Horticul- sible that if samples had been taken earlier in the growing turae 105, 29–44 season, the N concentration in the plant tissue would have [10] Christelle M. A., M. Ghislain, P. Bertin, M. Oufir, M.R. Her- been a better indicator of yield potential of potato. Manga- rera, L. Hoffmann, J. Hausman, Y. Larondelle and D. Evers (2007). Andean Potato Cultivars (Solanum tuberosum L.) as a nese concentration of the plant tissue was positively related Source of Antioxidant and Mineral Micronutrients. Journal of to yield and should be considered for future studies. We Agricultural and FoodChemistry 55 (2), 366-378 found no significant relationships between tuber yield and [11] Galdon, B.R., L.H. Rodriguez, D.R. Mesa, H.L. Leon, N.L. leaf K, S, Ca, Mg, Fe, Cu or Zn concentrations, but leaf Mn Perez, E.M.R. Rodriguez and C.D. Romero (2012). Differen- concentrations increased significantly with increasing tu- tiation of potato cultivars experimentally cultivated based on ber yield. Many investigators reported that manganese their chemical composition and by applying linear discrimi- treatment increased all plant characteristics relating to nant analysis. Food Chemistry 133, 1241–1248 yield and quality of potato crop [23,24,25,26]. In accord- [12] Lombardo, S., G. Pandino and G. Mauromicale (2013). The ance with these previous results, the findings of our study influence of growing environment on the antioxidant and min- eral content of ‘‘early’’ crop potato. Journal of Food Compo- suggest that especially N and Mn are important in the yield sition and Analysis 32, 28–35 of potato. [13] Bremner, J. M. (1965). Total nitrogen. In: Methods of Soil Analysis. Part 2: Chemical and Microbiological Properties. The authors have declared no conflict of interest. Edited by C. A. Black et al. Agron. Series 9. Amer. Soc. Agron. Madison, Wisconsin. pp. 1149-1178. [14] Murphy, J. and J.P. Riley (1962). A modified single solution method for the determination of phosphate in natural waters. Anal. Chim. Acta 27:31-36.
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[15] SPSS, 2002. SPSS for Windows release 11.0 versions, Copy- right SPSS inc., NY. [16] White, P.J., J.E. Bradshaw, M.F.B. Dale, G. Ramsay, J.P. Hammond, M.R. Broadley (2009). Relationships between yield and mineral concentrations in potato tubers. HortScience 44:6-11.
[17] Marsh, K.B. and L.A. Peterson (1990). Gradients in Mn accu- mulation and changes in plant form for potato plants affected by Mn toxicity. Plant and Soil 121,157-163
[18] Mousavi, S.R., M. Shahsavari and M. Rezaei (2011). A gen- eral overview on manganese (Mn) importance for crops pro- duction. Australian Journal of Basic and Applied Sciences, 5(9), 1799-1803. [19] Çelik, H., B.B. Aşık, S.Gürel and A.V. Katkat (2010). Effects of potassium and iron on macro element uptake of maize. Zemdirbyste-Agriculture, 97(1), 11–22 [20] Ekin, Z., (2011). Some analytical quality characteristics for evaluating the utilization and consumption of potato (Solanum tuberosum L.) tubers. African Journal of Biotechnology Vol. 10 (32), 6001-6010, [21] Hoover, T.R., A.D. Robertson, R.L. Cerny, R.N. Hayes, J. Im- perial, V.K. Shah and P.W. Ludden (1987). Identification of the V factor needed for synthesis of the iron-molybdenum co- factor of nitrogenase as homocitrate. Nature 329, 855-857.
[22] Ekin, Z., F. Oğuz, M. Erman, and E. Öğün (2009). The effect of Bacillus sp. OSU-142 inoculation at various levels of nitro- gen fertilization on growth, tuber distribution and yield of po- tato (Solanum tuberosum L.). African Journal of Biotechnol- ogy 8(18), 4418-4424 [23] Kelling, K.A. and P.E. Speth (2001). Effect of micronutrient on potato tuber yield and quality at Spooner, Department of soil Science University of Wisconson–Madis. [24] Crosier, C.R., N.G. Cremer and M.A. Cubeta (2004). Soil facts. Soil fertility management for Irish potato production in eastern North Carolina. North Carolina Cooperative Exten- sion. [25] Mousavi, S.R., M. Galavi and G. Ahmad (2007). Effect of zinc and manganese foliar application on yield, quality and en- richment of potato (Solanum tuberosum L.). Asian Journal of Plant Sciences, 6, 1256-1260.
[26] Ahmed, A.A.; M.M.H. Abd El- Baki; Y.I. Helmi and M.R. Shafeek (2013). Improvement of Potato Growth and Produc- tivity by Application of Bread Yeast and Manganese. Journal of Applied Sciences Research, 9(8), 4896-4906
Received: September 03, 2014 Revised: August 27, 2015 Accepted: September 08, 2015
CORRESPONDING AUTHOR
Kürşat Korkmaz Ordu University Faculty of Agriculture Department of Soil Science and Plant Nutrition 52200 Altınordu/Ordu TURKEY
E-mail: [email protected]
FEB/ Vol 24/ No 11c/ 2015 – pages 4108 - 4113
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DEVELOPMENT AND TESTING OF A DIATOM-BASED INDEX OF BIOTIC INTEGRITY FOR RIVER ECOSYSTEMS IMPACTED BY ACID MINE DRAINAGE IN GAOLAN RIVER, CHINA
Xiaoyu Dong1,2, Xinghuan Jia1, Wanxiang Jiang1, Naicheng Wu1, Tao Tang1,* and Qinghua Cai1,*
1State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, China 2University of Chinese Academy of Sciences, Beijing 100049, China
ABSTRACT 1. INTRODUCTION
Acid mine drainage (AMD) characterized by low pH, Acid mine drainage (AMD) is a major stressor to global high concentrations of dissolved metals and metal oxide aquatic ecosystems [1-6], and is often characterized by high deposits affects aquatic ecosystems severely. However, the acidity, high concentrations of dissolved metals (e.g. Fe, Al, quantitative impacts of AMD on lotic ecosystem, especially Zn, Mn), and metal oxide deposition [7]. The influences of the seasonal variation of impacts, are still unclear. We devel- AMD on freshwater ecosystems can be multiple: contami- oped a diatom-based index of biotic integrity (AMD-DIBI) nating both surface runoff and groundwater [8]; altering to evaluate AMD impairment on Gaolan River. Pollution nutrient cycles and abiotic factors [4]; impairing the struc- Tolerance Index (PTI), % No. of Cocconeis species, % No. tures and functions of communities including algae [6, 7, 9], of Pinnularia species, No. of Cymbella species, relative benthic invertebrates [10] and fish [11]. Moreover, AMD abundance of prostrate diatoms, and relative abundance of can impact food webs and relationships among organisms Achnanthes were selected to build AMD-DIBI. AMD-DIBI [12], resulting in changes to community succession or extinc- scores between reference and impaired sites displayed sig- tion of sensitive taxa [13]. Therefore, remediation measures nificant differences. On the basis of AMD-DIBI scores, the for river ecosystems impacted by AMD become more and ecological conditions of most reference sites were “excel- more urgent, and have been paid more and more attentions lent” or “good”, while the impaired sites were “Poor” or worldwide. “Moderate”. AMD impairments were more severe in dry Assessing and quantifying impacts of AMD are of season than that in wet season: about 20km lower reach great significance to make management plans and take re- were impaired in wet season, while 50km at least in dry mediation activities. Many advantages make benthic dia- season. AMD-DIBI is meaningful to assess the ecological tom ideal organisms to assess ecological conditions of conditions, quantify impairment of AMD and measure the streams. Stream benthic diatom is sessile, short-life cycle effectiveness of restoration. [14], and the base of aquatic food chain [15]. It has distinct taxonomic characters but also can respond sensitively to environmental changes, thus was widely used as indicators of ecological conditions in lotic waters [16]. Numerous di- KEYWORDS: Acid mine drainage (AMD); Diatom; Index of biotic integrity (IBI); Quantitative assessment; River health. atom indexes have been developed. For example, Diatom Assemblage Index (DAIpo) [17] was used to indicate or- ganic pollution, Pollution sensitivity Index (PSI) [18], and Trophic Diatom Index (TDI) [19, 20] was used to indicate
the trophic condition of water, Saprobic Index [14] and Bi- ABBREVIATIONS ological Diatom Index (BDI) were used to assess water quality [21]. Thus, diatoms are considered to be a good in- AMD, Acid Mine Drainage; IBI,Index of biotic integ- dicator of AMD. Many studies have examined the impacts rity; AMD-DIBI, a diatom-based index of biotic integrity of AMD on biodiversity, species composition, and produc- for river ecosystem impacted by AMD; ICP-AES,induc- tivity of benthic diatom communities [6, 7, 22, 23], and sev- tively coupled plasma atomic emission spectrometry; CoI, eral have proved that diatoms can be used to indicate the im- Correlation index; PTI,Pollution Tolerance Index pacts of AMD [24-26]. Furthermore, index of biotic integrity
(IBI) was commonly used in aquatic ecosystems impacted
by different factors [16, 27-29] as an effective tool for as- * Corresponding author sessing impacts and making restoration and conservation
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policy [26, 30], and the IBI based on macroinvertebrate has into Gaolan River almost without any treatment, and pol- been established to assess the impacts of AMD [31]. Nev- luted the water seriously [34]. ertheless, macroinvertebrates and diatoms differ in their Twelve sites (site 1-12) were investigated in Novem- sensitivity to AMD, and to our knowledge, IBI based on ber 2006, January and July 2007 respectively, and 9 sites diatoms used to assess the impacts of AMD on lotic eco- (site 1-9) in May 2007 because of flood. Consequently, system has been rarely built yet but Zalack et al. [28]. 45 samples were collected in total. The average distance Above all, the quantitative impacts of AMD on diatoms re- between adjacent sites was about 6 km. Site 1-4 (on the main poorly understood, especially on the seasonal varia- tributary named Xiayang River) with little anthropogenic tion of impacts. impact and no AMD inputs were designated as reference Gaolan River was severely polluted by AMD [32], sites according to the standards of reference sites [35], and providing an appropriate opportunity to study the impacts the remaining 8 sites located in the mainstream (site 5-12) of AMD on aquatic ecosystems. Former studies in Gaolan were designated as impaired sites (Fig. 1), among which River suggested that AMD had a great negative effect on site 5-7 and site 8-12 were before and after the confluence benthic communities [33, 34]. However, the quantitative of Xiayang and Gaolan River respectively. Pyrite was lo- impacts are still unclear. So this study focuses on (1) de- cated upstream of site 5, and site 5-12 were impaired by veloping a benthic diatom-based IBI for river ecosystems AMD of different severity. With a climate of subtropical impacted by AMD (AMD-DIBI) in a small catchment, (2) monsoon style, the Gaolan River undergoes dry season in assessing and quantifying the impairments of AMD on November and January when the precipitation was less Gaolan River in different seasons, to provide some sugges- than 30 mm per month; and wet season in May and July tions to aquatic resource management, and offer a meas- when the monthly precipitation was more than 150 mm. urement of remediation effectiveness and recovery infor- Benthic diatoms were collected from 3-5 randomly se- mation of rivers in the future. lected stones (with diameter range of 10-20 cm) in each section, and three sections were sampled repetitively in each site, to make sure as many as possible habitats being 2. MATERIALS AND METHODS included. Periphyton was sampled with a circular lid (with radius of 2.7 cm). The stone surface covered within the lid 2.1 Study area and field sampling was vigorously scrubbed by a nylon brush and flushed 3- Gaolan River is one of the three main tributaries of 4 times with distilled water. Each sample had 3 replicates, Xiangxi River, located in Xingshan County, Hubei Prov- and each replicate was divided into three parts: one was ince. Pyrite resources are rich in this region, with reserves preserved with 4% formalin for identification, one was of 1.274 million tons in which sulfur content reach up to used to measure the concentrations of chlorophyll a (Chl- 40 % [32]. Pyrite mining activity began from the 1980s and a), and the other one was prepared for ash free dry mass stopped in 2004. After exploitation, the rejects were flowed (AFDM) measurement [36].
FIGURE 1 - Location and distribution of the sampling sites.
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2.2 Identification other box, and a value of 0 when both medians were within Permanent diatom slides were prepared for diatom iden- the range of the other box. Final metrics were selected tification after acid digestion of sample. At least 300 valves based on separation power ≥2 and CV < 1. 4 and 0–100 were counted at 1000× magnification under oil immersion. scaling system was used to normalize the final metric [16]. Diatoms were identified to the lowest taxonomic level pos- For metrics that decreased at impaired sites, the standardi- sible following the identification reference of Patrick and zation formula was: Reimer [37, 38]; Qi [39]; Zhu and Chen [40]; Qi and Li [41]; Shi [42] and Hu and Wei [43]. The total number of diatoms were calculated and converted to densities in unit (1) area expressed as ind. /m2 [16, 29, 34]. and for metrics that increased at impaired sites, stand- 2.3 Measurement of environmental factors ardization formula was: Conductivity (Cond), total dissolved solid (TDS), dis- solved oxygen (DO), pH were measured in situ with a multi-parameter water quality meters (Horiba W-23XD, (2) US). Concentration of NH4-N, SiO2-Si, PO4-P, T-P, NO3- N, and T-N were measured using a segmented flow ana- where X95%= 95th percentile of the metrics, and X5%= ++ lyzer (Skalar San , Netherlands). Dissolved Al, Ca, Cd, 5th percentile of the metrics. And X’= 0 when X< X5%, Cr, Cu, Fe, Mg, Mn, Pb, and Zn were measured with a in- whileX’=100 when X> X95%. The total AMD-DIBI values ductively coupled plasma atomic emission spectrometry were the average of metric scores based on 0–100 scaling (ICP-AES) after filter and acidify the water with HNO3 to system. pH<2. The measure methods were introduced in detail by Jia et al. [34]. 2.4 Independent test of metrics and AMD-DIBI
2.3 Development of AMD-DIBI We tested the AMD-DIBI and its metrics using the testing dataset including 8 reference and 13 impaired sam- In order to establish a benthic diatom-based IBI appli- ples. The calculation and scaling system of the final met- cable throughout the year, data both in dry and wet season rics for AMD-DIBI were the same as used for the training were selected in the development and validation process. dataset. Four methods were used to evaluate the AMD-DIBI Our 45 samples were divided into two dataset: 24 samples in the testing samples: 1) separation power. 2) % sites cor- that sampled in January 2007 (dry season) and July 2007 rectly classified, that is whether AMD-DIBI developed from (wet season) were training dataset, and 21 samples sampled training dataset could distinguish reference sites from im- in November 2006 (dry season) and May 2007 (wet sea- paired ones at testing dataset with a critical value of the av- son) were testing dataset. erage of the 75th percentile of AMD-DIBI scores at im- Training dataset was used to develop the AMD-DIBI paired sites and the 25th percentile of AMD-DIBI scores at based on the methods described by Wang et al. [44], and the reference sites from the training dataset [29]. 3) Box-plot metrics according to related literatures were selected [7, 22, separation ratio, in which we divided the distance between 27-29]. 24 metrics were calculated preliminarily, including boxes (defined by the difference between the 25th percentile 7 categories: biomass index, biotic index of diatoms, diver- of reference sites and the 75th percentile of all sites in the sity index, growth forms, similarity, taxonomic composition, dataset) by the median AMD-DIBI score of reference sites sensitive species and tolerant species (Table 1). The metrics [29, 44]. 4) Correlation index (CoI) and Cumulative R2 [29]: were selected by four steps: 1) metrics with medians of 0 for both reference and impaired sites were eliminated due to the (3) prevention of identifying the differences between these two where rs is Spearman’s correlation coefficient between groups [44]. 2) Metrics with significant differences (p< 0.05) AMD-DIBI and a given environmental variable. between reference and impaired sites were selected, and the ability to separate reference and impaired sites was evalu- ated by non-parametric Kruskal–Wallis test, since many (4) metrics did not follow normal distributions (Kolmogorov– Smirnov test, p< 0.05) despite after several transfor- where S is the number of rs statistically significant at mations. 3) Separation power and coefficient of variation p< 0.05, and n is the number of environmental variables (CV) were used to determine the final metrics. Separation evaluated. The developed AMD-DIBI was considered ac- power was defined as the degree of overlap between boxes ceptable if there was no significant difference between val- (25th and 75th percentiles) in box plots of the metric values ues of the four criteria at training and testing dataset (ex- for reference and impaired sites [44]. We assigned a sepa- amined by paired t-test) [29, 45]. In our study, a sample ration power of 3 when boxes did not overlap between the was the average of three replicates. The descriptive statis- two site groups, a value of 2 when interquartile ranges tics, paired t-test, correlation, and nonparametric tests were overlapped but did not reach medians, a value of 1 when conducted with SPSS 16.0, and the statistical figures were only one median was within the interquartile range of the created with Origin7.0.
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TABLE 1 - Benthic diatom community metrics, taxonomic levels, and their descriptions
Metrics Taxonomic Description level Biomass index chlorophyll a The concentration of chlorophyll a of benthic diatoms AFDM Ash-free dry mass of benthic algae AI index The ratio of these AFDM: Chl a
Biotic index of diatoms KYDPTI Species Weighted average of average abundance and tolerance value [46] pH index Species Weighted average of average abundance and tolerance value [50]
Diversity index Shannon Diversity Species Measure of evenness and richness Species richness Species Number of species in the count No. of genera Genus Number of genera in the count
Growth form % prostrate individuals Genus Relative abundance of prostrate genera % erect individuals Genus Relative abundance of erect genera % stalked individuals Genus Relative abundance of stalked genera % unattached individuals Genus Relative abundance of unattached genera % mobile individuals Genus Relative abundance of motile genera
Similarity % reference species Species % of species found in reference sites that occurred in impaired sites No. of distinct reference species Species No. of species found primarily in reference sites not in impaired sites Average similarity to reference Species Mean Bray–Curtis similarity in species composition sites to reference sites
Taxonomic composition Total density Species Total number of diatoms Dominant species Species The density of dominant species % dominant species Species Relative abundance of dominant species % (genus) individuals a) Genus Relative abundance of (Achnanthes b), Cymbella c), Eunotia, Fragilaria, Nitzschia, Pinnu- laria, Synedra genus) No. of (genus) species Genus No. of (Achnanthes, Cymbella, Eunotia, Fragilaria, Nitzschia, Pinnularia, Synedra genus) species % No. of (genus) species Genus % No. of (Achnanthes, Cymbella, Eunotia, Fragilaria, Nitzschia, Pinnularia, Synedra ge- nus) species
Sensitive species and Tolerant species KY % sensitive individuals Species Relative abundance of KYDPTI sensitive species KY % tolerant individuals Species Relative abundance of KYDPTI tolerant species a) The genera that have been reported in AMD were calculated for this metrics [7]. b) Achnanthes species: include Achnanthes, Achnanthidium, Karayevia, Planothidium, Platessa and Rossithidium taxa. c) Cymbella species: include Cymbella, Delicata, Encyonema and Reimeria taxa.
2.5 Application of the AMD-DIBI to assess AMD impacts 3. RESULTS After testing, we applied AMD-DIBI to calculate the score of ecological condition for each site in different A total of 157 diatom taxa were identified in our study month. The ecological conditions were classified as excel- period. During dry season, 106 and 140 taxa were recorded lent, good, moderate, or poor by using the 90th, 50th and in reference and impaired sites, respectively; while 74 and 25th percentile of the total AMD-DIBI scores of all sites 102 during wet season. Community composition and the [28, 29, 44]. dominant species were different between dry and wet sea- son, so were between reference and impaired sites. Ros- Finally, we use One-way ANOVA in SPSS 16.0 to sithidium linearis (W.Sm.) Round & Bukht. was the most compare the difference of AMD-DIBI scores among refer- abundant species both in reference and impaired sites dur- ence sites group (site 1-4) and impaired sites groups of be- ing both seasons. However, in reference sites, the dominant fore (site 5-7) and after confluence (site 8-12). If equal var- species were Achnanthidium minutissimum (Kütz.) Czarn. iances assumed, LSD were used for multiple comparisons; and Eunotia spp. during both seasons, and were Cocconeis if not, then Tamhane’s T2 were conducted. placentula Ehren., Diatoma mesodon (Ehren.) Kütz., Fragi- laria capucina var. vaucheriae (Kütz.) Lange-Bert., Gom- phonema parvulum (Kütz.) Kütz., Gomphonema parvulum
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var. subelliptica Brand., and Gomphonema constrictum var. 3.1 Development and test of the AMD-DIBI capitatum (Ehren.) Grun. in dry season, and were Fragilaria Six metrics- Pollution Tolerance Index (PTI), % No. fragilarioides (Grun.) Choln. and Planothidium ellipticum of species of Cocconeis genus, % No. of species of Pinnu- (Cl.) Round & Bukht. in wet season. In impaired sites, the laria genus, No. of species of Cymbella genus (including communities were dominant by Cocconeis placentula Eh- Delicata, Encyonema and Reimeria), % prostrate individu- ren. during both seasons, and Achnanthidium minutissimum als, and % Achnanthes individuals (including Achnanthid- (Kütz.) Czarn., Eunotia spp., and Melosira varians C.Ag. in ium, Karayevia, Planothidium, Platessa and Rossithidium) dry season, and Gomphonema constrictum var. capitatum - were selected following the procedure mentioned above (Ehren.) Grun., Planothidium ellipticum (Cl.) Round & Bu- to build AMD-DIBI. The metric % No. of species of Coc- kht., and Reimeria sinuata (Gre.) Kociolek & Stoermer in coneis genus decreased at impaired sites, while the other 5 wet season. The relative abundance of the dominant species metrics increased. The AMD-DIBI scores at reference sites is shown in Table 2. were significantly higher than impaired sites (Mann-Whit- Among the 20 environmental variables, concentrations ney U=1, p<0.001; separation power=3, Fig. 2). of Al and Mn, conductivity, TDS, and pH showed signifi- cant differences between reference and impaired sites (Ta- ble 3).
TABLE 2 - The relative abundance ( % ) of dominant species (>5%) in reference and impaired sites during dry and wet season
Dry season Wet season Dominant species Reference site Impaired site Reference site Impaired site (n=8) (n=16) (n=8) (n=14) Achnanthidium minutissimum (Kützing) Czarnecki 5.19 7.32 17.79 Cocconeis placentula Ehrenberg 7.54 6.81 17.56 Diatoma mesodon (Ehrenberg) Kützing 8.35 Eunotia spp. 6.1 6.23 9.31 Fragilaria capucina var. vaucheriae (Kützing) Lange-Bertalot 8.09 Fragilaria fragilarioides (Grunow) Cholnoky 9.17 Gomphonema constrictum var. capitatum (Ehrenberg) Grunow 7.26 14.88 Gomphonema parvulum (Kützing) Kützing 6.25 Gomphonema parvulum var. subelliptica Brandy 5.4 Melosira varians C.Agardh 5.53 Planothidium ellipticum (Cleve) Round & Bukhtiyarova 15.59 5.39 Reimeria sinuata (Gregory) Kociolek & Stoermer 6.83 Rossithidium linearis (W.Smith) Round & Bukhtiyarova 11.93 44.02 13.87 31.08
TABLE 3 - Differences of environmental variables between reference and impaired sites
Environmental variables Reference site Impaired site p value (n=16, mean±SD) (n=29, mean±SD) Al (mg L-1) 0.06 ±0.04 3.69±6.35 0.005*a) Ca (mg L-1) 25.02±12.13 29.65±13.16 0.326 Cd (mg L-1) 0.00±0.00 0.00±0.00 0.632 Cr (mg L-1) 0.00±0.00 0.00±0.01 0.206 Cu (mg L-1) 0.01±0.00 0.01±0.01 0.973 Fe (mg L-1) 0.1±0.16 1.13±2.80 0.122 Mg (mg L-1) 9.08±4.64 11.96±5.40 0.131 Mn (mg L-1) 0.00±0.00 0.19±0.28 0* Pb (mg L-1) 0.00±0.00 0.01±0.02 0.941 Zn (mg L-1) 0.02±0.01 0.02±0.02 0.779 Conductivity (us cm-1) 174.48±66.76 261.45±119.80 0.002* TDS (mg L-1) 99.14±37.67 137.00±64.46 0.036* DO (mg L-1) 8.55±0.46 8.85±0.88 0.92 pH 8.12±0.53 5.71±1.63 0.012* NH4-N (mg L-1) 0.04±0.03 0.06±0.07 0.562 SiO2-Si (mg L-1) 6.75±4.57 7.54±6.23 1 PO4-P (mg L-1) 0.01±0.01 0.01±0.03 0.783 T-P (mg L-1) 0.03±0.02 0.03±0.05 0.119 NO3-N (mg L-1) 0.46±0.33 0.51±0.12 0.906 T-N (mg L-1) 1.20±1.06 0.93±0.91 0.244 a) *, Significant difference between reference and impaired sites.
