Stoeva Daniela, Gencheva Deyana, Radoslavov Georgi, Hristov Peter, Yordanova Rozalina, Beev Georgi
Department of Biological Sciences, Faculty of Agriculture, Trakia University, Students Campus, 6000 Stara Zagora, Bulgaria.
Department of Fundamental Sciences in Animal Husbandry, Faculty of Agriculture, Trakia University, Students Campus, 6000 Stara Zagora, Bulgaria.
Methods Protoc. 2025 Jul 9;8(4):78. doi: 10.3390/mps8040078.
spp. represent a critical threat to maize production and food safety due to their mycotoxin production. This study introduces a refined molecular identification protocol integrating four genomic regions-ITS1, IGS, , and -for robust species differentiation of spp. isolates from post-harvest maize in Bulgaria. The protocol enhances species resolution, especially for closely related taxa within the species complex (FFSC). A newly optimized multiplex PCR strategy was developed using three primer sets, each designed to co-amplify a specific pair of toxigenic genes: , , and . Although all five genes were analyzed, they were detected through separate two-target reactions, not in a single multiplex tube. Among 17 identified isolates, (52.9%) dominated, followed by , , , and . All isolates harbored at least one toxin biosynthesis gene, with 18% co-harboring genes for both fumonisins and zearalenone. This dual-protocol approach enhances diagnostic precision and supports targeted mycotoxin risk management strategies.
由于其产生霉菌毒素，[具体物种名称]对玉米生产和食品安全构成了重大威胁。本研究引入了一种精细的分子鉴定方案，整合了四个基因组区域——ITS1、IGS、[此处缺失一个基因区域名称]和[此处缺失一个基因区域名称]，用于对保加利亚收获后玉米中的[具体物种名称]分离株进行稳健的物种区分。该方案提高了物种分辨率，特别是对于[具体物种名称]复合种（FFSC）内亲缘关系密切的分类群。使用三个引物组开发了一种新优化的多重PCR策略，每个引物组设计用于共同扩增一对特定的产毒基因：[此处缺失产毒基因名称]、[此处缺失产毒基因名称]和[此处缺失产毒基因名称]。虽然对所有五个基因进行了分析，但它们是通过单独的双靶点反应检测到的，而不是在单个多重管中。在17个已鉴定的分离株中，[具体优势物种名称]（52.9%）占主导地位，其次是[此处缺失其他物种名称]、[此处缺失其他物种名称]、[此处缺失其他物种名称]和[此处缺失其他物种名称]。所有分离株都至少含有一个毒素生物合成基因，18%的分离株同时含有伏马菌素和玉米赤霉烯酮的基因。这种双方案方法提高了诊断精度，并支持有针对性的霉菌毒素风险管理策略。
