Screening and identification of antagonistic strains against Staphylococcus aureus

Abstract:

[Background] The secondary metabolites of endophytic bacteria are the important sources of new natural active substances. [objective] We aimed to isolate the bacterial strains and their secondary metabolites with antibacterial activity against Staphylococcus aureus from Paeonia lactiflora. [Methods] The antagonistic strains were screened against Staphylococcus aureus through the method of dual culture. The strains were indentified based on characteristics in morphology and DNA identification. The biosynthetic genes of lipopeptide were amplified using polymerase chain reaction, and the antibacterial activity of their fermentation and crude lipopeptides were detected using the method of Oxford cup. The antibacterial activity of the crude lipopeptides was isolated through Sephadex LH-20 gel chromatography, which was analyzed by MALDI-TOF mass spectrometry. [Results] A total of 13 antagonistic strains were screened from the endophytic bacteria isolates, which had different degrees of inhibition against Staphylococcus aureus. The isolates SY11 had the most obvious inhibition ability, and their fermentation and crude lipopeptides had strong inhibitory effect. The isolates SY11 was identified as Bacillus amyloliquefaciens based on characteristics in morphology and phylogenetic analysis of 16S rRNA gene sequences. Three biosynthetic genes of lipopeptide, fenA, ituD and srfkn were detected using polymerase chain reaction. The results suggested that the strain had the ability to synthesize lipopeptide. By further analysis of MALDI-TOF mass spectrometry. The main active substance was presumed as Bacillomycin D. [Conclusion] Bacillus amyloliquefaciens SY11 had significant inhibitory effect on Staphylococcus aureus, and their crude lipopeptides also showed inhibitory activity against Staphylococcus aureus in vitro, which laid the foundation for the further development and application of endophytes from Paeonia lactiflora.