Antibacterial Mechanism Of Bacillus From Shanxi Aged Vinegar And Its Inhibition Quorum Sensing Of Spoilage Bacteria

In this study,the antibacterial mechanism of Bacillus with antibacterial activity isolated from Shanxi aged vinegar and its inhibitory effect on the signal molecules of pathogenic bacteria were studied and applied to prevent the corruption of fresh-cut apple.The results were as follows:(1)The antibacterial activity of 75 strains of bacillus isolated from shanxi aged vinegar was screened.The bacteriostatic effects of Escherichia coli,Staphylococcus aureus and Aspergillus niger 710 were determined by microbroth or bacteriostatic circle method.The bacillus with high bacteriostatic rate was further screened by the bacteriostatic circle method for Escherichia coli and Staphylococcus aureus.The results showed that the cell-free extract of Bacillus atrophaeus CP 297 in 75 strains of Bacillus had the highest inhibition rate against Escherichia coli,which was 77.03%,and could produce 18.90 mm inhibition zone;The cell-free extract of Bacillus licheniformis CP 1671 had the highest inhibition rate against Staphylococcus aureus,up to 96.5%,and the inhibition zone diameter was 20.20 mm.The cell-free extract of B.amyloliquefaciens CP 2014 had obvious antibacterial effect on Aspergillus niger 710.(2)The antibacterial spectrum and physicochemical properties of the bacteriostatic products of Bacillus atrophaeus CP 297,Bacillus licheniformis CP 1671 and Bacillus amyloliquefaciens CP 2014 with high inhibition rate isolated from Shanxi aged vinegar were determined.The results showed that the cell-free extract of Bacillus atrophaeus CP 297 and Bacillus licheniformis CP 1671 could not only inhibit Gram-positive bacteria,but also Gram-negative bacteria.And they had a broad spectrum of inhibition against Escherichia coli and Shigella flexneri,Salmonella,Klebsiella pneumoniae,Enterobacter hormaechei,Pseudomonas aeruginosa;Bacillus amyloliquefaciens CP 2014 cell-free extract had obvious inhibition zones which against Aspergillus niger,Aspergillus flavus,Mucor,Fusarium oxysporum,Penicillium.The spectrum of inhibition was wider.The optimum p H for antibacterial activity of the antibacterial products of Bacillus atrophaeus CP 297,Bacillus licheniformis CP 1671 and Bacillus amyloliquefaciens CP 2014 were 7.0~8.0.Under the action of acid,alkali and high temperature,the antibacterial activity were reduced and it is sensitive to protease.(3)Through determining of potassium ion concentration,AKP content,conductivity,protein content,bacterial cells,hyphal morphology,etc,the antibacterial mechanism of Bacillus atrophaeus CP 297cell-free extract(CFE)against Escherichia coli,Bacillus licheniformis CP 1671 CFE against Staphylococcus aureus,Bacillus amyloliquefaciens CP 2014 CFE against Aspergillus niger 710 werestudied.The results showed that Bacillus atrophaeus CP 297 and Bacillus licheniformis CP 1671 CFE could destroy the integrity of the cell wall of Escherichia coli and Staphylococcus aureus,respectively.And it had a certain degree of damage to the cell membrane.Thus it played a suppressing role;Bacillus amyloliquefaciens CP 2014 CFE can inhibit the growth of mycelium and the germination of spore of Aspergillus niger 710,thus inhibiting Aspergillus niger.(4)By analyzing the crude extracts of Escherichia coli,Staphylococcus aureus,Shigella flexneri,Salmonella and Listeria monocytogenes,it was found that all 5 pathogens produced cyclo-(L-Pro-L)-Gly),cyclo-(L-Pro-L-Leu)and cyclo-(L-Pro-L-Phe)three types of DKPs signaling molecules.By studying the effects of Na Cl concentration,p H value,temperature and CFE on the signal molecule DKPs,it was found that with the increase of Na Cl concentration,the number of signal molecules decreased,and the relative content of various DKPs signal molecules decreased.When the p H and temperature rised,the relative content of each signal molecule increased,and then decreased.The cell-free extract of Bacillus could also inhibit the ability of the producing signal molecules DKPs of spoilage pathogens,and reduce the species and relative content of the signal molecules DKPs.(5)By studying the sensory evaluation,the number of microbial colonies and the changes of physical and chemical indicators of fresh-cut apple under different treatments of without bacterial pathogen,bacterial pathogen mixture and mold spore suspension,the results showed that compared with the fresh-cut apple after treatment with sterile water,fresh-cut apples treated with the addition of exogenous signal molecule DKPs accelerate the decay of apples significantly;the treatment with the Bacillus antibacterial product(CAE)had a certain effect of preservation,and had obvious antibacterial effect.The number of colonies of spoilage bacteria was reduced,indicating that CAE had a preventive effect on the corruption of fresh-cut apples;Treatment with the signal molecule DKPs accelerated the decay of apples significantly.