喀什帕米尔高原牦牛乳传统发酵生奶酪中微生物多样性及分子鉴定

Microflora Diversity and Molecular Identification from Traditional Fermented Raw Cheese of Yak Milk in Tashkorghan Pamirs

目的:研究喀什帕米尔高原塔县地区传统牦牛乳发酵生奶酪中微生物的多样性和菌群结构。方法:从帕米尔高原区不同牧民家庭中采集6份生奶酪样品,采用高通量测序技术分析其微生物多样性。通过纯培养方法分离传统牦牛乳发酵生奶酪样品中的菌种,采用传统分类与16S rDNA序列测定和26S rDNA D1/D2间隔区序列测定方法对分离的细菌和酵母菌进行种属鉴定。结果:通过细菌的16S rDNA基因V3-V4区测序,共获优化序列945432。通过真菌的ITS区域测序,共获优化序列1302962。在属水平上,优势细菌属为乳杆菌属和链球菌属,优势真菌属为克鲁维酵母属、有孢圆酵母属、伊萨酵母属和未分类的真菌类群。根据主坐标轴分析,6份样品的群落结构间既有相似性又有差异性。在MRS和MC培养基中共分离52株细菌,在YGC培养基上共分离46株酵母菌,通过细菌16S rDNA序列和真菌26S rDNA D1/D2隔间区序列分析,将52株细菌归为7个属11个种,分别是耐久肠球菌、粪肠球菌、热带芽孢杆菌、蜡样芽孢杆菌、类副粘杆菌、希腊假单胞菌、副干酪乳杆菌、鸡乳杆菌、霍氏肠杆菌、细黄链霉菌、表皮葡萄球菌。将46株酵母菌归为8个属9个种,分别是解脂耶氏酵母、诞沫假丝酵母、发酵毕赤酵母、库德毕赤酵母、普兰久浩酵母、异常威克汉逊酵母、马克思克鲁维酵母、东方伊萨酵母、酿酒酵母。本研究首次对帕米尔高原地区传统发酵乳制品中微生物资源进行分析,其结果可为该地区发酵乳制品的研究提供参考。需要进一步更系统的的深入研究挖掘。

Objective:To explored the microbial diversity and community structure of traditional fermented raw cheese of yak milk in Tashkorghan Pamirs Xinjiang.Methods:6 raw cheese samples were collected from different herdsmen's families.The microbial diversity was analyzed by high-throughput sequencing technology.At the same time,the traditional yak milk fermented cheese samples were selected and isolated by pure culture.The species of bacteria and yeast were identified by traditional classification,16 S rDNA sequencing and 26 S rDNA D1/D2 spacer sequencing.Results:The opti mized sequence 945 432 was obtained by sequencing the V3-V4 region of 16 S r DNA gene.The optimized sequence1 302 962 was obtained by sequencing the its region of fungi.At the genus level,Lactobacillus and Streptococcus belonging to Firmicutes were the dominant bacterium,Kluyveromyces,Torulaspora,Issatchenkia and unclassified dipodascacease belonging to Ascomycotea were the dominant fungi.According to PCo A analysis,the community structure of the six samples had both similarities and differences.52 strains of bacteria were isolated on MRS and MC medium and 46 strains of yeast were isolated on YGC medium.Through the analysis of bacterial 16 S rDNA sequence and fungal 26 S rDNA D1/D2 compartment sequence,52 strains of bacteria were classified into 7 genera and 11 species.They were Enterococcus durans,Enterococcus faecium,Bacillus tropicus,Bacillus cereus,Bacillus paramycoides,Pseudomonas helleri,Lactobacillus paracasei,Lactobacillus gallinarum,Enterobacter hormaechei,Streptomyces microflavus and Staphylococcus epidermidis.46 yeast strains were classified into 8 genera and 9 species.They were Yarrowia lipolytica,Candida zeylanoides,Pichia fermentans,Pichia kudriavzevii,Guehomyces pullulans,Wickerhamomyces anomalus,Kluyveromyces marxianus,Issatchenkia orientalis and Saccharomyces cerevisiae.In this study,the microbial resources in traditional fermented dairy products in Tashkoghan Pamirs were analyzed for the first time.There is diversity of microorganisms in yak milk fermented milk products in this area.Further in-depth research and excavate are needed.