ABSTRACT
Cellulolytic bacteria with probiotic functions play a crucial role in promoting the intestinal health in herbivores. In this study, we aimed to correlate the 16S rRNA gene amplicon sequencing and fiber-degrading enzyme activity data from six different herbivore feces samples. By utilizing the separation and screening steps of probiotics, we targeted and screened high-efficiency fiber-degrading bacteria with probiotic functions. The animals included Maiwa Yak (MY), Holstein cow (CC), Tibetan sheep (TS), Southern Sichuan black goat (SG), Sichuan white rex rabbit (CR), and New Zealand white rabbit (ZR). The results showed that the enzymes associated with fiber degradation were higher in goat and sheep feces compared to cattle and rabbit's feces. Correlation analysis revealed that Bacillus and Fibrobacter were positively correlated with five types of fiber-degrading related enzymes. Notably, the relative abundance of Bacillus in the feces of Tibetan sheep was significantly higher than that of other five herbivores. A strain TS5 with good cellulose decomposition ability from the feces of Tibetan sheep by Congored staining, filter paper decomposition test, and enzyme activity determination was isolated. The strain was identified as Bacillus velezensis by biological characteristics, biochemical analysis, and 16S rRNA gene sequencing. To test the probiotic properties of Bacillus velezensis TS5, we evaluated its tolerance to acid and bile salt, production of digestive enzymes, antioxidants, antibacterial activity, and adhesion ability. The results showed that the strain had good tolerance to pH 2.0 and 0.3% bile salts, as well as good potential to produce cellulase, protease, amylase, and lipase. This strain also had good antioxidant capacity and the ability to antagonistic Staphylococcus aureus BJ216, Salmonella SC06, Enterotoxigenic Escherichia coli CVCC196, and Escherichia coli ATCC25922. More importantly, the strain had good self-aggregation and Caco-2 cell adhesion rate. In addition, we tested the safety of Bacillus velezensis TS5 by hemolysis test, antimicrobial susceptibility test, and acute toxicity test in mice. The results showed that the strain had no hemolytic phenotype, did not develop resistance to 19 commonly used antibiotics, had no cytotoxicity to Caco-2, and did not have acute toxic harm to mice. In summary, this study targeted isolated and screened a strain of Bacillus velezensis TS5 with high fiber-degrading ability and probiotic potency. This strain can be used as a potential probiotic for feeding microbial preparations for ruminants.
ABSTRACT
Introduction: In recent years, a large number of studies have shown that Bacillus velezensis has the potential as an animal feed additive, and its potential probiotic properties have been gradually explored. Methods: In this study, Illumina NovaSeq PE150 and Oxford Nanopore ONT sequencing platforms were used to sequence the genome of Bacillus velezensis TS5, a fiber-degrading strain isolated from Tibetan sheep. To further investigate the potential of B. velezensis TS5 as a probiotic strain, in vivo experiments were conducted using 40 five-week-old male specific pathogen-free C57BL/6J mice. The mice were randomly divided into four groups: high fiber diet control group (H group), high fiber diet probiotics group (HT group), low fiber diet control group (L group), and low fiber diet probiotics group (LT group). The H and HT groups were fed high-fiber diet (30%), while the L and LT groups were fed low-fiber diet (5%). The total bacteria amount in the vegetative forms of B. velezensis TS5 per mouse in the HT and LT groups was 1 × 109 CFU per day, mice in the H and L groups were given the same volume of sterile physiological saline daily by gavage, and the experiment period lasted for 8 weeks. Results: The complete genome sequencing results of B. velezensis TS5 showed that it contained 3,929,788 nucleotides with a GC content of 46.50%. The strain encoded 3,873 genes that partially related to stress resistance, adhesion, and antioxidants, as well as the production of secondary metabolites, digestive enzymes, and other beneficial nutrients. The genes of this bacterium were mainly involved in carbohydrate metabolism, amino acid metabolism, vitamin and cofactor metabolism, biological process, and molecular function, as revealed by KEGG and GO databases. The results of mouse tests showed that B. velezensis TS5 could improve intestinal digestive enzyme activity, liver antioxidant capacity, small intestine morphology, and cecum microbiota structure in mice. Conclusion: These findings confirmed the probiotic effects of B. velezensis TS5 isolated from Tibetan sheep feces and provided the theoretical basis for the clinical application and development of new feed additives.
ABSTRACT
The multi-functional properties of lactic acid bacteria (LAB) on host health have been a popular research topic. The aim of present study was to assess the multi-functional potential of five LAB strains isolated from giant panda. In this study, we analyzed five giant panda LAB strains (Weissella confuse WJ202003 (W3), WJ202009 (W9), WJ202021 (W21), BSP201703 (X3); Lactiplantibacillus plantarum BSGP201683 (G83)) and found that they exhibited rapid growth as well as strong acid production capacity. The five LAB strains possessed high cell surface hydrophobicity to the four tested solvents (xylene, hexadecane, chloroform, ethyl acetate; except strain W9), auto-aggregation ability, co-aggregation ability with three pathogens (Escherichia coli, Enterotoxigenic Escherichia coli, Salmonella), adhesion ability to Caco-2 cell line, and strongly biofilm formation ability, suggesting an adhesion property. As investigated for their antioxidative potential, all the strains showed good tolerance to H2O2, high scavenging ability against 1, 1-diphenyl-2-picrylhydrazyl (DPPH), and hydroxyl (OH-), and reduction ability. Furthermore, the five LAB strains could produce multiple probiotic substances, including exopolysaccharide (EPS), gamma-aminobutyric acid (GABA), bile salt hydrolase (BSH), cellulase (only strain G83), and protease (except strain X3), which was the first to report the production of EPS, GABA, BSH, cellulase, and protease in giant panda-derived LAB strain. These results demonstrated that strains W3, W9, W21, X3, and G83 had multi-functional potential and could be utilized as potential probiotics for giant panda.
