شناسایی ژنوتیپی و بررسی تنوع درون گونه‌ا‌ی جدایه‌های بیماری¬زای زانتوموناس از میزبان‌های لوبیا و یونجه در استان‌های آذربایجان‌شرقی و غربی
فردین نصرت نژاد 1 , سویل نعمت اللهی 2 , کیومرث روح رضی 3
1 - آزاد تبریز
2 - دانشگاه
3 - دانشگاه آزاد
کلید واژه: آذربایجان شرقی, آذربایجان غربی, لوبیا, یونجه, زانتوموناس,
چکیده مقاله :
طی سال‌های 1398 و 1399 به منظور جداسازی باکتری‌های بیماری‌زا از میزبان‌های لوبیا و یونجه دارای علائم بیماری لکه‌برگی، پژمردگی و بلایت که از برخی مزارع و باغات استان‌های آذربایجان شرقی و غربی نمونه برداری شده بودند، در مجموع 140 جدایه باکتریایی روی محیط کشت نوترینت آگار به صورت جدایه‌های اولیه، کشت و خالص‌سازی گردیدند. براساس رنگ و شکل ظاهری کلنی‌ها جدایه‌هایی که به باکتری زانتوموناس مورد نظر ما نزدیک بودند انتخاب شده و پس از کشت دوباره و خالص سازی در مجموع 62 جدایه بدست آمد. برای اثبات بیماریزایی جدایه‌ها، هر یک از آنها به صورت جداگانه به میزبان خود تلقیح شده و در شرایط گلخانه‌ای به مدت دو هفته نگهداری شدند که 33 استرین توانستند روی میزبان خود بیماریزایی ایجاد کنند. در این استرین‌ها قطعه 239 جفت بازی ژن اختصاصی زانتوموناس که بخشی از ژن ITS است تکثیر شد. بنابراین این استرین‌ها به عنوان جنس زانتوموناس درنظر گرفته شدند. براساس انگشت نگاری ژنتیکی به روش rep-PCR استرین‌های جداشده از لوبیا و یونجه نشان داد که استرین‌های لوبیا به چهار خوشه و استرین‌های یونجه به سه گروه تقسیم بندی شدند که نتایج حاکی از تنوع ژنتیکی بین استرین‌ها می‌باشد. براساس توالی یابی ژن های 16S rRNA، gyrB و rpoD،، استرین‌های یونجه X. alfalfae subsp. alfalfa و استرین‌های جداشده از میزبان لوبیا به عنوان گونه X. phaseoli pv. pahseoli شناسایی شدند. با بررسی منابع موجود، تحقیق حاضر اولین گزارش از جداسازی، شناسایی و بررسی تنوع ژنتیکی این استرین‌ها از میزبان‌های لوبیا و یونجه در استان‌های آذربایجان شرقی و غربی می‌باشد.
In order to isolate pathogenic bacteria from the hosts of bean and alfalfa, 140 bacterial isolates were cultured and purified on NA medium in East and West Azerbaijan provinces. Based on the color and appearance of the colonies, the isolates that were close to the desired Xanthomonas bacteria were selected. After re-cultivation and purification, a total of 62 isolates were obtained. To perform the pathogenicity test of the isolates, each of them was inoculated individually to its host and kept in greenhouse conditions for two weeks. 33 strains were able to cause pathogenicity on the host. The strains were able to amplify the 239 bp fragment of Xanthomonas specific gene, which is part of the ITS gene. Therefore, these strains were considered as Xanthomonas bacteria. Based on genetic fingerprinting by rep-PCR method, the strains isolated from bean and alfalfa was divided into four and three groups, respectively, which results indicated genetic diversity between the strains. Based on the sequencing of 16S rRNA, gyrB and rpoD genes, the strains isolated from alfalfa were identified as X. alfalfae subsp. alfalfae and bean strains were identified as X. phaseoli pv. pahseoli. By reviewing the available sources, the present research is the first report of the isolation, identification and genetic diversity of these strains from bean and alfalfa hosts in East and West Azerbaijan provinces.
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