Dataset Information

Broad Spectrum Antimicrobial Activity of Forest-Derived Soil Actinomycete, Nocardia sp. PB-52.

ABSTRACT: A mesophilic actinomycete strain designated as PB-52 was isolated from soil samples of Pobitora Wildlife Sanctuary of Assam, India. Based on phenotypic and molecular characteristics, the strain was identified as Nocardia sp. which shares 99.7% sequence similarity with Nocardia niigatensis IFM 0330 (NR_112195). The strain is a Gram-positive filamentous bacterium with rugose spore surface which exhibited a wide range of antimicrobial activity against Gram-positive bacteria including methicillin-resistant Staphylococcus aureus (MRSA), Gram-negative bacteria, and yeasts. Optimization for the growth and antimicrobial activity of the strain PB-52 was carried out in batch culture under shaking condition. The optimum growth and antimicrobial potential of the strain were recorded in GLM medium at 28°C, initial pH 7.4 of the medium and incubation period of 8 days. Based on polyketide synthases (PKS) and non-ribosomal peptide synthetases (NRPS) gene-targeted PCR amplification, the occurrence of both of these biosynthetic pathways was detected which might be involved in the production of antimicrobial compounds in PB-52. Extract of the fermented broth culture of PB-52 was prepared with organic solvent extraction method using ethyl acetate. The ethyl acetate extract of PB-52 (EA-PB-52) showed lowest minimum inhibitory concentration (MIC) against S. aureus MTCC 96 (0.975 ?g/mL) whereas highest was recorded against Klebsiella pneumoniae ATCC 13883 (62.5 ?g/mL). Scanning electron microscopy (SEM) revealed that treatment of the test microorganisms with EA-PB-52 destroyed the targeted cells with prominent loss of cell shape and integrity. In order to determine the constituents responsible for its antimicrobial activity, EA-PB-52 was subjected to chemical analysis using gas chromatography-mass spectrometry (GC-MS). GC-MS analysis showed the presence of twelve different chemical constituents in the extract, some of which are reported to possess diverse biological activity. These results confirmed that the presence of bioactive constituents in EA-PB-52 could be a promising source for the development of potent antimicrobial agents effective against wide range of microbial pathogens including MRSA.

SUBMITTER: Sharma P

PROVIDER: S-EPMC4796592 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Broad Spectrum Antimicrobial Activity of Forest-Derived Soil Actinomycete, Nocardia sp. PB-52.

Sharma Priyanka P Kalita Mohan C MC Thakur Debajit D

Frontiers in microbiology 20160318

A mesophilic actinomycete strain designated as PB-52 was isolated from soil samples of Pobitora Wildlife Sanctuary of Assam, India. Based on phenotypic and molecular characteristics, the strain was identified as Nocardia sp. which shares 99.7% sequence similarity with Nocardia niigatensis IFM 0330 (NR_112195). The strain is a Gram-positive filamentous bacterium with rugose spore surface which exhibited a wide range of antimicrobial activity against Gram-positive bacteria including methicillin-re ...[more]

PMID: 27047463

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Project description:Emergence of antimicrobial resistance coupled with the slowdown in discovery of new antimicrobial compounds points to serious consequences for human health. Therefore, scientists are looking for new antimicrobial compounds from unique and understudied ecosystems such as tropical peat swamp forests. Over the course of isolating antimicrobial producing bacteria from North Selangor tropical peat swamp forest, Malaysia, a Gram variable, rod shaped, endospore forming, facultative anaerobic novel strain MSt1(T) that exerts potent and broad spectrum antimicrobial activity was isolated. Phylogenetic analysis using 16S rRNA gene sequences showed that strain MSt1(T) belonged to the genus Paenibacillus with the highest similarity to Paenibacillus elgii SD17(T) (99.5%). Whole genome comparison between strain MSt1(T) with its closely related species using average nucleotide identity (ANI) revealed that similarity between strain MSt1(T) with P. elgii B69 (93.45%) and Paenibacillus ehimensis A2 (90.42%) was below the recommended threshold of 95%. Further analysis using in silico pairwise DDH also showed that similarity between strain MSt1(T) with P. elgii B69 (55.4%) and P. ehimensis A2 (43.7%) was below the recommended threshold of 70%. Strain MSt1(T) contained meso-diaminopilemic acid in the cell wall and MK-7 as the major menaquinone. The major fatty acids of strain MSt1(T) were anteiso-C15:0 (48.2%) and C16:0 (29.0%) whereas the polar lipid profile consisted of phosphatidylglycerol, phosphatidylethanolamine, diphosphatidylglycerol, one unknown lipid, two unknown glycolipids, and one unknown phospholipid. Total DNA G+C content of strain MSt1(T) was 51.5 mol%. The extract from strain MSt1(T) exerted strong antimicrobial activity against Escherichia coli ATCC 25922 (MIC = 1.5 μg/mL), MRSA ATCC 700699 (MIC = 25 μg/mL) and Candida albicans IMR (MIC = 12.5 μg/mL). Partially purified active fraction exerted a strong effect against E. coli ATCC 25922 resulting in cell rupture when viewed with SEM. Based on distinctive taxonomic differences between strain MSt1(T) when compared to its closely related type species, we propose that strain MSt1(T) represents a novel species within the genus of Paenibacillus, for which the name Paenibacillus tyrfis sp. nov. (= DSM 100708(T) = MCCC 1K01247(T)) is proposed.

