基于16S rRNA基因的不同基原、不同产地甘草微生物群落特征分析

Microbial community characteristics analysis of licorice from different provenances and different habitats based on 16S rRNA gene

目的探究4批不同基原、不同产地的甘草微生物群落信息。方法参照《中国药典》2020年版四部,对甘草药材中的需氧菌总数、霉菌和酵母菌总数、耐热菌总数以及控制菌进行检查,并对致病菌平板上的菌落进行革兰染色,同时进行甘草药材的16S rRNA基因测序及数据分析。结果甘草药材中的需氧菌总数、霉菌和酵母菌总数、耐热菌总数对数值分别为4.15~4.95、2.40~4.85、1.67~3.15;经热处理(100℃、30 min)后发现其需氧菌总数对数值的下降百分比分别为100.00%、24.10%、40.20%、66.26%;各甘草中耐胆盐革兰阴性菌可能总数较高,但均未检测到大肠埃希菌和沙门菌。16S rRNA基因测序共获得19 phylum、40 class、68 order、137 family、236 genus,其中GC-2获得最多的phylum(18)、class(36)、order(58),GC-1和GC-3获得最多的family(121),GC-1获得最多的genus(162)。多样性分析结果表明,甘草药材中污染微生物主要分布于3个门、3个属,不同基原、不同产地的甘草药材中优势菌属同时具有相似性和差异性,优势菌属均为布丘氏菌属(Buttiauxella)、假单胞菌属(Pseudomonas)和藻类(Streptophyta)。结论甘草药材中微生物污染较为严重,并随基原、产地的不同而存在差异。

Objective To explore the microbial community information of 4 batches of licorice from different provenances and different habitats.Methods According to the Chinese pharmacopoeia 2020 edition four parts,the total number of aerobic bacteria,mold and yeast,thermophilic bacteria and control bacteria in licorice were checked,and the bacterial colonies on the pathogenic bacteria plate were subjected to Gram staining.At the same time,16S rRNA gene sequencing and data analysis of licorice were performed.Results The logarithmic values of the total number of aerobic bacteria,fungal and yeast,and thermophilic bacteria in licorice were 4.15‒4.95,2.40‒4.85,and 1.67‒3.15,respectively.After heat treatment(100℃,30 min),it was found that the decreasing percentages of the log value of the total number of aerobic bacteria were 100.00%,24.10%,40.20%,66.26%,respectively.The total number of bile salt resistant Gram-negative bacteria in licorice may be high,but Escherichia coli and Salmonella were detected in all of them.A total of 19 phylum,40 class,68 order,137 family and 236 genus were obtained by 16S rRNA gene sequencing,among which GC-2 obtained the most phylum(18),class(36)and order(58).GC-1 and GC-3 obtained the most families(121),and GC-1 obtained the most genus(162).The results of diversity analysis showed that the contaminating microorganisms in licorice were mainly distributed in 3 phyla and 3 genera.The dominant bacteria in licorice from different provenances and regions had similarities and differences at the same time.The dominant bacterial genera were all Buttiauxella,Pseudomonas and Streptophyta.Conclusion The microbial contamination of licorice is serious and varies with the provenances and habitats of licorice.