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ney U=13, p=0.003; separation power=3, Fig. 3a). Except % No. of species of Cocconeis genus and % prostrate individ- uals, the other 4 metrics had separation power ≥2 (Fig. 3b- g). The critical value to classify reference and impaired sites was 51.27, and % sites classified correctly in the training and testing dataset were 95.8% and 85.7% respectively. In the training and testing dataset, the box-plot separation ratios were 0.041 and 0.259. AMD-DIBI scores were significantly correlated with Al, Mn, pH and Mg, Mn, TN respectively,
FIGURE 2 - Box plots of total AMD-DIBI scores of reference and and Spearman’s correlation coefficients between AMD- impaired sites in training dataset. Boxes show interquartile ranges DIBI scores and environmental variables (rs) are shown in (25th and 75th percentiles), middle lines are medians, points in the Table 4, and cumulative R2 values were 3.747 and 2.337; CoI box are mean value, the upper and below lines are 95th and 5th per- were 0.035 and 0.018. Paired t-test showed no difference be- centiles. tween values of training and those of testing dataset (p= We tested the AMD-DIBI and its metrics using the test- 0.42), in terms of separation powers, % sites classified cor- ing dataset. The AMD-DIBI scores showed significant dif- rectly, box-separation ratios, Cumulative R2 and CoI (Table ference between reference and impaired sites (Mann-Whit- 5), indicating that the AMD-DIBI was reliable.
FIGURE 3 - Box plots of total AMD-DIBI scores (a) and the six metrics (b. PTI; c. percentage of the species number of Cocconeis genus; d. percentage of the species number of Pinnularia genus; e. the species number of Cymbella species; f. relative abundance of prostrate diatoms; g. relative abundance of Achnanthes) for the reference and impaired sites in testing dataset. Boxes show interquartile ranges (25th and 75th percentiles), middle lines are medians, points in the box are mean value, the upper and below lines are 95th and 5th percentiles
4119 © by PSP Volume 24 – No 11c. 2015 Fresenius Environmental Bulletin
TABLE 4 - Spearman rank correlation coefficients (rs) between November 2006. For the impaired sites before the conflu- AMD-DIBI and environmental variables in training and testing da- ence, the ecological conditions were “poor”, but in May taset 2007 were “moderate”. The ecological conditions of sites Environmental variables AMD-DIBI downstream of the confluence of an un-impacted and the Training dataset Testing dataset impaired tributary improved to “moderate” or “good”, es- (n=24) (n=21) -1 a) pecially in May 2007 when the condition can reach to “ex- Al (mg L ) -1.000* -0.390 cellent”; nevertheless, there were still some sites classified Ca (mg L-1) -0.217 -0.040 Cd (mg L-1) -0.224 0.012 as “poor” in November 2006 (Table 7). Cr (mg L-1) — b) -0.221 Cu (mg L-1) — -0.066 Fe (mg L-1) -0.490 -0.275 Mg (mg L-1) -0.238 -0.569* Mn (mg L-1) -0.846* -0.487* Pb (mg L-1) -0.143 -0.298 Zn (mg L-1) -0.250 -0.366 Conductivity (us cm-1) -0.372 -0.406 TDS (mg L-1) -0.374 -0.690 DO (mg L-1) 0.459 -0.103 pH 0.888* 0.003 -1 NH4-N (mg L ) -0.076 0.339 -1 SiO2-Si (mg L ) -0.405 -0.121 -1 PO4-P (mg L ) -0.235 -0.330 T-P (mg L-1) -0.012 -0.149 -1 NO3-N (mg L ) 0.227 -0.340 T-N (mg L-1) -0.002 0.493* a) *, Significant difference between AMD-DIBI and environmental varia- bles. b) —, Data absent.
TABLE 5 - Separation powers, % sites correctly classified, box-sepa- ration ratios, Cumulative R2 and CoI of AMD-DIBI from training and testing dataset
Training data set Testing data set separation power 3 3 % sites correctly classified 0. 958 0.857 box-plot separation ratio 0.041 0.259 Cumulative R2 3.747 2.337 CoI 0.035 0.018 Paired t-test df=4, t=0.898, p=0.42
3.2 Assessing AMD impacts with the AMD-DIBI The AMD-DIBI scores ranged from 13.42 to 85.19 among study sites. The mean AMD-DIBI scores of refer- ence sites, sites before and after the confluence of Xiayang River and Gaolan River were 62.83, 25.80 and 42.37 re- spectively (Fig. 4c). Impairment of AMD varied among seasons. During the wet season (May 2007 and July 2007), the average AMD-DIBI score of sites after the confluence was 50.38, which was high and close to that of reference sites (63.19, Table 6, Fig. 4b), while during the dry season (November 2006 and January 2007), the average score of sites after the confluence was 36.76, significantly lower than that of reference sites (62.48) though better than sites before the confluence (24.28, Table 6, Fig. 4a). Sites with scores greater than 78.38 were categorized as excellent group, 42.01 to 78.38 as good group, 28.23 to 42.01 as moderate group, and below 28.23 as poor group. Based on FIGURE 4 - The AMD-DIBI scores of the study area in wet and dry the classification, the ecological conditions of most refer- season (a. AMD-DIBI scores in dry season; b. AMD-DIBI scores in ence sites were “excellent” or “good”, except two sites in wet season; c. Average AMD-DIBI scores of the study area).
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TABLE 6 - Descriptive statistics and the results of multiple comparisons of AMD-DIBI scores in reference sites, sites before and after conflu- ence
AMD-DIBI Scores Test of Homogeneity R a) Bb) Ac) of variances d) Wet season 63.19±10.41 a 27.31±11.21 b 50.38±16.43 a 0.871 Dry season 62.48±17.76 a 24.28±2.42 b 36.76±8.28 c 0.006 a) R, reference sites. b) B, sites before the confluence of Xiayang and Gaolan River. c) A, sites after the confluence of Xiayang and Gaolan River. d) In wet season, we use LSD to make multiple comparisons; In dry season, we use Tamhane’s 2 to make multiple comparisons.
TABLE 7 - The ecological conditions of study sites in different months
Site group Site November 2006 January 2007 May 2007 July 2007 Average Reference group 1 M a) Ec) E G G 2 G b) G G G G 3 G E G G G 4 M G G G G impaired group before the confluence 5 P d) P M P P 6 P P M P P 7 P P M P P impaired group after the confluence 8 M M G M G 9 M G G G G 10 M M —e) G M 11 P M — P M 12 P G — G G a) M, moderate b) G, good c) E, excellent d) P, poor e) —, Data absent
4. DISCUSSION tula Ehren., and C. placentula var. euglypta (Ehren.) Grun. Although Cocconies is a genera with little species diver- The developed AMD-DIBI was an effective indicator sity, but % No. of Cocconies taxa had significant difference of the impairments caused by AMD and its metrics were between reference and impaired sites. C. placentula var. suitable and reliable for assessing ecological conditions in euglypta were just observed once and C. placentula wide- this area. Among the 6 metrics finally selected for building spread distributed among every site during both dry and AMD-DIBI, pollution Tolerance Index (PTI) can reflect wet season. C. pediculus preferred to attach on filamentous the tolerance degree of benthic diatom communities [46], algae which decreased in impaired sites [48, 49], so that so PTI value of impaired sites is higher than the reference they were mainly found in the habitats where pH>7 [50]. sites (Fig. 3b). But in the wet season, as the dilution effect of In addition, the taxa in reference were much less than that rain on the pollution, PTI value of some sites downstream in impaired site, the absence of C. pediculus could make decreased to very low value, such as site 8 in May 2007, PTI great contributions to differences of the metric % No. of value was 1.778. The genera Pinnularia and Achnanthes Cocconies taxa between reference and impaired sites. Pros- were more frequently observed in impaired sites (especially trate diatoms always adhere to the substrate with the entire Pinnularia), implying that these diatoms are more tolerant to valve surface [51], but some prostrate genera were highly AMD. Similar findings were also reported by other research- motile, such as Cymatopleura, Gyrosigma, Hantzschia, ers [6, 7, 47]. Thereinto, genus Pinnularia is particularly Nitzschia, and Surirella [44]. They can move through sed- useful in indicating low pH [22, 24, 34]. In our study, the iments and reach the surface for light, hence can survived number of Cymbella species (including Delicata, Encyo- at impaired sites, where fine oxide particulates settled out nema and Reimeria) in impaired sites was more than that in onto the stream bed. It seemed that the impairments of reference sites. In reference sites, the Cymbella species were AMD were a combination of low pH, heavy metal contam- mainly Cymbella affinis, Encyonema minutum, Delicata del- ination and metal oxide deposition, and this was demon- icatula and Reimeria sinuata. While in impaired sites, ex- strated by many researches [7, 52, 53], and our metrics can cept the species mentioned above, some acid-tolerant spe- reflect the impacts comprehensively. cies bloomed such as C. tumida, E. microcephala and E. In our study, ecological conditions of reference sites muelleri [22, 24]. For Cocconies genera, three taxa were were generally better than that of impaired sites (Fig. 4). found in our study: Cocconeis pediculus Ehren., C. placen- Significant differences were observed between reference
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sites and impaired sites both in the wet and dry season. varied and gradually eased as the river flowed to lower However, the AMD-DIBI scores of reference sites didn’t reach, which indicated the impacts can be correlated with vary much between the wet and dry season (the mean val- water flow. So we can restore the impaired reaches by in- ues were 63.19 and 62.48 respectively, Table 6), while creasing water flow to neutralize low pH and dilute the pol- scores of the impaired sites in dry season was substantially luted water with high concentrations of metal oxide depo- lower than that in wet season (the mean values were 40.57 sitions. But how much water was needed for restoration? in wet season and 32.08 in dry season, Table 6, Fig. 4). It Further study of quantitative relation between water flow seems that AMD impairments seasonally varied and the and the reach of AMD impairments is still needed to make. impairments were more severe in dry season (Fig. 4). To determine how severe the impairments were, impaired sites were divided into two groups based on the locations before 5. Conclusions and after confluence (site 5-7 as group 1 and site 8-12 as group 2) and multiple comparisons of scores were made In conclusion, we developed AMD-DIBI composed of between groups (Table 6). During the wet season, the 6 diatom metrics that are sensitive to AMD impairments, AMD impacts degraded after the confluence, and no sig- i.e. Pollution Tolerance Index (PTI), % No. of species of nificant difference showed between the reference sites and Cocconeis genus, % No. of species of Pinnularia genus, the sites after the confluence (Table 6). It meant that the No. of species of Cymbella genus, % prostrate individuals, AMD impaired approximately 20 km reach (distance be- and % Achnanthes individuals. It can be used as an effec- tween Pyrite and site 7). On the contrary, during the dry tive indicator to assess ecological conditions of river im- season, significant difference between reference sites and pacted by AMD, and a quantitative measurement method sites after confluence can be observed (Table 6), which in- of the impairments. The ecological conditions of reference dicate that AMD impaired 50 km at least (distance between sites were better than that of impaired sites. AMD impair- Pyrite and site 12), although the impairments alleviated af- ments were varied between seasons: about 20 km lower ter confluence, and there were also significant difference reach were impaired in the wet season, while the influence between group 1 and 2. The possible reason is that AMD distance reached 50 km during the dry season. We recom- contamination is correlated with precipitation. Although mend an effective remediation by increasing water flow of AMD loadings were higher in wet season due to more rain- river. While further research would focus on how much fall, river flow was also higher which can dilute metal ionic water is needed for restoration from AMD impairments. concentrations in the mine drainage and reduce water acid- ity as well, resulting in alleviation of pollution. However few reference sites in November 2006 were ACKNOWLEDGMENTS classified as unhealthy sites, which might due to the lack of necessary supply of nutrients in mountain streams, and dia- This project was funded by Ministry of Environmental tom density and species richness were very low. Moreover, Protection of the People’s Republic of China (2012ZX07104- there were differences on structure and composition of ben- 002), and the Administrative Bureau of Shennongjia National thic diatom communities among seasons, so if we assess the Nature Reserve (2012SNJ002). ecological conditions of the same site in different months us- ing the AMD-DIBI, the results differ. Besides, the AMD- The authors have declared no conflict of interest. DIBI scores of lower reach (site 10, 11) were greatly various.
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[48] Jahn, R., Kusber, W.H. and Romero, O.E. (2009) Cocconeis pediculus Ehrenberg and C. placentula Ehrenberg var. placen- tula (Bacillariophyta): Typification and taxonomy. Fottea 9, 275–288. [49] Urrea-Clos, G. and Sabater, S. (2009) Comparative study of algal communities in acid and alkaline waters from Tinto, Odiel and Piedras river basins (SW Spain). Limnetica 28, 261- 272. [50] Van Dam, H., Mertens, A. and Sinkeldam, J. (1994) A coded checklist and ecological indicator values of freshwater diatoms from the Netherlands. Netherland Journal of Aquatic Ecology Received: October 07, 2014 28, 117-133. Revised: January 27, 2015 [51] Berthon, V., Bouchez, A. and Rimet, F. (2011) Using diatom Accepted: February 16, 2015 life-forms and ecological guilds to assess organic pollution and trophic level in rivers: A case study of rivers in south-east- ern France. Hydrobiologia 673, 259-271. CORRESPONDING AUTHOR [52] Ferreira da Silva, E., Almeida, S.F., Nunes, M.L., Luís, A.T., Borg, F., Hedlund, M., de Sá, C.M., Patinha, C. and Teixeira, P. (2009) Heavy metal pollution downstream the abandoned Prof. Qinghua Cai Coval da Mó mine (Portugal) and associated effects on epi- Institute of Hydrobiology lithic diatom communities. Science of the Total Environment Chinese Academy of Sciences 407, 5620-5636. No. 7 Donghu South Road [53] Battarbee, R.W., Charles, D.F., Dixit, S.S. and Renberg, I. Wuchang District (1999) Diatoms as indicators of surface water acidity. The di- 430072 Wuhan, Hubei Province atoms: applications for the environmental and earth sciences. P.R. CHINA Cambrige University Press, Cambrige, pp. 98-121. E-mail: [email protected] [54] Biggs, B. (1996) Patterns in benthic algae of streams, In Algal ecology: Freshwater Benthic Ecosystems, Academic Press, Dr. Tao Tang New York, pp. 31-56. Institute of Hydrobiology [55] Huang, L., Bai, J., Xiao, R., Gao, H. and Liu, P. (2012) Spatial Chinese Academy of Sciences distribution of Fe, Cu, Mn in the surface water system and their effects on wetland vegetation in the Pearl River Estuary of No. 7 Donghu South Road China. Clean–Soil, Air, Water 40, 1085-1092. Wuchang District 430072 Wuhan, Hubei Province P.R. CHINA E-mail: [email protected]
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DEVELOPING A CLIMATE-BASED RECREATION MANAGEMENT SYSTEM FOR A MEDITERRANEAN ISLAND
Hossein G.T. Olya1,* and Habib Alipour2
1Faculty of Tourism, British University of Nicosia, Girne, Turkey 2Faculty of Tourism, Eastern Mediterranean University, Gazimagusa, Turkey
ABSTRACT damage to the ecosystem. Hall conducted a survey that found that tourist visits damaged the vegetation cover of Weather and climate have a crucial influence on tour- Sherwood Forest in England [1]. Similarly, Filimonau et ism activities. As a result, tourism is vulnerable to the ef- al., in an assessment of the carbon impact of short-haul fects of climate change. One important category of such tourism using life-cycle analysis, revealed that tourism sig- effects is the change in climatic suitability of tourist desti- nificantly escalates the global carbon footprint [2]. Tour- nations. This study proposes a model for landscape man- ism is one of the main contributors toward global warming agement based on climate attractiveness for tourists within and climate change. Such adverse impacts of tourism could the context of the Theory of Ecological Modernization be minimized through a knowledge-based management [3- (TEM). The modelling process conforms to Ackoff’s wis- 6], the application of technology and financial sources [7] dom hierarchy, which transforms data into information, and the participation of stakeholders [8, 9]. This notion is knowledge and wisdom. Data were obtained from a mete- corroborates the principles of Theory of Ecological Mod- orological station in Northern Cyprus where the Mediterra- ernization (TEM), a school of thought in social sciences nean Sea and its favorable climate are key resources for the praised for addressing environmental issues [10]. tourism sector. The monthly Tourism Climate Index (TCI) The environmental impact of tourism is a foregone con- was used at all stations and was digitized in Geographical clusion [11]. However, the level of impact differs regionally Information System (GIS) setting. Spatial analysis was per- and is highly dependent on the concentration of tourist flow formed using Geostatistical methods, and tourism climate over time and space [12]. For instance, water consumption maps were produced for a 12 month period, aiming to iden- by the tourism sector, has become a great challenge, espe- tify temporal variations of TCI values on the island. A cli- cially in the Mediterranean but even more so in the case of mate-based Recreation Management System (RMS) is pro- water-stressed Northern region [13]. Overall, international posed to determine three variables that are both spatial and tourism water consumption may be less than 1% of na- temporal drivers for tourism activities: climate, environ- tional water use; however, in the case of Northern Cyprus ment and social factors. The main application of this pro- the rate increases to 4.8% due to incoming tourists [12]. It cess is the mitigation of problems of “seasonality”, as they is anticipated that chronic water shortage will occur in always pose a challenge for the managers of different des- Northern Cyprus and many other island states by the year tinations. 2050. Potential conflicts between residents and tourists
over water consumption are addressed by numerous au-
KEYWORDS: Climate change; GIS; seasonality; Ecological Mod- thors [12, 14-16]. ernization Theory; Northern Cyprus. In the context of climate change scenarios, conflicts
are expected to worsen due to “forecasted impacts of global climate change on the spatial and temporal varia- 1. INTRODUCTION bility of precipitation-particularly as it relates to recharge of surface and groundwater in regions all around the Tourism provides remarkable contributions such as world” [16, p2]. If adequate adaptation policies are not put economic gain, job creation, infrastructure development, into place to mitigate the conflicts, direct consequences on cultural exchange, improvement of the livelihood of local environmental quality, socio-economic wellbeing and sus- communities and funding of conservation projects. On the tainability of the tourist destinations, especially in island other hand, crowding or deterioration of the environment states are to be expected [15, 17]. negatively affect the social acceptance level of the hosts as Tourism has negative environmental impacts on Med- well as the attitude of the tourists and can cause irreversible iterranean islands, which are intensified by the fragility of the environment, human pressure and lack of spatial plan- * Corresponding author ning [18, 19]. Vehbi and Doratli [20] in their assessment of
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environmental impacts of tourism on coastal cities of the leakage and land mismanagement, there is hesitation about northern part of the island witnessed deterioration and re- the pro-poor role of tourism in improving the livelihood duction of green fields, loss of natural landscape, loss of and welfare of residents on the island [21]. Moreover, sea- open space, sea water and air pollution, noise pollution, sonality results in large differences in tourist arrivals and waste and visual pollution. The negative impact of tourism revenues in the winter and the summer seasons in Mediter- development is particularly strong in the coastal region ranean islands [22, 23]. To address this problem, an adap- where soils and water are highly susceptible to erosion and tive management system is needed to cover multifaceted pollution [21]. There is significant evidence of a trend in interactions between humans and the environment with re- environmental degradation in Northern Cyprus, whose spect to political, social, ecological, and geographical char- economy is highly dependent on tourism, especially na- acteristics of the island [18]. In other words, a comprehen- ture-based tourism. Figure 1 highlights an example of lack sive perspective is required to develop the RMS, which in landscape planning and spatio-temporal recreation mis- should involve the opinions and visions of local communi- management in the study area. Approximately 75 % of the ties, planners and decision makers, business sector and sci- island hotels have been built on the northwest shoreline. entists. Therefore, tourism scholars can effectively contrib- Vehbi and Doratli [20] note that natural beauty, historical ute to tourism management through generating knowledge heritage and traditional urban patterns explain why the and sharing innovative ideas. The gap between theory and northwestern cities (e.g. Kyrenia) are leading tourist desti- practice can be reduced by proposing technical approaches nations in the Mediterranean Basin since the 1930s. Fur- that are developed by application of scientific tools (e.g. thermore, Kyrenia possesses a well-developed infrastruc- GIS). ture, easy access to the airport and capital city and access In fact, the qualities of tourism development are highly to the harbor and shipping lines. These attract investment correlated with land management strategies considering the especially in luxury hotels construction. principals of sustainability, conservation, cooperation, learn- The concentration of tourism activities, regardless of ing, appreciation, responsibility and respect for the re- the carrying capacity issue, is associated with excessive ex- sources. To prepare a scientific framework for a RMS, at- ploitation of natural resources, water pollution and water tributes of different resources can be digitized in a GIS set- scarcity, solid and hazardous wastes, erosion and soil deg- ting to produce materials for spatial analysis that provide radation, air pollution and loss in biodiversity. The conse- graphical guidance for policy makers. Turner recommended quences of this improper Recreation Management System this procedure for landscape planning which acted as an in- (RMS) are combined with a complex set of political, social, terface between social and environmental issues [25]. and cultural issues. For example, as a result of economic
FIGURE 1 - Lack of a proper recreation management system in Northern Cyprus (spatial imbalance in tourism development).
