Diversity Of Lactic Acid Bacteria In Yak Milk Cheese Of Western Sichuan Plateau And The Adaptive Laboratory Evolution Under Antibiotic Stress

Based on the diversity,and reveal the phylogenetic relationship of the natural and genetic diversity of lactic acid bacteria on the classification of lactic acid bacteria resources and the breeding of excellent strains.On this basis,the adaptive molecular mechanism of the lactobacillus Lactobacillus casei YA8 in the antibiotic environment.YA8 was grown for 2000 consecutive passages in medium containing sulfamethomethazine.During culture,the phenotype,growth curve,enzyme activity,fermentation performance and drug resistance were monitored,and the adaptive evolution of lactic acid bacteria under antibiotic stress was resolved by molecular means such as transcriptomics and whole genome resequencing.The main study results are presented as follows:1.Screening samples from Hongyuan County,Ruoergai County,Kangding County,and Rangtang County and other areas,a total of 70 strains of lactic acid bacteria,among which,4 strains are acid resistant,15 strains of osmotic pressure resistant,3 strains of bile salt resistance,3 strains of strong acid production,4 strains of high temperature resistance,3strains of pathogen resistance strong dominant strains;The results of BOXA1R-PCR diversity analysis showed that lactobacillus was distributed in Lactobacillus,Enterococcus and Lactococcus,among which Lactobacillus accounted for 50%,which was the dominant genus of yak lactobacillus in western Sichuan.According to the comprehensive phenotypic determination results,we obtained a potential lactic acid bacteria YA8 with excellent performance,which was identified as Lactobacillus casei by 16S rRNA.This study laid the foundation for the related research on the resource diversity of lactobacillus in western Sichuan Plateau,and contributed to the screening of excellent lactic acid bacteria strains in traditional fermentation products.2.We determined the resistance of YA8 to 10 commonly used antibiotics and found that YA8 had the highest resistance to sulfonethomethazine,reaching 1280 μg/m L.The transcriptome was determined by placing YA8 in the semi-inhibitory concentration of sulfonamide,and sequencing indicated 17 differentially expressed genes after antibiotic treatment,of which 5 genes were upregulated and 12 were downregulated.Lactobacillus casei YA8 may resist environmental intermethacine stress mainly through the pyrimidine metabolism pathway and alanine,aspartate and glutamate metabolism pathways.This study provides a molecular theoretical basis for further research on the resistance and control measures of lactic acid bacteria in fermented dairy products.3.YA8 was grown for 2000 consecutive passages in medium containing semi-inhibitory concentrations of sulfamidazine,There is no change in its phenotype;Log growth time,four enzymes activities and fermentation nutrient quality were suppressed first,With the improvement of YA8,it gradually recovered or even more than 0 generations;The minimum inhibitory concentration of sulfonethomethazine was increased by 2-fold over generation 0 and remained unchanged;Whole-genome resequencing showed that,T: A> C:G and C: G> T: A conversion are two common types of mutation,Significantly higher than the switching frequency,YA8 possesses unique metabolic features during adaptive evolution,Complete adaptation to stress stress through its own metabolism,To maintain their own stable development.This study provides a molecular theory for the deep study of the adaptive evolution of lactic acid bacteria in fermented dairy products.In conclusion,this study analyzed the diversity of lactobacillus in traditional fermented dairy products in western Sichuan Plateau,and revealed the molecular mechanism from phenotype and molecular level,and provided new insights and basis for the utilization of wild Lactobacillus resources and its continuous response under antibiotic stress.