معرفی روش درون‌ شیشه‌ای آزمون بیماری‌زایی قارچ ascochyta rabiei روی برگ‌های جدا شده ارقام نخود مقاوم و حساس

شناسایی منابع مقاومت به بیماری برق‌زدگی با انجام آزمایش‌های به گزینی در شرایط آلودگی مصنوعی عامل بیماری یکی از روش‌های مرسوم در اصلاح نباتات به‌شمار می‌رود. هزینه‌بر بودن و مخاطرات ظهور و گسترش پاتوتیپ‌های جدید در اثر استفاده از پلات‌های آلوده، از جمله محدودیت های مطالعات به گزینی محسوب می‌شوند. فراهم شدن یک روش به گزینی قابل اعتماد در شرایط درون شیشه ای می‌تواند علاوه‌بر سرعت بخشیدن به برنامه‌های اصلاحی، بسیاری از این محدودیت‌ها را برطرف نماید. بدین منظور، ابتدا ماندگاری و احتمال رشد آلودگی‌های سطحی و درون زاد، روی برگ‌های جدا شده از گیاهچه‌های رشد یافته در شرایط گلخانه مربوط به دو ژنوتیپ نخود به‌نام‌های mcc133 (مقاوم) و ilc1929 (حساس) پس از ضدعفونی سطحی و کشت در شرایط درون شیشه ای بدون بستر کشت مغذی، مورد بررسی قرار گرفت. همچنین شدت و سرعت بروز علائم بیماری ناشی از مایه‌زنی قارچ ascochyta rabiei به دو روش قطره‌ای و غوطه‌وری مورد مقایسه قرار گرفت. به‌علاوه امکان تمایز ژنوتیپ‌های مقاوم و حساس در پاسخ به پاتوتیپ pi بررسی شد. در مرحله پایانی، امکان تمایز پاتوتیپ‌های مختلف قارچ بر اساس قدرت بیماری‌زایی روی رقم مقاوم، ارزیابی شد. نتایج نشان داد که برگ‌های جدا شده از گیاه تا پس از گذشت 10 روز در شرایط درون شیشه بدون مواد مغذی و بدون بروز علائم کمبود قابل نگهداری هستند و روش آماده‌سازی برگ‌ها از بروز هرگونه آلودگی سطحی جلوگیری نموده است. نتایج آزمون بیماری‌زایی نشان داد که تلقیح به‌روش غوطه‌ورسازی در مقایسه با روش قطره‌ای با راندمان بالاتری سبب بروز علائم می‌شود. در مجموع، این نتایج نشان می‌دهد که روش آزمون بیماری‌زایی درون شیشه ای ابداع شده به‌خوبی قادر است، ارقام را بر اساس سطح مقاومت از یکدیگر متمایز نماید و بر این اساس، می‌توان از این روش در مطالعات به ‌گزینی و ارزیابی سطح مقاومت نمونه‌های گیاهی در مقابل آلودگی با جدایه‌های قارچ a. rabiei استفاده نمود.

introduction of an in vitro pathogenicity test of ascochyta rabiei on chickpea leaves detached from resistant and susceptible cultivars

introductionchickpea (cicer arietinum l.) is considered as one of the rich sources of plant protein in human diet and an important factor in soil fertility which has of special position in the crop rotation programs. in iran, chickpea is grown in spring and autumn. ascochyta blight is currently considered as the most important limiting factor of winter cultivation of chickpea in iran. identifying sources of resistance to ascochyta blight by performing screening tests using artificial inoculation is one of the common methods in plant breeding. the cost and risks of spreading the diversity of pathogenic fungi due to the use of infected plots are among the limitations of alternative studies. providing a reliable in vitro screening method besides speeding up the breeding programs can overcome many of these limitations. materials and methodstwo chickpea seed samples were selected in the presented study which were including; mcc133 and ilc1929, respectively, as resistant and susceptible lines to ascochyta blight. the seed samples were provided by the seed bank of the research center for plant sciences of ferdowsi university of mashhad, iran. the fungal isolates called pi (pathotype no. 1), related to ascochyta rabiei with strain number of fum 1001 was obtained from the microorganisms collection of ferdowsi university of mashhad (wdcm 1207), iran. the germinated chickpea seeds were planted in pots containing equal proportions of leached sterilized coco peat and perlite. the pots were kept in a growth chamber at 24 ±2 °c and 16:8 h light/dark photoperiod and fertilized weekly by zista nutritional plant solution up to 5 leaves. three leaves were detached from each seedling and placed under sterile conditions in test tubes. the durability and growth potential of surface and endogenous infections on chickpea leaves detached from the seedlings grown in greenhouse condition were investigated after surface disinfection and culture under in vitro condition without nutrient medium. moreover, the severity and rate of disease symptoms development as a result of applying two inoculation methods were compared on detached leaves separated from susceptible cultivar. in addition, the possibility of differentiation of resistant and susceptible genotypes was investigated in response to a virulent pathotype. in the final stage, the possibility of differentiation of different fungal pathotypes was evaluated based on pathogenicity on resistant cultivar. results and discussionour daily visual observation showed that, no signs of fungal growth or symptoms of infection related to saprophytes were observed on detached leaves inside the test tube until the tenth day. the leaves were completely fresh until the tenth day and there were no symptoms of wilting on them. on the tenth day, a number of leaf samples had slightly reduced their greenness, but were perfectly healthy. the results showed that using this method the detached leaves can be stored safely in the test tube for at least 10 days. in addition, the method of preparing the leaves has prevented any surface infection. the results of pathogenicity test showed a clear difference between the disease symptoms in the two inoculation methods. symptoms of the disease were visible on the leaves of susceptible cultivars in the immersion method from the third day in the form of very small spots resulting from the destruction of the cuticular layer on the leaf surface. these symptoms were strengthened on the fifth day. disease development was increased rapidly in the whole leaf and the wounds were quite visible on the eighth day. in the drip method, no visible symptoms were visible until the eighth day, and on the tenth day, the wounds were observed at the site of inoculation. the results indicate that the immersion method induces quicker symptom development on leaflets, with a more rapid and widespread effect across the entire leaf.