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This research focuses on the nexus between tourism RMS as an implementable approach cannot only mitigate and climatology by using GIS to provide RMS guidelines negative environmental and social impacts of tourism by for the island. Notwithstanding the environmental fragility reducing the density of tourism locations, it also contrib- of the study area, the tourism industry is highly dependent utes to sustainability through diversifying tourism prod- on a favorable climate, which can be mapped as a useful ucts/activities, improvement of infrastructure, and poverty factor in tourism planning. There are two bases for the se- alleviation, especially in remote areas [42] where mass lection of climate as a main resource for RMS: Firstly, cli- tourism has not infiltrated yet. mate is an effective factor in RMS that affects human ac- In addition to the RMS’s environmental advantages tivities and the ecosystem [26-29]. Secondly, climate is fre- through creation of spatio-temporal decentralization, RMS’s quently reported as one of the most valuable natural re- social and environmental implications are also in line with sources in tourism development [30, 31] that affects tourist the TEM [10]. Fisher and Freudenburg [10] argued that decision-making in the selection process of a destination TEM becomes functional as a practical interface between so- [24, 32]. Furthermore, climate does not solely contribute to cial (i.e. cooperation of decision-makers and stakeholders), the suitability of resorts but it also significantly influences and environmental (i.e. RMS as spatio-temporal climate their supplementary natural resources, which can be con- data/map). Correspondingly, Burns [43] proposed TEM as a sidered unique tourist assets as well [33]. model for addressing ecological problems in “tourism mas- Mapping climate changes (using spatio-temporal analy- ter-planning” to justify how ecological problems should be sis) raises awareness of tourists and helps stakeholders to integrated into democratic politics. Belle and Bramwell [44] properly plan tourism development and management [34] by employed TEM to examine tourism managers’ and policy- providing a realistic image of the destination’s weather for makers’ perspectives on climate change in a small island. tourists [35-37]. Incorporation of the RMS in tourism holi- Since there is a comprehensive consensus among research- day package and advertisements will also have implications ers [e.g., 23, 45, 46] about the key role of climate in tour- for tourism marketers. Therefore, tourism planners and the ism, climate data can be a useful tool for tourism manage- business sector need to take notice “as tourists are becoming ment in the context of TEM. more educated and increasingly sophisticated in their con- sumption patterns; and they demand better quality tourism 1.1 Contribution to the current knowledge base products and experiences. [Therefore], travel and tourism This study contributes to tourism climate literature in firms will need to respond with creative marketing strategies several ways: Firstly, there is a paucity of empirical re- based on better quality products and more informative and search on the application of TEM in the tourism climate trustworthy advertising messages” [38, p 87]. nexus [47]. This study fills this gap by developing a model In addition to the advantage of a favorable climate in that functions in the context of TEM on the basis of a land the selection of a tourist destination, it also positively management system performed to integrate socio-ecologi- boosts tourists’ loyalty to revisit a place [30]. Empirical ev- cal issues (tourism and climate) and technology (GIS). The idence from Scandinavia revealed that the relationship be- aim is to mitigate environmental pressures through the spa- tween tourist’s perception and his/her intention to return tio-temporal distribution of tourism activities. Furthermore, was influenced by lack of access to clear information re- it opens a new venue for researchers to share knowledge and garding the weather [36]. Therefore, a tourism climate map cooperation, which are pivotal elements in TEM. In Gid- that illustrates the temporal and spatial variability of the dens’ words, “Ecological modernization implies a partner- climate-related well-being for tourists is one approach that ship in which governments, businesses, moderate environ- can mitigate the probable gap between tourist expectation mentalists, and scientists cooperate in the restructuring of and actual experience. Using the same method, Wan et al., the capitalist political economy along more environmentally linked the spatial variation of water quality and land man- defensible lines” [48, p. 57]. Giddens [48] reference to agement [39]. Hence, a tourism climate map will assist ‘moderate environmentalists’ was meant to be a reference planners and decision-makers to design RMS that not only to proponents of TEM, who are neither market fundamen- controls tourist access to vulnerable natural areas, but also talists (i.e., who believe nature has restorative properties addresses the seasonality of tourist activities. This will also that go well beyond any impact human beings might have facilitates the equitable distribution of tourism expenditure on the environment), nor radical ecologists (i.e. whose ide- throughout the island by dispersing tourists to for instance, ologies are deployed in pursuit of ‘de-industrialization and the hinterland and away from coastal zones which are un- de-marketing) [49]. In a way, the moderate environmental- der stress. Furthermore, it will also decrease local depend- ist attitude, as a proponent of TEM, is a reaction to the anti- ency on the natural resources [40]. modernist views of fundamentalists and radical ecology in Concentration of tourism activities in coastal cities managing the processes of production and consumption (i.e., based on S3 tourism), has led to adverse environmen- without dismantling the market system and opting for mit- tal impacts such as land deterioration due to land use igating environmental impact [49, 50]. change, soil and beach erosion, water scarcity, air, water, Following this, the study is built on Imran et al.’s state- and noise pollutions, degradation of flora and fauna habi- ment [9], who noted that the participation and cooperation tats, and exploitation of fragile natural resources [20, 41]. of stakeholders in the process of tourism development is
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not a straightforward issue. In this respect, Gouldson and [48, 54]. Such a transformation, Giddens noted, is reflected Sullivan revealed that mitigating actions to combat climate by the fact that the “countries most influenced by the idea change can be implemented within the context of TEM of ecological modernization are the cleanest and greenest [51]. In other words, the proposed RMS plan based on spa- of the industrial nations”. In Eder’s [53, p. 210] words, “Its tio-temporal analysis of the Tourism Climate Index (TCI) transformation into a new ideological master frame pro- is an example of stakeholders’ cooperation in the tourism vides the possibility of a way out, legitimating social insti- industry as well as an embodiment of TEM. tutions by means of environment-related ethical frames.” Secondly, to the best of the authors’ knowledge, this re- The focus of this study is to propose RMS based on spatio- search is the first to develop a model built on a knowledge- temporal variations of climate for tourism actions on a based management system, proposed by Ackoff [3], in tour- Mediterranean island in the context of TEM. ism climate literature. The proposed model conforms to the TEM has captured the attention of numerous scholars transformation of “data” into “wisdom” in tourism manage- around the world. More so in Western Europe (i.e., Ger- ment. In other words, meteorological data are used to create many, the Netherlands, the UK, Denmark, and Sweden), the information (TCI value), followed by the production of tour- United States and more recently in newly industrialized ism climate maps through spatial analysis in GIS software. countries such as Malaysia and Thailand where the concept A synthesis of the spatial patterns of TCI over 12 months has been molded into an epistemological/paradigmatic (temporal analysis) contributes to proposing a guideline for framework [54, 55]. Anyhow, “Joseph Huber (from Ger- RMS based on climate attractiveness and other considera- many) should be acknowledged as the father of ecological tions. This guideline is the RMS schedule/calendar. Ac- modernization theory due to his theoretical contributions to cordingly, planners and decision-makers will have access the environment and society from the 1980s onward” [56, p. to sufficient knowledge to manage tourism activities wisely 2]. in regions where climate is one of the most important tour- TEM has also been applied to justify the shifting of in- ism resources. stitutional structures for solid waste management and im- Thirdly, the results of this study will offer useful ap- proving its system in Malaysia [57] and/or reforming the plications for tackling seasonality issues of tourism in planning system in Australia's island-state [58]. Neverthe- Mediterranean regions including innovative tourism mar- less, the paradigmatic structure of TEM is rooted in a process keting by raising tourist awareness, restructuring spatial of production and consumption (i.e., the decoupling or de- and temporal patterns of tourism activities, reducing envi- linking of material flows from economic flows) and institu- ronmental degradation and achieving equitable income dis- tional transformation, especially in the public sphere [53, tribution. This model has the appropriate flexibility to con- 59]. The crux of the theory was initially established when: sider adaptive strategies against climate change. The social dynamics behind these changes that are the Finally, climate is the most important tourism resource emergence of actual environment-induced transformations in Northern Cyprus, which attracts thousands of tourists of institutions and social practices in industrialized socie- from European countries. Surprisingly, no empirical re- ties are encapsulated in the ecological modernization the- search has been conducted on the spatio-temporal variations ory. This theory tries to understand, interpret and concep- of tourism climate on this Mediterranean island. Changes of tualize the nature, extent and dynamics of this transfor- TCI are mapped in Europe by Perch-Nielsen et al. [52] as mation process [59, p. 93]. part of PESETA (Projection of Economic impacts of climate In the case of Northern Cyprus, TEM has not yet been change in Sectors of the European Union based on boT- recognized as a policy framework due to lack of commit- tom-up Analysis). However since the accuracy of the index ment and political will towards establishing political infra- at the continental scale is far too low, only one value is rec- structures essential for the implementation of a TEM orded and projected for the whole island. This means that agenda. One of the pillars of TEM is close cooperation and the TCI map cannot reflect any pattern of variation of cli- consensus among numerous stakeholders [60], especially mate tourism across the island. the involvement of knowledge-based institutions for the purpose of innovation. This aspect of TEM requires struc- tural changes in institutions and social practices [59], 2. THEORETICAL BACKGROUND which is still absent in Northern Cyprus. TEM is highly conducive to integrating new tourism policy towards adap- 2.1. Theory of Ecological Modernization (TEM) tation to climate change. As a backdrop to this study, TEM is initiated as a dis- The main purpose of this research is to provide a cli- course in response to ecological problems. This is because mate-based Recreation Management System (RMS) based sustainable development, notwithstanding its grand goals, on TEM. This objective is the implementation of RMS is perceived as vague and difficult to operationalize [53]. (Figure 2) based on knowledge generation and knowledge However, this is not meant to undermine the credibility of sharing by scientists (i.e. transforming meteorological data the sustainable development paradigm. In fact, it has been to a RMS calendar) embedded in the contribution of all a major force in the transformation of environmentalism stakeholders (action plan in the context of tourism master into the ecological discourse resulting in the birth of TEM plan) in accordance with the concept of TEM.
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Thus, RMS is a loud call to planners and politicians who prus accommodating over 40,000 international students have promoted 3S (Sea, Sand and Sun) tourism solely for (http://www.studyinnorthcyprus.org/?page_id=3652). short term economic benefits without putting any meas- Tourism remains a crucial source of GDP as well as ure/policy in place towards protection of coastal zones and the main source of job creation. The tourism sector em- reduction of environmental impacts in Northern Cyprus [20]. ployed over 12,000 people in the year 2013. The ratio of net tourism income to the trade balance is 39% and the net 2.2. Tourism Climate Index income from tourism reached 616 million $US in the year Two main approaches for the estimation of climate fa- 2013. The number of arrivals reached 1,366.077 in the year vorability for tourism activities are firstly, expert-based in- 2014. In comparison to 30 and 10 years ago, tourist arrivals dices related to the TCI proposed by Mieczkowski [32] and n increased by 2815% and 94%, respectively [68]. The secondly, user-based indices with relation to the Climate main markets for Northern Cyprus’s tourism are: Turkey, Index for Tourism (CIT) developed by De Freitas et al. the UK, Russia, Azerbaijan, Kazakhstan, Syria, Iran and [61]. Furthermore, the Beach-Users Climate Index, pro- Poland. posed by Morgan et al. [62] is based on tourist response. In Regardless of historical, cultural, and growth of edu- the context of the TCI, Mieczkowski weighted five mete- cational tourism in recent years, 3S tourism is still the dom- orological parameters (temperature, relative humidity, pre- inant attraction over the whole island, more so in Northern cipitation, wind speed, and sunshine hours) to estimate a Cyprus (i.e., main destination for coastal tourism in the categorical tourism climate index ranging from unfavora- Mediterranean) [69]. Nevertheless, determinants of 3S ble (-20) to excellent (100) [32]. tourism, also known as ‘beach’ or ‘coastal’ tourism has its The TCI has several drawbacks, such as ignoring non- own complexity including the ‘climate’ variable. In the thermal aspects of weather and climate [63], overriding the meantime, demand for 3S tourism differs according to the effect of precipitation and wind [61] and the lack of empir- type of tourists (i.e. international or domestic). ical validation due to its expert-based approach [52]. Nev- For the case of Northern Cyprus, both domestic and ertheless, it has been frequently employed to estimate cli- international tourists have different expectations from so- mate favorability for tourism activities and to investigate called coastal tourism. Such expectations are highly rele- climate change [23, 45, 52, 64]. Georgopoulou et al. used vant to the climate/ tourism nexus. With the perspectives the TCI to calculate the physical risks of climate change in of climate change and changing relations between climate the banking sector (with the hospitality industry regarded and tourism demand, it is generally agreed that climate as a subsector). They also used the TCI as an indicator of change will affect tourist behavior spatially by shifting it to attractiveness for tourists to change their location in a re- higher altitudes and latitudes [69, 70]. This will have seri- gression model that considered other non-meteorological ous implications for destination such as Northern Cyprus parameters (e.g., infrastructure and economic growth) [65]. which is highly dependent on coastal tourism. Further- more, attributes of coastal tourism have various function- 2.3. Study area alities depending on climate, environment, biodiversity, as Northern Cyprus was an active tourism destination well as susceptibility to anthropogenic pressures [70, 71]. long before the explosion of mass tourism in the 1960s In the case of a Mediterranean Island State, anthropogenic [66]. However, in the aftermath of the Turkish intervention pressure, which is manifested in coastal development and in 1974, the tourism trajectory took a dramatic twist as the cultural activities, demands specific policies/strategies for island was partitioned into the Turkish and Greek enclaves. coastal protection. Nevertheless, both sides are blessed by a suitable climate It is in this context that environmental degradation is that generates calm seas and stable beaches. Long and dry most probable as the pressure of tourism impact is concen- summers with a concentration of precipitation over a few trated on the coastal regions away from the hinterland. months during the mild winter renders perfect conditions Such pressure has been noticed by tourism planners in for so- called 3S tourism [67]. southern Cyprus, which resulted in a moratorium on new Geçitkale and Nicosia are two populated cities inland tourism projects in coastal areas and a shift to the hinter- that are not considered part of the coastal zones (See also land through incentives for rural tourism [72, 73]. Katir- Figure 1). The land use that dominates the island is dense cioglu et al. [73, p. 638] explored the idea that “tourist ar- to sparse forest, sparse forest and brush, cultivated and gar- rivals are a catalyst for energy consumption and therefore den crops and irrigated fields. The main agricultural and climate change in the long term of the Cyprus economy. horticultural productions on the island are wheat, barley, When the Cypriot government sets measurements for envi- vegetable, olive and citrus fruits. Northern Cyprus’s econ- ronmental protection (controlling climate changes), the in- omy is composed mainly of small and medium-size enter- ternational tourism sector should be seriously taken into prises (SMEs) in the tourism and agricultural sector. How- consideration”. ever, over the last two decades policy makers have focused The issue of rural tourism, agro-tourism, and ecotour- on making this part of the island a university hub and re- ism are raised in the context of sustainable tourism which freshing attention to restructuring the tourism sector. Pres- is a dominant concept and concern for the relationship be- ently, there are more than 10 universities in Northern Cy- tween tourism and environment. These forms of tourism
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FIGURE 2 - Climate-based Recreation Management System (RMS).
are categorized under the so called ‘alternative’ tour- and politically. In the case of Northern Cyprus, ecotourism ism as opposed to mainstream/mass tourism criticized for projects have been established in different rural areas and being inconsiderate environmentally, socially, ethically remote villages.
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These projects can be restructured in order to allow illustrates how climate as a unique, free, and non- substi- collective participation of all stakeholders from local pub- tutable resource, can be used in tourism planning (Figure lic sector institutions, business, the commercial sector, 2). It also means that the RMS calendar is a technical as- non-profit organizations and ecotourism entities. Applica- sessment that can be a guideline for the preparation of a tion of these projects will be effective if a collective ap- tourism master plan. In other words, comprehensive tech- proach is practiced. However the effectiveness of manage- nical assessment including evaluation results of socio-eco- ment and stakeholder participation is undermined when nomic, hydrologic, watershed management and geomor- there is either a lack of collective approach or dominant phology should be incorporated in the master planning pro- centralized decision-making. This is not just the case for cess. Hence, outputs of other assessments (e.g. information Northern Cyprus but unfortunately also for most develop- regarding water scarcity, land degradation and soil erosion) ing economies. As Larson and Poudyal [74, p. 924] stated: have been used as inputs/feedbacks for monitoring and “A heavy emphasis on centralized decision-making orches- evaluation steps in the model. Thus, monitoring and evalu- trated by powerful government elites, not governance that ation ensures proposed flexibility in approach. involves a full range of invested individuals and organiza- The application of a knowledge-based management tions, has been a major constraint for developing countries system is an important part of the methodological ap- trying to promote community participation in the tourism proach. According to Ackoff [3] and Reed et al. [75], mak- industry”. ing the right decision in a complex system (e.g. destination management), allows planners/decision makers to use knowledge instead of raw data (Figure 2). As mentioned 3. METHODOLOGY earlier, the proposed model can be implemented in the con- text of TEM [51], which emphasizes knowledge generation 3.1. A descriptive and analytical approach and sharing [6], as well as the cooperation of decision-mak- The research model (Figure 2), illustrates RMS devel- ers and stakeholders [9]. There is a consensus among pro- opment and operationalization processes based on the well- ponents of TEM that researchers are required to produce being of tourism climate – climate being the most im- knowledge and share it with stakeholders [6, 75]. Further- portant tourism resource on the island. Two approaches, more, there is some evidence that postulates the application namely, a knowledge-based management system and the of TEM in the integration of climate change and develop- TEM, are utilized as theoretical framework for designing ment criteria under similar circumstances [51, 57, 58]. Ac- the model. The model also depicts the process of develop- cordingly, meteorological data are inserted into the TCI ment and application of RMS in tourism planning by a flow equation to calculate TCI values for each meteorological chart that begins with calculation and ends with action/im- station (Table 1). Then, TCI values are digitized in a GIS plementation. This process is embedded in the context of setting using Arc GIS software. Spatial analysis of TCI is data calculation and production of wisdom. Both flow pro- performed using geo-statistical techniques to create a TCI cesses correspond to each other, as shown in Figure 2. map. The utility of the TCI value and TCI map with the final An interpolation technique which encompasses meas- production of the RMS calendar, are paramount to the de- ured TCI values is weighted to derive a predicted value for sign of RMS and a tourism master plan. In fact, this model an unmeasured location on the island. Weights are based
TABLE 1 - Location and meteorological parameters of the stations
Meteorological station Geographical location* A (m) Meteorological parameters X Y AT MAT ARH MARH P AWS SH Kyrenia 35.3339 35.3339 170 20.2 24.1 69.2 54.3 470.5 8.1 2.5 Nicosia 35.1948 35.1948 256 19.2 26.3 61.3 35.0 305.8 8.7 3.4 Ercan 35.1506 35.1506 182 19.2 26.3 61.3 35.0 305.8 8.7 3.4 Geçitkale 35.2507 35.2507 97 19.4 26.1 64.2 39.3 328.4 2.7 3.4 Famagusta 35.1332 35.1332 137 19.7 24.9 70.9 50.8 330.2 8.2 2.9 Yeni Erenkoy 35.5164 35.5164 55 19.6 23.8 68.2 50.0 460.5 3.1 2.9 Guzelyurt 35.1833 35.1834 162 20.2 24.1 69.2 54.3 470.5 8.1 2.5 Çamlibel 35.3001 35.3001 138 18.1 22.8 69.2 48.8 443.4 1.4 2.5 Tatlisu 35.3835 35.3835 164 20.0 24.9 68.8 52.1 456.6 6.3 2.7 Lefke 35.1001 35.1001 347 19.5 23.2 69.4 51.3 466.9 8.4 2.6 Note: *: The unit is decimal degrees. Coordinate system for the data source is WGS 1984. A is altitude, AT is average temperature (°C), MAT is maximum average temperature (°C), ARH is average relative humidity (%), MARH is minimum average relative humidity (%), P is precipitation (mm), AWS is average wind speed (m/s), and SH is Sunshine hours. These numbers were obtained by calculation of parameters average for a period of 33 years (1980-2013).
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on the distance between the meteorological stations, the tails continuous monitoring of climate change and patterns prediction locations, and the overall spatial arrangement of tourists who visit the destination. Since the input to the among the measured points. Arc GIS software provides system is meteorological data, the model ought to recalcu- geo-statistical tools to create surfaces from sample data late new data in order to design accurate recreation man- (TCI value of stations) using interpolation methods such as agement calendar. The proposed model covers cross-cul- inverse weighted distance, radial-based functions, global tural properties of tourism climate by monitoring the quan- and local polynomials, kriging, co-kriging, and isotropical tity and cultural origin of visitors. For instance, an optimal or anisotropical models. Kriging is known as the best linear climate is perceived differently by Russian compared to unbiased estimator amongst all interpolation methods. It is Chinese tourists. As observed by the authors, the swim- also a popular geostatistical method in the interpolation of ming season for Russian tourists in Northern Cyprus al- natural elements [76]. It provides an easy method for char- ready begins in March whereas the locals will wait until acterizing the variance or the precision of predictions. This early July. Therefore, the RMS should be modified accord- method uses variograms to express the spatial variation and ing to changes in visitor pattern (geographic segment) [83] minimizes the error of predicted values which are esti- to avoid mismatch between the expectation and experience mated by spatial distribution of the predicted TCI values. of a destination with respect to climate change. As noted by Olya and Alipour [77] the Kriging technique Nevertheless, the use of long term data ensures reliabil- functions adequately for the interpolation of tourism cli- ity of the results of the proposed model. Three factors can be mate risk caused by excessive precipitation as a destructive considered in the recalculation of the proposed model: factor related to the well-being of tourists. Therefore it is firstly, changing climatic parameters, secondly changing applied as geostatistical technique for the interpolation of tourist pattern in terms of cross-cultural characteristics and TCI value in this study. The process of TCI calculation and thirdly, as elaborated earlier, the implementation of a RMS interpolation are conducted for 12 months (January to De- as an important principle for the Tourism Master Plan. Since cember). Hence, a temporal analysis of TCI is produced Master Plans are normally revised only every five years, it is based on the interpretation of monthly TCI maps. suggested that the TCI should be recalculated for all stations As climate is one of the most important resources for and the RMS calendar be prepared every five years consid- tourism development on the island a RMS pattern based on ering all changes in parameters. spatio-temporal variations of TCI is developed. Notwith- More details about data and the procedure are elabo- standing the existence of sophisticated approaches such as rated in the following sections. the Recreational Opportunity Spectrum [78] and Limits of Acceptable Change [79] the proposed model has great po- 3.2. Data collection and analysis tential to be used in regions where climate is the major Data was collected from the meteorological stations of tourism resource. Northern Cyprus in the Mediterranean, covering a data Based on the results, the five practical managerial im- base from 1980 to 2013. The data was obtained after a plications are as follows: lengthy discussion with the head of the meteorological or- Overcoming the seasonality problem ganization in Lefkosa, the capital city of Northern Cyprus. Equity in distribution of the economic benefits Permission to use the data was granted after the purpose of the research was explained. Northern Cyprus is located in Reduction of pressure on the ecosystems the Eastern flank of the Mediterranean Sea, whose climatic Proposing an adaptive approach to climate change characteristics include warm and dry summers, mild win- Tourism marketing ters, more than 3,300 hours of annual sunshine and light-to- moderate winds. Such climatic characteristics create typical The aforementioned process is in line with Wong et al. 3S tourism that attracts a large number of tourists. Sun, Sea who emphasize the importance of evaluating the policy en- and Sand (3S) tourism is the main product that motivates vironment for climate change adaptation in tourism, as well mass tourism. In this context tourists seek a warm climate to as continuous monitoring imbedded in the proposed model indulge in sun tanning on sizzling beaches [84, 85]. This [80]. For evaluation and monitoring of adaptive coastal form of tourism has been dominant since the 1950s and management Jacobson et al. [81, p. 51] stated that ‘moni- 1960s with strong seasonal concentration, environmental de- toring involves activities that measure the effectiveness of struction and unsustainable development along the coastal actions, whereas evaluating involves the interpretation of areas of the Mediterranean but also in the Adriatic, Aegean, that information’. Caribbean, and many islands in the Pacific, to name a few Furthermore, updating the outputs of the model with [86, 87]. Now-a-days, with the support of its international new meteorological datasets enables destination communi- institutional agencies the tourism industry has embraced a ties to better adapt to forthcoming climate change. At the moral agenda to redirect tourism planners and developers same time this is a practical approach for moving the tour- towards a so called ‘New tourism’ as an alternative to 3S ism-and-climate-change nexus from a descriptive problem- tourism (mass tourism). “Whereas Mass Tourism might be based issue toward a more affirmative, action-oriented dis- characterized by sameness, crudeness, destructiveness, and course [82]. The functionality of the proposed model en- modernity, New Moral Tourism can be viewed as difference
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and sensitivity as well as constructive and critical of moder- with the TEM theory (Figure 2). With this regard, GIS is nity. There are several categories of travel that might be de- used as an integration technology to project spatial and scribed as New Moral Tourism, including ecotourism, com- temporal variations of TCI [1]. Kriging is used as a popular munity tourism, agro-tourism, and sustainable tourism” geostatistical method in the interpolation of natural ele- [as cited in 85]. ments [76, 77]. A spatial analysis of TCI is performed for 3S tourism is also subject coastal areas to overdevel- each month to enable temporal analysis of the variations of opment. Though economically lucrative in the short run the TCI on the island. About 12 TCI maps illustrate the some destinations, especially Mediterranean islands have temporal and spatial attractiveness/appropriateness of cli- realized that the long-term viability of this dominant mode mate for tourism (i.e. by month and location) (Figure 3). of tourism has engendered a malaise in the tourism system. Based on this rational justification, the proposed RMS cal- For instance, in Northern Cyprus, mass tourism has not endar can be applied as a scientific guideline in destination only resulted in degradation of the coastal environment, it planning and management. has also subjected the island to a seasonality syndrome. “Hence, the image of Cyprus as a sun-lust destination can be deemed a critical cause of seasonality. This is also true 4. RESULTS about Malta, which is struggling with extreme seasonality” [87, p. 704-711]. TCI values varied ranging from 41 (acceptable) in Yeni Erenkoy to 98 (excellent) in Çamlibel and Kyrenia. Yeni 3.1.1. Tourism climate index Erenkoy is a flat shoreline characterized by dense to sparse forest. Land use mainly consists of bush which is also the The TCI index is formulated based on the monthly habitat of famous Cyprus donkey. Çamlibel has a special to- means of seven meteorological factors: maximum daily pography that varies from coastal (altitude=0 m) to moun- temperature, mean daily temperature, minimum daily rela- tainous (altitude >1000 m). Similar to the Yeni Erenkoy and tive humidity, daily relative humidity, precipitation, daily Karpaz region, this area is also covered with dense to sparse duration of sunshine, and wind speed. To measure thermal forest and bush. This area also has cultivated fields and gar- comfort, Mieczkowski developed two sub-indices named den crops. Kyrenia and Çamlibel have similar topograph- CID (Daytime Comfort Index – which combines the varia- ical and geomorphological conditions. Apart from forest bles of maximum daily temperature and minimum daily and cultivated fields and garden crops, irrigated farms are relative humidity) and CIA (Daily Comfort Index – which also a dominant land use. The average TCI values for 12- combines the variables of mean daily temperature and daily months ranges from 70 (very good) in Nicosia to 85 (ex- relative humidity) [32]. Accordingly, coefficients of each cellent) in Kyrenia, except for Yeni Erenkoy which is 56. variable were weighted, and the formula (1) was used as an Other districts have “very good” (70-80) climate condi- index to estimate the level of climatic well-being for tour- tions for tourism activities. During spring and fall, higher ists: TCI scores are found throughout the island. Similar find- 8 2 4 4 2 (1) ings are reported in Amelung and Viner’s [23], Deniz’s [45], and Perch-Nielsen et al.’s [52] studies, which re- Where CID and CIA are two sub-indices represented vealed that variations in average TCI scores present a bi- as a daytime comfort index; R is precipitation; S is hours modal distribution in the Mediterranean regions. This means of sunshine per day; and W is wind speed. The details of that climate in the spring and fall is more suitable for the the variable rating system for the calculation of the TCI are well-being of tourists than that in the winter and summer. described in Mieczkowski’s paper [32]. The TCI scores clas- Researchers have recognized that increasing temperatures sified in five mapping categories are outlined in Table 2. caused by climate change will result in a decreased climate
TABLE 2 - A classification scheme for mapping the TCI quality in the Mediterranean during the summer. Perch- Nielsen et al. [52] in their comparison of climate change No. Numerical value of TCI Mapping Category for a period of 110 years found that the number of accepta- ble (TCI>40), good (TCI>60), and excellent (TCI>80) 1 80-100 Excellent days in summer for Mediterranean region will decrease by 2 70-79 Very good 7, 10, and 8 days, respectively. 3 60-69 Good 4.1. Tourism climate index map 4 40-59 Acceptable According to the calculated TCI score for January (Fig- 5 -20-39 Unfavorable ure 3a), the climate is good (60-70) for tourism. The climates Source: [23]. of Nicosia, Lefke and Guzelyurt are more favourable (68, 65, and 64, respectively) for tourists during this month (see 3.1.2. Recreation management system Table 2 for score classification). In Çamlibel the TCI scores In the knowledge-based management system developed remain acceptable (59) (Figure 3b). In February Nicosia has by Ackoff [3], techniques and technology help to transform a very good index (72). Eastern and south-western areas data into information and knowledge, which is consistent have good climates for tourist’ well-being. During the month
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of March, a “very good” climate (70-80) prevails over the The diversity of climate well-being for leisure in- whole island (Figure 3c). However, the climate is more suit- creases in July when the TCI score fluctuates between 53 able in the eastern parts of the island. (acceptable) in Nicosia to 80 (excellent) in Çamlibel (Fig- During the winter there are no excellent climate scores ure 3g). Famagusta (70), Guzelyurt (69), have “good” cli- (80-100) on the island, with the exception of Geçitkale in mates in June, while Lefke (77), Yeni Erenkoy (78) and March. Geçitkale is a flat inland area (altitude= 100-300 Kyrenia (74) have “very good” climates. Variations of TCI m) that is mainly covered by irrigated and dryland farms. scores in August are similar to July, demonstrating “ac- ceptable” and “good” conditions in the central, south eastern The TCI values improve to an excellent category in and south-western areas. Meanwhile, Çamlibel (80) and April and May (Figure 3d and 3e), with the exception of Yeni Erenkoy (68) are climatically “very good” for tourists Çamlibel in April (73). The TCI reaches the highest score (Figure 3h). In the last month of summer, all areas have a (≥90) in Yeni Erenkoy and Çamlibel in May (Figure 3e). TCI rating below 75 except for Çamlibel (80) and Geçitkale The TCI scores drop from “excellent” to “very good” (75). In September, Çamlibel and Geçitkale have “excellent” during the first two months of spring due to increasing tem- and “very good” climates, respectively (Figure 3i). perature. However, Çamlibel has an excellent (84) climate for recreation purposes in June (Figure 3f).
