مجله بیوتکنولوژی کشاورزی

مجله بیوتکنولوژی کشاورزی

فیلوژنی قارچ Aspergillus clavatus جداشده از مدارس ابتدایی عراق و بررسی توانایی آن در تولید پاتولین

نوع مقاله : مقاله پژوهشی

نویسندگان
میسون م. الجبوری 1
احسان علی الزاملی 2
ماجد کاظم الشبلی 3
1 اداره آموزش استان مثنی، عراق.
2 گروه زیست‌فناوری کاربردی، دانشکده زیست‌فناوری، دانشگاه سبز القاسم، بابل 51013، عراق
3 گروه زیست‌شناسی، دانشکده علوم تربیتی، دانشگاه القادسیه، عراق
10.22103/jab.2026.26476.1817
چکیده
هدف: مدارس از مهم‌ترین محیط‌های زندگی کودکان به شمار می‌روند و کودکان مدت زمان قابل‌توجهی را در مدرسه سپری می‌کنند. با این حال، آلودگی هوای داخل مدارس می‌تواند خطرناک باشد. اثرات بهداشتی خاص قارچ‌ها در محیط‌های بسته می‌تواند منجر به مشکلات حاد سلامت، به‌ویژه در میان دانش‌آموزان، شود. مطالعه حاضر در مدارس ابتدایی شهر سماوه انجام شد که در آن چندین مدرسه به‌صورت تصادفی انتخاب شدند و جداسازی قارچ‌ها از کف، هوا، دستگیره درها و نیمکت‌ها به روش جداسازی مستقیم انجام گرفت. سپس شناسایی قارچ‌ها با استفاده از روش‌های سنتی و همچنین تکنیک PCR صورت پذیرفت. هدف این مطالعه، ثبت Aspergillus clavatus به‌عنوان عامل اتیولوژیک در مدارس عراق و بررسی وجود ژن PatA به‌عنوان ژن مسئول تولید سم پاتولین بود.
مواد و روش‌ها: نمونه‌برداری از کف، دستگیره‌ها، نیمکت‌ها و هوا انجام شد. نمونه‌ها با استفاده از سوآپ‌های پنبه‌ای و به روش جداسازی مستقیم جمع‌آوری گردیدند. به‌منظور خالص‌سازی محصولات تکثیر مورد نیاز برای تعیین توالی، از سیستم Prep-a-Gene شرکت Bio-Rad Laboratories و آگاروز با نقطه ذوب پایین استفاده شد. تعیین توالی DNA با روش دیدئوکسی انجام گرفت.
نتایج: قارچ‌های جداشده به پنج جنس شامل Aspergillus، Penicillium، Alternaria، Rhizopus و Cladosporium تعلق داشتند که جنس Aspergillus غالب بود و شامل ۲۰ ایزوله و پنج گونه Aspergillus niger، A. flavus، A. candidus، A. terreus و A. clavatus می‌شد. جنس Penicillium در رتبه دوم قرار داشت و با هفت ایزوله شامل Penicillium chrysogenum، Penicillium digitatum و P. candidus شناسایی شد. جنس Alternaria با چهار ایزوله شامل A. alternata و A. oryzae مشاهده گردید. سپس جنس Cladosporium با گونه‌های C. cladosporioides و C. herbarum و در نهایت Rhizopus stolonifer شناسایی شدند. شناسایی مولکولی برای ایزوله مشکوک انجام شد و تعلق آن به A. clavatus تأیید گردید. همچنین درخت فیلوژنتیکی برای پشتیبانی از یافته‌ها ترسیم شد. علاوه بر این، وجود ژن PatA که مسئول تولید پاتولین در این گونه است، مورد بررسی قرار گرفت.
نتیجه‌گیری: تحلیل درخت ژنتیکی همگرایی آشکاری را بین توالی‌های قارچ مورد مطالعه و سایر گونه‌های ثبت‌شده در پایگاه داده ژن‌بانک NCBI نشان داد.
کلیدواژه‌ها
پاتولین
درخت فیلوژنتیکی
مدرسه ابتدایی
Aspergillus clavatus
PatA

عنوان مقاله English

Phylogeny of Aspergillus clavatus isolated from primary schools of Iraq and detection of its ability for production of Patulin

نویسندگان English

Maysoon M. AL-Jobouri 1
Ihsan Ali Alzamily 2
Majid Kadhim Al-Shibly 3
1 Muthanna Education Directorate, Iraq.
2 Department of Applied Biotechnology, College of Biotechnology, Al-Qasim Green University. Babylon 51013, Iraq.
3 Department of biology, College of Education, Al-Qadisiyah University, Iraq.
چکیده English

