| In recent years,probiotics for pets have developed rapidly,but most of the probiotics on the market are Bacillus,yeast or their mixed bacteria.These probiotics come from a wide range of sources,and most of them have not been functionally evaluated.As a pet intestinal flora regulator,the effect of use is mixed,which is far from meeting the needs of the rapidly developing pet market.The purpose of this study was to isolate and screen lactic acid bacteria with the function of relieving canine diarrhea from healthy dog feces and canine milk,and lay a foundation for the development of functional canine microecological preparations.The strains isolated from healthy canine feces and canine milk were identified by Gram staining and 16 S rDNA sequence analysis,and the bacteriostatic ability,antibiotic sensitivity,acid and bile salt resistance and cell adhesion ability of the strains were evaluated.The best strains J40,J48,X28 and X53 were used as treatment groups to conduct in vivo experiments on Shigella-challenged puppies and to detect serum immune factors(Ig G/Ig A/Ig M/IL-6/IFN-α/TNF-α)and The feces were collected for gut microbiota diversity analysis.Gram staining and 16 S rDNA sequence analysis showed that a total of 185 canine fecal-derived lactic acid bacteria and 31 canine milk-derived lactic acid bacteria were isolated,including J40(Pediococcus acidilactici),J48(Pediococcus acidilactici),X28(Lactobacillus sake)and X53(Pediococcus pentosaceus)etc.;Bacteriostatic test results showed that 11 strains of lactic acid bacteria such as J40 and J48 were more effective against Escherichia coli,Shigella,Salmonella and Campylobacter jejuni.The antibacterial effect was good(all antibacterial diameters were ≥ 13 mm),among which J40 had the strongest antibacterial effect;Further antibiotic susceptibility tests showed that the 45 strains of lactic acid bacteria obtained from the antibacterial test were highly sensitive to Penicillin,Ampicillin and Cefotaxime(S),which proves that the strain does not carry the above-mentioned drug resistance genes,and has certain safety and controllability;The top 25 strains screened by the above tests with better performance were subjected to the cell adhesion test,and the results showed that the 25 strains were resistant to Caco-2 The adhesion number of cells was all >6,and J40 had the best adhesion performance.The 8 strains with strong adhesion were cultured for 2 h under the conditions of p H 2 and bile salt concentration of 0.1%,and the results showed that the survival rate of the strains was all ≥50%.,indicating that the screened bacteria have good acid resistance and bile salt resistance.The results of animal experiments show that the selected strains J40,J48,X28 and X53 can effectively reduce the diarrhea rate of puppies,and the J40 treatment group has the best effect;The concentration of IL-6/TNF-α was down-regulated to improve the immune function of the body;And the results of the diversity analysis of intestinal flora showed that the relative abundance of Lactobacillus in the treatment group was significantly higher than that in the control group,while the Escherichia-Shiga in the control group was significantly higher than that in the control group.The relative abundance of genus was significantly higher than that in the treatment group.To sum up,the four probiotic strains J40,J48,X28 and X53 screened in this study can effectively treat canine bacterial diarrhea,improve the body’s immune function,and improve the distribution of intestinal flora,and can be used as potential probiotics for pets.Probiotic strains,and the experimental data of this study provide data support for the industrial application of these four strains. |