FIGURE 3 - Map of spatio-temporal variations of the tourism climate index over the 12 months of the year in Northern Cyprus (a-m)
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Compared to the summer, TCI values are high across Tourism businesses can arrange exhibitions, festivals and most of the island in the fall. The TCI values for Çamlibel, events to accompany the other functions of the capital city Lefke, Yeni Erenkoy, and Geçitkale are greater than 85, (e.g. trade, politics). Coincidentally, Nicosia has a “very which represent an “excellent” climate in October (Figure good” to “excellent” climate for tourism during this period 3k). Lefke is a coastal area with irrigated and cultivated (Figure 4). An International theatre festival and similar social fields and garden crops including wheat, vegetables, pota- events are held in Nicosia in September. These events could toes, and citrus fruits. In November, the central regions pre- be shifted to October when the climate is “excellent” (81). sent excellent (>80) climates for tourism activities. Never- Geçitkale is a destination with an excellent climate in theless, other regions still have “very good” climates dur- February, March, April, September, and November. It has ing this month (Figure 3l). It can be clearly seen that the a remarkable potential for the development of rural tour- tourism climates in Famagusta (81) and Nicosia (80) are ism, agro-tourism, ecotourism, and other types of alterna- excellent in December (Figure 3m). Famagusta is a coastal tive tourism. For instance, the Hellim festival is a well- area with a low altitude (<200 m) composed of irrigated known food festival that is organized in Geçitkale in April. farms and garden crops. Nicosia is an inland area whose Tourist flow to Yeni Erenkoy and Karpaz where the sandy altitude varies between 100 to 300 m with a dominant land beaches and panoramic views are abundant could be en- use of cultivation of agriculture and garden products. couraged during the months of March, May, July and Au- gust. The Karpaz region is also home to the flower festivals 4.2. Recreation management system calendar (e.g. Medoş and Lale Festivalsi) and presentation of local Results of the spatio-temporal pattern of TCI reveal handicrafts in March. that there are viable opportunities to reorganize tourism ac- Iskle can be visited by tourists in August and Septem- tivities in the northwest part of the island, where two-thirds ber when regional and national food festivals (e.g. Yeni of the hotels are located (Figure 1). A Pearson correlation Boğaziçi Pulya Festivali) are held to revive the local food test (an inferential statistical analysis) was performed to in- culture. Birdwatching is also popular in Iskele during the vestigate the correlation between number of the hotels and months of March and April. favorability level of the climate for tourism activities. Ac- cording to the statistical results, the number of hotels does Kyrenia attracts tourists who enjoy the pleasant not correlate significantly to the level of TCI (r=-262, weather, social events, historical attractions, and trekking P>.05). Such results confirm that climate is ignored in rec- during the months of March, April, June, and July. Kyrenia reation management despite it being the most important is also home to Alagadi Beach where the unique kinds of tourism resource in the study area. Hamilton et al. [46], turtles (green and the loggerhead turtles) have their nesting Amelung and Viner [23], and Saarinen [31], observed that grounds. This is an attraction for nature lovers during the climate as a key determinant of tourist demand, especially months of June and July. Kyrenia is also venue for a social in the Mediterranean islands, can be considered just as im- event called Lepta Orchid, which takes place during the portant a factor in spatial and temporal tourism planning. month of March, and home to several museums as well as Besides the importance of the climate factor for tourists, an ancient shipwreck. the need to propose adaptive strategies for climate change Famagusta is a coastal city and home to a major uni- and seasonality issues provides a rational incentive to re- versity. Over time it has suffered anthropogenic pressure distribute tourism activities (through RMS) based on the due to overdevelopment and poor infrastructural amenities. favorability of climate in terms of spatial and temporal It has access to several beaches that are crowded by domes- changes. This logic is supported by Ma et al. [26] and Li et tic and international tourists between April and October. It al. [27], who draw attention to the inevitable role of climate is also a venue for numerous social events e.g. Children’s in RMS. Hence, based on spatio-temporal changes in the festival, Sand sculpting, musicals and spring festivals or- TCI, the RMS calendar is developed to determine the loca- ganized by the municipality and university. tion and duration of tourism activities on the island (Figure 4). However, environmental and social concerns/potentials Çamlibel is a mountainous landscape that provides a must be considered within the development procedure of favorable climate during the hot season (May to October) the RMS calendar with respect to the principles of TEM. as shown in Figure 4. Apart from its climate well-being For example, based on the spatio-temporal patterns of TCI, benefits, it hosts an exhibition in June which provides an January and December are suitable months in Guzelyurt opportunity for local people to sell their organic products and Lefke while simultaneously coincide with the citrus as well as handicrafts. It is also venue for strawberry festi- fruit harvesting season. Such coincidences enhance and in- val during the harvest. fluence visitors’ perceptions and experiences. In addition, According to the TCI results, high temperature and rel- the orchid flower festival held in Guzelyurt and Lefke every ative humidity negatively affects the suitability of the cli- February (named Orkide festivali) is another attractivity in mate for tourism activities. Based on climate change sce- this region. Surfing is also a popular activity in the Guzelyurt narios, estimated temperature will rise in the Mediterra- and Lefke coastal area in October due to optimal wind con- nean regions with negative impacts expected on tourist ditions. Nicosia, capital of the island, mainly attracts tourists comfort. Currently, spring and autumn are the most appre- in January, February, November and December (Figure 4). ciated seasons for travel to the island. Nevertheless, it is
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FIGURE 4 - Calendar of tourism activities based on the recreation management system (RMS) in Northern Cyprus.
expected that the favorability of the climate will decline in prevent irreparable effects and carry out spatio-temporal early fall and late spring due to climate change. In contrast, analyses to reduce environmental degradation. As such, the winter could become a suitable future season for tourists. practicality of RMS based on spatial analysis in a GIS set- Furthermore, the period of the 3S tourism will expand due ting was demonstrated by Turner [25] and Wan et al. [39]. to temperature increase in the study area. Thus, climate One should keep in mind that the successful implementa- change might generate new opportunities on the island by tion of such a project relies on the participation of research- attracting tourists in spring instead of summer. ers, decision-makers, stakeholders, and local communities. According to the proposed model, stakeholders have var- Contributions and achievements of the project should meet ious options in terms of time and space to develop and rear- their satisfaction [9, 27]. In other words, a package and ad- range their business. This will also allow decision-makers to vertisement calendar as a part of Master Plan requires the apply knowledge-based approaches to their planning pro- participation of all stakeholders. For instance, an adjust- cesses. Therefore, this model meets the criteria of TEM [7, ment of the date and place of social events/festivals that do 58, 59]. Moreover, this model follows the funneling process not match the results of RMS in terms of climate suitability from “data to wisdom” that was suggested by Ackoff [3]. requires cooperation of local communities and business sectors. Similarly, deconcentration and diffusion of tour- 4.3. Policy implications ism activities from one location to another would require 4.3.1. Reducing environmental degradation the development of certain types of rules to accelerate the The proposed model provides a strategy for tourism process. decision makers to adapt tourism activities to the capacity and tolerance of the resources that are available in Northern 4.3.2. Seasonality and tourism marketing Cyprus. Redistribution of tourism activities based on the The results of seasonal variation of TCI on the island RMS calendar would not only decrease human pressures are depicted in Figure 5. As illustrated, the island has a (social carrying capacity) but also reduce the negative im- “very good” (70-79) to “excellent” (80-100) climate during pact of tourism on the environment. Based on the RMS cal- all seasons. Nevertheless, spring is the best season in terms endar, each area is matched by tourist demand in terms of of favorability of the climate with all stations reporting an climatic attractivity/ favorability. According to Perkins et “excellent” climate (>80) with the exception of Ercan (79). al. [6] and Cantasano and Pellicone [18], planners should In autumn, Nicosia (83), Lefke (80), and Geçıtkale (80) be sensitive to proper recreation management systems to have an “excellent” climate whilst other areas achieve a
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100 Win. Spr. Sum. Aut.
80
60
40
20
0 Ercan Nicosia Geçıtkale Çamlibel Guzelyurt Tatlisu Famagusta Yeni Lefke Kyrenia Erenkoy Seasonal TCI
FIGURE 5 - Seasonal variation of TCI in the study area
“very good” climate (>70). During the winter, the tourism fusing tourism activities and their benefits temporally and climate in Niocsia (72) and Geçıtkale (70) is “very good”. spatially throughout the island. One should bear in mind Other areas present a “good” climate for tourist activities that the concentration of tourism in Kyrenia is accompa- (>70). Nicosia (57), Guzelyurt (58), and Famagusta (57) nied by luxury hotels and casinos which are prone to for- offer an “acceptable” climate for tourist well-being during eign investment, unsustainable modes of imports as well as the summer. However, other destinations present a “good” leakages [83]. In the context of TEM [48], the proposed climate for tourists (>60). According to these results, there model paves the way for innovative ideas that will not only is no unfavorable climate (<40) in Northern Cyprus and tour- contribute to environmental quality but also benefit remote ists can chose locations with “excellent” and “very good” cli- areas. As described earlier, most of the remote areas have mate during all seasons. As for the findings of Amelung and potentials for alternative forms of tourism without requir- Viner [23], TCI values undergo possible shifts in climatic fa- ing high capital or foreign investment. vorability for various areas during different seasons. The is- sue here is how the RMS calendar can address the spatial 4.3.4. Adaptive strategy to climate change and temporal distribution of touristic activities throughout In the proposed model, evaluating and monitoring are the island and respective districts. embedded as the key processes that enable the RMS calen- More importantly, advertising and marketing the cli- dar to adapt to climate change. Feeding the model with new matic attractiveness of all the districts during different sea- meteorological datasets will produce updated information sons is a significant attribute for boosting the destination that can be translated into knowledge for policy-makers to image [22]. Implementation of the RMS calendar, which is apply a new strategy to climate change. It will also allow positively linked to tourist loyalty will enable tourism de- monitoring of activities that measure the effectiveness of cision makers to minimize the gap between the expecta- actions [75, 79]. As most of the climate change patterns tions and experiences of tourists [30, 36]. The proposed estimate temperature increase [88], especially in the Med- model, which functions based on variations in the suitabil- iterranean region, TCI will vary in the near future. Hot ity of the climate for tourists, addresses seasonality issues. summer weather in Northern Cyprus will adversely affect This claim matches Ridderstaat et al.’s study [24] who re- the favorability of climate for tourists. Increasing tempera- iterated that seasonality is strongly associated with climate tures will be exacerbated by high relative humidity on the attractiveness. Therefore, the current study tackles this island, thereby decreasing tourist comfort. The situation is problem by proposing the RMS calendar that identifies likely to escalate in inland areas (e.g. Nicosia) where there which district renders the most favorable climate. The re- are no resources for 3S tourism. sult of this study is a practical example of the application This model considers Jacobson et al. [81] and Reed et of TEM in tourism climate literature, which coherent with al.’s [75] advice that it is not adequate to apply only adap- Belle and Bramwell’s study [44]. tive management. Information, monitoring, evaluation and knowledge must also inform future decisions in coastal ar- 4.3.3. Distribution of tourism benefits (social achievement) eas. Similarly, Wong et al. [82] emphasized the importance The concentration of tourism enterprises in Kyrenia re- of monitoring and evaluating steps in adjusting environmen- gion is not necessarily a benefit for the rest of the island. tal policies to climate change adaptation. The multicultural As Eagles [40] and Truong et al. [22] reported, this type of differentiation of tourism climate, which is often overlooked development tends to ignore the pro-poor role of tourism in [61, 89], enhances the practicality of the RMS calendar. In poverty alleviation. This research, by proposing a knowledge- other words, tourists who come from various places express based management system [3], provides a guideline for dif- different perceptions/sensations about climate. Hence, the
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proposed model helps managers to match those sensations communities) to translate the spatio-temporal patterns of with the RMS calendar, which is extracted from the spatio- TCI into practical strategies. Ultimately, this study attempts temporal patterns of the TCI. to strengthen the link between theory and practice by de- veloping the RMS calendar, which is clearer and more un- derstandable for non- experts in this field. 5. CONCLUSION 5.1. Limitations and future research directions This study develops a RMS model based on climatic There are four limitations that can be considered for attractiveness for tourism on the Mediterranean island of future research. Northern Cyprus. The districts that do not have a favorable Firstly, this research proposes a model that is triggered climate are not considered as destinations in RMS calen- by meteorological data. It would be beneficial if data were dar. In contrast, tourism activities are distributed to regions available for the entire island of Cyprus. This study focuses where a more favorable climate exists during a specific pe- on Northern Cyprus. The RMS process needs to examine riod of time. Another functionality of the RMS calendar is the system as a whole by scrutinizing the effective dimen- its consideration of environmental and human pressures in sions of sustainable tourism development pertinent to po- the context of recreational management. Furthermore, tour- litical, socio-cultural, environmental, technological, and ism specific resources (i.e. climate and environment) have economic dimensions. In future studies, it is recommenda- been considered for developing RMS calendar. This ap- ble to apply the proposed model in more sophisticated ap- proach not only reduces negative environmental and social proaches within tourism recreation management, such as impacts that are caused by over-concentration of touristic the Recreational Opportunity Spectrum and the Limits of activities in one location. It also addresses the suitability Acceptable Change. Furthermore, it is worth examining the of numerous locations with a favorable climate for redistri- extent to which these sophisticated approaches are compat- bution of tourists over time and space in view of improved ible with TEM in tourism recreation management. marketing as well as for solving the seasonality problem. Secondly, the RMS calendar was based on a lengthy The proposed approach is supported by the TEM the- data set (1980-2013); however, a comparison of TCI maps ory through generating and sharing knowledge, using tech- derived from two long term scenarios would provide more nology as well as considering the cooperation of policy accurate information for developing the RMS calendar. makers, stakeholders and other contributors. In other Thirdly, from a technical point of view and consonant words, this model integrates environmental concerns with with other studies, the Kriging method was used for the spa- economic benefits by utilizing GIS as an interface between tial analysis of TCI values. However future studies should environmental and social issues. Spatio-temporal patterns identify appropriate geostatistical approaches for the inter- of TCI are considered as the base for proposing the RMS polation of TCI values amongst the meteorological stations. calendar that follows both environmental (i.e. reducing hu- Fourthly, due to the popularity and capabilities of TCI man pressure on the environment, reducing excessive ex- in the projection of tourism climate variations, it is used to ploitation of natural resources and managing biodiversity) perform spatio-temporal analysis in the study area. It is sug- and socio-economic concerns. These objectives can be gested to devise a mechanism in future that is capable of ex- achieved by valorizing different locations, adapting re- tracting and applying tourists’ attitudes about climate condi- sources to demand, innovating marketing strategies based tions. In other words, instead of expert-based indices such as on favorability of the climate, raising tourist awareness, de- TCI, it would perhaps be beneficial to cover demand spec- veloping alternative forms of tourism and avoiding the pro- trum considerations using CIT or fuzzy-based CIT. poor role of tourism by enhancing the livelihood of local communities. Furthermore, assessment of the multicultural charac- teristics of tourism climate and risk assessment of negative The process of the transformation of data to knowledge climate impacts on tourism activities are two areas of fu- and wisdom is based on a knowledge-based management sys- ture research that could improve the functionality of the tem that recommends decision making built on knowledge in- current model. stead of raw data. In fact, the RMS calendar provides practical knowledge for managers on how to manage recreation ac- The authors have declared no conflict of interest. tivities over time and space. 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[76] Park, N. W., and Jang, D. H. (2014). Comparison of Geosta- tistical Kriging Algorithms for Intertidal Surface Sediment Fa- cies Mapping with Grain Size Data. The Scientific World Jour- nal, DOI: 10.1155/2014/145824.