Objective
Schools are among of the most essential environments of children. Children spend an extensive period of time at school. However, indoor air pollution in schools might be dangerous. Fungi's specific health impacts in restricted areas can lead to acute health problems, particularly among school students. The current study carried out in the primary schools of Samawa city that included the selection of several schools randomly for isolation of fungi from the floors, air, door handles and seats in a direct isolation manner then diagnosed by traditional methods in addition to PCR technique. The current study aimed to record Aspergillus clavatus as an etiological agent in schools of Iraq and investigating existence of PatA gene which is responsible of production of Patulin toxin.
Materials and Methods
The samples collection included floors, handles, seats and air. They were collected in a direct isolation method by cotton swabs. Using Bio-Rad Laboratories' Prep-a-Gene system, low-melting-point agarose was purified to get the amplification products needed for sequencing. The dideoxy technique was used to DNA sequencing.
Results
The isolated fungi belonging to five genera, Aspergillus, Penicillium, Alternaria, Rhizopus and Cladosporium with the predominance of the genus Aspergillus where it represents 20 isolates and five species, Aspergillus niger, A. flavus, A. candidus, A. terreus and A. clavatus. The fungus Penicillum came second represented by seven isolates, Penicillium chrysogenum, Penicillium digitatum and P. candidus. The genus Alternaria represented by four isolates, A. alternata and A.oryzae. Then the genus Cladosporium with C. cladosporioides and C. herparum and finally Rhizopus stolonifer. Molecular identification was done for the suspected isolate and confirmed that is belonging to A. clavatus and the phylogenetic tree was drawn to support the findings. Also, the study included the determination of PatA gene that is responsible for producing Patulin in this species.
Conclusion
Phylogenetic tree analysis showed a clear convergence between the sequences of fungus under study and other species registered on the NCBI gene bank.

