Microbiological and Molecular studies on Antibiotic producing Rare Actinomycetes

Abstract

Fifty rare actinomycetes were successfully isolated from Egyptian soils and showed significant antimicrobial activities against some bacterial pathogens. Among which, Micromonospora Rc5 and Streptomyces Ru87 reported the highest spectrum of antimicrobial activities. Morphological, chemotaxonomic and DNA sequencing studies showed that Rc5 and Ru87 belonged to genera Micromonospora, Streptomyces, respectively. Phylogenetic analysis indicated that both strains were distinct from the already known related species. Optimization, extraction and Partial characterization of the active bands were performed. We confirmed that the active compound produced by Micromonospora sp. Rc5 was phthalate derivative and aromatic peptides in case of Streptomyces sp. Ru87. Molecular studies confirmed that both strains recorded positive hits of NPRS and PKS genes. Fosmid genomic libraries were successfully constructed and integrated with NRPS and PKS genes. Neighbor-Joining phylogenetic tree of PKS gene fragments of both Micromonospora sp. Rc5 and Streptomyces sp. Ru87 were highly distinctive with low bootstrap value (28%, 42%, respectively). This is to confirm that Micromonospora sp. Rc5 and Streptomyces sp. Ru87 strains are unique. They both harbor phylogenetically divergent PKS gene cluster that is responsible for distinct biochemical pathways of novel antibiotics. To our knowledge, this is first attempt to tackle phylogenic analysis of NRPS and PKS genes and discover uncommon and novel types of actinomycetes isolated from Egyptian soils. These novel isolates may contain unexpected genes, and subsequently, offers a significant contribution to alternative novel antimicrobial compounds in drug discovery. Furthermore, Illumina whole genome sequencing was conducted and the genomic features of each strain were recorded. Gene annotation using antibiotics and Secondary Metabolite Analysis SHell (antiSMASH) predicted the presence of valuable biosynthetic clusters in both strains. Based on antiSMASH results and physiochemical analysis results, we identified the active band produced by Micromonospora sp. Rc5 as tetrocarcin and Labyrinthopeptins in case of Streptomyces sp. Ru87.