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[86] Trias, M. C., Molina, D. J., Santacreu, D. A., and Rosselló, J. G. (2014). Enhancing “Places” Through Archaeological Her- itage in Sun, Sand, and Sea Touristic Coastal Areas: A Case Study From Mallorca (Spain). The Journal of Island and Coastal Archaeology, 9(3), 341-363. [87] Koutra, C., and Karyopouli, S. (2013). Cyprus' image—a sun and sea destination—as a detrimental factor to seasonal fluc- tuations. Exploration into motivational factors for holidaying in Cyprus. Journal of Travel & Tourism Marketing, 30(7), 700-714. Received: October 16, 2014 [88] Meehl GA, Stocker TF, Collins WD, Friedlingstein P, Gaye Revised: April 13, 2015, July 15, 2015 AT, Gregory JM, Kitoh A, Knutti R, Murphy JM, Noda A, Accepted: September 16, 2015 Raper SCB, Watterson IG, Weaver AJ, Zong-Ci Z (2007) Global climate projections. In: Solomon S, Qin D, Manning M, Chen Z, Marquis M, Averyt KB, Tignor M, Miller HL (eds) Climate change 2007: the physical science basis. Contribution CORRESPONDING AUTHOR of working group I to the Forth Assessment Report of the In- tergovernmental Panel on Climate Change. Cambridge Uni- versity Press, Cambridge, pp 747–845. Hossein G.T. Olya Faculty of Tourism [89] Olya H., and Alipour H. (2015). Modelling Tourism Climate Indices through Fuzzy Logic, Climate Research. 66(1), 49-63. British University of Nicosia Via Mersin 10 Girne/KKTC, 80 TURKEY
E-mail: [email protected] Webpage: www.olyah.com
FEB/ Vol 24/ No 11c/ 2015 – pages 4125 - 4141
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QUANTIFICATION OF THE HEALTH EFFECTS OF
EXPOSURE TO AIR POLLUTION (NO2) IN TABRIZ, IRAN
Mohammad Ghanbari Ghozikali1,2, Alessandro Borgini3, Andrea Tittarelli3, Abdeltif Amrane4, Kazem Naddafi5,6, Mahmoud Mohammadyan7, Gholamreza Goudarzi8, Roberto Bono9 and Behzad Heibati7,*
1Environmental Health Engineering, Department of Environmental Health Engineering, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran 2 Department of Environmental Health Engineering, School of Public Health and Center for Air Pollution Research (CAPR), Institute for Environmental Research (IER), Tehran University of Medical Sciences, Tehran, Iran 3 Fondazione IRCCS Istituto Nazionale dei Tumori, Environmental Epidemiology and Cancer Registry Unit, Via Venezian 1, Milan, Italy 4 Ecole NationaleSupérieure de Chimie de Rennes, Université de Rennes 1, CNRS, UMR 6226, Rennes, Avenue du Général Leclerc, CS 50837, 35708 Rennes Cedex 7, France 5Department of Environmental Health Engineering, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran 6Center for Air Pollution Research (CAPR), Institute for Environmental Research (IER), Tehran University of Medical Sciences, Tehran, Iran 7Health Sciences Research Center, Faculty of Health, Mazandaran University of Medical Sciences, Sari, Iran 8Ahvaz Jundishapur University of Medical Sciences, Department of Environmental Health Engineering, Health Faculty, Ahvaz, Iran 9Department of Public Health and Pediatrics University of Torino, Torino, Italy
ABSTRACT tion of nitrogen oxide (NO) by atmospheric oxidants such as O3. Regarding atmospheric pollution, released NO is Epidemiological studies show that long-term exposure rapidly oxidized by ozone and then converted to NO2. The to NO2/NOx is associated with an increased risk of cancer. toxicity of NO2 is higher than that of NO [1]. New contam- The mortality and morbidity rates due to NO2 in the Tabriz inants can be formed from photochemical reactions when city was determined considering the relative risk and base- organic aerosol and nitrate aerosols sulfate are used; they line incidence provided by the World Health Organization contributed significantly in PM10 and PM2.5 aerosols and in (WHO) / European Center for Environment Health, Bilt- their risk properties [2]. Human activities represent the main hoven Division using the AirQ2.2.3 software to analyze the sources of NO2, from automobile exhaust emissions to sta- data. The results of this study showed that 0.85% (CI 95% tionary sources such as fossil fuels, power plants, industrial 0.36 - 0.96) of acute myocardial infarction, 0.47% (CI 95% boilers, waste incinerators and heating household appli- 0 - 0.94) of cardiovascular mortality and 9.28% (CI 95% ances; but the main source of nitrogen dioxide in the urban 0.1 - 2.19) of hospital admissions for chronic obstructive area is the gas emitted by the public transports. NO2 con- pulmonary disease (COPD) can be attributed to NO2 con- centration varies from morning to night. The main sources 3 centrations over 10 μg/m . According to this model, cases of NO2 in indoor environment are the natural gas cookers of cardiovascular deaths and acute myocardial infarction and the smoke produced by cigarettes. Acute short-term (one caused by NO2 in Tabriz were about 0.23 and 0.10% of the hour) effects and low concentrations have been observed in total mortalities, respectively. In addition, about 0.9% of animals [3-5]. Observational data are derived from outdoors the total hospital admissions related to chronic obstructive studies where NO2 is one component among a complex mix- pulmonary disease (HA COPD) was caused by NO2. ture of different pollutants found in ambient air, and also Therefore, due to this pollution, preventive measures and from studies of NO2 indoors exposure where its sources in- effective strategies for implementing policies related to re- clude unvented combustion appliances. Epidemiological duce air pollution should be undertaken in Tabriz city. studies over the past decade in Europe and worldwide showed increased mortality and illness associated with NO2 exposure [6-9]; it was also estimated that 5% – 7% of lung KEYWORDS: cancers in non-smokers and ex-smokers could be attributed Quantification, Air pollution, NO2, AirQ2.2.3 software. to exposure to high levels of air pollution, including NO2, or vicinity to heavy-traffic roads [10]. Large meta-analyses of 1. INTRODUCTION studies on the short-term health effects of NO2 indicate a positive association between daily increases of NO2 and Nitrogen dioxide (NO2) is a gas with oxidant proper- natural, cardiovascular, and respiratory mortality [11, 12]. ties capable of contaminating ambient air in many urban NO2 could be a more relevant health-based exposure indica- and industrial contexts, as well as indoor air of homes with tor than PM [13, 14]. Undesirable effects of NO2 on the chil- combustion appliances. NO2 is mainly derived from oxida- dren health in urban areas with high concentrations of NO2 have been identified [15-18]. An annual average for Air 3 * Corresponding author quality guideline values for NO2 gas is 40 μg/m , which is
4142 © by PSP Volume 24 – No 11c. 2015 Fresenius Environmental Bulletin
lower than the toxicological threshold and represents a monitored stations were Abrasan (residential-commercial strong confirmation of the negative effects of outdoor area in the eastern part of the city), Baqshomal (the down- NO2.Owing to the continuous increase of health effects due town of the city), Health center (residential area in the to air pollution, European governments and the World northern part of the city), Hakim-nezami (residential area Health Organization as well as other groups and associa- in the southern part of the city), Rah-ahan (Commercial In- tions, using the data collected during various studies have dustrial area in the western part of the city) and Raste- begun shaping method and environmental policy to assess kouche (commercial area in the downtown). In Abrasan the effects of air pollution on public health [19, 20]. Tabriz station the HORIBA model AP360 was used, while the En- city is located in northwest of Iran in East Azerbaijan prov- viron tech model M200 was used in the other stations; these ince between Eynali and Sahand mountains in a fertile area systems continuously monitored atmospheric NO2 concen- in shores of Aji River and Ghuri River. It has a semi-arid trations and involved a cross-flow modulated semi decom- climate with regular seasons. With a population of over one pression chemiluminescence method. Raw data, such as and a half million peoples, it is the fourth most populous daily average concentrations of NO2, has been saved in a city in Iran after Tehran, Mashhad and Isfahan. Tabriz is Microsoft Office Excel spread sheet and all processing considered the second largest contaminated city after Teh- mechanisms for conversion between volumetric and grav- ran due to the presence of many large industrial plants and imetric units and etc, such as correction of temperature and the high number of cars. Modern industries in this city in- pressure, coding, averaging and filtering were performed. clude the manufacturing of machinery, vehicles, chemicals Annual and seasonal average, maximum and annual 98th and petrochemical materials, refinery, cement, electrical percentiles of NO2 were obtained and concentrations were and electronic equipment, home appliances, textiles and divided into 10 μg/m3 categories. Then, the association be- leather, nutrition and dairy factories and woodcraft [21]. tween daily average concentrations of NO2 and health im- Acute myocardial infarction is a common disease with se- pacts was estimated for one year by using AirQ2.2.3 Soft- rious consequences in mortality, morbidity, and cost to the ware. society. Coronary atherosclerosis plays a pivotal part as the underlying substrate in many patients [22]. A link between 2.2 Statistical analysis ambient air pollution and cardiovascular disease, in partic- The approach proposed by the World Health Organi- ular myocardial infarction, has been suggested by a number zation (WHO) was applied using the AirQ2.2.3 software de- of epidemiological studies and has attracted substantial at- veloped by the WHO European Centre for Environment tention within the cardiological community, as attested by and Health, Bilthoven Division. This model is used to esti- the recent American Heart Association scientific statement mate the impacts of exposure to specific Air pollutants on air pollution and cardiovascular disease [23]. There is (confounders in any case) on the health of people living in convincing evidence of a relation between overall and car- a given period and area.This software consists of two quan- diovascular mortality and the degree of air pollution on the tification models and lifetime tables. Quantification model previous day [24]. Chronic obstructive pulmonary disease was used in this study, which was used to determine the (COPD) is a chronic inflammatory airway disease that is Attributable Proportion (AP), the Relative Risk (RR) and described by hyper secretion, cough and inflammatory cell the Based Incidence (BI) used for each health conse- influx. COPD is a major cause of morbidity and mortality quences. Relative risks (RR) with 95% confidence interval worldwide and continues to cause a heavy health and eco- (CI) for each 10μg/m3 increase in daily mean concentra- nomic burden around the world, especially in the industri- tions of NO2 pollutants have been reported for hospital ad- alized and the developing countries [25]. Air pollution has missions due to chronic obstructive pulmonary disease been certified as a trigger for exacerbation of COPD for (COPD) 1.0038 (1.0004 to 1.0094) by WHO. Also, base- more than fifty years and has lead to the expansion of air line incidences (BI), namely the incidence expected for a quality standards in many countries worldwide, leading to specific disease, per hundred thousand people have been a substantial decrease of the level of variation of air pollu- presented: 132 for acute myocardial infarction (MI), 497 tants derived from the burning of different fossil fuels [25]. for cardiovascular disease and 101.4 for COPD by WHO. The main objective of the present study was the assessment The assessment is based on the attributable proportion of health outcome due to NO2 in Tabriz air. (AP), defined as the fraction of the health outcome in a given population attributable to exposure to a given atmos- pheric pollutant, assuming a proven causal relation be- 2. MATERIALS AND METHODS tween exposure and health outcome and no major con- founding effects in that association. The AP can be easily 2.1 Study population and exposure assessment calculated through the following general formula [26]: This study was conducted during one year (from 21th AP =∑{ [RR(c) -1]×p(c)}/ [RR(c) ×p(c)] (1) Mars 2011 to 21th Mars 2012) in Tabriz city which has a population of 1 545 491 inhabitants. NO2 concentration (in Where AP is the attributable proportion of the health volumetric unit) in Tabriz was measured by the Eastern outcome, RR is the relative risk for a given health outcome, Azerbaijan Department of Environment (EADoE), using in category "c" of exposure, obtained from the exposure– fix stations in six areas during 365 days in 2011. The six response functions derived from epidemiological studies
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and P(c) is the proportion of the population in category "c" 34.2 / 1 545 491 = 2.2 deaths per 100000 people of exposure. If the baseline frequency of the health outcome in the population under investigation is known, the rate at- tributable to the exposure can be calculated as follows [26]: 3. RESULTS AND DISCUSSION IE = I AP (2) In the table there are the days of exposure associated Where IE is the rate of the health outcome attributable with the concentrations of NO and the annual average of to the exposure and I is the baseline frequency of the health 2 all stations (measured only in winter and in summer) and outcome in the population under investigation. Finally, for the stations that have shown the highest and lowest con- knowing the size of the population, the number of cases centrations. Accordingly, the average and maximum annu- attributable to the exposure can be estimated as follows: als of NO2 in the considered stations of Tabriz were 33.54 NE = IE N (3) and 78.52 µg/m3 respectively. The maximum amount was Where NE is the number of cases attributed to the ex- related to Raste-khoche station located in the downtown posure and N is the size of the population investigated. RR and the minimum was related to Rah-ahan station that is in gives the increase in the probability of the adverse effect the western part of the city (Table 1). associated with a given change in the exposure level, and 3 comes from time-series studies where day-to-day changes TABLE 1 - Highest and lowest concentrations of NO2 (μg/m ) corre- sponding to the stations for use in the AirQ2.2.3 model in air pollutants over long periods are related to daily mor- tality, hospital admissions and other public health indica- Raste-kouche Rah-ahan Total Parameter tors. This program is used to estimate the impact of expo- (Maximum) (minimum) stations sure to specific atmospheric pollutants on the health of peo- average annual 68.07 9.25 33.54 ple living in a given period and area. The assessment is average summer 34.06 11.75 29.92 based on the attributable proportion (AP), defined as the Average winter 38.3 8.52 24.82 fraction of the health outcome in a given population at- 98 percentiles annual 166.7 16.25 65.35 maximum annual 200.83 20.52 78.52 tributable to exposure to a given atmospheric pollutant, as- Maximum summer 200.83 20.1 78.52 suming a proven causal relation between exposure and Maximum winter 82.91 20.52 47.34 health outcome and no major confounding effects in that association. RR and estimated attributable proportions (AP) for acute myocardial infarction (MI) attributable to NO2 are 2.3 Health End Point shown in Table 2. According to the model default, the base- Using the results of data processing and recorded data line incidence (BI) of this health endpoint for NO2 was 132 regarding the pollutants monitored in the considered sta- per 100000 people; so, the number of excess cases was es- tions of Tabriz the following information can be drawn: timated for 16 persons at centerline of relative risk In this studied city with the population equivalent to 1 (RR=1.0036 and AP=0.8504). Moreover, the RR and esti- 545 491 people, the total cumulative of cardiovascular mated AP are presented in table 2 for cardiovascular mor- mortality due to NO2 per hundred thousand can be calcu- tality. BI obtained from model default was 497 per 100000 lated as follows: people for this health endpoint. Therefore, estimated num- ber of excess cases were calculated to be 34 persons at cen- (Mct/ Pt) x 100000 (4) terline of relative risk (RR=1.002) and attributable propor- M : The number of cardiovascular deaths from expo- ct tions (AP=0.4742). Amounts of RR, AP and hospital ad- sure to NO2 mission chronic obstructive pulmonary disease (COPD) at- Pt: Total Population tributable to NO2 using the model in moderate limits were Unit: Number of deaths per 100000 persons estimated to be 1.0038, 0.8972, respectively. BI of this health
TABLE 2 -Relative risk, Attributable proportion and number of persons suffering from type of health end point due to NO2 exposure
Index Estimated Estimated number of excess cases Health End Point RR Estimate AP (%) (persons) Lower 1.0015 0.3561 6.8 Acute myocardial infarc- central 1.0036 0.8504 16.3 tion(BI=132 per 105 persons) upper 1.0084 1.962 37.6 Lower 1 0 0 Cardiovascular mortality central 1.002 0.4742 34.2 (BI=497 per 105 persons) upper 1.004 0.944 68 Lower 1.0004 0.0952 1.4 COPD central 1.0038 0.8972 13.2 (BI=101.4 per 105 persons) upper 1.0094 2.1905 32.2
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endpoint was 101.4 per 100000, the estimated number of ex- model default for assessing health effects of NO2. Further- cess cases were 13 persons. Most of the resulting health con- more, baseline incidence (BI) values were also taken from sequences had been related to concentrations in the range of model default. Charts drawing based on the cumulative 20 to 60 µg/m3 of pollutants (Table2 and Figure 2). Table 2 number of each Health End Point showed number of cases represents the estimated Attributable proportion (AP) and in three modes, 5, 50 and 95% relative risks, leading there- number of excess cases in comparison to the cases giving fore to three curves (related to lower, central and upper rela- a pollutant concentration of 10μg/m3. The numbers of ex- tive risks) in each graph, middle curve corresponds to a rel- cess cases regarding acute myocardial infarction, cardio- ative central risk, lower curve corresponds to a relative risk vascular mortality and chronic obstructive pulmonary dis- of 5% (underestimate) and the upper curve corresponds to a ease (COPD) are estimated based on the different Relative 95% relative risk (overestimate). We found about 0.9% of Risk (RR) and Attributable proportion (AP). total hospital admissions related to the chronic obstructive pulmonary disease (HA COPD) was caused by NO2. More- Figures 1-3 illustrates the NO2 concentrations versus the over, the results of this study showed that 0.85% (CI 95%; cumulative of acute myocardial infarction, cardiovascular 0.36%; 0.96%) of acute myocardial infarction, 0.47% (CI mortality and chronic obstructive pulmonary disease 95%; 0; 0.94%) of cardiovascular mortality and 9.28% (CI (COPD). The short-term health impacts of exposure to NO2 95%; 0.1%; 2.19%) of cardiovascular admissions for above a reference value of 10 μg/m3 are shown in these fig- chronic obstructive pulmonary disease (COPD) can be at- 3 ures. Three ranges of relative risk were considered based on tributed to NO2 concentrations over 10 μg/m .
40 35 MI 30
Acute 25
of 20 rr=1.0015 (persons) 15 10 rr=1.0036
Cumulative 5 0 rr=1.0084
No2 concentration (microgram/cubic meter)
FIGURE 1 - Relationship between Cumulative number of acute myocardial infarction (MI) and NO2 concentration
80
70 60 50 (persons) cardiovascular
40 of
30 rr=1
mortality 20 rr=1.002 10 Cumulative rr=1.004 0
No2 concentration (microgram/cubic meter)
FIGURE 2 - Relationship between Cumulative number of cardiovascular mortality and NO2 concentration
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35 30 COPD
25 HA 20 of
15 rr=1.0004 (persons) 10 rr=0038
Cumulative 5 rr=1.0094 0
NO2 concentration (microgram/cubic meter)
FIGURE 3 - Relationship between Cumulative number of hospital admission for chronic obstructive pulmonary disease (COPD) and NO2 concentration
Figures 1 to 3 illustrate the NO2 concentration versus out of 38 people in a year. The NO2 effect on cardiovascu- related to the health end point. Indicators of Relative Risk lar mortality was an excess of about 68 cases in a year as (RR) and Attributable Proportion (AP) for acute Myocar- well as the excess of hospital admissions for chronic obstruc- dial Infarction (MI) to nitrogen dioxide defined on the tive pulmonary disease (COPD) of about 1 out of 32 people Baseline Incidence (BI) were 132 from 105 persons accord- in a year. The number of excess cases was calculated for ing to the AirQ model from the WHO. The results showed concentrations above 10 μg/m3; so assuming a relatively low three ranges of relative risks based on the model default, reference value, particularly compared to air standards. which were considered for assessing health effects of NO2. However, this was justifiable because the aim was to assess Furthermore, baseline incidence (BI) values were also taken the human health impact of NO2 as the air pollutant and not from the model default. For a population of one and half whether any particular air quality standards or guidelines millions people and based on BI of 497 per one hundred were exceeded. As Figures 1 to 3 show, health end point thousand peoples each year, some 7 455 cardiovascular mor- (number of outcomes) approximately under 70 μg/m3 of tality cases can be expected annually; out of this number, NO2 has highly increased. This indicates that most days of 34 cases can be attributed to NO2 concentrations above exposure have been lower than of this concentration. In 20μg/m3. Seventy percent of acute cardiovascular mortality other words, the number of days of people exposure has cases occurred in days with pollution not exceeding 50 μg/m3, been higher in this concentration interval. Gudarziet al. 97.7% of cardiovascular mortalities occurred in days with [5]exploited AirQ model to estimate the NO2 hygienic ef- pollutant not exceeding 70 μg/m3 and 87.9% of COPD cases fects in Tehran. Based on their results, almost 3.4% of all occurred in days with pollutant not exceeding 60 μg/m3. Al- cases of the whole cardiovascular and respiratory mortali- though cardiovascular mortality had the lowest relative risk ties are attributed to NO2 concentrations, greater than 60 (RR) in centerline compared to acute cardiovascular mor- μg/m3. The number of cases of myocardial infarction caused tality and chronic obstructive pulmonary disease, a high by exposure to NO2 was reduced to zero when health out- number of this health end point case was due to its higher BI come was closed to 5% range. In addition, if no appropriate value (497 per 105 peoples). On the other hand, a greater measures to control air pollution and reduce health effects number of acute cardiovascular mortality cases compared to were taken, the corresponding RR was close to 95% range acute cardiovascular mortality (MI) ones were related to and the number of cases of myocardial infarction was esti- higher baseline incidence for MI. The comparison between mated to be more than 2-fold increase in exposure to NO2. acute cardiovascular mortality and obstructive pulmonary This study has a number of limitations: one of the limita- disease showed that the baseline incidence (BI) had more tions of this approach is that the health impact focuses on importance than relative risk (RR). This paper offers a individual compounds without considering the simultane- study case using the WHO approach to assess the impact ous exposure to several ones, which is what actually oc- of atmospheric pollution on human health for people living curs. The health effects of atmospheric pollution are indeed in Tabriz, one of the most densely populated and industri- the consequence of interactions between different air con- alized areas in Iran, where the geographical features make taminants, and between these ones and other compounds of the air quality among the worst in the world. Considering natural origin. Generally, in quantitative assessments of health short-term effects of NO2 on the 1 500 000 inhabitants, effects, the interactions between different contaminants are causing an excess of acute myocardial infarction (MI) of 7 not investigated as it would require a good knowledge of
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the mechanisms of toxicity for the different compounds, REFERENCES which is rarely available. A further limitation is due to the RR estimates derived in studies of different populations in [1] Defense, H. (1996) Health effects of outdoor air pollution. comparison to that under investigation. Furthermore, this American journal of respiratory and critical care medicine, pp.477-498. model does not consider intra-individual differences due to different behaviors inside the population considered (i.e [2] Traversi, D., Degan, R., De Marco, R., Gilli, G., Pignata, C., active or passive smoking, mobility during the day, prox- Ponzio, M., Rava, M., Sessarego, F., Villani, S. and Bono, R. (2008) Mutagenic properties of PM2.5 air pollution in the imity to major roads, etc.). That’s because the approach is Padana Plain (Italy) before and in the course of XX Winter ecological and not epidemiological. Finally, another limi- Olympic Games of “Torino 2006”. Environment international, tation related to the exposure assessment is that the ap- pp.966-970. proach assumes that concentrations measured in specific [3] Al-Hurban, A. E. and Al-Ostad, A. N. (2010) Textural charac- sampling points are representative of the average exposure teristics of dust fallout and potential effect on public health in suffered from people living in Tabriz. Having said that, the Kuwait City and suburbs. Environmental Earth Sciences, pp.169-181. following actions should be performed in order to reduce health effects caused by air pollution, so that better quanti- [4] Fung, K., Krewski, D., Burnett, R. and Dominici, F. (2005) fication studies are recommended: 1- Restricting the use of Testing the harvesting hypothesis by time-domain regression analysis. I: baseline analysis. Journal of Toxicology and Envi- underground resources and fossil material; measures are ronmental Health, Part A, 68, pp.1137-1154. necessary to reduce the high urban traffic, which implies [5] Goudarzi, G., Mohammadi, M. J., Angali, K. A., Neisi, A. K., the raise of the level of urban public transport systems and Babaei, A. A., Mohammadi, B., Soleimani, Z. and Geravandi, correct traffic management, as well as the implementation S. (2011) Estimation of Health Effects Attributed to NO2 Ex- of effective strategies to improve the quality of industrial posure Using AirQ Model. Archives of Hygiene Sciences, productions. 2- The use of this model or other models is pp.59-66. recommended to investigate and evaluate the effects of air [6] Gehring, U., Heinrich, J., Krämer, U., Grote, V., Hochadel, pollution on health of people in other metropolitan country, M., Sugiri, D., Kraft, M., Rauchfuss, K., Eberwein, H. G. and and comparisons should be made. 3- Because the determi- Wichmann, H.-E. (2006) Long-term exposure to ambient air pollution and cardiopulmonary mortality in women. Epidemi- nation of the impact of air pollution on the related diseases ology, pp.545-551. and estimation of the health effects of air pollutants is a [7] Krzyzanowski, M. (2008) WHO air quality guidelines for Eu- prerequisite, epidemiological indicators based on expert rope. Journal of Toxicology and Environmental Health, Part calculations are needed. A, pp.47-50.
[8] Miri, A., Ahmadi, H., Ghanbari, A. and Moghaddamnia, A. (2007) Dust storms impacts on air pollution and public health 4. CONCLUSION under hot and dry climate. Int. J. Energy Environ, pp.101-105 [9] Samet, J. M., Zeger, S. L., Dominici, F., Curriero, F., Coursac, This study applied the AirQ software and the approach I., Dockery, D. W., Schwartz, J. and Zanobetti, A. (2000) The proposed by the WHO to provide quantitative data on the national morbidity, mortality, and air pollution study. Part II: morbidity and mortality from air pollution in the United States impact of NO2 on the health of people living in a given area and this was the first attempt to assess health impacts of air Res Rep Health Eff Inst, pp.75-79. pollution in Tabriz, Iran. Although the results of this study [10] Vineis, P., Hoek, G., Krzyzanowski, M., Vigna-Taglianti, F., are in line with results of other researches around the Veglia, F., Airoldi, L., Overvad, K., Raaschou-Nielsen, O., Clavel-Chapelon, F. and Linseisen, J. (2007) Lung cancers at- world, despite the fact that the demographic, geographic tributable to environmental tobacco smoke and air pollution in and climate characteristics are different, there is still a high non-smokers in different European countries: a prospective need to further studies to specify local RRs and BIs instead study. Environmental Health, pp.1-7. of WHO defaults. As mentioned above, air quality daily [11] Touloumi, G., Katsouyanni, K., Zmirou, D., Schwartz, J., affects health conditions dramatically. Accordingly, cost- Spix, C., de Leon, A. P., Tobias, A., Quennel, P., Rabczenko, effective measures and management schemes should be D. and Bacharova, L. (1997) Short-term effects of ambient ox- considered to reduce air pollution concentrations and/or re- idant exposure on mortality: a combined analysis within the APHEA project. American journal of epidemiology, pp.177- duce exposure of people to air pollutants. The results are in 185. line with those of other investigations and, despite the lim- [12] Stieb, D. M., Judek, S. and Burnett, R. T. (2003) Meta-analysis itations, which are in common to similar studies, indicate of time-series studies of air pollution and mortality: update in that this method offers an effective and easy tool, helpful relation to the use of generalized additive models. Journal of in decision-making. the Air & Waste Management Association, pp.258-261. [13] Kan, H. and Chen, B. (2003)A case-crossover analysis of air The authors have declared no conflict of interest. pollution and daily mortality in Shanghai. Journal of occupa- tional health, pp.119-124. [14] Schwartz, J., Dockery, D. W., Neas, L. M., Wypij, D., Ware, J. H., Spengler, J. D., Koutrakis, P., Speizer, F. E. and Ferris Jr, B. G. (1994) Acute effects of summer air pollution on res- piratory symptom reporting in children. American journal of respiratory and critical care medicine, pp.1234-1242.
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[15] Barnett, A. G., Williams, G. M., Schwartz, J., Best, T. L., Nel- ler, A. H., Petroeschevsky, A. L. and Simpson, R. W. (2006) The effects of air pollution on hospitalizations for cardiovas- cular disease in elderly people in Australian and New Zealand cities. Environmental health perspectives, pp.1018-1023 [16] Sunyer, J., Basagana, X., Belmonte, J. and Anto, J. (2002) Ef- fect of nitrogen dioxide and ozone on the risk of dying in pa- tients with severe asthma. Thorax, pp.687-693. [17] Seaton, A., Godden, D., MacNee, W. and Donaldson, K. (1995) Particulate air pollution and acute health effects. The Lancet, pp.176-178. [18] Krzyzanowski, M., Kuna-Dibbert, B. and Schneider, J. (2005) Health effects of transport-related air pollution. WHO Re- gional Office Europe, pp.205. [19] Krzyzanowski, M., Cohen, A. and Anderson, R. (2002) Quan- tification of health effects of exposure to air pollution. Occu- pational and environmental medicine, pp.791-793. [20] KuÈnzli, N., Kaiser, R., Medina, S., Studnicka, M., Chanel, O., Filliger, P., Herry, M., Horak Jr, F., Puybonnieux-Texier, V. and Quenel, P. (2000) Public-health impact of outdoor and traffic-related air pollution: a European assessment. The Lan- cet, pp.795-801.
