| Naturally fermented milk has been produced and consumed for thousands of years and has become an important part of the daily diets of pastoralists around the world,and the microbial resources contained in it have undergone thousands of years of natural selection and evolution to form a stable and heritable habitat microecology.However,the current research on the microbial genome in naturally fermented dairy products is not comprehensive enough,and it is urgent to elucidate the functional diversity between different isolated source strains from a genomic perspective.In this study,10 samples of naturally fermented kefir collected from Pakistan were used as the research object,and lactic acid bacteria were isolated and identified by pure culture.The dominant strain,Streptococcus thermophilus,and the whole genomics sequence of the Streptococcus thermophilus strain published in the NCBI database was compared for genomic analysis.While elucidating the microbiota structure of lactic acid bacteria in naturally fermented milk in Pakistan,the genomic diversity of Streptococcus thermophilus from different isolates was revealed from the perspective of functional genes.The results are as follows:1.A total of 154 strains of lactic acid bacteria were isolated from 10 samples of fermented milk by pure culture method,and the species identification was carried out by using the homology comparison and phylogenetic relationship of 16 S r RNA gene sequence.The results showed that 154 lactic acid bacteria belonged to 5 genera and 16 species.They are divided into Streptococcus,Lactobacillus,Lactococcus,Bifidobacterium and Staphylococcus.Among them,the dominant strains was Streptococcus thermophilus,accounting for 19% of the total isolates.2.The average genome size of the 73 strains of Streptococcus thermophilus was 1.81±0.29 Mb,the average G+C content was 38.92±0.54%,and the predicted number of genes was1884±398.There were significant differences in genome size,G+C content and predictive coding genes of different source isolates.The core gene-pan-genset showed that the genome of this species belongs to the open genome,and the functions of the core gene set are mainly carbohydrate metabolism,protein translation and nucleotide metabolism.3.The phylogenetic tree of 73 strains of Streptococcus thermophilus was constructed based on 178 core genes,and Average Nucleotide Identity(ANI)was calculateded,and the results showed that 73 strains of Streptococcus thermophilus were divided into 5 evolutionary branches.The ANI value was higher than 95% among them,which showed that the species had high homology.It was known that the evolutionary relationship of Streptococcus thermophilus had geographical correlation and isolated source correlation according to the ANI cluster heat map and the core gene phylogenetic tree.4.The results of functional genomics analysis showed there were significant differences in functional genes between the strains from different isolated sources.The significant differences among fermented milk source isolates and the other three source isolates were mainly in terms of predicted gene number and carbohydrate active enzyme utilization.The genomic function prediction of cultures showed that the genes numbers in fermented milk isolated strains were higher than the other three source isolates s in terms of amino acid metabolism,signal transduction,exopolysaccharides and other related functional genes.The differences in specific genes of different isolated source strains were reflected in amino acid metabolism,specific transport of various monosaccharides and CRISPR-Cas defense mechanism.In conclusion,in this study we analyzed the community of lactic acid bacteria in naturally fermented milk in Pakistan by pure culture technology,and systematically explored the genomic diversity of Streptococcus thermophilus,which provided a theoretical basis for the development and utilization of Streptococcus thermophilus. |
