基于高通量测序技术分析四川两个奶牛场乳汁的菌群差异

Analysis of the differences in the microflora of milk from two dairy farms in Sichuan province based on high-throughput sequencing technology

【目的】本试验测定了两个奶牛场健康乳汁和乳房炎乳汁中微生物菌群的变化,以揭示不同奶牛场之乳汁菌群的异同,评估其对乳汁代谢的影响是否相同。【方法】采用16S rRNA高通量测序技术,分别测定两个奶牛场6头健康奶牛和6头乳房炎奶牛乳汁中微生物16S rRNA V4区序列,并对菌群群落结构和多样性进行比较,分析场内及场间的乳汁菌群差异。【结果】四组乳汁样本共获得4013234条原始序列,经过滤后获得2887024条优化序列。Alpha多样性Chao指数、Ace指数、Shannon指数、Simpson指数差异均不显著(P>0.05);Beta多样性四组样本均分别聚类;在场1和场2中,引起奶牛乳房炎的优势菌属分别是克雷伯氏菌属和埃希氏菌属;在2个奶牛场的健康乳汁中,场2的埃希氏菌属、葡萄球菌属的丰度显著高于场1;在2个奶牛场的乳房炎乳汁中,场2的埃希氏菌属、乳球菌属的丰度显著高于场1;2个奶牛场健康乳汁中的嗜冷菌总丰度分别为31.87%和38.72%;关联分析及功能预测分析表明,2个奶牛场健康乳汁与乳房炎乳汁优势物种之间的关系差异较大;场1无论是Level 1还是Level 2水平,均发现显著性差异的代谢通路,而场2均未发现显著性差异的代谢通路。【结论】本试验研究了两个奶牛场健康乳汁和乳房炎乳汁微生物菌群之间的异同,为两个奶牛场在乳房炎的预防工作以及原料奶在冷链运输过程中质量控制提供理论依据。

[Objective]This experiment studied the microbial flora in healthy milk and mastitis milk from two dairy farms to explore the similarities and differences of milk flora in different dairy farms,and to assess whether their effects on milk metabolism are the same.[Methods]The 16 S rRNA high-throughput sequencing technology was used to determine the 16 S rRNA V4 region sequence of microorganisms in the milk of 6 healthy cows and 6 mastitis cows in two dairy farms.The structure and diversity of bacterial community were compared,and the differences in milk flora within and between the two farms were analyzed.[Results]A total of 4013234 Raw Reads were obtained from the four groups of milk samples,and 2887024 Clean Reads were obtained after filtering.No significant differences were found in Chao index,Ace index,Shannon index and Simpson index of Alpha diversity(P>0.05);Four groups of samples with Beta diversity are clustered respectively;In farm 1 and farm 2,the dominant bacteria causing cow mastitis were Klebsiella and Escherichia,respectively;the abundances of Escherichia and Staphylococcus in the healthy milk in farm 2 were significantly higher than those in farm 1;the abundance of Escherichia and Lactococcus in the mastitis milk in farm 2 were significantly higher than those in farm 1;the abundance of Psychrophilic bacteria in the healthy milk of the two dairy farms were 31.87%and 38.72%respectively;correlation analysis and functional prediction analysis showed that there was a significant difference in the relationship between the healthy milk and the dominant species of mastitis in the two dairy farms;significant differences in metabolic pathways were found in both Level 1 and Level 2 in farm 1,while no significant differences in metabolic pathways were found in farm 2.[Conclusion]This experiment studied the similarities and differences of microflora between healthy milk and mastitis milk in two dairy farms,and provided a theoretical basis for the prevention of mastitis in the two dairy farms and the quality control of raw milk during cold chain transportation.