Paramasamy Gunasekaran

Strategies for accessing soil metagenome for desired applications.

Most of the microorganisms in nature are inaccessible as they are uncultivable in the laboratory. Metagenomic approaches promise the accessibility of the genetic resources and their potential applications. Genetic resources from terrestrial environments can be accessed by exploring the soil metagenome. Soil metagenomic analyses are usually initiated by the isolation of environmental DNAs. Several methods have been described for the direct isolation of environmental DNAs from soil and sediments. Application of metagenomics largely depends on the construction of genomic DNA libraries and subsequent high-throughput sequencing or library screening. Thus, obtaining large quantities of pure cloneable DNA from the environment is a prerequisite. This review discusses the recent developments related to efficient extraction and purification of soil metagenome highlighting the considerations for various metagenomic applications.

Production and structural characterization of crystalline silver nanoparticles from Bacillus cereus isolate.

To date, biosynthesis of metal nanoparticles has been intensively studied using bacteria and fungi. We have isolated and identified metal resistant bacterial strains from electroplating industries, they produce silver nanoparticles. The reduction reaction of aqueous silver nitrate with bacterial biomass was carried out for 120 h. Bacteria produced metallic nanoparticles showed a strong absorbance at surface plasmon resonance wavelength around 420 nm. The size and morphology of these nanoparticles were typically imaged using high resolution transmission electron microscopy, the particles size ranges between 4 and 5 nm and are spherical in shape. The crystal structure of the particles was characterized by X-ray diffraction pattern. The full width half maxima from X-ray diffraction measurements indicated that the particles exhibited face-centered cubic phase.

Antimicrobial Peptides: Versatile Biological Properties

Antimicrobial peptides are diverse group of biologically active molecules with multidimensional properties. In recent past, a wide variety of AMPs with diverse structures have been reported from different sources such as plants, animals, mammals, and microorganisms. The presence of unusual amino acids and structural motifs in AMPs confers unique structural properties to the peptide that attribute for their specific mode of action. The ability of these active AMPs to act as multifunctional effector molecules such as signalling molecule, immune modulators, mitogen, antitumor, and contraceptive agent makes it an interesting candidate to study every aspect of their structural and biological properties for prophylactic and therapeutic applications. In addition, easy cloning and recombinant expression of AMPs in heterologous plant host systems provided a pipeline for production of disease resistant transgenic plants. Besides these properties, AMPs were also used as drug delivery vectors to deliver cell impermeable drugs to cell interior. The present review focuses on the diversity and broad spectrum antimicrobial activity of AMPs along with its multidimensional properties that could be exploited for the application of these bioactive peptides as a potential and promising drug candidate in pharmaceutical industries.

Production of ethanol from liquefied cassava starch using co-immobilized cells of Zymomonas mobilis and Saccharomyces diastaticus

Co-immobilized cells of Saccharomyces diastaticus and Zymomonas mobilis produced a high ethanol concentration compared to immobilized cells of S. diastaticus during batch fermentation of liquefied cassava starch. The co-immobilized cells produced 46.7 g/l ethanol from 150 g/l liquefied cassava starch, while immobilized cells of yeast S. diastaticus produced 37.5 g/l ethanol. The concentration of ethanol produced by immobilized cells was higher than that by free cells of S. diastaticus and Z. mobilis in mixed-culture fermentation. In repeated-batch fermentation using co-immobilized cells, the ethanol concentration increased to 53.5 g/l. The co-immobilized gel beads were stable up to seven successive batches. Continuous fermentation using co-immobilized cells in a packed bed column reactor operated at a flow rate of 15 ml/h (residence time, 4 h) exhibited a maximum ethanol productivity of 8.9 g/l/h.

Optimization of media for β-fructofuranosidase production by Aspergillus niger in submerged and solid state fermentation

Abstract The kinetics of β-fructofuranosidase (Ffase) production by Aspergillus niger in submerged (SmF) and solid-state fermentation (SSF) systems was investigated. The maximum productivity of Ffase (81.8 U/litre/h) was obtained in SSF for 72 h while it was 18.3 U/litre/h in SmF for 120 h. The productivity of extra cellular Ffase produced in SSF was five-fold higher than in SmF. Optimization of fermentation medium for Ffase production was carried out using De Meos fractional factorial design with seven components such as (NH 4 ) 2 SO 4 , KH 2 PO 4 , FeSO 4 , MgSO 4  · 7H 2 O, sucrose, urea and yeast extract. The media designed for SmF after two steps of optimization supported the growth of A. niger and higher productivity of Ffase (58.3 U/litre/h) than with the medium before optimization. The optimized medium of SmF when used in SSF, did not improve the Ffase productivity and therefore the medium for SSF was optimized independent of SmF. After two optimization steps, the media was defined for SSF which supported the growth and high level of Ffase productivity (149.1 U/litre/h) in SSF compared to the medium before optimization (81.8 U/litre/h) and optimized medium for SmF (58.3 U/litre/h). The results suggested that the optimized media for SmF and SSF for the production of Ffase have to be different.

