基于高通量测序的5种中药饮片微生物多样性研究

Study on Microbial Diversity of Five Traditional Chinese Medicine Decoction Pieces Based on High-Throughput Sequencing

摘要: 目的：运用16S rRNA与ITS高通量测序技术对中药饮片污染微生物种群进行分析。方法：根据《中国药典》2025版对车前子、菊花、桑叶、茵陈及郁金5种中药饮片的控制菌进行检查，采用质谱进行鉴定，同时借助16S rRNA与ITS高通量测序技术进行测序，获得细菌和真菌的物种组成、物种丰度、群落比较等信息。结果：1) 通过质谱鉴定发现，5种中药饮片检出肠杆菌属、克雷伯氏菌属、芽孢杆菌属及克罗诺杆菌属等条件致病菌，车前子和桑叶各有1批次、郁金3批次检出大肠埃希菌。2) 利用16S rRNA与ITS高通量测序技术检测得出，5种中药饮片细菌共计435属296种，车前子和菊花优势菌属为芽孢杆菌属，桑叶优势菌属为鞘氨醇单胞菌属，茵陈优势菌属为欧文氏菌属，郁金优势菌属为欧文氏菌属和肠杆菌属；真菌共计438属680种，郁金、车前子、菊花优势菌为帚状曲霉和热带节担菌，桑叶、茵陈优势菌为帚状曲霉。结论：高通量测序方法结合传统培养方法，进行中药饮片微生物多样性研究，对饮片污染微生物风险评估及控制具有指导意义。

Abstract: Objective: Using 16S rRNA and ITS high-throughput sequencing technology, analyze the microbial population contaminated with five TCM decoction pieces. Methods: According to the 2025 edition of the Chinese Pharmacopoeia, the control bacteria of five TCM decoction pieces, namely Plantaginis semen, Chrysanthemi flos, Mori folium, Artemisiae scopariae Herba, and Curcumae radix, were examined. Mass spectrometry was used for identification, and 16S rRNA and ITS high-throughput sequencing techniques were used for sequencing to obtain information on the species composition, species abundance, and community comparison of bacterial and fungal. Results: 1) Through mass spectrometry identification, pathogenic bacteria such as Escherichia coli, Klebsiella, Bacillus, and Cronobacter were detected in five TCM decoction pieces. Escherichia coli was detected in 1 batch of Plantaginis semen, 1 batch of Mori folium, and 3 batches of Curcumae radix. 2) 16S rRNA and ITS high-throughput sequencing detected a total of 435 genera and 296 species of bacteria in five TCM decoction pieces. Among them, Bacillus was the dominant genus for Plantaginis semen and Chrysanthemi flos, Sphingomonas was the dominant genus for Mori folium, Erwinia was the dominant genus for Artemisiae scopariae Herba, and Erwinia and Escherichia were the dominant genera for Curcumae radix. There are a total of 438 genera and 680 species of fungi. The dominant fungi in Curcumae radix, Plantaginis semen, and Chrysanthemi flos are Aspergillus niger and Fusarium oxysporum, while the dominant fungi in Mori folium and Artemisiae scopariae Herba are Aspergillus niger. Conclusion: The high-throughput sequencing method combined with traditional cultivation methods is used to study the microbial diversity of traditional Chinese medicine decoction pieces, which has guiding significance for the risk assessment and control of microbial contamination in decoction pieces.