Genome-Guided Isolation of Novel Antibiotics Encoded by Nonribosomal Peptide Synthetase Pathways in Flavobacteria

Abstract

The emergence and spread of multidrug-resistant pathogens has made the development of novel antibiotic compounds an important global health concern. Many existing drugs previously used as first-line treatments for bacterial and fungal infections are becoming less effective due to antibiotic resistance, a problem significantly exacerbated by the dramatic decline in research and development of new antimicrobial drugs over the past 25 years. Importantly, bacteria are rich resources of bioactive small molecule compounds, encoded by biosynthetic gene clusters (BGCs), with great therapeutic potential. Actinobacteria have traditionally been thought to be the best producers of these and other drug-like molecules. However, here we use genome mining to identify Flavobacteria strains rich in nonribosomal peptide synthetase (NRPS)-containing gene clusters, which are predicted to encode lipopeptide antibiotics even though no known lipopeptides have been previously isolated from Flavobacteria or any other members of the Bacteriodetes phylum. We performed both chemical and genetic analyses of metabolite compounds extracted from nine Flavobacteria isolates to provide experimental support for these genomic predictions, and found various degrees of antimicrobial activity against indicator strains representing a spectrum of Gram-negative and Gram-positive bacteria and fungal pathogens. To identify the active components, we employed bioassay-guided fractionation and chromatographic separation. In addition, we attempted to develop genetic tools to manipulate these Flavobacteria strains for use in gene knockout and molecular cloning studies, which may be used to analyze these and other related BGCs in the future. More work will be required to understand the bacterial genetics of these environmental isolates. Our findings establish an important link between the gene sequence, chemical structure, and biological activity of genetically encoded compounds isolated from Flavobacteria, and provide a basis for further investigations into their promising antibiotic potential.