کلیدواژه‌ها English

Aspergillus clavatus
PatA
Patulin
phylogenetic tree
primary school
Abdel-Nasser, A., Fathy, H. M., Badr, A., Hathout, A., & Barakat, O. S. (2022). Prevalence of aflatoxigenic fungi in cereal grains and their related chemical metabolites. Egyptian Journal of Chemistry, 65(10), 455-470. https://doi.org/10.21608/ejchem.2022.122494.5487
Ahsani, M. R., Mohammadabadi, M. R., & Shamsaddini, M. B. (2010). Clostridium perfringens isolate typing by multiplex PCR. Journal of Venomous Animals and Toxins Including Tropical Diseases, 16(4), 573-578. https://doi.org/10.1590/S1678-91992010000400006
Alshibly, M. K., Al Zamily, I. A., & Fazil, N. (2019). Survey of fungi found in books on the shelves of the libraries of the University of Qadisiyah—Iraq. IOP Conference Series: Materials Science and Engineering, 571(1), Article 012042. https://doi.org/10.1088/1757-899X/571/1/012042
Bérdy, J. (2005). Bioactive microbial metabolites. The Journal of Antibiotics, 58(1), 1-26. https://doi.org/10.1038/ja.2005.1
Cai, G. H., Hashim, J. H., Hashim, Z., Ali, F., Bloom, E., Larsson, L., ... Norbäck, D. (2011). Fungal DNA, allergens, mycotoxins and associations with asthmatic symptoms among pupils in schools from Johor Bahru, Malaysia. Pediatric Allergy and Immunology, 22(3), 290-297. https://doi.org/10.1111/j.1399-3038.2010.01127.x
Charles, B., Freddy, P., Minati, D., & Jay, P. (2009). Alternaria and Cladosporium fungal allergen epitopes are denatured by sodium hypochlorite. World Allergy Organization Journal, 2(12), 296-302. https://doi.org/10.1097/WOX.0b013e3181c4c98f
Daisey, J. M., Angell, W. J., & Apte, M. G. (2003). Indoor air quality, ventilation and health symptoms in schools: An analysis of existing information. Indoor Air, 13(1), 53-64. https://doi.org/10.1034/j.1600-0668.2003.00153.x
Davis, P. J. (2001). Molds, toxic molds and indoor air quality. California Research Bureau, California State Library.
Diba, K., Mirhendi, H., Kordbacheh, P., & Rezaie, S. (2014). Development of RFLP-PCR method for the identification of medically important Aspergillus species using single restriction enzyme MwoI. Brazilian Journal of Microbiology, 45(2), 503-507. https://doi.org/10.1590/S1517-83822014000200018
Dziadzio, L., & Bush, R. K. (2001). Assessment and control of fungal allergens. Current Allergy and Asthma Reports, 1(5), 455-460. https://doi.org/10.1007/s11882-001-0033-3
Ejdys, E., Dynowska, M., Biedunkiewicz, A., & Sucharzewska, E. (2013). An overview of the species of fungi occurring in school rooms—A four-year study. Polish Journal of Environmental Studies, 22(6), 1665-1672.
Fouillaud, M., & Dufossé, L. (2022). Microbial secondary metabolism and biotechnology. Microorganisms, 10(1), Article 123. https://doi.org/10.3390/microorganisms10010123
Fouladi Fard, R., Hosseini, M. R., Faraji, M., & Omidi Oskouei, A. (2018). Building characteristics and sick building syndrome among primary school students. Sri Lanka Journal of Child Health, 47(4), 332-337. https://doi.org/10.4038/sljch.v47i4.8595
Fouladi-Fard, R., Vaezi, N., Mahvi, A. H., & Beigi, M. Z. (2023). Airborne culturable fungi in primary schools: Building characteristics and environmental factors in Qom, Iran. Journal of School Administration Research and Development, 8(2), 97-108. https://doi.org/10.32674/jsard.v8i2.4319
Grinn-Gofroń, A. (2011). Airborne Aspergillus and Penicillium in the atmosphere of Szczecin, (Poland) (2004-2009). Aerobiologia, 27(1), 67-76. https://doi.org/10.1007/s10453-010-9177-8
Hajji, M., Kanoun, S., Nasri, M., & Gharsallah, N. (2007). Purification and characterization of an alkaline serine-protease produced by a new isolated Aspergillus clavatus ES1. Process Biochemistry, 42(5), 791-797. https://doi.org/10.1016/j.procbio.2007.01.011
Harris, J. L. (2001). [Review of the book Atlas of clinical fungi (2nd ed.), by G. S. de Hoog, H. Gene, & M. J. Figueras. Centraalbureau voor Schimmelcultures]. Mycopathologia, 152(2), 159-160. https://doi.org/10.1023/A:1013183715057
Hu, J., Li, N., Zou, S., Yoshino, H., Yanagi, U., Yu, C. W., & Qu, H. (2019). Indoor environmental conditions in schoolchildren’s homes in central-south China. Indoor and Built Environment, 29(7), 956-971. https://doi.org/10.1177/1420326X19875185
Jasim, A. A., Abaed, S. A., & Hassoni, A. A. (2021). Isolation and identification of the air fungi present inside the schools’ buildings in Babylon province. Biochemical & Cellular Archives, 21(2), 4201-4206. https://connectjournals.com/03896.2021.21.4201
Khabiri, A., Toroghi, R., Mohammadabadi, M., & Tabatabaeizadeh, S. E. (2023). Introduction of a Newcastle disease virus challenge strain (sub-genotype VII.1.1) isolated in Iran. Veterinary Research Forum, 14(4), 221-227. https://doi.org/10.30466/vrf.2022.548152.3373
Khabiri, A., Toroghi, R., Mohammadabadi, M., & Tabatabaeizadeh, S. E. (2025). Whole genome sequencing and phylogenetic relative of a pure virulent Newcastle disease virus isolated from an outbreak in northeast Iran. Letters in Applied Microbiology, 78(4), Article ovaf049. https://doi.org/10.1093/lambio/ovaf049