[21] Badescu, V. and Cathcart, R. (2011) Macro-engineering Sea- water in Unique Environments: Arid Lowlands and Water Bodies Rehabilitation. Springer, pp. 323-329.
[22] Antman, E. M., Anbe, D. T., Armstrong, P. W., Bates, E. R., Green, L. A., Hand, M., Hochman, J. S., Krumholz, H. M., Kushner, F. G. and Lamas, G. A. (2004) ACC/AHA guidelines for the management of patients with ST-elevation myocardial infarction—executive summary: a report of the American Col- lege of Cardiology/American Heart Association Task Force on Practice Guidelines (Writing Committee to Revise the 1999 Guidelines for the Management of Patients With Acute Myo- cardial Infarction). Journal of the American College of Cardi- ology, pp.671-719. [23] Brook, R. D., Franklin, B., Cascio, W., Hong, Y., Howard, G., Lipsett, M., Luepker, R., (2004) Mittleman, M., Samet, J. and Smith, S. C. Air pollution and cardiovascular disease A state- ment for healthcare professionals from the expert panel on population and prevention science of the American Heart As- sociation. Circulation, pp.2655-2671.
[24] Sarnat, J. A., Schwartz, J. and Suh, H. (2001) Fine particulate air pollution and mortality in 20 US cities. N Engl J Med, pp.1253-1254. Received: January 19, 2015 [25] Ghozikali, M. G., Mosaferi, M., Safari, G. H. and Jaafari, J. Revised: March 25, 2015 (2014) Effect of exposure to O3, NO2, and SO2 on chronic ob- Accepted: April 09, 2015 structive pulmonary disease hospitalizations in Tabriz, Iran. Environmental Science and Pollution Research, pp.1-7.
[26] Fattore, E., Paiano, V., Borgini, A., Tittarelli, A., Bertoldi, M., CORRESPONDING AUTHOR Crosignani, P. and Fanelli, R. (2011) Human health risk in re- lation to air quality in two municipalities in an industrialized area of Northern Italy. Environmental research, pp.1321-1327. Behzad Heibati Health Sciences Research Center Faculty of Health, Student Research Committee Mazandaran University of Medical Sciences Sari IRAN
Phone: +98 9389489389 Fax: +98 2188950188 E-mail: [email protected]
FEB/ Vol 24/ No 11c/ 2015 – pages 4142 - 4148
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IMPACT OF COPPER AND ITS Ca MIXTURES ON THE GLUTATHIONE S-TRANSFERASE ACTIVITY, GLUTATHIONE LEVEL AND METAL ACCUMULATION IN TISSUES OF FRESHWATER FISH Oreochromis niloticus
Gülbin Firidin1,* and Ferit Kargın2
1Life Science Application and Research Center, Gazi Univercity, 06830 Ankara, Turkey 2Department of Biology, Faculty of Art and Science, Cukurova University, 01330 Adana, Turkey
ABSTRACT tissues, may become extremely toxic for aquatic animals including fish as its concentration in water increases [2]. Ac- This study was designed to evaluate the effects of dif- cumulation of Cu in tissues of fishes leads redox reactions ferent concentrations of copper (Cu) and copper+calcium generating free radicals and, hence, may cause changes in (Cu+Ca) mixtures in the liver, kidney, and brain tissues of certain physiological processes [3]. Oreochromis niloticus. The fish were exposed to 0.1, 0.5, Multiple factors including temperature, salinity, alkalin- 1.0 mg/L Cu and 0.1 mg/L Cu+1.0 mg/L Ca, 0.5 mg/L ity, hardness, pH and interaction with other metals strongly Cu+5.0 mg/L Ca, 1.0 mg/L Cu+10.0 mg/L Ca mixtures for influence metal toxicity in water and, consequently, its bio- 7, 14, and 28 days to observe Cu accumulation, total gluta- availability for absorption by fish [4-5]. Ca is the major cat- thione (tGSH) level and glutathione S-transferase (GST) ion which is responsible protective effects of hard water. It activity. The alterations in these parameters were deter- has been reported to have a direct ameliorative effect against mined. Cu accumulation increased in the tissues with in- Cu toxicity [6- 7]. creasing concentrations of Cu in the exposure medium and O. niloticus, native fish species of Egypt, is a widely used with increasing duration of exposure. Cu levels in the tis- species in aquaculture for food source and has been suggested sues, exposed to Cu + Ca mixtures were significantly lower as a bioindicator of water contamination, so that easy to breed than exposed to the single Cu. GST activity increased sig- and grow in a variety of aquaculture systems [8]. nificantly in the liver on the contrary of decreasing in the kidney and brain under Cu exposure. Cu treatment caused Liver and kidney tissues are known to play an im- a depletion of GSH level in the brain, while increasing in portant role in detoxification and excretion of toxicants, the kidney and liver. In conclusion, Ca exposure seemed to which explains the high accumulation of Cu in these organs reduce the severity of Cu-induced toxicity in the tissues of [9]. Brain is the center of the nervous system in all verte- O. niloticus. brates. As an organ in which homeostasis must be strictly maintained, brain tissue contains large amounts of polyun-
saturated fatty acids, which are particularly vulnerable to KEYWORDS: free radical attacks [10]. The glutathione S-transferases Oreochromis niloticus, copper, calcium, accumulation, antioxidant (GST, EC 2.5.1.18), are a multi-gene family of enzymes involved in the detoxification which catalyze conjugation reaction with the tripeptide glutathione to a variety of com- 1. INTRODUCTION pounds with an electrophilic group [11]. Glutathione (GSH, γ-glutamylcysteinylglycine), a low molecular- Environmental pollution by heavy metals has become weight sulphur-containing compound, is a tripeptide that one of the most important problems in the world due to ex- easily oxidized and can be regenerated rapidly. Its func- tensive use of them in industrial, agricultural and anthro- tions are synthesis of protein and DNA, amino acid pogenic activities that are becoming threats to living organ- transport, maintenance of the thiol-disulfide status, free isms. Aquatic ecosystems which receive a wide range of radical scavenging, signal transduction, as an essential co- pollutants are the most important final receptacles [1] thus factor of several enzymes, as a non-toxic storage form of aquatic organisms are exposed to a significant amount of cysteine, and as a defense against oxidizing molecules and these pollutants. Cu, a trace metal essential for all living potentially harmful xenobiotics such as metals [12, 13].
Fish cells usually try to remove metals by direct conjuga- * Corresponding author tion with GSH [14].
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The main purpose of this study was to assess Ca could 9500×g (Sigma (2-16)) for 30 min to obtain supernatants protect against Cu-induced toxicity in tissues of O. nilot- which were used for GST activity, GSH and total protein icus. It was investigated the changes in Cu accumulation, levels. All process was carried out at + 4°C. Chemicals GST activity and GSH level in liver, kidney, and brain tis- used in this study were obtained from Sigma or Merk (Ger- sues in response to Cu and Cu + Ca exposure. many). GSH was determined according to the method of An- derson [16]. The reaction medium contained 143 mM so- 2. MATERIALS AND METHODS dium–potassium buffer (Na-K Phosphate (contained 6,3 mM EDTA, pH 7,5)), 0,3 mM NADPH, 6 mM DTNB, and 2.1. Test animals and experimental protocol 50 units of GR. DTNB formation was monitored continu- O. niloticus (mean weight 28.85±0.68 g, mean length ously at 412 nm for 3 min. Absorbance was converted to 12.10±0.24 cm) were obtained from Çukurova University concentration by using standard graph that was prepared Fish Culture Farm and transferred to the laboratory. Before with GSH. the experiments, the fish were acclimatized to laboratory GST activity was determined by the method described conditions in 120 L glass tanks filled with aerated and de- in Habig et al. [17] using 1-chloro-2.4-dinitrobenzene chlorinated tap water for 1 month. Throughout the exper- (CDNB) as a substrate. The reaction mixture consisted of iments, the mean measured water quality parameters were: 100 mM Tris buffer (pH 7.4), 1 mM GSH, 1 mM CDNB. temperature 21.14±0.18, dissolved oxygen 7.35±0.11 mg/L The conjugation rate of GSH with CDNB was monitored (constant aeration), pH 8.19±0.009, total hardness 352.63± at 340 nm for 2 min. Enzyme activity calculated by using 5.14 mg/L CaCO3, and alkalinity 201.5±5.871 mg/L as a molar extinction coefficient of 9.6×103 M−1 cm−1. CaCO3 at acceptable levels. The fish were maintained under Protein concentrations of the supernatants for both at 12-h light/12-h dark illumination period. Both experi- GST analysis and GSH assay were measured using a col- mental and control fish were fed with commercial pellet orimetric method of Lowry et al. [18] using bovine serum feed at a rate of 3% of their body weight per day (Pinar albümin as a standard. Protein content was expressed in Feed, Turkey). mg/g wet weight of the tissues. After the period of acclimatization, experimental fish stock have been randomly divided into three groups each 2.4. Statistical analysis containing 18 fish. The first group was held in tap water as All parameters were expressed as means standard ± er- control and other groups were exposed to Cu (CuSO .5H O) 4 2 ror. Data analyses were carried out using Student Newman and Cu + Ca (CaSO .2H O) mixtures for 7, 14, and 28 days. 4 2 Keul’s Test (SNK) in SPSS statistical package programs. Three replicate groups of fish were used for each exposure. Data shown with different letters are significant at the P < All the experimental solutions were completely replaced 0.05 level. every other day from a newly prepared stock solution to reduce the ammonia content in water. There was no fish mortality during the experiments. 3. RESULTS Prior to analysis, six fish were taken out from the aquaria and their liver, kidney, and brain tissues were dis- Metal levels in the tissues of fish were shown in Tables sected separately on an ice cold plate as well as control 1-3. Cu accumulated significantly in liver, kidney, and fish. Tissues were then washed in ice cold physiological brain tissues duration of exposure at all concentration. saline solution, blotted dry, weighed and stored frozen at However, Cu levels in brain were below the detection limit −80 °C until metal and biochemical analyses. for the instrument in control groups. The highest metal ac- cumulation was observed in liver, it was followed by kid- 2.2. Metal analysis ney and brain (P<0.05). Cu accumulation in tissues of fish Tissue samples were dried at 105 oC for 48 h. Samples exposed to Cu + Ca mixtures were significantly lower than were transferred into a digestion flask containing 2 ml in fish exposed to the single Cu. When compared with 1.0 o HNO3 and 1 ml HCIO4 and digested at 120 C for 3 h. [15]. mg/L Cu exposure, metal levels decreased by 39, 19, 8% The digest was diluted with distilled water appropriately in (7 days); 25, 39, 15% (14 days) and 38, 31, 7% (28 days) the range of standards which were prepared from stock in tested 1.0 mg/L Cu + 10.0 mg/L Ca group in brain, kid- standard solution of the metals. After dilution Cu levels of ney and liver respectively (P<0.05) (Tables 1,2,3). Statisti- tissues measured on a Perkin Elmer AS 3100 atomic ab- cal difference was determined to be important between Cu sorption flame spectrophotometer and metal concentration and Cu + Ca mixtures in liver and kidney tissues in all treat- in a tissue was presented as μg metal/g dry weight. ment durations. Accumulation in tissues increased signifi- cantly in Cu-exposed fish (Tables 1.2.3) and the highest in- 2.3. Biochemical analysis crease was found at 1.0 mg/L Cu concentration. The tissues were homogenized (1:10, w/v) in 0.05 M Significant changes were recorded in kidney, liver, and Phosphate buffer (pH 7.4) containing 0.25 M sucrose at brain GST activity with respect to control groups. Kidney 9500 rpm for 2 min. Homogenates were centrifuged at and brain GST activity decreased in all treatments whereas
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TABLE 1 - The Effects of Ca on Cu accumulation in tissues of O. niloticus at 7 day exposures (µg Cu/g k.a.).
Metal concentrations (mg/L) Brain Kidney Liver
0.0 N.A 0.25±0.009 a 1.26±0.006 a 0.1 Cu 1.26±0.17 ax 20.83±0.29 by 199.8±0.95 bz 0.1 Cu +1.0 Ca 0.80±0.15 ax 18.17±0.39 cy 164.1±1.31 cz
0.0 N.A 0.25±0.009 a 1.26±0.006 a 0.5 Cu 2.56±0.22 ax 28.83±0.10 by 317.5±1.34 bz 0.5 Cu+5.0 Ca 2.50±0.10 ax 19.33±0.64 cy 302.3±1.20 cz
0.0 N.A 0.25±0.009 a 1.26±0.006 a 1.0 Cu 3.41±0.22 ax 41.69±0.87 by 398.2±1.49 bz 1.0 Cu+10.0 Ca 2.09±0.37 bx 33.86±0.87 cy 367.3±1.45 cz NA: not available Values are expressed as mean ± Standard error (N=6). Letters a, b and c indicate differences among concentrations and letters x, y and z indicate statistical differences among tissues at the P<0.05 level.
TABLE 2 - The Effects of Ca on Cu accumulation in tissues of O. niloticus at 14 day exposures (µg Cu/g k.a.).
Metal concentrations (mg/L) Brain Kidney Liver
0.0 N.A 0.23±0.012 a 1.25±0.009 a 0.1 Cu 3.01±0.42 ax 32.72±0.15 by 260.6±2.71 bz 0.1 Cu + 1.0 Ca 2.24±0.35 abx 23.32±0.73 cy 248.6±1.85 cz
0.0 N.A 0.23±0.012 a 1.26±0.009 a 0.5 Cu 3.42±0.10 ax 47.72±0.49 by 358.6±1.73 bz 0.5 Cu+5.0 Ca 3.05±0.01 bx 24.50±0.45 cy 326.2±2.48 cz
0.0 N.A 0.23±0.012 a 1.26±0.009 a 1.0 Cu 4.26±0.44 ax 62.20±0.64 by 433.9±1.22 bz 1.0 Cu+10.0 Ca 3.21±0.14 bx 38.24±0.67 cy 370.6±1.50 cz NA: not available Values are expressed as mean ± Standard error (N=6). Letters a, b and c indicate differences among concentrations and letters x, y and z indicate statistical differences among tissues at the P<0.05 level.
TABLE 3 - The Effects of Ca on Cu accumulation in tissues of O. niloticus at 28 day exposures (µg Cu/g k.a.).
Metal concentrations (mg/L) Brain Kidney Liver
0.0 N.A 0.26±0.003 a 1.25±0.015 a 0.1 Cu 5.11±0.71 ax 41.50±0.60 by 306.2±2.92 bz 0.1 Cu + 1.0 Ca 4.83±0.44 ax 30.41±0.24 cy 252.1±1.37 cz
0.0 N.A 0.26±0.003 a 1.25±0.015 a 0.5 Cu 6.22±0.37 ax 59.02±0.69 by 405.9±1.81 bz 0.5 Cu+5.0 Ca 4.68±0.10 bx 38.99±0.48 cy 369.0±0.92 cz
0.0 N.A 0.26±0.003 a 1.25±0.015 a 1.0 Cu 7.79±0.10 ax 87.21±0.85 by 498.7±1.17 bz 1.0 Cu+10.0 Ca 4.85±0.43 bx 59.78±0.30 cy 464.8±1.22 cz NA: not available Values are expressed as mean ± Standard error (N=6). Letters a, b and c indicate differences among concentrations and letters x, y and z indicate statistical differences among tissues at the P<0.05 level.
liver GST activity increased. GST activity in kidney miti- an additive effect in brain GST activity except at 1.0 mg/L gated at all Cu-exposed during exposure period, this was Cu + 10.0 mg/L Ca concentration for a period of 28 days essential statistically except at 0.5 mg/L Cu concentration compared to Cu-exposed. Exposure of the highest Cu and for 17 days (Fig. 1). The combination of Cu + Ca exhibited Cu + Ca concentrations caused a decline 30%, 10% (7 days)
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and 43%, 46% (28 days) in brain GST activity in relation GST activity in liver increased 14 and 5% with respect to control values (P<0.05) (Fig. 3). It was not significantly to control at highest concentration of Cu and Cu + Ca groups altered at all concentration of Cu and Cu + Ca mixtures for a period of 28 days (P<0.05). All treatments of Cu and between exposure periods (P>0.05) (Fig. 2) although sta- Cu + Ca mixtures induced GST activity in liver whereas de- tistical difference was observed between Cu and Cu + Ca creased in brain and kidney as compared to control groups, mixtures at exposure concentration. during exposure periods. Results showed that Cu and Cu + Ca mixtures altered GST activity inducing oxidative stress.
Control 0.1 mg/L Cu 45 * * 0.5 mg/L Cu 40 * * * * * 1.0 mg/L Cu * * * 0.1+1.0 mg/L Cu+Ca 35 * * 0.5+5.0 mg/L Cu+Ca 30 * 1.0+10.0 mg/L Cu+Ca 25 20 15 10 5 0 GST activitiy GST activitiy (U/mg protein) 71428
Exposure time (in days)
FIGURE 1 - Kidney GST activity (U mg-1 of protein) in O. niloticus exposed to metals for 7, 14, and 28 days. Data are expressed as mean±standard error (N=6). * shows significant differences between time for the same exposure group (P<0.05).
Control 60 0.1 mg/L Cu 50 0.5 mg/L Cu 1.0 mg/L Cu 40 0.1+1.0 mg/L Cu+Ca 0.5+5.0 mg/L Cu+Ca 30 1.0+10.0 mg/L Cu+Ca 20 10
GST activitiy GST activitiy (U/mg protein) 0 71428
Exposure time (in days)
FIGURE 2 - Liver GST activity in (U mg-1 of protein) O. niloticus exposed to metals for 7, 14, and 28 days. See Fig. 1 for details.
Control 0.1 mg/L Cu 7 0.5 mg/L Cu
6 * 1.0 mg/L Cu * 0.1+1.0 mg/L Cu+Ca 5 * 0.5+5.0 mg/L Cu+Ca 1.0+10.0 mg/L Cu+Ca 4 *
3 2
1
GST activitiy GST activitiy (U/mg protein) 0
71428
Exposure time (in days)
FIGURE 3 - Brain GST activity(U mg-1 of protein) in O. niloticus exposed to metals for 7, 14, and 28 days. See Fig. 1 for details.
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Exposure to Cu and Cu + Ca caused an increase in 1.0 mg/L Cu + 10.0 mg/L Ca concentrations respectively GSH level in kidney and liver when compared to control in kidney when compared to control values (P<0.05). Con- levels (Figs. 4, 5, 6) (P<0.05). This increase was higher for siderable differences did not occur in GSH contents of kid- exposure to Cu alone than in presence of Ca with Cu, at the ney at 0.1 and 0.5 mg/L Cu concentrations between the ex- end of the period. After the first period of 7 days, GSH posure times (P>0.05), however at 1.0 mg/L Cu, it showed level was increased by 22%, 25% at 0.1 mg/L Cu and 0.1 a significant reduction after 28 days according to that at mg/L Cu + 1.0 mg/L Ca; 33%, 17% at 0.5 mg/L Cu and 0.5 14 days (P<0.05) (Fig.4). mg/L Cu+5.0 mg/L Ca and 40%, 35% at 1.0 mg/L Cu and
Control 0.1 mg/L Cu 0,2 * * * 0.5 mg/L Cu 1.0 mg/L Cu 0,15 0.1+1.0 mg/L Cu+Ca 0.5+5.0 mg/L Cu+Ca 0,1 1.0+10.0 mg/L Cu+Ca
0,05
GSH level (µM/mg protein) GSH level 0 71428
Exposure time (in days)
FIGURE 4 - Kidney GSH level (µM/mg protein) in O. niloticus exposed to metals for 7, 14, and 28 days. See Fig. 1 for details.
Control 0.1 mg/L Cu 1,4 * 0.5 mg/L Cu 1,2 * 1.0 mg/L Cu * * 0.1+1.0 mg/L Cu+Ca 1 * * * * * * 0.5+5.0 mg/L Cu+Ca 0,8 * 1.0+10.0 mg/L Cu+Ca * 0,6 0,4 0,2 GSH level GSH level (µM/mg protein) 0 71428
Exposure time (in days)
FIGURE 5 - Liver GSH level (µM/mg protein) in O. niloticus exposed to metals for 7, 14, and 28 days. See Fig. 1 for details.
Control 0,9 0.1 mg/L Cu 0,8 * * 0.5 mg/L Cu * 1.0 mg/L Cu 0,7 * * * * * 0.1+1.0 mg/L Cu+Ca 0,6 * * * 0.5+5.0 mg/L Cu+Ca 0,5 * 1.0+10.0 mg/L Cu+Ca 0,4 0,3 0,2 0,1 GSH level GSH level (µM/mg protein) 0 71428
Exposure time (in days)
FIGURE 6 - Brain GSH level (µM/mg protein) in O. niloticus exposed to metals for 7, 14, and 28 days. See Fig. 1 for details.
