Screening And Diversity Revelation Of PKS And NRPS Gene In Sponge Associated Microbes

Sixteen strains with ketosynthase (KS) domain genes and fifteen strains with nonribosomal peptide synthetase (NRPS) genes were obtained by PCR screening among 109 isolates from four South China Sea sponges, Stelletta tenuis, Halichondria rugosa, Dysidea avara and Craniella australiensis. Meanwhile, strains with PKS and NRPS gene fragments were tested for their antimicrobial potential against 6 indicator strains.By BLAST and phylogenetic analysis of 16S rDNA, the 26 strains were grouped into three bacterial groups including Firmicutes (22/26), Proteobacteriu (2/26), and Actinomycetes (2/26). Most of the strains were grouped into phylum Firmicutes especially Bacillus sp. except Staphylococcus lentus A75. DA20 and DA23 were identified as Actinomycetes. A05 and A11 matched Alcaligenes sp. of phylum Proteobacterium.The obtained 16 KS domain sequences were 690 bp in length. The BLAST result showed that the KS domains were most closely related to the KS sequences of Bacillus subtilis with relatively high confidence (above 91%). Phylogenetic analysis demonstrated the KS domains belong to trans-AT type I PKSs, which were commonly involved in the biosynthesis of pharmacologically active polyketides.The obtained 15 NRPS sequences were 1000 bp in length. The BLAST result indicated that they were related to adenylation (A) domain of NRPS and most of them(10/15) showed low similarity (<70%)to the known sequence. Base on the phylogenetic analysis results of 16S rDNA and adenylation (A) domain fragments, it can be deduced that six NRPS fragments are novel.Based on antibiotic activity assay, the 26 isolates with PKS and NRPS genes exhibited broad-spectrum antimicrobial activities against fungi, gram-positive bacteria and gram-negative bacteria.These findings demonstrate:(1)NRPS and PKS gene cluster widely exist in the bacteria associated with the four South China Sea marine sponges and distribute in different phylum including Firmicutes,Proteobacteriu, and Actinomycetes. (2) These results reveal the great diversity and novelty of both PKS and NRPS genes in the South China Sea. It will be helpful to find out valuble pharmacological strain which will produce novel natural compounds in the further study.Our study suggests the role of sponge-symbiosis bacteria played in the host's chemical defense and provides further proof for the hypothesis that sponge-associated bacteria are the true producers of many novel bioactive compounds.