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@#Objective To isolate and identify Lactobacillus paracasei and Bacillus subtilis from feces of northern white geese.Methods Lactobacillus paracasei and Bacillus subtilis,numbered S00834-8 and IS00439-2 respectively,were isolated from feces of 30 52-week-old northern white geese and cultured in specific medium,and identified by 16S DNA sequencing. The two strains were cultured for different time duration(0,2,4,6,8,10,12,14,16,18,20,22,24,36 and 48 h),at different temperature(34,36,37,38 and 40 ℃)and pH(5. 5,6. 0,6. 5,7. 0,7. 5 and 8. 0). The growth characteristics of the strains were evaluated according to the A600values measured by UV-Vis spectrophotometer. At the same time,the acid and bile salt tolerance(pH 2. 0,2. 5,3. 0,3. 5 and 7. 0,bile salt concentration 0,0. 3%,0. 5%,1. 0% and 1. 5%)and the antibiotic tolerance were tested. Results The strains S00834-8 and IS00439-2 were Lactobacillus paracasei subspecies tolerance H and Bacillus subtilis KA9,respectively. The strain S00834-8 cultured under the optimum condition of the culture medium with pH 6. 0,at 37 ℃,for 12 h,had good bile resistance and gastric acid resistance,which was moderately susceptible to piperacillin,norfloxacin,amikacin,gentamicin and tetracycline,and susceptible to levofloxacin,streptomycin and chloramphenicol. The strain IS00439-2 best cultured in the culture medium with pH 7. 5,at 36 ℃,for 8 h,showed good bile resistance and gastric acid resistance,which was moderately sensitive to levofloxacin and tetracycline,while sensitive to piperacillin,norfloxacin,streptomycin and chloramphenicol. Conclusion Goose-derived Lactobacillus paracasei subspecies tolerance H and Bacillus subtilis KA9 have good acid resistance and bile salt resistance,which are expected to be candidate strains for goose-derived microecological preparations.
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Background: Bacillus subtilis is a probiotic strain that is widely used as a feed supplement for ruminants. In this study, one B. subtilis strain isolated from the ruminal fluid of Holstein dairy cows was used for an ex vivo study with ruminal tissue explants. The main goal was to assess the potential endosymbiotic links between B. subtilis and the ruminal epithelium using molecular analyses and amino acid profiling. The explant culture protocol was first optimized to determine the ideal conditions in terms of tissue viability before performing the actual experiments involving active and inactive bacteria with or without protein synthesis inhibitors, such as LY294002 (phosphatidylinositol 3-kinase inhibitor) or rapamycin [mammalian target of rapamycin (mTOR) inhibitor]. Results: The mRNA levels of phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit beta (PIK3CB), serine/threonine kinase (AKT), mTOR, P70S6K1, and eukaryotic translation initiation factor 4E binding protein 1 were the highest (p < 0.01), while those of programmed cell death 4 were the lowest when the tissue was incubated with 107 of B. subtilis. Compared with the inactivated bacteria, the expression levels of PIK3CB and AKT, and overall changes in mTOR and P70S6K1 were greater in rumen explants with living bacteria (p < 0.05). With an increase in B. subtilis concentration, the trends of protein and corresponding gene changes were consistent. There were differences in the concentrations of individual amino acids in the supernatants of living and inactivated bacterial culture groups, with most amino acids enriched in pathways, such as aminoacyl tRNA biosynthesis, cyanoamino acid metabolism, monobactam biosynthesis, or glycine, serine, and threonine metabolism. The addition of psilocybin upregulated the expression levels of PIK3CB and AKT. A significant decrease (p < 0.05) in PIK3CB and mTOR protein expression levels was detected after the addition of LY294002 and rapamycin. In addition, These responses were associated with the downregulation (p < 0.05) of AKT and P70S6K protein expression levels. Conclusions: We confirmed that the in vivo ruminal tissue culture system is a suitable model for studying probiotic-induced alterations in tissue function. As such, this study provides a means for future mechanistic studies related to microbial regulation and the dietary supply of proteins. In addition, living and inactivated B. subtilis can promote protein synthesis in ruminal tissue explants by altering the expression levels of related factors in the PIK3CB-AKT-mTORC1 pathway, which could further aid in optimizing the feed efficiency and increasing the use of inactivated bacteria as additives in dairy cow farming.
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Weighted gene co-expression network analysis (WGCNA) was used to understand the pathogenesis of subacute ruminal acidosis (SARA) and identify potential genes related to the disease. Microarray data from dataset GSE143765, which included 22 cows with and nine cows without SARA, were downloaded from the NCBI Gene Expression Omnibus (GEO). Results of WGCNA identified highly correlated modules of sample genes, and Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses allowed further biological insights into SARA-related modules. The protein-protein interaction (PPI) network, modules from the PPI network, and cistron annotation enrichment of modules were also analyzed. A total of 14,590 DEGs were used for the WGCNA. Construction of a protein-protein network identified DCXR, MMP15, and MMP17 as hub genes. Functional annotation showed that DCXR mainly exhibited L-xylulose reductase (NADP+) activity, glucose metabolic process, xylulose metabolic process, and carbonyl reductase (NADPH) activity, which are involved in the pentose and glucuronate interconversion pathways. MMP15 and MMP17 mainly have a collagen catabolic process. Overall, the results of this study aid the clarification of the biological and metabolic processes associated with SARA at the molecular level. The data highlight potential mechanisms for the future development of intervention strategies to reduce or alleviate the risk of SARA.
