Discovery,Isolation,Identification and Characterization of Novel Non-ribosomal Peptide Antibacterial Active Substances in Bacillus

The aim of this study was to search for novel non-ribosomal peptide antimicrobial substances based on the screening of bacterial secondary metabolites.The bacteria isolated from soil,sea water and common marine organisms in Yantai coastal area were isolated and purified.E.coli ATCC 25922 and Staphylococcus aureus ATCC 29213 were selected as indicator bacteria,and E.coli B2 (bla_NDM-5+mcr-1) and methicillin-resistant Staphylococcus aureus (MRSA) T144 were used as indicator for secondary screening.The genome was extracted and the PCR products were sequenced to determine the active species.The secondary metabolites of bacteria were extracted by organic extraction,purified by gel chromatography and preparative liquid chromatography,and purity was detected by analytical liquid chromatography.The results of sequencing showed that the active strain belonged to Bacillus amyloliquefaciens sp.,and was named as Bacillus amyloliquefaciens 9-14 (active bacterium 9-14).The results of antibacterial test showed that the metabolites of active bacterium 9-14 had high inhibitory effect on Staphylococcus aureus ATCC 29213,MRSA T144,E.coli ATCC 25922 and E.coli B2.The metabolites of active bacterium 9-14 were cyclic lipopeptides composed of amino acid chains,which belonged to the derivatives of ibuprofen.The biological characteristics and antibacterial spectrum of the metabolites of active bacterium 9-14 were studied.The results showed that the metabolites of active bacterium 9-14 had good thermal stability and acid-base stability.And the antibacterial activity of the antibacterial substance treated with trypsin,pepsin,protease K and papain was not significantly weakened and had good stability.The antibacterial substance also had inhibitory effect on Staphylococcus aureus and E.coli,but had no activity against Pseudomonas aeruginosa,Klebsiella pneumoniae,Enterococcus faecalis and Bacillus cereus.In this study,a new antibacterial substance was obtained,which could be used as the precursor of antibacterial drugs,and could provide certain reference for food safety and disease control.