Functional Evolution of Biological Systems Team, Institute of Plant Genetics, Polish Academy of Sciences, Poznan, Poland.
J Appl Microbiol. 2014 Jun;116(6):1607-20. doi: 10.1111/jam.12488. Epub 2014 Mar 18.
We propose and test an efficient and rapid protocol for the detection of toxigenic Fusarium isolates producing three main types of Fusarium-associated mycotoxins (fumonisins, trichothecenes and zearelanone).
The novel approach utilizes partially multiplexed markers based on genes essential for mycotoxin biosynthesis (fumonisin--fum6, fum8; trichothecenes--tri5, tri6; zearalenone, zea2) in Fusarium spp. The protocol has been verified by screening a collection of 96 isolates representing diverse species of filamentous fungi. Each Fusarium isolate was taxonomically identified through both molecular and morphological techniques. The results demonstrate a reliable detection of toxigenic potential for trichothecenes (sensitivity 100%, specificity 95%), zearalenone (sensitivity 100%, specificity 100%) and fumonisins (sensitivity 94%, specificity 88%). Both presence and identity of toxin biosynthetic genes were further confirmed by direct sequencing of amplification products.
The cross-species-specific PCR markers for key biosynthetic genes provide a sensitive detection of toxigenic fungal isolates, contaminating biological material derived from agricultural fields.
The conducted study shows that a PCR-based assay of biosynthetic genes is a reliable, cost-effective, early warning system against Fusarium contamination. Its future use as a high-throughput detection strategy complementing chemical assays enables effective targeted application of crop protection products.
我们提出并测试了一种高效快速的方法，用于检测产生三种主要类型镰刀菌相关霉菌毒素（伏马菌素、单端孢霉烯族毒素和玉米赤霉烯酮）的产毒镰刀菌分离株。
该新方法利用基于霉菌毒素生物合成必需基因（伏马菌素 - fum6、fum8；单端孢霉烯族毒素 - tri5、tri6；玉米赤霉烯酮，zea2）的部分多重标记物。该方案已通过筛选代表丝状真菌不同种的 96 个分离株进行了验证。通过分子和形态技术对每个镰刀菌分离株进行了分类鉴定。结果表明，该方法能够可靠地检测出产毒潜力的单端孢霉烯族毒素（敏感性 100%，特异性 95%）、玉米赤霉烯酮（敏感性 100%，特异性 100%）和伏马菌素（敏感性 94%，特异性 88%）。毒素生物合成基因的存在和身份进一步通过扩增产物的直接测序得到证实。
关键生物合成基因的种间特异性 PCR 标记物提供了对污染农业领域生物材料的产毒真菌分离株的敏感检测。
该研究表明，基于 PCR 的生物合成基因分析是一种可靠、具有成本效益的镰刀菌污染预警系统。其作为补充化学分析的高通量检测策略的未来应用，能够有效靶向应用作物保护产品。
