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A polyketide biosynthesis gene cluster is required for production of bactericidal activity by Burkholderia contaminans strain MS14.

Shien Lu: Mississippi State University

<div><em>Burkholderia contaminans </em>MS14 produces the antifungal glycopeptide occidiofungin via a nonribosomal peptide synthase mechanism. MS14 also possesses a broad range of antibacterial activities against Gram negative bacteria, such as <em>Erwinia amylovora</em> and <em>Xanthomonas citri</em>. Random mutagenesis and gene complementation revealed that four genes are required for production of MS14 antibacterial activity. However, sequence-based function prediction indicated that none of them are directly involved in biosynthesis of the bactericidal activity. In order to identify the biosynthetic gene cluster for the unknown antibacterial compound production, RNA-Seq was conducted to analyze the transcriptomes of the MS14 wild type and its mutants that lack antibacterial activity. A polyketide synthases (PKS) gene cluster was predicted to be directly involved in antibacterial activity. Mutants of the two PKS genes were generated using kanamycin cassette insertion and CRISPR-Cas9 system. Plate bioassays showed that the mutations in the two PKS genes resulted in undetectable antibacterial activity against <em>E. amylovora</em>. As expected the mutants defective in antibacterial activity retained the wild-type production level of occidiofungin. The results suggest the PKS gene cluster is directly responsible for the production of MS14 bactericidal activity.</div>