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研究半夏曲不同发酵时间点的微生物种类、数量变化以及优势菌种的分离鉴定,为探讨半夏曲炮制机制奠定基础。用选择性培养基对半夏曲发酵过程中5个时间点(0,18,36,54,72 h)样品中的细菌、霉菌、酵母菌分别进行培养、菌落计数、分离纯化优势菌种;采用16S rDNA,26S rDNA序列分别对优势细菌、优势真菌进行鉴定。结果表明,半夏曲发酵过程中细菌数量少、变化平缓,而酵母菌和霉菌数量发酵至54 h时迅速增加,至发酵结束时达1×10~7CFU·mL~(-1)以上;通过NCBI同源性比对及构建系统发育树,鉴定出半夏曲炮制过程中的优势细菌为Streptomyces sp.,Bacillus pumilus,B.subtilis,B.aryabhattai,Bacillus sp.;优势酵母菌为Meyerozyma guilliermondii;优势霉菌为Paecilomyces variotii,Byssochlamys spectabilis,Aspergillus niger。提示半夏曲的发酵过程涉及多种微生物共同参与,其中以酵母菌和霉菌为主。M.guilliermondii,P.variotii,P.variotii,A.niger和Bacillus sp.等优势菌种可能在半夏曲炮制中起重要作用。
In order to explore the mechanism of Pinellia tuber cultivation, the types and quantity of microorganisms and the isolation and identification of dominant species at different fermentation time were studied. Bacteria, molds and yeasts in 5 samples (0, 18, 36, 54 and 72 h) during the fermentation process of Pinellia ternate were cultured in selective medium and their colonies were counted. The dominant strains were isolated and purified. The 16S rDNA and 26S rDNA sequences were used to identify dominant bacteria and dominant fungi respectively. The results showed that during the fermentation of Pinellia ternata, the number of bacteria was small and the variation was gentle. However, the quantity of yeast and mold rapidly increased after fermentation to 54 h and reached to 1 × 10 ~ 7CFU · mL -1 at the end of fermentation. NCBI homology comparison and phylogenetic tree construction, the dominant bacteria in the process of Pinellia ternata were identified as Streptomyces sp., Bacillus pumilus, B. subtilis, B.aryabhattai and Bacillus sp .; dominant yeasts Meyerozyma guilliermondii; The dominant molds are Paecilomyces variotii, Byssochlamys spectabilis, Aspergillus niger. Tip Pinellia process involves a variety of microorganisms involved in the fermentation process, of which yeast and mold-based. The dominant species M. cuilliermondii, P. variotii, P. variotii, A. niger and Bacillus sp. May play an important role in Pinellia tuber production.