Screening And Molecular Identification Of Bacteriostatic Bacillus Pumilus4D-14, Probiotics Development And Application Research

First, this paper established a rapid screening method of laboratory96pores plate based onminimum inhibitory concentration and screened Bacillus strains from farm soil and probioticproducts for animals. Then selected Bacillus strains using Salmonella typhimurium ATCC14028as the indicator bacteria to antibacterial ability of Bacillus, obtained7antibacterial Bacillusstrains from farm soil. The antibacterial effect of7Bacillus strains from farm soil and Bacillusstrains from probiotic products were evaluated with4strains of different serotypes were furtherused animal source pathogenic bacteria Escherichia coli E1, E3, E4, Salmonella gallinarum(Salmonella gallinarum) SC1, swine Salmonella (Salmonella choleraesuis) and SP1ofPseudomonas aeruginosa, found that the antibacterial effect of Bacillus strains4D-14, J-14from the soil and Bacillus strains Y14, Y16from the market products analogous to20ppmcolistin sulfate in vitro.Bacillus cereus, Bacillus pumilus and Bacillus mycoides highlighted the Bacillus srtains in farmsoil and Bacillus subtilis highlighted the Bacillus srtains in market animal probiotic products,according to the bacteriostasis Bacillus identification with the use of molecular16SrRNA andgyrB gene method. The characters of Bacillus supernatant of the strain Bacillus subtilis Y16,Bacillus lincheniformis Y14and Bacillus cereus J14, Bacillus pumilus4D-14was observedthrough the changes of antibacterial activity of protease, temperature, different pH, and foundthat properties of antibacterial activity of different strains were different. Antibacterial activityof B. lincheniformis Y14fluctuations was affected by protease and temperature, antifungalactivity in the supernatant may be provided by proteins. B. subtilis Y16, B. cereus J14, B.pumilus4D-14antibacterial activity is not affected by protease and temperature, especially the B.cereus J14, B. pumilus4D-14antibacterial material has thermal stability, show its antibacterialactivity may be provided by small peptides. Considering the restrictive provision of microbial strains used for animal feed and breeding ofChina’s agriculture ministry and the research status of animal probiotics, selected the strain B.pumilus4D-14to do further research in application effect in this paper. At the same time, thestrain B. subtilis Y16which screened from market animal probiotic product was selected as thecontrol study.The probiotic samples of B. pumilus4D-14and B. subtilis Y16were prepared with20Lfermentation tank. Finally, prepared B.pumilus4D-14culture2.35kg, bacteria quantity was1.47×1010CFU/g; B. subtilis Y16culture2.7kg, bacteria quantity was6.8×109CFU/g.In the application research, using Avilamycin as positive control, B. pumilus4D-14, B. subtilisY16and market representative Bacillus probiotics as treatment on broiler breeding test.Throughing the analysis of the Ross308male broilers production performance, immune indexand intestinal morphology, found that Bacillus pumilus (B. pumilus4D-14) can effectivelyreduce the late breeding ratio of feed and meat and to promote the intestines recess is shorter,the epithelial cell survival rate increased; and can replace Avilamycin, used as a growthpromoting agent used in broiler feed.