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The highest Cu and Cu + Ca treatment caused more [25, 26]. GST is important for xenobiotic metabolism and significant elevation in liver GSH during exposure period antioxidative protection in aquatic organisms [27, 28]. In (103%, 41% at 7 days; 69%, 57% at 14 days; 59%, 54% at this study, all GST activity, except in kidney, elevated sig- 28 days, respectively) (P<0.05) (Fig.5). GSH level of brain nificantly in treated groups with respect to control in brain was lower in all Cu–exposed than in control fish during expo- and liver. Basha and Rani [29] demonstrated that sublethal sure time and significant reduction was recorded at 0.1 mg/L Cd (cadmium) exposure lead to increase in the activity of Cu and 0.1 mg/L Cu + 1.0 mg/L Ca concentrations as 40%, liver GST in O. mossambicus significantly. A similiar re- 18% (7 days) and 52%, 11% (28 days), respectively, in re- lation had been detected in our study. lation to control values (Fig.6) (P<0.05). Activity of GST, seems to be more affected at metal Treatment of Cu + Ca mixtures exerted antagonistic exposure which may be a result of changing the activity of effects with respect to Cu alone in GST activity and GSH enzymes either by binding to their functional groups level each time of exposure. Observed changes on these (sulfhydryl, carboxyl, imidazole, etc.) or by displacing the parameters were generally tissue-specific and dose-de- metal associated with the enzyme [30]. GST activity could pendent. be induced owing to neutralize the metal toxicity as form- ing part of an adaptive response to Cu toxicity. GSH is considered as a first line of cellular defense 4. DISCUSSION system against metals by chelating and detoxifying [31]. In the present investigation, GSH level in kidney and liver in- In the present study, accumulation of Cu in whole tis- creased under effects of Cu and Cu + Ca especially at the sues of O. niloticus decreased in the presence of Ca. This highest concentrations. Firat et al. [32] have suggested that is probably due to interactions at sites binding both metals GSH level increased in liver of O. niloticus exposed to Cd in tissues. In this study, an antagonistic effect of Ca on Cu and zinc (Zn). GSH induction in different fish species ex- accumulation was determined in the O. niloticus. Many posed to metals was also observed in other studies. Liver studies of Cu-Ca interaction in aquatic animals have con- GSH level was elevated during the exposure period in tis- centrated on the antagonistic effet of Ca, in other words, on sues of Brachydanio rerio which exposed to Cu concentra- the protective effect of Ca against the accumulation and tions (40 and 140 µg/L) [33]. 3.0 mg/L Cd exposure caused toxicity of Cu [19-20-21]. The researchers have suggested induction of GSH status in liver of O. niloticus [34]. In- that Ca has an antagonistic effect on Cu accumulation ei- creasing GSH level may be due to predictably through in- ther by the inhibition of Cu uptake from the gills or by the terference with GSH metabolism [35]. acceleration of the transport of the ingested Cu to the inter- As explained previously, GST and GSH are found vir- nal organs. The antagonistic effect of Ca on the Cu accu- tually in all tissues of vertebrates but their high level rep- mulation in O. niloticus tissues can be partially explained resent in liver which is a major organ for metal uptake and by the competition of these two metals for the binding sites enzymatic transformation of ROS (reactive oxygen spe- on the protein carriers such as metallothionein, or at the cies) [27]. However, GST activity and GSH level were not uptake sites on the gills. Other similar studies on heavy altered significantly in some Cu + Ca medium, due to the metal interaction in fish have observed that Ca reduces the protector effects of Ca against metal toxicity [36, 37]. uptake of Cd from water [22, 23]. Cu had an oxidative stress potential via alteration of Liver and kidney are highly active in fish metabolism GST activity and GSH level in O. niloticus tissues. Fluctua- so that they can accumulate metals higher than brain, as tion in antioxidant defense system in fish could be due to shown in this study. Cu accumulation in all tissues of fish increase in the production of ROS because of Cu accumula- increased significantly with increasing concentrations of tion. These changes could be reduced in the presence of cal- Cu, particularly in long treatment durations. In all concen- cium. Results of the present study showed that the effects of tration tested, highest level of Cu was found in liver of O. metals on GSH level and GST activity in tissues of O. nilot- niloticus followed by kidney and brain. Increased liver icus were higher or lower in Cu concentrations than Cu + Ca concentration of Cu may be related to elevated metal-bind- mixtures as compared to control groups. Ca exposure had ing protein synthesis. These findings were in accordance the capacity to decrease Cu toxicity in O. niloticus. with findings of Cogun and Kargin [5] which metal accu- mulation was the lowest in brain and the highest in liver and kidney. Kim and Kang [24] who reported that Sebastes 5. CONCLUSION schlegeli exposed to Cu showed more accumulation in liver than intestine, kidney, and muscle respectively. Cu accu- As is well known Cu is toxic to fish when exceeded the mulation was found to be time and concentration depend- lethal-dose. In this study, significant of Cu accumulation ent and in all tissues of fish elevated with increasing con- was determined among liver, kidney, and brain of O. nilot- centrations of Cu and exposure period [5]. icus. All tissues showed an alteration in antioxidant status Enzyme activities and GSH contents under the effects at different time intervals and exposures. This situation in- of metals indicate an increase or a decrease depending on dicated that Cu could be responsible for oxidative stress in the species, organ specificity, dose and route of exposure tissues. Furthermore mixture exposure of Cu + Ca produced
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antagonistic effect in comparation with Cu in GST activity, [13] Elia, A.C., Galarini, R., Taticchi, M.I. and Dorr, A.J.M. (2003) GSH level and Cu accumulation in all tissues during expo- Mantilacci L. Antioxidant responses and bioaccumulation in Ictalurus melas under mercury exposure. Ecotoxicol Environ, sure time. Results suggested that toxicity of Cu on tissues of 55, 162-167. fish can be reduced significantly with the application of Ca. [14] Monteiro, D.A., Rantin, F.T. and Kalinin, A.L. (2010) Inor- ganic mercury exposure: toxicological effects, oxidative stres biomarkers and bioaccumulation in the tropical freshwater fish ACKNOWLEDGMENTS matrinxa, Brycon amazonicus (Spix and Agassiz, 1829). Eco- toxicol, 19, 105–123. The research presented in this article was supported by [15] Muramoto, S. (1983) Elimination of copper from Cu-contam- inated fish by long-term exposure to EDTA and freshwater. J Research Fund (FEF2007D20) from Cukurova University Environ Sci Health A, 19 (3), 455-461. (Turkey). [16] Anderson, M.E. (1985) Determination of glutathione and glu- tathione disulfide in biological samples. Methods in Enzymol, The authors have declared no conflict of interest. 113, 548-555. [17] Habig, W. H., Pabst, M.J. and Jakoby, W.B. (1974) Glutathi- one S- transferases the first enzymatic step in mercapturic acid REFERENCES formation. J Biol Chem, 249, 7130-7139. [18] Lowry, O.H., Rosenbrough, N.J., Farr, A. and Randall, R.J. [1] Prego R. and Cobelo-Garcia, A. (2003) Twentieth century (1951) Protein measurement with the folin-phenol reagent. J overview of heavy metals in Galician Rias (NW Iberian Pen- of Biochem, 193, 265-277. insula). Environ Pollut, 121, 425-452. [19] Pagenkopf, GK. (1983) Gill surface ınteraction model for trace - metal toxicity to fishes: Role of complexation, pH, and water [2] Pelgrom S.M.G.J., Lock, R.A.C., Balm, P.H.M. and Wen- hardness. 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Aquat Toxicol, 70, 63–81. [4] Tao, S., Wen, Y., Long, A., Dawson, R. and Cao, J. (2001) Simulation of acid–base condition and copper speciation in [22] Wicklund, A., Runn, P. and Norrgren, L. (1988) Cadmium and fish gill microenvironment. Comp Chem, 25, 215–223. zinc interactions in fish: Effects of zinc on the uptake, organ distribution and elimination of Cd in the zebrafish, Branchy- [5] Cogun, H.Y. and Kargin, F. (2004) Effects of pH on the mor- danio rerio. Arch Environ Contam Toxicol, 17, 345-354. tality and accumulation of copper in tissues of Oreochromis niloticus. Chemosphere, 55, 277–282. [23] Kaviraj, A. and Dutta, T.K. (2000) Use of quick lime (CaO) as a means to reduce cadmium toxicity in common carp, Cypri- [6] Spry, D.J. and Wiener, J.G. (1991). Metal bioavailability and nus carpio. J of Applied Aquaculture, 10 (1), 87–95. toxicity to fish in low-alkalinity lakes: a critical review. Envi- ron Pollut, 71, 243–304. [24] Kim, S.G. and Kang, J.C. (2004) Effect of dietary copper ex- posure on accumulation, growth and hematological parameters [7] Wurts, W.A. and Perschbacher, P.W. (1994) Effects of Bicar- of the juvenile rockfish, Sebastes schlegeli. Marine Environ bonate Alkalinity and Calcium on the Acute Toxicity of Cop- Research, 58, 65–82. per to Juvenile Channel Catfish (Ictalurus punctatus). Aqua- culture, 125, 73-79. [25] Sanchez, W., Palluel, O., Meunier, L., Coquery, M., Porcher, J.M. and Aissa, S.A. (2005) Copper-induced oxidative stress [8] El-Sayed, A.M. (2006) Tilapia culture in salt water: Environ- in three-spined stickleback: relationship with hepatic metal mental requirements, nutritional implications and economic levels. Environ Toxicol and Pharmacol, 19, 177-183. potentials. Eighth Symposium on Advances in Nutritional Aq- uaculture. November, 15–17, Nuevo Leon, Mexico [26] Ali, M., Parvez, S., Pandey, S., Atif, F., Kaur, M., Rehman, H. and Raisuddin, S. (2004) Fly ash leachate induces oxidative [9] Mazon, A.F. and Fernandes, M.N. (1999) Toxicity and tissue stress in freshwater fish Channa punctata (Bloch). Environ accumulation of copper in the tropical freshwater fish, Proch- Int, 30, 933–938. ilodus scrofa (Prochilodontidae). Bull Environ Contam Toxi- col, 63, 797-804. [27] Lopes, P.A., Pinheiro, T., Santos, M.C., Mathias, M.L, Col- lares-Pereira, MJ. and Viegas-Crespo, A.M. (2001) Response [10] Sahin, E. and Gumuslu. S. (2004) Alterations in brain antioxi- of antioxidant enzymes in freshwater fish populations (Leucis- dant status, protein oxidation and lipid peroxidation in re- cus alburnoides complex) to inorganic pollutants exposure. sponse to different stress models. Behavioural Brain Research, Sci Total Environ, 280, 153-163. 155, 241–248. [28] Blanchette, B.N. and Singh, B.R. (2002) Isolation and charac- [11] Eaton, D.L. and Bammler, T.K. (1999) Concise review of the terization of Glutathione S-Transferase Isozyme Q3 from the glutathione S-transferases and their significance to toxicology. Northern Quahog, Mercinaria mercinaria. J of Protein Chem- Toxicol Sci, 49, 156–164. istry, 21, 151-159. [12] Pena-Llopis, S., Pena, J.B., Sancho, E., Fernandez-Vega, C. [29] Basha, P.S. and Rani, A.U. (2003) Cadmium-induced antioxi- and Ferrando, M.D. (2001) Glutathione-dependent resistance dant defense mechanism in freshwater teleost Oreochromis of the European eel Anguilla anguilla to the herbicide moli- mossambicus (tilapia). Ecotoxicol and Environ Safety, 56, nate. Chemosphere, 45, 671–681. 218–221.
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[31] Sies, H. (1999) Glutathione and its role in cellular functations. Free Radical Biol Medicine, 27, 916-921.
[32] Firat, O., Gok, G., Cogun, H.Y., Yuzereroglu, T.A. and Kargin, F. (2008) Concentrations of Cr, Cd, Cu, Zn and Fe in crab Charybdis longicollis and shrimp Penaeus semisulcatus from the Iskenderun Bay, Turkey. Environ Monitor and As- sess, 147, 117-123. [33] Paris-Palacios, S., Biagianti-Risbourg, S. and Vernet, G. (2000) Biochemical and (ultra) structural hepatic perturbations of Brachydanio rerio (teleostei, cyprinidae) exposed to two sublethal concentrations of copper sulfate. Aquat Toxicol, 50, 109-124.
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[37] Barron, M.G. and Albeke, S. (2000) Calcium Control of Zinc Uptake in Rainbow Trout.; Aquat Toxicol, 50, 257-264.
Received: January 24, 2015 Revised: June 11, 2015 Accepted: July 09, 2015
CORRESPONDING AUTHOR
Gülbin Firidin
Life Science Application and Research Center
Gazi University
06830 Ankara
TURKEY
Phone: +90 3124842157
Fax : +90 312 484 62 71
E-mail: [email protected]
FEB/ Vol 24/ No 11c/ 2015 – pages 4149 - 4156
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ANTIMONY DISTRIBUTION AND EFFECT FACTORS IN THE SURFACE SEDIMENT OF NANSI LAKE, CHINA
Tongguang Shi1, Lu Zhang2, Yuanxin Cao2 and Shiliang Wang2,*
1 Research Center of Remote Sensing and Geographic Information System, Shandong Jianzhu University, Jinan 250101, China 2 School of Geography and Tourism, Qufu Normal University, Rizhao 276826, China
ABSTRACT hazardous substances on the Basel Environment Conven- tion [1]. Antimony and its compounds were also listed as The objectives of this study were to determine the con- pollutants controlled with priority by European Union tent and chemical forms of antimony (Sb) and to investi- (EU) and the Environmental Protection Agency of the gate the effect of some mineral phases on its distribution in United States [5]. surface sediments of Nansi Lake. The results showed that In the past decades, Sb was usually overlooked as an the content of the total Sb (TSb) was in the range of 1.75- element of environmental concern and its study has been 6.29 mg/kg averaging 4.05 mg/kg. Results of a selective largely neglected, probably due to its lower abundance and sequential extraction of Sb in the sediment showed that the the relative insolubility of most of its compounds [6]. How- amorphous and well-formed crystalline Fe and Al hydrous ever, the Sb demand in the international market rose sharply oxides bound Sb comprised about 50% of the TSb. More- in recent years. In 2008, the world production of Sb was es- over, the level of amorphous Fe and Al hydrous oxides timated to 165,000 tones [2]. In fact, antimony has been bound Sb increased with the TSb content. In addition, the widely applied in numerous products including flameproof average percentage content of the Sb in residual phase was retardants, batteries, antifriction alloys, small arms and tracer also high (31.7%). The sediment in the upper lakes con- bullets, and cable sheathing [1]. Therefore, investigations on tained relatively high-level Sb, thus suggesting that anthro- the wide distribution of Sb in the environmental matrix have pogenic activities might be the main contributor of the con- received larger attention from the scientific community in tamination of Sb in the sediment of Nansi Lake. Regression recent years. For example, studies on the distribution of Sb analysis showed that TSb content and that of Fe and Al hy- in water [7], sediment or soil [8, 9, 10], atmosphere [11], drous oxides bound Sb content were positively and signif- and biota [8, 10, 12] had been conducted systematically. icantly correlated with the content of clay, Fe, and Al of the sediment, which indicated some significant effects of Natural processes and human activities induced the oc- the mineral phases containing clay, Fe and Al on the distri- currence of antimony in environment [4]. However, the Sb bution and environmental behavior of Sb in the sediments pollution originates mainly from mining and industrial of the lake. emission sources [13]. High levels of Sb pollution have been frequently detected around mining area [7, 10]. For
example, Sb concentrations in rivers around the antimony KEYWORDS: Antimony, Fractionation, Selectively sequential ex- mine of Xikuangshan in China were up to 6384 µg/L in traction, Sediment, Nansi Lake water samples and to 7316 mg/kg in sediment samples [9].
Beauchemin et al. [14] reported that the highest Sb concen- trations in an adit pool at a gold deposit in Canada could 1. INTRODUCTION reach 199 µg/L in water samples and 105 mg/kg in surface sediment samples. However, available data about the con- Antimony (Sb) is a metalloid belonging to group 15 of tents of Sb in soils/sediments away from the antimony min- the Periodic Table. It occurs naturally at trace levels in en- ing area are still scarce. vironment with various oxidation states (-III, 0, III, V) [1]. As the largest freshwater lake in the southwest Shandong The various speciation of Sb presents different toxicities, province, Nansi Lake (34°27′–35°20′N, 116°34′–117°21′E) which are following the general order of: antimonites (III) > consists of four smaller lakes Nanyang, Duanshan, Zhaoyang antimonates (V) > organoantimonials (e.g. methylated spe- and Weishan (Figure 1). It has received considerable amount cies) [2]. Studies showed that Sb is categorized as a non-es- of anthropogenic pollutants originating from industrial, agri- sential element for plants, animals, and humans but is toxic cultural and urban sources. Rich coal resource around Nansi to most organisms [3, 4]. In 1999, antimony was listed as Lake promoted the development of coal-related industry, such as smelting and coal-fired power plants. Large quan- * Corresponding author tities of wastewater from these industrial activities were
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FIGURE 1 - Location map of sampling sites in Nansi Lake.
discharged through rivers into Nansi Lake. Therefore, Nansi 2.2 Samples analysis Lake has been under serious contamination pressures in re- The total organic carbon (TOC) content of sediment cent years. The pollution status of metals [15, 16] and or- was determined with an elemental analyzer. The pH values ganic pollutants [17] in the sediments of Nansi Lake is re- of sediment were determined using a combined pH elec- ported by recent studies. However, there was no investiga- trode (Orion, USA) in a 1:10 solid: liquid ratio suspension. tion on the pollution by Sb in the sediments of Nansi Lake. The grain size distribution of the sediment samples was an- Therefore, the main goals of this study were: (1) to de- alyzed using a LS 230 laser diffraction particle analyzer termine the content and fractionation of Sb in the sediments (Beckman Coulter). The percentages of clay (< 2 μm), silt of Nansi Lake; and, (2) to investigate the effects of some (2-20 μm), and sand fractions (20-2,000 μm) were deter- mineral phases on the distribution and environmental be- mined. havior of Sb in the sediments of Nansi Lake. About 1.00 g of sediment sample was digested with a mixture of HNO3–HF–HClO4 (5:5:1) and the content of Fe, Al, Mn, and Ca was determined by ICP-AES (IRIS Intrepid 2. MATERIALS AND METHODS II, Thermo Electron Co., USA). Another aliquot of sedi- ment sample was digested by the mixed solution of aqua 2.1 Samples collection regia, 1% (w/v) KMnO4 and 1% (w/v) oxalic acid. Hydride In this study, twenty surface sediment samples were generation - atomic fluorescence spectroscopy (HG-AFS) collected (0-15 cm) in April 2013 from various sites in (HG-AFS230, Beijing Haiguang Instruments, Inc., China) Nansi Lake (Figure 1). At each site, the samples were taken was used to measure the total Sb (TSb) concentration in the from five different locations using cable operated sediment digested solutions. samplers (Van Veen grabs, Eijkelkamp). They were mixed The accuracy of the Fe, Al, Mn, and Ca concentrations together and placed in dark-colored polyethylene bags. Then was checked by conducting an analysis of three kinds of they were taken to the laboratory immediately, freeze-dried reference materials (GSS1, GSS2, and GSS9) obtained (FD-1A, China), slightly crushed, passed through a two-mil- from the Institute of Geophysical and Geochemical Explo- limeter sieve, and stored at 4 oC in glass bottles before anal- ration, Chinese Academy of Geological Science. The aver- ysis. age relative errors of Fe, Al, Mn, and Ca concentrations in
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these reference materials were -2.12~1.67%, -0.72~1.43%, of Sb were 3.56, 5.16, 2.48, and 4.27% for F1, F2, F3, and -0.58~1.65%, and 1.47~2.14%, respectively. The preci- F4, respectively. sions were evaluated by the variation of coefficients (CVs) of two replicates from six sediment samples. Average CVs were 2.15, 1.25, 1.42, and 1.53% for Fe, Al, Mn, and Ca, 3. RESULTS AND DISCUSSION respectively. The accuracy of the TSb was checked by con- ducting an analysis of a certified reference material (BCR- 3.1 Basic properties of the sediment CRM 277b). The differences between the certified and the- Basic physico-chemical properties of the sediment in- oretical values were under 7%. cluding the content of Fe, Al, Mn, Ca, P, and TOC, the val- ues of pH, and the percentages of clay, silt, and sand are 2.3 Sequential extraction analysis of Sb in the sediment summarized in Table 2. The TOC content ranged from 0.1 In this study, the five-step sequential extraction proce- to 1.19% in the sediment. The percentages of clay, silt, and dure for arsenic (As) described by Wenzel et al. [18] was sand ranged from 1.34 to 17.37%, from 9.13 to 51.68%, adopted for the chemical operationally-defined forms ex- and from 36.7 to 88.42%, respectively, with average values traction of Sb. The specific extraction procedures are of 9.39, 32.31 and 54.03%, with no large difference among shown in Table 1. The four chemical forms including non- the four lakes. The average content of Al, Fe, Ca, P, and Mn specifically sorbed Sb (F1), specifically-sorbed Sb (F2), in the sediment was 67.48, 27.24, 11.92, 0.53, and 0.63 g/kg, amorphous and poorly-crystalline Fe and Al hydrous ox- respectively, which were higher than that of Mn (0.53 g/kg), ides bound Sb (F3), and well-formed crystalline Fe and Al but lower than that of Al (77.44 g/kg), Fe (30.89 g/kg), Ca hydrous oxides bound Sb (F4) were sequentially and selec- (29.45 g/kg), and P (0.66 g/kg) in the upper continental tively extracted by (NH4)2SO4, NH4H2PO4, NH4-oxalate, crust [19]. and NH4-oxalate + ascorbic acid solutions, respectively (Table 1). The content of residual Sb (F5) was determined 3.2 Total Sb content in the sediment by calculating the difference between the TSb content and The TSb content in the sediments of Nansi Lake ranged the sum contents of the four fractions described above. The from 1.75 to 6.29 mg/kg, with an average value of 4.05 mg/kg suspensions were centrifuged at 10,000 rpm for 10 min at (Table 3, Figure 2). The TSb content of the stations located in room temperature using a Xiang Yi centrifuge in each step. the Dushan Lake was generally higher than that of stations at All the above supernatants were filtered through a 0.45 µm other lakes. For example, the sediment contained the high- membrane and analyzed by HG-AFS. All reagents were est level of TSb (6.29 mg/kg) at site S10 located at the Du- analytical grade. The precision of the sequential extraction shan Lake. In addition, the TSb levels of the sediment at method was evaluated by the variation of coefficients (CV) sites S8, S9, S11, and S12 were much higher than those of of ten replicates from five sediment samples. Average CVs other sites. The sediment at site S18 located at the Weishan
TABLE 1 - Steps of the selective sequential extraction procedure [18].
Step Fraction Extractant Experimental conditions
1 Non-specifically sorbed Sb 0.05 M (NH4)2SO4 4 h shaking (20 °C)
2 Specifically-sorbed Sb 0.05 M NH4H2PO4 16 h shaking (20 °C)
3 Amorphous and poorly-crystalline 0.2 M NH4-oxalate (pH 3.25) 4 h shaking in the dark (20 °C) Fe and Al hydrous oxides bound Sb wash step: 0.2 MNH4-oxalate 10 min shaking (20 °C)
o 4 Well-crystallised Fe and Al hydrous 0.2 M NH4-oxalate+0.1M ascorbic acid (pH 3.25) 30 min shaking in water bath at 96 C oxides bound Sb wash step: 0.2 M NH4-oxalate 10 min shaking (20 °C)
TABLE 2 - General properties of the sediments in Nansi Lake.
Site TOC Clay Silt Sand Al Fe Ca P Mn TSb % % % % g/kg g/kg g/kg g/kg g/kg mg/kg Average 0.65 9.39 32.31 54.03 67.48 27.24 11.92 0.53 0.63 4.05 Stda 0.38 4.28 10.51 14.05 11.90 11.73 3.25 0.14 0.25 1.48 CV%b 59 46 33 26 18 43 27 26 39 36 Minimum 0.10 1.34 9.13 36.70 37.74 6.65 7.49 0.23 0.26 1.75 Maximum 1.19 17.37 51.68 88.42 79.68 41.91 20.23 0.67 0.97 6.29 a Standard deviation b Coefficient of variance
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Lake contained the lowest level of TSb (1.75 mg/kg). More- by the effluents. After sedimentation and retention in the up- over, the TSb levels of the sediment at the site S19 and S20 per lake, sediments in the lower lake are consequently at a were relatively low. The spatial distribution tendencies corresponding lower contamination level. Moreover, the showed that the TSb level of the sediment was relatively faster development of aquaculture in Dushan Lake in com- higher in the upper lakes including Nanyang, Zhaoyang, and parison to other lakes might also contribute to higher level Dushan than that in the lower lake Weishan. Annual large of contaminants. Therefore, the TSb level of sediments in amount of domestic sewage and industrial wastewater from the upper lake is higher than that of the lower lake. In addi- the Jining, Zaozhuang, and Heze cities discharged into the tion, there were some large smelting plants and coal-fired lake through the rivers flowing into Nanyang and Dushan power plants nearby Nansi Lake, which might produce Lakes. Water flows from the upper lake to the lower lake emission of Sb pollutants. Thus, the deposition of atmos- under natural conditions in Nansi Lake. Therefore, some pheric Sb from coal-consuming industries may be the po- inorganic and organic pollutants are deposited firstly in the tential source of the Sb in the sediments of Nansi Lake. upper lake. The sediments of the upper lake are affected first
TABLE 3 - Basic statistical parameters of various Sb chemical forms in the sediments of Nansi Lake.
Sb concentration (mg/kg) Sb proportions (%) Sites F1 F2 F3 F4 F5 TSb F1 F2 F3 F4 F5 Average 0.06 0.84 1.33 0.69 1.13 4.05 1.43 19.11 31.73 16.07 31.67 Stda 0.03 0.47 0.62 0.35 0.21 1.48 0.59 5.35 4.78 4.50 12.54 CV% 49 56 46 51 19 36 41 28 15 28 40 Minimum 0.01 0.20 0.45 0.15 0.73 1.75 0.66 11.15 21.41 6.89 17.39 Maximum 0.11 1.57 2.41 1.26 1.56 6.29 2.87 28.55 38.30 23.28 53.88 a Standard deviation b Coefficient of variance
7 (a) F1 F2 F3 F4 F5 6
5
4
3
Sb conc. (mg/kg) Sb conc. 2
1
0 S18 S17 S19 S20 S16 S15 S1 S14 S13 S2 S3 S7 S5 S8 S4 S6 S12 S11 S9 S10 Sampling sites (b) 100% 90% 80% 70% 60% 50% 40%
Sb fraction (%) 30% 20% 10% 0% S18S17S19S20S16S15 S1 S14S13 S2 S3 S7 S5 S8 S4 S6 S12S11 S9 S10 Sampling sites
FIGURE 2 - Absolute partition patterns (a) and relative partition patterns (b) of Sb among sediment fractions in the sediments of Nansi Lake.
