Characterization and phylogenetic analysis of novel polyene type antimicrobial metabolite producing actinomycetes from marine sediments:Bay of Bengal India

Objective:To isolate and indentify the promising antimicrobial metabolite producing Streptomyces strains from marine sediment samples from Andraprudesh coast of India.Methods:Antagonistic aetinomycetes were isolated by starch casein agar medium and modified nutrient agar medium with 1%glucose used as a base for primary screening.Significant antimicrobial metabolite producing strains were selected and identified by using biochemical and 16S rDNA level.Minimum inhibitory concentrations of the organic extracts were done by using broth micro dilution method.Results:Among the 210 actinomyeetes,64.3%exhibited activity against Gram positive bacteria,48.5%showed activity towards Cram negative bacteria,38.8%exhibited both Cram positive and negative bacteria and 80.85%isolates revealed significant antifungal activity.However,five isolates AP-5,AP-18,AP-41 and AP-70 showed significant antimicrobial activity.The analysis of cell wall hydrolysates showed the presence of LL-diaminopimelic acid and glycine in all the isolates.Sequencing analysis indicated that the isolates shared 98.5%-99.8%sequence identity to the 16S rDNA gene sequences of the Streptomyces taxons.The antimicrobial substances were extracted using hexane and ethyl acetate from spent medium in which strains were cultivated at 30X for five days.The antimicrobial activity was assessed using broth micro dilution technique.Each of the culture extracts from these five strains showed a typical polyenelike property.The lowest minimum inhibitory concentrations of ethyl acetate extracts against Escherichia coli and Cumularia lunula were 67.5 and 125.0μg/mL,respectively.Conclusions:It can be concluded that hexane and ethyl acetate soluble extracellular products of novel isolates are effective against pathogenic bacteria and fungi.

Cytotoxic (A549) and antimicrobial effects of Methylobacterium sp.isolate (ERI-135) from Nilgiris forest soil,India

Objective:To assess the antimicrobial and cytotoxic effects of Methylobacterium sp.isolated from soil sample of Doddabetta forest,Nilgiris,Western Ghats of Tamil Nadu.Methods:Isolation of Methylobacterium was performed from soils by serial dilution plate technique.The strain was grown in modified nutrient gulucose agar(MNGA)medium to study the morphology and biochemical characteristics.Methylobacterium sp.was screened for its antimicrobial activity against pathogenic bacteria and fungi.The strain was subjected to 16S rRNA analysis and was identified as Methylohacterium sp.The nucleotide sequence of the 16S rRNA gene of the isolate exhibited close similarity with other Methylobacterium sp.and has been submitted to Genbank.The antibacterial substances were extracted using chloroform and ethyl acetate from MNGA medium in which ERI-135 had grown for 5 d at 30℃.Cytotoxic effect was also studied.GC-MS analysis was carried out.The antimicrobial activity was assessed using broth micro dilution technique.Results:Ethyl acetate extract showed activity against bacteria such as Bacillus subtilis,Klebsiella pneumoniae(K.pneumoniae),Pseudomonas aeruginosa,Salmonella typkimurium,Shigella flexneri,Enterobacter aerogenes,Staphylococcus aureu and Staphylococcus epidermidis(S.epidermidis)and fungi such as,Candida albicans and Trichophyton rubrum.The lowest minimum inhibitory concentrations were:250μg/mL against 5.epidermidis and 250μg/mL against K.pneumonia.The isolate had the ability to produce enzymes such as protease.The exyract showed cytotoxic effect in human adenocarcinoma cancer cell line(A549).GC-MS analysis showed the presence of isovaleric acid(3.64%),2-Methylbulanoic acid(5.03%),isobutyramide(5.05%),N,Noimethylformamide-di-t-butylacetal(9.79%),benzeneacetamide(15.56%),octyl butyl phthalate(3.59%)and diisooctyl phthalate(5.79)in the extract.Conclusions:Methylobaeterium sp.(ERI-135)showed promising antibacterial and cytotoxic activity.This is the first report in the antimicrobial and cytotoxic effect of Methylobaeterium sp.