Inhibition Effect Of Bovine Bacillus Amyloliquefaciens And Its Antibacterial Protein On Salmonella

Salmonella can cause gastroenteritis in human,livestock and poultry.It not only causes enormous economic losses to the farming industry,but also endangers global public health.Antibiotics were once commonly used to prevent Salmonella infection.However,with the increasing frequency of antibiotic use,Salmonella has developed serious resistance to antibiotics.In this context,there is an urgent need for new antiseptic products to prevent and treat Salmonella infection.Bacillus can synthesize a wide range of secondary metabolites,including Surfactants,Ituin and Fengycin such as ibuprofen and Cardamine,proteins such as chitosanase and volatile organic compound,it has good antagonism to animal and plant pathogens.The aim of this study was to explore the role of Bacillus and its antibacterial protein in the prevention and control of Salmonella infection of bovine origin,and to lay a foundation for the development of new anti-salmonella drugs.In this experiment,26 strains of Bacillus were isolated from the fresh feces of healthy cattle and sheep,among which 4 strains showed strong antagonism against Salmonella,a resistant strain Ba4-8 was obtained and identified as Bacillus amyloliquefaciens by Gram staining,spore staining,molecular biology and biochemical tests.Using acid precipitation to precipitate the protein substance in the fermentation broth of Ba4-8,PBS was resuspended to obtain the crude extract of the protein antibacterial substance.The antifungal substances were isolated and purified by Sephadex G-75 gel chromatography column and QAE-Sephadex A-25 ion exchange column,SDS-PAGE showed that the size of the antifungal protein was about 11 k Da.Salmonella infection model was established in mice,the transcription levels of Occludin,Claudin-1 and ZO-1 genes in mouse ileum were detected by RT-q PCR.The changes of jejunum villi were observed,and the levels of IL-6 and TNF-α in serum and the amount of bacteria in spleen and jejunum were detected.It was found that Ba4-8,Ba4-8 antibacterial protein and Ba4-8 lysates could protect intestinal epithelial barrier integrity by up-regulating the levels of Occludin,Claudin-1 and ZO-1,and decrease the content of pro-inflammatory factors and Salmonella load.To sum up,Bacillus amyloliquefaciens and its antibacterial protein have inhibitory effect on Salmonella in vitro and in vivo,which lays a foundation for further research and application of Bacillus amyloliquefaciens in antibacterial field.