Kaushik Bhattacharjee

Diversity of Streptomyces spp. in Eastern Himalayan region - computational RNomics approach to phylogeny

Isolation and characterization of actinomycetes from soil samples from altitudinal gradient of North-East India were investigated for computational RNomics based phylogeny. A total of 52 diverse isolates of Streptomyces from the soil samples were isolated on four different media and from these 6 isolates were selected on the basis of cultural characteristics, microscopic and biochemical studies. Sequencing of 16S rDNA of the selected isolates identified them to belong to six different species of Streptomyces. The molecular morphometric and physico-kinetic analysis of 16S rRNA sequences were performed to predict the diversity of the genus. The computational RNomics study revealed the significance of the structural RNA based phylogenetic analysis in a relatively diverse group of Streptomyces.

NEMiD: a web-based curated microbial diversity database with geo-based plotting.

The majority of the Earths microbes remain unknown, and that their potential utility cannot be exploited until they are discovered and characterized. They provide wide scope for the development of new strains as well as biotechnological uses. The documentation and bioprospection of microorganisms carry enormous significance considering their relevance to human welfare. This calls for an urgent need to develop a database with emphasis on the microbial diversity of the largest untapped reservoirs in the biosphere. The data annotated in the North-East India Microbial database (NEMiD) were obtained by the isolation and characterization of microbes from different parts of the Eastern Himalayan region. The database was constructed as a relational database management system (RDBMS) for data storage in MySQL in the back-end on a Linux server and implemented in an Apache/PHP environment. This database provides a base for understanding the soil microbial diversity pattern in this megabiodiversity hotspot and indicates the distribution patterns of various organisms along with identification. The NEMiD database is freely available at www.mblabnehu.info/nemid/.

A selective medium for recovery and enumeration of endolithic bacteria

The study of lithic microbial communities, inhabiting rock substrates has been gathering momentum due to a growing attention of their wide importance as model systems in ecological studies and for their community structure. It is generally accepted that the success of cultivation-based technique is primarily based on suitable culture medium for isolation. The media available for enumeration and recovery of endolithic bacteria are mainly specific to particular type of rock which may not be suitable to isolate endolithic bacterial community from diverse lithobiontic niches. In this study, a new unoptimized medium was formulated, designated LM10 (unoptimized) for enumeration and recovery of endolithic bacteria by addition and/or omission of media components to the basal medium R2G, which was selected after experimental evaluation of five different existing media. The endolithic bacterial count in LM10 medium (unoptimized) was significantly higher than the R2G medium (t=-12.57, p<0.0001). The culture and nutritional parameters associated with unoptimized LM10 medium were optimized using statistical approach to maximize the recovery and enumeration of endolithic bacteria. The first phase of the study comprised of a Plackett-Burman (PB) design experiment conducted to screen thirteen medium components and two culture parameters as variables with effect on bacterial enumeration and recovery. Out of these, Yeast extract, Casein hydrolysate, Glucose, Starch and Sodium thiosulphate were found to be significantly affecting the bacterial count (p<0.05) based on PB design. On keeping rest of the media components and culture conditions at fixed value as per the PB design analyses (p>0.05 and coefficients), further optimization was carried out for significant factors using Box-Behnken design (BBD) of response surface methodology (RSM). Optimized media components obtained by BBD were Yeast extract, Casein hydrolysate, Glucose and Starch in 0.05g/l each and Sodium thiosulphate in 0.047g/l concentrations. The composition of optimized LM10 medium formulated (per litre) is 0.05g Yeast extract, 0.05g Casein hydrolysate, 0.05g Glucose, 0.05g Starch, 0.01g K2HPO4, 0.02g Sodium pyruvate, 0.2g MgSO4, 0.001g FeSO4·7H2O, 0.285g NH4Cl, 0.039g CaCl2·2H2O, 0.047g Na2S2O3·5H2O, 0.002g NaHCO3 and 11g Gellan gum (pH=7.4). Validation of optimized LM10 medium using nine different rock samples from Meghalaya clearly indicated that optimized LM10 medium was better suited for higher recovery and enumeration of endolithic bacteria under both aerobic and anaerobic conditions.

Interaction of caffeine and sulfadiazine with lysozyme adsorbed at colloidal metal nanoparticle interface: influence on drug transport ability and antibacterial activity

The modulated bioactivity of proteins immobilized on nanoparticle (NP) interfaces is of tremendous interest toward designing better therapeutic and diagnostic tools. In this work, binding behavior and the antibacterial activity of free lysozyme (LYS) as well as its non-covalent assembly with silver (Ag) and gold (Au) colloidal NPs were compared in presence of two model drugs, viz. sulfadiazine (SDZ) and caffeine (CAF). Intrinsic protein fluorescence was found to quench due to the formation drug–protein complex in case of CAF resulting a linear Stern–Volmer (SV) plot with KSV = 1.83 × 103 M−1.On the other hand, a positive deviation beyond [SDZ] ~0.15 mM is explained due to the formation of a fluorophore – quencher sphere with radius of 13.85 ± 1.80 Å that results almost one order of magnitude higher KSV (1.75 × 104 M−1). Molecular docking calculation also predicts relatively more stabilized complex of SDZ with LYS in comparison to CAF (ΔE ~ 3 kJ mol−1). Synchronous fluorescence results corresponding to Trp and Tyr residues as well as FTIR spectra in the amide I region of LYS confirms minimal deformation in the LYS secondary structure on adsorption to spherical NP surface. Although the nature of LYS–drug interaction remains invariant, the extent of quenching interaction as well as the drug binding ability is strongly modulated in presence of NPs. Further, the antibacterial activity of LYS in presence of the investigated drugs shows 9–14% upsurge with AuNP, in sharp contrast to ca. 31–34% decrease in AgNP.

Precursor-directed combinatorial biosynthesis of cephalosporin analogue by endolithic actinobacterium Streptomyces sp. AL51 by utilizing thiophene derivative

Natural products or their derivatives provide a reliable resource for new drugs. The multi-step chemical reaction to produce new drug is not only expensive but also release pollutants. The precursor-based combinatorial biosynthesis (PCB) is, however, a better option to produce novel natural products with potential pharmaceutical applications. The present work is an attempt to synthesize an antibacterial compound by transforming thiophene precursor using endolithic Streptomyces sp. AL51. The Streptomyces sp. AL51 was isolated from a granite rock sample collected from Mylliem, Meghalaya, India. The isolate was identified as Streptomyces sp. based on its cultural, morphological, biochemical and molecular characteristics. The bioactive compound CAx1 was extracted from the fermentation broth. The compound was characterized by bioactivity-guided fractionation and identified by infrared, UV–visible, nuclear magnetic resonance and mass spectrometry data and identified as 7-[1-(thiophene-5-yl)-1-formamido]-3-propylenyl-3-cephem-4-carboxylic acid with molecular formula C15H14N2O4S2. The purified compound showed considerable in vitro antibacterial activity against both Gram-positive and Gram-negative bacteria showing its broad spectrum property. The obtained results provide promising baseline information for the potential use of endolithic actinobacterium for semisynthetic drug discovery. This is the first report on PCB of broad range antibacterial compound by endolithic Streptomyces strain.