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

نوع مقاله : مقاله پژوهشی

نویسندگان

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

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

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

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مجله بیوتکنولوژی کشاورزی
دوره 18، شماره 2
خرداد 1405
صفحه 173-188

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