Bioprospecting, Isolation And Identification Of The Verrucosispora Strains

Development and application of molecular biology, bioinformatics and "omics" technologies provides not only new technologies but also new strategies for exploring microbial resources. The primary aim of this study was to isolate members of the genus Verrucosispora that has been proved to be a rich resource of new bioactive compounds, using the strategy of’bioprospecting-biodiversity-selective isolation’, highlighting the biotechnological potential and ecological significance of the members of this taxon.A set of genus-specific oligonucleotide primers was generated based on signatural nucleotides sequences of the16S rRNA genes of Verrucosispora strains deposited in GenBank. In silico and pure culture studies showed that the primers matched perfectly with target sequences of16S rRNA gene of representatives of the genus Verrucosispora. The primers, designated S-G-Verr-0195-a-S-20and S-G-Verr-1152-a-A-18, amplified an≈960bp stretch of16S rRNA genes of Verrucosispora strains, but not those of representatives of other genera classified in the family Micromonosporaceae. Genus-specific amplicons were detected in17out of20community DNA samples prepared from diverse marine sediments and coastal soils. Phylogenetic analysis of over40%of clones derived from5of the samples belonged to novel Verrucosispora species. The primers were also used to confirm the identity of794Verrucosispora-like strains isolated from10positive samples and3negative samples using9selective media. Twenty one isolates,18from the Megas Gialos sediment and3from the Wenchang mangrove sample, were provisionally assigned to the genus Verrucosispora on the basis of colony morphology gave the genus-specific amplification product with primers S-G-Verr-0195-a-S-20and S-G-Verr-1152-a-A-19.Metabolites of5Verrucosispora strains inhibited Staphylococcus aureus strain SO14, and3out of5strains inhibited MRSA (Methicillin Resistant Staphylococcus aureus) strain9551. The novel candidate species2603PH03was found significant active against these two pathogen strains with inhibition ratio of91.87%and74.78%, respectively. Moreover, this strain was different from other members of the genus Verrucosispora on patterns of PKSs and NRPSs genes, which showed a potential for further research on drug discovery against MRSA.Polyphasic taxonomic study of strain234402and strain2603PH03gave result in two novel candidate species, near to V. lutea and V. gifhornensis, respectively. Strain232076and strain234606were assigned to the species V. sediminis.