Liu, B. H., Wu, T. S., Yu, F. Y., & Su, C. C. (2007). Induction of oxidative stress response by the mycotoxin patulin in mammalian cells. Toxicological Sciences, 95(2), 340-347. https://doi.org/10.1093/toxsci/kfl156
Liu, Z., Li, A., Hu, Z., & Sun, H. (2014). Study on the potential relationships between indoor culturable fungi, particle load and children respiratory health in Xi'an, China. Building and Environment, 80, 105-114. https://doi.org/10.1016/j.buildenv.2014.05.029
Lopez-Diaz, T. M., & Flannigan, B. (1997a). Production of patulin and cytochalasin E by Aspergillus clavatus during malting of barley and wheat. International Journal of Food Microbiology, 35(2), 129-136. https://doi.org/10.1016/S0168-1605(96)01211-1
Lopez-Diaz, T. M., & Flannigan, B. (1997b). Mycotoxins of Aspergillus clavatus: Toxicity of cytochalasin E, patulin, and extracts of contaminated barley malt. Journal of Food Protection, 60(11), 1381-1385. https://doi.org/10.4315/0362-028X-60.11.1381
Mendell, M. J., & Heath, G. A. (2005). Do indoor pollutants and thermal conditions in schools influence student performance? A critical review of the literature. Indoor Air, 15(1), 27-52. https://doi.org/10.1111/j.1600-0668.2004.00320.x
Mohammadabadi, M. R., Shaikhaev, G. O., Sulimova, G. E., & Rahman, O. (2004). Detection of bovine leukemia virus proviral DNA in Yaroslavl, Mongolian, and Black Pied cattle by PCR. Cellular and Molecular Biology Letters, 9(4A), 766-768.
Mohammadabadi, M. R., Soflaei, M., Mostafavi, H., & Honarmand, M. (2011). Using PCR for early diagnosis of bovine leukemia virus infection in some native cattle. Genetics and Molecular Research, 10(4), 2658-2663. https://doi.org/10.4238/2011.October.27.2
Mohammadabadi, M., Afsharmanesh, M., Khezri, A., Kheyrodin, H., Babenko, O. I., Borshch, O., Kalashnyk, O., Nechyporenko, O., Afanasenko, V., Slynko, V., & Usenko, S. (2025). Effect of mealworm on GBP4L gene expression in the spleen tissue of Ross broiler chickens. Agricultural Biotechnology Journal, 17(2), 343-360. https://doi.org/10.22103/jab.2025.25277.1714
Mohammadabadi, M., Babenko, O. I., Borshch, O., Kalashnyk, O., Ievstafiieva, Y., & Buchkovska, V. (2024). Measuring the relative expression pattern of the UCP2 gene in different tissues of the Raini Cashmere goat. Agricultural Biotechnology Journal, 16(3), 317-332. https://doi.org/10.22103/jab.2024.24337.1627
Moon, K. W., Huh, E. H., & Jeong, H. C. (2014). Seasonal evaluation of bioaerosols from indoor air of residential apartments within the metropolitan area in South Korea. Environmental Monitoring and Assessment, 186(4), 2111-2120. https://doi.org/10.1007/s10661-013-3521-8
Odebode, A., Adekunle, A., Stajich, J., & Adeonipekun, P. (2020). Airborne fungi spores’ distribution in various locations in Lagos, Nigeria. Environmental Monitoring and Assessment, 192(2), Article 87. https://doi.org/10.1007/s10661-019-8038-3
Park, M., Do, E., & Jung, W. H. (2013). Lipolytic enzymes involved in the virulence of human pathogenic fungi. Mycobiology, 41(2), 67-72. https://doi.org/10.5941/MYCO.2013.41.2.67
Pepper, I. L., & Gerba, C. P. (2015). Aeromicrobiology. In Environmental microbiology (pp. 89-110). Academic Press. https://doi.org/10.1016/B978-0-12-394626-3.00005-3
Puel, O., Galtier, P., & Oswald, I. P. (2010). Biosynthesis and toxicological effects of patulin. Toxins, 2(4), 613-631. https://doi.org/10.3390/toxins2040613
Sanger, F., Nicklen, S., & Coulson, A. R. (1977). DNA sequencing with chain-terminating inhibitors. Proceedings of the National Academy of Sciences, 74(12), 5463-5467. https://doi.org/10.1073/pnas.74.12.5463
Sciortino, C. V., Jr. (2017). Atlas of clinically important fungi. John Wiley & Sons. https://doi.org/10.1002/9781119069720
Shahdadnejad, N., Mohammadabadi, M. R., & Shamsadini, M. (2016). Typing of Clostridium perfringens isolated from broiler chickens using multiplex PCR. Genetics in the Third Millennium, 14(4), 4368-4374.
Tomee, J. F. C. M., & Kauffman, H. F. (2000). Putative virulence factors of Aspergillus fumigatus. Clinical & Experimental Allergy, 30(4), 476-484. https://doi.org/10.1046/j.1365-2222.2000.00796.x
Waksman, S. A., Horning, E. S., & Spencer, E. L. (1943). Two antagonistic fungi, Aspergillus fumigatus and Aspergillus clavatus, and their antibiotic substances. Journal of Bacteriology, 45(3), 233-248. https://doi.org/10.1128/jb.45.3.233-248.1943
Watanabe, T. (2010). Pictorial atlas of soil and seed fungi: Morphologies of cultured fungi and key to species (3rd ed.). CRC Press. https://doi.org/10.1201/EBK1439804193
White, P. L., & Barnes, R. A. (2008). Aspergillus PCR. In Aspergillus fumigatus and aspergillosis (pp. 373-388). ASM Press. https://doi.org/10.1128/9781555815523.ch29
مجله بیوتکنولوژی کشاورزی
دوره 18، شماره 2
بهار 1405
صفحه 173-188