PDF(Pubmed)
Abstract：
Mushroom poisoning contributes significantly to global foodborne diseases and related fatalities. Amanita mushrooms frequently cause such poisonings; however, identifying these toxic species is challenging due to the unavailability of fresh and intact samples. It is often necessary to analyze residues, vomitus, or stomach extracts to obtain DNA sequences for the identification of species responsible for causing food poisoning. This usually proves challenging to obtain usable DNA sequences that can be analyzed using conventional molecular biology techniques. Therefore, this study aimed to develop a DNA mini-barcoding method for the identification of Amanita species. Following the evaluation and optimization of universal primers for DNA mini-barcoding in Amanita mushrooms, we found that the internal transcribed spacer (ITS) gene sequence primer ITS-a was the most suitable DNA barcode primer for identifying Amanita species. Forty-three Amanita samples were subsequently amplified and sequenced. The sequences obtained were analyzed for intra- and inter-species genetic distances, and a phylogenetic tree was constructed. The findings indicated that the designed primers had strong universality among the Amanita samples and could accurately identify the target gene fragment with a length of 290 bp. Notably, the DNA mini-barcode accurately identified the 43 Amanita samples, demonstrating high consistency with the conventional DNA barcode. Furthermore, it effectively identified DNA from digested samples. In summary, this DNA mini-barcode is a promising tool for detecting accidental ingestion of toxic Amanita mushrooms. It may be used as an optimal barcode for species identification and traceability in events of Amanita-induced mushroom poisoning. KEY POINTS: • Development of a DNA mini-barcoding method for Amanita species identification without fresh samples. • The ITS-a primer set was optimized for robust universality in Amanita samples. • The mini-barcode is suitable for screening toxic mushroom species in mushroom poisoning cases.
摘要：
蘑菇中毒是全球食源性疾病和相关死亡的重要原因。Amanita蘑菇经常引起这种中毒；然而，由于无法获得新鲜和完整的样品，因此识别这些有毒物种具有挑战性。通常需要分析残留物，呕吐物,或胃提取物以获得DNA序列，用于鉴定导致食物中毒的物种。这通常证明获得可用的DNA序列具有挑战性,所述可用的DNA序列可以使用常规分子生物学技术进行分析。因此,这项研究旨在开发一种DNA迷你条形码方法，用于鉴定天牛物种。在对Amanita蘑菇中DNA迷你条形码的通用引物进行评估和优化之后，我们发现内部转录间隔区（ITS）基因序列引物ITS-a是鉴定天牛物种最合适的DNA条形码引物。随后扩增并测序了43个Amanita样品。对获得的序列进行了种内和种间遗传距离分析，并构建了系统发育树。结果表明，所设计的引物在天牛样品中具有很强的普适性，可以准确鉴定出长度为290bp的目的基因片段。值得注意的是,DNA迷你条形码准确识别了43个天牛样本，证明与常规DNA条形码的高度一致性。此外，它有效地从消化样品中鉴定出DNA。总之,这种DNA迷你条形码是一种有前途的工具，用于检测意外摄入有毒的鹅膏菌。它可以用作最佳条形码，用于在天牛引起的蘑菇中毒事件中进行物种识别和可追溯性。关键点:•开发用于无新鲜样品的天牛物种鉴定的DNA迷你条形码方法。•ITS-a引物集经优化以在天牛样品中实现稳健的通用性。•迷你条形码适用于在蘑菇中毒情况下筛选有毒蘑菇物种。