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The average content of Sb in the upper continental forms followed the order of F3 (31.73%) > F5 (31.67 %) > F2 Earth’s crust is 0.4 mg/kg [20]. The background content of (19.11%) > F4 (16.07%) > F1 (1.43%). The average percent- Sb in the world sediment [22], in Chinese soil [21], and in age of Fe and Al hydrous oxides bound Sb (F3 and F4) Shandong province soil [21] is 1.2, 1.06, and 0.9 mg/kg, comprised about 50% the TSb content. The Sb content in F3 respectively. Therefore, the TSb concentrations at all the of the sediment increased with the TSb content (Figure 2). sampling stations were higher than the values mentioned These results confirm the crucial role of Fe and Al hydrous above. It can be concluded from these results that anthro- oxides in immobilizing Sb in superficial environments. Fe pogenic activities might have been the main factor contrib- and Al hydrous oxides have an important effect on the en- uting to the high level of Sb in the sediments of Nansi Lake. vironmental behavior of Sb in sediment, due to their abun- The sediments around the mining regions generally have dance, large specific surface area, and strong binding affin- high levels of Sb. For example, the contents of Sb range ity [2, 24, 25, 29]. Anthropogenic sources of Sb might from 57 to 7316 mg/kg in sediments and from 100 to 5045 firstly bind to amorphous and poorly-crystalline Fe and Al mg/kg in soil in the Xikuangshan area in China, respec- hydrous oxides. These results were consistent with that of tively [8, 9]. In abandoned antimony mine, the contents of an element of the same group As. Tighe et al. [24] reported Sb in soils range from 19 to 4400 mg/kg in Italy and from that 30-47% of total Sb was bind to non-crystalline Al and 5 to 40 mg/kg in Spain, respectively [7, 10]. However, Fe hydrous oxides in soil collected from Australia. Müller studies on the content of Sb in non-mine area were scarce. et al. [25] reported that an average of 90% of TSb is bound However, the sediments contain relatively low content of to amorphous and crystalline Al and Fe oxides in soil col- Sb away from the mining region. For example, Lin et al. lected from Scotland. [22] showed that the Sb content was in the range of 0.13- The residual fraction of soils and sediments is usually 1.77 mg/kg in the sediments of the Daliao River system. determined with a strong acid digestion and generally rep- The content of Sb ranged from 0.50 to 2.70 mg/kg in the resents the portion of the unavailable metal or metalloid in sediment of the Yangtze River system [13]. soil or sediment [2]. This fraction could possibly contain Sb attached to resistant sulfides or pyrite. There are many 3.3 Chemical forms of Sb in the sediment factors influencing the content of this fraction, such as con- Single or sequential chemical soil extractions are often taminant source, pH, redox, and competing ions. Various used to deduce the Sb association with specific phases such levels of this fraction were reported by some previous stud- as minerals or organic matter [2]. The information contrib- ies. For example, He et al. [8] reported that the majority utes to the comprehensive understanding of potential met- (88.2-97.9%) of Sb was in residual fraction in the soil from alloid availability or mobility under specific environmental the Xikuangshan mine whereas Wang et al. [9] reported conditions. The extraction procedure of the Sb forms in the that 5.7-53.5% of the TSb was in the residual fraction in sediments of Nansi Lake is shown in Table 3. the sediment around the Xikuangshan mine. In addition, As shown in Table 3, the Sb content in operationally- exchangeable or weakly bound Sb (F1) contributes only to defined fractions F1, F2, F3, F4, and F5 were in the range a small proportion of the TSb in sediment, which is in ac- of 0.01-0.11, 0.20-1.57, 0.45-2.41, 0.15-1.26, and 0.73- cordance with previous studies [8, 9]. 1.56 mg/kg, respectively. The average percentage of Sb in five
TABLE 4 - Pearson correlation coefficients between the TSB and various forms of Sb and the mineral phases in the sediments of Nansi Lake.
TOC Clay Silt Sand Al Fe Ca P Mn TSb F1 F2 F3 F4 F5 TOC 1 Clay 0.712** 1 Silt 0.221 0.465* 1 Sand -0.450* -0.727** -0.886** 1 Al 0.892** 0.566** 0.135 -0.310 1 Fe 0.980** 0.650** 0.204 -0.408 0.948** 1 Ca -0.119 -0.178 -0.212 0.237 -0.368 -0.211 1 P -0.034 -0.039 0.257 -0.210 0.069 0.020 -0.318 1 Mn -0.141 -0.110 0.362 -0.213 0.049 -0.068 -0.336 0.829** 1 TSb 0.871** 0.788** 0.350 -0.592** 0.842** 0.872** -0.274 0.032 -0.011 1 F1 0.597** 0.650** 0.340 -0.461* 0.608** 0.607** -0.322 0.372 0.352 0.722** 1 F2 0.866** 0.752** 0.238 -0.531* 0.819** 0.865** -0.256 0.021 -0.060 0.965** 0.703** 1 F3 0.808** 0.801** 0.390 -0.631** 0.768** 0.794** -0.316 0.093 0.041 0.981** 0.758** 0.937** 1 F4 0.792** 0.756** 0.456* -0.626** 0.790** 0.798** -0.380 -0.005 0.073 0.932** 0.681** 0.869** 0.933** 1 F5 0.396** 0.143 -0.028 -0.002 0.419 0.431 0.253 -0.138 -0.231 0.319 -0.005 0.234 .0.185 0.093 1 **.Correlation is significant at the 0.01 level. *.Correlation is significant at the 0.05 level
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7 (a) (b)7 y = 0.1097x + 1.0583 6 y = 0.1044x - 3.0023 6 R² = 0.7102 R² = 0.7609 5 5
4 4
3 3 TSb (mg/kg) TSb (mg/kg) 2 2
1 1
0 0 0 204060801000 1020304050 Al (g/kg) Fe (g/kg) 7 7 (c) y = 3.3541x + 1.8666 (d) y = 0.2713x + 1.4986 6 R² = 0.7596 6 R² = 0.6211
5 5
4 4
3 3 TSb (mg/kg) TSb (mg/kg) 2 2
1 1
0 0 0 0.2 0.4 0.6 0.8 1 1.2 1.4 0 5 10 15 20 TOC (%) Clay (%) FIGURE 3 - Correlations between the content of TSb and the content of Al, Fe, TOC, and clay in the sediments of Nansi Lake: (a) TSb vs. Al, (b) TSb vs. Fe, (c) TSb vs. TOC, (d) TSb vs. clay.
3.4 Correlations analysis between mineral phases and Sb in [30]. For instance, Newton et al. [31] observed that both the sediment lead (Pb) and arsenic (As) could bind tightly to soil humic The nature on the mineral phases was one of the most acid molar mass fractions in soil of a contaminated apple important factors impacting the retention of Sb in soils or orchard by employing size exclusion chromatography-in- sediments [2]. Understanding the association of Sb with ductively coupled plasma mass spectrometry SEC-ICP- mineral phase is fundamental for investigating the distribu- MS. It was shown that Sb was bound to soil-derived humic tion and environmental behavior of Sb. Therefore, the cor- acid mass fractions in a shooting range soil also by employ- relations between TSb, the chemical forms of Sb and the ing SEC-ICP-MS [32]. In addition, Steely et al. [33] re- mineral phases in the sediments of Nansi Lake are pre- ported the importance of soil humic acids in converting in- sented in Table 4 and Figure 3. organic Sb (III) species to a less toxic species [i.e., Sb (V)] The results showed that TOC, clay, Fe, and Al are pos- as well as limiting mobile antimony in contaminated soils itively and significantly correlated with the TSb and the by complexing with it. each forms of Sb in the sediments of Nansi Lake, respec- These investigations suggest that Fe, Al, and Mn ox- tively. Therefore, the basic mineral phases of sediment ides and hydroxides and TOC have an important effect on strongly affect the distribution and behavior of Sb in the the motivation and transformation of Sb in soil and sedi- sediments of Nansi Lake. ment. Previous investigation has certified that Sb was bound to Fe hydroxides by employing X-ray absorption and pho- toelectron spectroscopy [26]. Fe and Al oxides and hydrox- 4. CONCLUSIONS ides could sorb Sb in soil or sediment [2]. Formation of edge-sharing inner-sphere sorption complexes at the goe- This study determined the content of Sb in the sedi- thite surface in the case of Sb (V) and the formation of a ments of Nansi Lake. The total content ranged from 1.75 bidentate, corner-sharing complex for Sb (III) were found to 6.29 mg/kg, and averaged 4.05 mg/kg. The average pro- [27]. Thanabalasingam et al. [28] reported that Sb (III) sorb portion of Sb in five operationally-defined chemical forms to oxides and hydroxides, with the sorption order of: followed the order of: F3 (31.73%)> F5 (31.67 %)> F2 MnOOH > Al (OH)3> FeOOH. In addition, Belzile et al. (19.11%)> F4 (16.07%)> F1 (1.43%). Antimony was [29] reported that amorphous Fe and Mn oxyhydroxides mostly associated with Al and Fe hydrous oxides (47%), could oxidize Sb (III) to Sb (V). followed by the association with a residual and refractory Organic matter such as fulvic and humic acids could fraction (31.67%). The sediment in the upper lakes con- affect the environmental behavior of most trace elements tained relatively higher level of Sb than that in the lower
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lake. Industrial emissions might be the main contributor of [12] Okkenhaug, G., Zhu, Y.G., Luo, L., Lei, M., Li, X. and the high content of Sb in the sediments of Nansi Lake. The Mulder, J. (2011) Distribution, speciation and availability of antimony (Sb) in soils and terrestrial plants from an active Sb mineral phases of sediments including the content of Fe, mining area. Environmental Pollution 159, 2427-2434. Al, clay, and TOC have an important effect on the distribu- [13] He, M.C., Wang, X.Q., Wu, F.C. and Fu, Z.Y. (2012) Anti- tion and environment behavior of Sb in the sediments of mony pollution in China. Science of the Total Environment Nansi Lake. 421, 41-50.
[14] Beauchemin, S., Kwong, Y.T., Desbarats, A.J., MacKinnon, T., Percival, J.B., Parsons, M.B. and Pandya, K. (2012) Down- ACKNOWLEDGMENTS stream changes in antimony and arsenic speciation in sedi- ments at a mesothermal gold deposit in British Columbia, Can- This study is supported by the Encouraging Founda- ada. Applied Geochemistry 27, 1953-1965. tion for Outstanding Youth Scientists of Shandong Prov- [15] Cao, X.Z., Shao, Y., Deng, W.J., Wang, H. and Wang, S. ince (No.BS2012HZ006), the National Nature Science (2014) Spatial distribution and potential ecologic risk assess- Foundation of China (No.41301532), and the First Class ment of heavy metals in the sediments of the Nansi Lake in China. Environmental Monitoring and Assessment 186, 8845- General Financial Grant from the China Postdoctoral Sci- 8856. ence Foundation (No.2013M540103). [16] Wang, S.L., Lin, C.Y., Cao, X.Z. and Zhong, X. (2012) Arse- nic content, fractionation, and ecological risk in the surface The authors have declared no conflict of interest. sediments of lake. International Journal of Environmental Sci- ence and Technology 9, 31-40. [17] Zhang, G., Pan, Z., Bai, A., Li, J. and Li, X. (2014) Distribu- tion and bioaccumulation of organochlorine pesticides (OCPs) REFERENCES in food web of Nansi Lake, China. Environmental Monitoring and Assessment 186, 2039-2051. [1] Filella, M., Belzile, N. and Chen, Y.W. (2002) Antimony in the environment: a review focused on natural waters: I. Occur- [18] Wenzel, W.W., Kirchbaumer, N., Prohaska, T., Stingeder, G., rence. Earth-Science Reviews 57, 125-176. Lombi, E. and Adriano, D.C. (2001) Arsenic fractionation in soils using an improved sequential extraction procedure. Ana- [2] Wilson, S.C., Lockwood, P.V., Ashley, P.M. and Tighe, M. lytica Chimica Acta 436, 309-323. 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[29] Belzile, N., Chen, Y.W. and Wang, Z. (2001) Oxidation of an- timony (III) by amorphous iron and manganese oxyhydrox- ides. Chemical Geology 174, 379-387.
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Received: March 18, 2015 Revised: July 20, 2015 Accepted: August 12, 2015
CORRESPONDING AUTHOR
Dr. Shiliang Wang School of Geography and Tourism Qufu Normal University Rizhao 276826 P.R. CHINA
Phone: +86 633 3980719 Fax: +86 633 3980719 E-mail: [email protected]
FEB/ Vol 24/ No 11c/ 2015 – pages 4157 - 4164
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EVALUATION OF CONTAMINATION DATA WITH NON-DETECTS USING CENSORED DISTRIBUTIONS
Michal Fusek1,*, Jaroslav Michálek2, and Milada Vávrová3
Brno University of Technology, Faculty of Electrical Engineering and Communication, Technická 2848/8, 616 00 Brno, Czech Republic University of Defence, Faculty of Economics and Management, Šumavská 4, 662 10 Brno, Czech Republic Brno University of Technology, Faculty of Chemistry, Purkyňova 464/118, 612 00 Brno, Czech Republic
ABSTRACT Generally, musk compounds can be divided into four groups: linear, macrocyclic, polycyclic and nitro musk When measuring concentration of chemical compounds, compounds. The last two groups are used most frequently we often have to deal with a situation when the resulting val- as substitutes for natural musks in fragrances and personal ues are found below the limit of detection or limit of quanti- hygiene products [2]. Galaxolide (HHCB) and tonalide fication of the determination method. In order to statistically (AHTN) are examples of the most important polycyclic evaluate such data, the method of maximum likelihood con- musk compounds. Musk xylene, musk ketone and musk sidering doubly left-censored samples is applied. As a model ambrette are well-known nitro musk compounds (i.e. com- distribution of measured concentrations, Weibull distribu- pounds containing one or more nitro groups in a molecule). tion is considered. Moreover, considering the asymptotic The production of nitro musk compounds, that are gener- properties of maximum likelihood estimates, concentra- ally included in a group of substances posing a risk to the tions of chemicals can be compared using Wald’s test environment, has decreased over the last years [3, 4]. By based on the expected Fisher information matrix. Here we contrast, production of polycyclic synthetic aromatic sub- show that the described statistical method allows for a bet- stances, which are less toxic, has increased because of their ter evaluation of the obtained experimental data than com- frequent use as additives in many personal care products, monly used methods where all values below the detection e.g. soaps, shampoos, deodorants, body lotions, perfumes, limits are replaced by a constant. These methods are used cleaning and disinfecting agents, air fresheners and indus- for an analysis of the worldwide commonly used synthetic trial cleaning agents (see e.g. [5]). Synthetic aromatic sub- musk compounds (nitro and polycyclic) which were ex- stances were also detected in samples of air and dust col- tracted from the fish samples caught upstream (Group 1) and lected in indoor environments [6]. They often penetrate downstream (Group 2) from a high-capacity wastewater into the environment through wastewater because of their treatment plant. ineffective removal in the wastewater treatment plant (WWTP), see e.g. [7] and references inside. Accumulation of these substances in the environment (surface water, sed- iment) results in their occurrence in food chain, especially KEYWORDS: Doubly left-censored sample, maximum likelihood, in aquatic ecosystems. A number of studies revealed the musk compound, Wald’s test, Weibull distribution. presence of musk compounds in tissues of aquatic animals.
These compounds can also be found in human body, for
example in fat tissue, human milk and blood plasma (see 1. INTRODUCTION e.g. [8, 9]), as a consequence of fish consumption. This contribution is focused on introduction of ad- Synthetic aromatic substances or musk compounds are vanced statistical methods and their use for analyzing musk lipophilic contaminants able to accumulate in various com- compound concentrations in fish tissue. These methods are ponents of the environment. They represent a group of per- then compared with a standard approach. Moreover, the in- sistent pollutants, and may occur in environmental matri- fluence of the WWTP on concentration of musk com- ces and human tissues. Synthetic aromatic substances were pounds in fish muscle is assessed. launched on the market in the early 20th century and the volume of their production has significantly increased over the last 50 years [1]. Since they have potential carcinogenic 2. MATERIALS AND METHODS properties, efforts are currently being made to limit or pro- hibit their use in many regions worldwide. 2.1 Material and Equipment The sample of aquatic biota consists of 60 fish from the * Corresponding author carp family, specifically of the European chub (Leuciscus
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cephalus), which were caught in the Svratka River, Czech LOD is the lowest concentration of a substance that can be Republic, near the WWTP Brno-Modřice by Morava River distinguished from the absence of that substance in a sam- Basin Administration employees. Fish were caught on 10th ple, and LOQ is the lowest concentration at which we can November 2009; half of them came from a watercourse up- reasonably tell the difference between two different values stream (Group 1), and half of them from a watercourse of concentration. In this paper, detection limits were calcu- downstream (Group 2) from the WWTP. The fish were lated using calibration curves of particular analytes, see transported to the laboratory of the Institute of Veterinary [12] for more information. In the following text, the term Hygiene and Ecology of Veterinary and Pharmaceutical “detection limits” refers to both LOD and LOQ. Since two University in Brno, and examined by a veterinarian. Rele- fixed detection limits are present and known (see Table 1), vant characteristics were noted and then muscle, skin and it is necessary to work with doubly left-censored samples guts were separated. Muscle tissue was selected for the and type I censoring. musk compound analysis because it is considered to be rep- resentative of all of the body. The muscle tissue was ho- mogenized (using a blender), subsequently frozen at -20 ◦C 0.04 and kept frozen until the analysis. Fish of approximately 0.035 the same age were chosen for the analysis. For each analysis, 3 grams of defrosted muscles were 0.03 used, triturated and mixed with 5.5 g of PSE matrix. This 0.025 mixture was quantitatively transferred into extraction ves- sel and the analyte was extracted by pressurized solvent ex- 0.02 traction under optimized conditions (cyclohexane : ethyl 0.015 ◦ acetate [25:75] extraction agent, temperature 120 C, pres- Normalized frequency sure 140 bar, 4 min static phase, 3 cycles, 15 s solvent flush 0.01
[used twice], 1 min drying by nitrogen), see [10] for more 0.005 details. This type of extraction is commonly used when an- alyzing pesticides and other organic pollutants contamina- 0 0 LOD LOQ tion in sediments [11]. The obtained extract was reduced to Concentration [μg/kg] 2 ml on rotary evaporator. Pressurized solvent extraction was followed by adsorption column chromatography to pu- FIGURE 1 - Histogram of galaxolide concentration (upstream the WWTP) with Weibull density and parameters λ = 23.98, τ = 1.61, rify samples using Florisil as the adsorbent and ethyl ace- LOD = 8.95 µg/kg, LOQ = 29.83 µg/kg. tate as the eluent. The volume of ethyl acetate was 80 ml and the flow rate was 6 drops per minute. Afterwards, the Various censoring techniques and statistical analyses sample volume was reduced on rotary evaporator. The final of censored data are described in more details in many sample was concentrated by 30 µl of undecan, and 770 µl monographs, e.g. [13, 14]. In many environmental studies, of cyclohexane was added; therefore, the final volume of left censoring is based on normal distribution [15, 16]. sample transferred to a vial was 800 µl. Analysis by GC- However, the distribution of variable such as concentration MS on GC Agilent 6890N followed. The capillary column is positive and skewed to the right (see Fig. 1 for an exam- DB-5MS (20 m × 0.18 mm × 0.18 µm) was used with he- ple), thus the normal distribution is not the most suitable lium as carrier gas, 1 min splitless time and transfer line choice. On that account, methods for dealing with doubly temperature 285 ◦C. Temperature program started at 50 ◦C, left-censored samples from exponential distribution [17] held for 1 minute, followed by 15 ◦C-per-minute ramp up and Weibull distribution [18] considering type I censoring to 110 ◦C and 5 ◦C-per-minute ramp up to 165 ◦C, at which were developed. A novelty of these methods lies primarily it was held for 8 minutes. Finally, 15 ◦C-per-minute ramp in the analytically determined (expected) Fisher information up to 285 ◦C, at which it was held for 2 minutes, followed. matrix (FIM) which is necessary for calculating variability As a result of the analysis, four nitro (musk ambrette [AMB], of the estimated parameters by the maximum likelihood (ML) musk xylene [MX], musk tibetene [TIB], musk ketone [MK]), method and determination of the Wald’s test statistic (7). At and four polycyclic musk compounds (phantolide [PH], trase- first the musk compound concentration was modeled using olide [TR], galaxolide [HHCB], tonalide [AHTN]) were de- the exponential distribution [19]. It was shown that use of tected. exponential distribution (which is a special case of Weibull distribution) is sufficient in some cases (musk ambrette, 2.2 Data and Censoring musk tibetene, phantolide, traseolide). Therefore, these com- When analyzing musk compounds and chemical data pounds will be omitted from the analysis. However, it was in general, we often have to deal with a situation when the impossible to use exponential distribution for modeling con- substance is either absent, or exists at such a low concen- centrations of the remaining musk compounds. Therefore, tration that it is not present above the detection limit level. doubly left-censored Weibull distribution will be considered Chemical analyses do not provide precise determination of because of its flexibility. In this paper, the one-sample the concentrations when values are found below the limit Weibull model based on [18] will be enhanced to the two- of detection (LOD), or limit of quantification (LOQ). The sample model. The suitability of a particular distribution
4166 © by PSP Volume 24 – No 11c. 2015 Fresenius Environmental Bulletin
TABLE 1 - Pearson’s χ2 goodness-of-fit test for Weibull distribution; H denotes the null hypothesis.
Compound Upstream the WWTP Downstream the WWTP LOD LOQ H p-value H p-value [µg/kg] [µg/kg] HHCB not rejected 0.10 not rejected 0.31 8.95 29.83 AHTN not rejected 0.99 rejected 0.01 1.98 6.62 MX not rejected 0.87 not rejected 0.97 0.75 2.50 MK not rejected 0.52 not rejected 0.29 0.57 1.90
can often be derived from earlier studies. Here the suitabil- ML estimates and ̂ of parameters and were obtained ity of Weibull distribution was tested using Pearson’s by maximization of the log-likelihood function (2) using goodness-of-fit test in Matlab (version 8.1, R2013a). It can Nelder-Mead simplex algorithm, which is implemented in be seen from Table 1, that Weibull distribution can be used Matlab environment (version 8.1, R2013a). An optimiza- for modeling of galaxolide, musk xylene and musk ketone tion procedures based on the EM algorithm (see e.g. [20, concentration because the null hypothesis is not rejected at 21]) can also be used, however, it may converge slowly, a significance level of 0.05. It is well known that good- especially in case of a high censoring level. ness-of-fit test has quite a low power and the smaller the In order to determine variances of the estimated pa- size of the sample, the lower the power is. However, in case rameters and ̂, the expected FIM which is derived in [18] the distribution of data is not far from Weibull distribution is used. The FIM of the doubly left-censored Weibull dis- (which is possible considering the test results), it is possible tribution is to utilize the flexibility of Weibull distribution and proceed from Weibull approximation. Moreover, a correspondence , , (3) between the histogram and Weibull density suggests suita- bility of Weibull distribution for modeling of musk com- where the analytical forms of FIM elements are rather pounds concentration (see Fig. 1 for an example). complicated and can be found in [18]. 2.3 Statistical Processing Method Using the asymptotic properties of ML estimator ( ̂ respectively), according to [22], 2.3.1 Maximum Likelihood Estimation of the One-Sample Weibull Model √ (√ ̂ respectively) has asymptoti- cally normal distribution N 0, (N 0, respectively), Let ,…, be a type I doubly left-censored random sample from Weibull distribution with scale parameter where ( respectively). Considering the 0, shape parameter 0, and cumulative distribution gamma function Γ, the expected value , Γ 1 function (cdf) 1/ and variance σ λ, τ Γ 1 2/ λ, τ of the Weibull distribution can be calculated. Since λ, τ 1 exp , 0, , , (1) and σ λ, τ are continuous functions, the parameters in 0, 0. both formulas are replaced by their ML estimates , ̂ ob-
Furthermore, let ,…, be the ordered sample of tained from the censored samples. Considering the asymp- totic normality of , the lower (LCL) and upper ,…, . For simplicity, in all the formulas, detection lim- , ̂ its will be denoted as LOD , LOQ , and we put (UCL) 1 α % confidence limits for estimate μ can be d 0. Moreover, is the number of observations below calculated as the , is the number of observations in the interval , , and is the number of uncensored observations UCL ̂ , ̂ Var μ λ,τ , ,…, . LCL ̂ , ̂ Var μ λ,τ , Using the results from [13], the log-likelihood function of the censored sample is given by where / is the 1 /2 quantile of the standard