设施辣椒中茄腐镰刀菌致伏马毒素污染的发生规律与分子调控机制研究

项目名称： 设施辣椒中茄腐镰刀菌致伏马毒素污染的发生规律与分子调控机制研究

项目编号： No.31471661

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 农业科学

项目作者： 武爱波

作者单位： 中国科学院上海生命科学研究院

项目金额： 89万元

中文摘要： 设施辣椒是我国特种优势农产品，占膳食消费比重也越来越大。国内外关于其污染真菌毒素的相关基础研究很少，给食品安全及相关产业带来严重隐患。课题组已确定我国设施辣椒中以茄腐镰刀菌为优势菌，以伏马毒素污染为主；且该茄腐镰刀菌在不同基质中均产伏马毒素，与国外报道明显不同。本项目拟采用产毒高、中、低菌株接种辣椒，在不同环境条件下培养，阐明伏马毒素污染的发生规律；针对产毒最高的茄腐镰刀菌进行de novo测序分析，揭示伏马毒素产生的分子基础；采用双向电泳、MALDI-TOF-MS、LC-TOF-MS、高分辨质谱orbi-trap、NMR等挖掘差异蛋白与代谢小分子；采用RT-PCR与构建基因缺失株、抗体制备与western blot、化学合成与体外评估等分别验证1-2个关键基因、差异蛋白与小分子功能。最终揭示设施辣椒中茄腐镰刀菌致伏马毒素污染的发生规律与分子调控机制，为后期相关安全评价与控制奠定理论基础。

中文关键词： 设施辣椒；茄腐镰刀菌；伏马毒素；发生规律；分子调控机制

英文摘要： Greenhouse pepper is highly specialized and economically important agricultural products in China, which also contributes to more and more proportions in the whole dietary consumption.Till now, very few research targeting production pattern and molecular regulatory mechanism of mycotoxins contaminated in greenhouse pepper are performed worldwide, which leads serious menace to food safety issue and related industry development. In our previous studies, Fusarium solani presented as dominant species causing rot disease in greenhouse pepper, within in the population of almost 500 Fusarium isolates originating from various geographic locations in China, where fumonisions occurred as main mycotoxin contaminations there. More interestingly, the Fusarium. solani strains were capable of producing fumonisions in liquid culture, inoculated greenhouse pepper, as well as maize and rice cultures. As summarized, the above mentioned results are obviously different to those previously reported in other countries. In this proposal three Fusarium solani strains with high, medium and low fumonisions-producing capabilities identified via the well-established LC-MS/MS method are first inoculated with greenhouse pepper under different incubation time and conditions (temperatures, light time, and fungicide carbendazim treatment), in order to elucidate the production pattern of fumonisins contamination caused by Fusarium solani in greenhouse. In parallel, the Fusarium solani isolate with the highest fumonisions production capability is selected for full-genome analysis via de novo sequencing and related comparative genomic analysis, in order to unveil the candidate genes and related expression patterns responsible for fumonisions production, Moreover, the separately incoculated samples with the isolates of highly producing and non-producing of fumonisins are prepared intowith halves for protemoic anlysis via 2-D electrophoresis and MALDI-TOF-MS, and metabolic analysis by LC-TOF-MS, HRMS (orbi-trap) and NMR, respectively. More specifically, one or two key genes responsible for fumonisions biosynthesis via RT-PCR and gene-knock out analysis are selectively confirmed. Also, the functions of one or two differential proteins and metabolites of small molecules are individually verified by the approaches of antibody preparation and western blot, chemical synthesis and in vitro evaluation. Finally as resulted, the interaction and molecular regulatory mechanisms of the confirmed elements of genes, proteins and metabolites of small molecules are unveiled to elucidate the production patterns and molecular regulartory mechanisms of fumonisions contaminations caused by Fusarium solani in greenhouse pepper. This would be valuable theoretical references to subsequent studies on risk assessment and control of mycotoxins contaminated in greenhouse vegetables concerning food safety.

英文关键词： Greenhouse pepper;Fusarium solani;Fumonisins;Production pattern;Molecular regulatory mechanism