Project description: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 Methylobacterium 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 °C. 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 typhimurium, 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: 250 µg/mL against S. 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-Methylbutanoic acid (5.03%), isobutyramide (5.05%), N,N-oimethylformamide-di-t-butylacetal (9.79%), benzeneacetamide (15.56%), octyl butyl phthalate (3.59%) and diisooctyl phthalate (5.79) in the extract. CONCLUSIONS: Methylobacterium sp. (ERI-135) showed promising antibacterial and cytotoxic activity. This is the first report in the antimicrobial and cytotoxic effect of Methylobacterium sp.

Project description:The Actinomycetia isolate PBR11 was isolated from the forest rhizosphere soil of Pobitora Wildlife Sanctuary (PWS), Assam, India. The isolate was identified as Streptomyces sp. with 92.91% sequence similarity to their closest type strain, Streptomyces atrovirens NRRL B-16357 DQ026672. The strain demonstrated significant antimicrobial activity against 19 test pathogens, including multidrug-resistant (MDR) clinical isolates and dermatophytes. Phenol, 2,5-bis(1,1-dimethylethyl), is the major chemical compound detected by gas chromatography-mass spectrometry in the ethyl acetate extract of PBR11 (EtAc-PBR11). The presence of the PKS type II gene (type II polyketide synthases) and chitinase gene suggested that it has been involved in the production of antimicrobial compounds. Metabolic profiling of the EtAc-PBR11 was performed by thin-layer chromatography and flash chromatography resulted in the extraction of two bioactive fractions, namely, PBR11Fr-1 and PBR11Fr-2. Liquid chromatography-tandem mass spectrometry analysis of both the fractions demonstrated the presence of significant antimicrobial compounds, including ethambutol. This is the first report on the detection of antituberculosis drug in the bioactive fractions of Streptomyces sp. PBR11. EtAc-PBR11 and PBR11Fr-1 showed the lowest MIC values (>0.097 and >0.048 μg/mL, respectively) against Candida albicans MTCC 227, whereas they showed the highest MIC values (>0.390 and >0.195 μg/mL, respectively) against Escherichia coli ATCC BAA-2469. The effects of PBR11Fr-1 were investigated on the pathogens by using a scanning electron microscope. The results indicated major morphological alterations in the cytoplasmic membrane. PBR11Fr-1 exhibited low cytotoxicity on normal hepatocyte cell line (CC-1) and the percent cell viability started to decline as the concentration increased from 50 μg/mL (87.07% ± 3.22%) to 100 μg/mL (81.26% ± 2.99%). IMPORTANCE Novel antibiotic breakthroughs are urgently required to combat antimicrobial resistance. Actinomycetia are the principal producers of antibiotics. The present study demonstrated the broad-spectrum antimicrobial potential of an Actinomycetia strain Streptomyces sp. strain PBR11 isolated from the PWS of Assam, India, which represents diverse, poorly screened habitats for novel microorganisms. The strain displayed 92.4% sequence similarity with genes of the closest type strain, indicating that the strain may represent a novel taxon within the phylum Actinomycetota. The metabolomics studies of EtAc-PBR11 revealed structurally diverse antimicrobial agents, including the detection of the antituberculosis drug ethambutol, in the bioactive fraction of Streptomyces sp. PBR11 for the first time. The PBR11 strain also yielded positive results for the antibiotic synthesis gene and the chitinase gene, both of which are responsible for broad-spectrum antimicrobial activity. This suggests that the untouched forest ecosystems have a tremendous potential to harbor potent actinomycetia for future drug discovery.

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Dataset Information

Broad Spectrum Antimicrobial Activity of Forest-Derived Soil Actinomycete, Nocardia sp. PB-52.

Publications

Broad Spectrum Antimicrobial Activity of Forest-Derived Soil Actinomycete, Nocardia sp. PB-52.

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