Identification and characterization of a novel antimicrobial peptide compound produced by Bacillus megaterium strain isolated from oral microflora.

In recent years, the decreased efficacy of existing antibiotics toward management of emergent drug-resistant strains has necessitated the search for novel antibiotics from natural products. In this regard, Bacillus sp is well known for producing variety of secondary metabolites of potential use. Therefore, we performed an investigation to isolate and identify Bacillus sp from oral cavity for production of novel antimicrobial compounds. We extracted, purified, and identified a novel bioactive compound by B. megaterium (KC246043.1). The optimal production of compound was observed on de Man Rogosa and Sharpe broth by incubating at 37 °C, and pH 7.0 for 4 days. The bioactive compound was extracted by using n-butanol (2:1 v/v), purified on TLC plates with detection at Rf 7.8 cm; further characterized and identified as a cyclic ploypeptide sharing structural similarity with bacitracin. Minimum inhibitory concentration of bioactive compound was found to be 0.25, 0.5, 1.0, 3.125 and 6.25 µg/ml against Micrococcus luteus ATCC10240, Salmonella typhi ATCC19430, Escherichia coli ATCC35218. Pseudomonas aeruginosa ATCC27853 and Staphylococcus aureus ATCC25923 respectively, with no activity against Candida albicans ATCC10231. Our findings have revealed a novel cyclic peptide compound from B. megaterium with broad spectrum antimicrobial activity against both Gram positive and Gram negative bacteria.

Novel Strategies for Combating Pathogenic Biofilms Using Plant Products and Microbial Antibiosis.

Microorganisms prefer to live in three-dimensional self-organized communities (biofilms), and this behavior provides microbial pathogens inhabiting various sites in the human body or on medical devices with survival advantages. In fact, pathogens in the biofilm stage exhibit up to a thousandfold more tolerance to conventional antimicrobial agents, and thus, they are difficult to eradicate and biofilms generated during acute infections become persistent, chronic, and recurrent. Consequently, novel strategies are being sought to control biofilm associated infections. The developmental strategies used include improved drug delivery and the penetration of biofilm matrices, and in particular, natural products that interfere with virulence and cross talk between microbial cells are being investigated as potential anti-biofilm agents. This article provides an overview of existing and promising biofilm control strategies based on plant and microbial products. Control strategies like quorum sensing inhibition, microbial antibiosis, and the uses of phages and probiotics are reviewed along with current developments in high throughput screening and in our understanding of structure activity relationships related to the regulation of biofilms by small molecules.

Sub-MICs of Carum copticum and Thymus vulgaris influence virulence factors and biofilm formation in Candida spp.

BACKGROUND: Emergence of drug-resistant strains of Candida and inefficiency of conventional antifungal therapy has necessitated the search for alternative and new antifungal agents. Inhibition of virulence and biofilm are the potential drug targets. In this study, the oils of Carum copticum, Thymus vulgaris and their major active compound thymol as revealed by Gas chromatography and gas chromatography-mass spectrometry (GC-GC/MS) analysis were tested for their inhibitory activity against growth to determine sub-MIC values against 27 drug-resistant strains of Candida spp. METHODS: Brothmacrodilution method was used for determination of MIC of test oils against Candida strains. The spectrophotometric methods were used for detection and inhibition assays for virulence factors in Candida spp. Light and electron microscopy was performed to observe morphological effects of oils on biofilms. GC-GC/MS were used to evaluate the major active compounds of test oils. RESULTS: Virulence factors like proteinase and haemolysin were detected in 18 strains, both in solid and liquid media. A 70% of the test strains exhibited hydrophobicity and formed moderate to strong biofilms (OD280 0.5- > 1.0). Test oils exhibited MICs in the range of 45-360 µg.mL(-1) against the majority of test strains. All the oils at 0.25× and 0.5× MICs induced >70% reduction in the cell surface hydrophobicity, proteinase and haemolysin production. At 0.5× MIC, thymol and T. vulgaris were most inhibitory against biofilm formation. At sub-MICs electron microscopic studies revealed the deformity of complex structures of biofilms formed and cell membranes appeared to be the target site of these agents. CONCLUSIONS: Therefore, our findings have highlighted the concentration dependent activity of oils of C. copticum and T. vulgaris against virulence factors and biofilms in proteinase and haemolysin producing drug-resistant strains of Candida spp. The above activities of test oils are supposed to be mainly contributed due to their major active compound thymol. Further mechanism involving anti-proteinase, anti-haemolysin and anti-biofilm activities of these oils and compounds are to be explored for possible exploitation in combating Candida infections.