گیاه پزشکی

گیاه پزشکی

فعالیت ضد میکروبی تاناتین نوترکیب علیه برخی از بیمارگرهای باکتریایی گیاهان در شرایط آزمایشگاهی و گلخانه‌ای

نوع مقاله : علمی پژوهشی -انگلیسی

نویسندگان
1 استادیار، گروه زراعت و اصلاح نباتات، دانشکده کشاورزی، دانشگاه صنعتی شاهرود، شاهرود، ایران
2 دانشیار، گروه گیاه‌پزشکی، دانشکده کشاورزی، دانشگاه فردوسی مشهد، مشهد، ایران
3 مربی، گروه گیاه‌پزشکی، دانشکده کشاورزی، دانشگاه فردوسی مشهد، مشهد، ایران
چکیده
کنترل بیماری‌های قارچی و باکتریایی گیاهان عمدتاً مبتنی بر کاربرد قارچ­کش­ها و باکتری­کش­های معدنی و شیمیایی است که از آلاینده‌های محیطی و سمّی برای انسان محسوب می‌شوند. علاوه بر این، استفاده طولانی مدّت از ترکیبات فوق و آنتی­بیوتیک­ها منجر به ایجاد مقاومت در بین بیمارگرها گردیده و همچنین موجب افزایش مخاطرات زیست­محیطی و بهداشتی شده­است. از این رو، توسعه تیمارهای غیر­سمّی و غیر­آلاینده برای کنترل بیماری­های گیاهی، هدف پژوهش­های گسترده­ای در علوم کشاورزی بوده است. پپتیدهای ضدمیکروبی نوترکیب اخیراً به دلیل فعالیت ضد میکروبی با طیف وسیع و قوی خود به عنوان جایگزین‌های بالقوه برای سایر روش‌های کنترلی مرسوم مورد توجه گسترده‌ای قرار گرفته‌اند. سنجش‌های آزمایشگاهی روی پپتیدهای مختلف ساخته شده شیمیایی نشان می­دهد که پپتید تاناتین منشأ گرفته از سن شکارگرPodisus maculiveventris با طیف عمل گسترده‌ بیشترین پتانسیل را در از بین بردن قارچ‌های مولد آفلاتوکسین دارا می­باشد. با این حال، اثرات ضد باکتریایی تاناتین علیه بیمارگر‌های باکتریایی گیاهی تاکنون کمتر مورد بررسی قرار گرفته است. در این مطالعه، توالی کد کننده تاناتین برای بیان در ناقل Pichia pastoris بهینه سازی گردید. سپس این توالی کدکننده در وکتور بیانی pPICZαA همسانه­سازی شد و برای ساخت تاناتین نوترکیب مورد استفاده قرار گرفت. فعالیت ضد باکتریایی پپتید ساخته شده در شرایط in vitro و in vivo مورد بررسی قرار گرفت. نتایج نشان داد که تمامی مراحل ساخت، شبیه سازی و بیان در مخمر با موفقیت انجام شده­است. نتایج آزمون­های تعیین حداقل غلظت بازدارندگی (MIC) و حداقل غلظت باکتری­کشی(MBC) نشان داد که رشد اکثر بیمارگر‌های باکتریایی گیاهی اعم از گرم منفی و مثبت توسط تاناتین نوترکیب در شرایط آزمایشگاهی مهار شد. میزان MIC ارزیابی شده برای جدایه­های مختلف باکتریایی بین 016/0 تا 048/2 میکروگرم در میلی لیتر بود در حالی که مقادیر MBC برای جدایه­های باکتریایی مشابه بین 064/0 تا 096/4  میکروگرم در میلی لیتر تعیین شد. این مقادیر بسیار کمتر از مقادیر MIC و MBC بدست­آمده با استفاده از اکسی­کلرید ­مس بود. کاربرد 09/4 میکروگرم بر میلی لیتر تاناتین نوترکیب علیه بیماری باکتریایی سوختگی برگ برنج، این بیماری را در شرایط گلخانه ای به خوبی کنترل کرد. با این حال، این پپتید ضد میکروبی امیدوارکننده نیاز به بررسی بیشتر برای توسعه مولکول‌های جدید برای کنترل بیمارگرهای گیاهی دارد.
کلیدواژه‌ها
موضوعات

عنوان مقاله English

Antimicrobial activity of recombinant thanatin against some of the major bacterial plant pathogens under in vitro and greenhouse conditions

نویسندگان English

A. Tanhaeian 1
M. Mamarabadi 2
Y. Ramezany 3
1 Assistant professor, Department of Agronomy and Plant Breeding, Faculty of Agriculture, Shahrood University of Technology, Shahrood, Iran
2 Associate Professor, Department of Plant Protection, Faculty of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran
3 Instructor, Department of Plant Protection, Faculty of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran
چکیده English

The control of fungal and bacterial plant diseases mainly relies on the application of mineral and chemical fungicides/bactericides, which are considered to be environmental pollutants and toxic to humans. Moreover, the prolonged use of bactericides and antibiotics has led to the development of resistance among pathogens, as well as an increase in environmental and health threats. Therefore, the development of non-toxic and non-polluting treatments to control plant diseases has been the focus of extensive research in agriculture. Synthetic antimicrobial peptides have recently received extensive attention as the potential alternatives to other conventional methods in terms of their strong broad-spectrum antimicrobial activity. In vitro assays using various chemically synthesized peptides showed that the broad-spectrum peptide thanatin derived from the spined soldier bug (Podisus maculiventris (Say)) had the greatest potential for eliminating aflatoxigenic fungi. However, the antibacterial effect of thanatin against bacterial plant pathogens was less studied so far. The thanatin encoding sequence was codon optimized for expression in Pichia pastoris. This coding sequence cloned into P. pastoris expression vector pPICZαA and used for synthetizing the recombinant thanatin. Then, antibacterial activities of the constructed peptide were studied under in vitro and in vivo condition. The result showed that, all construction, cloning and expression processes were successfully performed in yeast. The results of the MIC and MBC tests showed that the growth rate of the majority of bacterial plant pathogens including gram-negative and gram-positive bacteria was inhibited by recombinant thanatin under in vitro conditions.
MIC values for different bacterial isolates ranged from 0.016 to 2.048, while the MBC values for the same bacterial isolates were determined to be between 0.064 and 4.096 μg/mL. These amounts were significantly lower than the MIC and MBC values obtained by applying copper oxychloride. The application of 4.09 μg/mL of recombinant thanatin against rice leaf blight splendidly controlled this bacterial disease under greenhouse conditions. However, this promising antimicrobial peptide requires further investigation for the development of novel molecules for the control of plant pathogens.

کلیدواژه‌ها English

Antimicrobial peptides
Pichia pastoris
rice leaf blight
Xanthomonas oryzae pv. oryzae
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