中国芒果炭疽菌多样性及致病力变异分子机制的研究

Mango anthracnose is one of the important diseases on mango causing significant yield losses and affecting fruits quality seriously at maturity. Occurrence of mango anthracnose is closely associated with species and virulence of pathogens, however, recent research showed that the population distribution, virulence differentiation, and differentiation mechanism of mango Colletotrichum spp. in China still remain unclear. On the basis of confirming the species, virulence, and genetic diversity of mango Colletotrichum spp. in Guangxi of China, the research project plans to carry out systematic morphological identification, sets of tests for determination of pathogenicity, and analysis of fungicides resistance to mango Colletotrichum spp. isolates from different areas of China, and confirm these isolates species using molecular identification with multiple genes of ITS, ACT, CHS-I, GAPDH, and TUB2. In this project, whole-genome sequencing, assembly, and gene prediction will be conducted to isolates of dominant species with different virulence using Solexa sequencing technique, and using comparative genomics, the genomic differences related to different virulent isolates will be clearly analyzed, the key functional genes affecting virulent differentiation will be successfully excavated, and distribution of these key genes located genome of different virulent isolates will be revealed. Completion of the research project can help to clear the species and dominant populations of mango Colletotrichum spp. in China, reveal the molecular mechanism of virulent differentiation of mango Colletotrichum spp., and not only understand the pathogenic mechanism and distribution regularities of genotype of mango Colletotrichum spp. among different areas, but also afford theoretical basis for the mango breeding in resistance to anthracnose, exploitation and design of control strategy aiming at new disease, and field control to mango anthracnose.

炭疽病是芒果的重要病害之一，严重影响芒果的产量和品质。该病的发生与炭疽菌种类及其致病力关系密切，而我国芒果炭疽菌的种群分布、致病力分化及变异机制尚不明确。本项目在明确广西芒果炭疽菌种类、致病力及基因组多样性的基础上，拟对我国不同地区芒果炭疽菌进行系统的形态学鉴定、致病力测定和抗药性分析，结合ITS、ACT、CHS-I、GAPDH和TUB2多基因分子鉴定明确其种类。采用Solexa测序技术对芒果炭疽菌优势种菌株进行全基因组测序、组装和基因预测，通过比较基因组学关联分析不同致病力菌株的基因组差异，挖掘影响致病力分化的关键基因，明确其在不同致病力菌株基因组上的分布。项目的完成可明确我国芒果炭疽菌的种类及优势种群，揭示芒果炭疽菌致病力变异的分子机制，不仅可探究芒果炭疽菌的致病机制和不同地区间基因型分布规律，而且为芒果抗炭疽病育种、新病害防治策略的开发设计和田间防控提供理论依据。

芒果炭疽菌是我国南方的重要的热带水果之一，主要种植于海南、广西、云南、四川攀枝花、贵州等地。本研究对采自中国7省份的芒果炭疽病样本进行了炭疽菌的分离，共获得270个芒果炭疽菌菌株。选择其中165个代表菌株，通过形态学观察和多基因分子系统学分析，共鉴定出13个种：亚洲炭疽菌，暹罗炭疽菌，果生炭疽菌，胶孢炭疽菌，喀斯特炭疽菌， 香蕉炭疽菌，内生炭疽菌，斯高维尔炭疽菌，柯氏炭疽菌，热带炭疽菌，长直孢炭疽菌，剪炭疽菌，辽宁炭疽菌。其中，亚洲炭疽为中国芒果炭疽病菌的优势种，广泛地分布于中国的7个省，占比32.12%，其次是果生炭疽，占26.06%和暹罗炭疽菌，占24.85%。在海南省、云南省和贵州省分别发现了11个中国新记录种，其中香蕉炭疽菌、内生炭疽菌、柯氏炭疽菌、长直孢炭疽菌、辽宁炭疽菌和斯高维尔炭疽菌为世界芒果上首次发现的新记录种。采用离体叶片和果实接种测定了136个芒果炭疽菌菌株的致病力，结果表明，不同菌株在芒果果实和叶片上的致病力均出现明显分化，且致病力强弱与地理来源无显著相关性。测定了136个不同种芒果炭疽菌对咪鲜胺、苯醚甲环唑和吡唑醚菌酯的敏感性，发现不同种芒果炭疽病菌对吡唑醚菌酯、咪鲜胺和苯醚甲环唑较为敏感，且不同地理来源的菌株间存在显著差异。采用ISSR-PCR分析了73个不同种芒果炭疽菌的多样性，分析后发现我国的芒果炭疽菌遗传多样性极为丰富。采用Solexa测序技术对44株不同毒力的炭疽菌株进行全基因组测序，通过比较基因组学关联分析不同致病力菌株的基因组差异。结果表明，芒果炭疽菌不同菌株基因组大小均在56Mb左右，GC含量在49.97%-60.69%之间。在基因功能注释的基础上进一步分析了一些潜在很可能影响基因功能的SNPs在基因组上的分布情况。本项目研究结果明确我国芒果炭疽菌的种类及优势种群，深入研究芒果炭疽菌的致病机制和不同地区间基因型分布规律，为芒果抗炭疽病育种、新病害防治策略的开发设计和田间防控提供理论依据。