High molecular weight cellulase-free xylanase from alkali-tolerant Aspergillus fumigatus AR1

Fungi producing xylanases are numerous but alkali-tolerant fungi producing cellulase-free xylanases are rare. Presently, an alkali-tolerant Aspergillus fumigatusAR1 producing xylanases was isolated from a paper mill effluent. A. fumigatus produced xylanases in a wide range of growth pH (30 U/ml at pH 9 and 135 U/ml at pH 5) and cellulase was detected at negligible level (<0.4 U/ml) at all the growth conditions. It is strictly non-cellulolytic since it did not grow on carboxymethyl cellulose (CMC) and avicel substrates. The pH-dependent induction of A. fumigatus xylanase by xylan (substrate for xylanase catalysis) and xylose (product of xylanase catalysis) is contrasting; the ratio of xylanase expression at pH 9 to that at pH 5 was 2.1 in xylose medium and 0.38 in xylan medium. The xylanase exhibited optimum pH of 6.0–6.5 and temperature of 60 ◦ C for activity. It was stable in the pH range 5–9 and up to 50 ◦ C. At 5 mM concentration, HgCl2 inhibited xylanase activity (32%) while MnSO4 and CuSO4 enhanced xylanase activity (17–33%). Activity staining indicated that A. fumigatus AR1 produced multiple xylanases of high molecular weight ranging from 212 to 253 kDa contrasting to other two alkali-tolerant fungal xylanases, which are less than 35 kDa.

Biodegradation of tannic acid by Citrobacter freundii isolated from a tannery effluent.

A bacterial strain capable of utilizing tannic acid as sole carbon source was isolated from the effluent of a tannery and was identified as Citrobacter freundii. This organism could grow at concentrations as high as 5% (w/v) of tannic acid and produced extracellular tannase to hydrolyze the same. When grown in minimal medium containing 1% tannic acid (w/v) at 30 °C, this strain produced 1.87 U/ml of tannase at 6 h. At that time, tannic acid degradation products, namely glucose and gallic acid, were detectable in the culture filtrate; the other intermediate metabolites formed were pyrogallol (extra‐cellular) and pyruvate (intracellular). 2‐hydroxymuconic acid is presumed to form as a result of ortho‐cleavage of pyrogallol. The proposed biochemical pathway for the degradation of tannic acid by Citrobacter freundii is: Tannic acid → [Glucose + Gallic acid] → Pyrogallol → 2‐hydroxymuco‐nic acid → [?] → Pyruvate.

Influence of medium components and fermentation conditions on the production of bacteriocin(s) by Bacillus licheniformis AnBa9

Recently, antibacterial peptides are gaining more attention as an alternative therapeutics and food and other products from spoilage and deterioration. Antibacterial peptide producing strains were isolated from sediments of slaughterhouse sewage wastes. One among them, identified as Bacillus licheniformis inhibited the growth of several gram positive bacteria. Response surface methodology with central composite rotary design was used for optimization of fermentation medium and conditions for antibacterial peptide production. Lactose, NH(4)NO(3), yeast extract and NaCl and environmental factors such as pH, temperature and incubation period were selected as variables. Among ingredients, high concentration of yeast extract and NaCl had a positive effect on antibacterial peptide production and specific activity, respectively. Alkaline pH and high temperature favoured the production of antibacterial peptide by B. licheniformis AnBa9. Under optimized condition, B. licheniformis AnBa9 produced 25-fold higher production of antibacterial peptide than the un-optimized condition. Biochemical characteristics of the antibacterial peptides of B. licheniformis AnBa9 revealed that they are of bacteriocin type.

Elevated Levels of Circulating DNA in Cardiovascular Disease Patients: Metagenomic Profiling of Microbiome in the Circulation

Cardiovascular diseases (CVDs) are the leading cause of death worldwide. An expanding body of evidence supports the role of human microbiome in the establishment of CVDs and, this has gained much attention recently. This work was aimed to study the circulating human microbiome in CVD patients and healthy subjects. The levels of circulating cell free DNA (circDNA) was higher in CVD patients (n = 80) than in healthy controls (n = 40). More specifically, the relative levels of circulating bacterial DNA and the ratio of 16S rRNA/β-globin gene copy numbers were higher in the circulation of CVD patients than healthy individuals. In addition, we found a higher circulating microbial diversity in CVD patients (n = 3) in comparison to healthy individuals (n = 3) by deep shotgun sequencing. At the phylum level, we observed a dominance of Actinobacteria in CVD patients, followed by Proteobacteria, in contrast to that in healthy controls, where Proteobacteria was predominantly enriched, followed by Actinobacteria. The circulating virome in CVD patients was enriched with bacteriophages with a preponderance of Propionibacterium phages, followed by Pseudomonas phages and Rhizobium phages in contrast to that in healthy individuals, where a relatively greater abundance of eukaryotic viruses dominated by Lymphocystis virus (LCV) and Torque Teno viruses (TTV) was observed. Thus, the release of bacterial and viral DNA elements in the circulation could play a major role leading to elevated circDNA levels in CVD patients. The increased circDNA levels could be either the cause or consequence of CVD incidence, which needs to be explored further.

Recent biotechnological interventions for developing improved penicillin G acylases

Penicillin G acylase (PAC; EC 3.5.1.11) is the key enzyme used in the industrial production of beta-lactam antibiotics. This enzyme hydrolyzes the side chain of penicillin G and related beta-lactam antibiotics releasing 6-amino penicillanic acid (6-APA), which is the building block in the manufacture of semisynthetic penicillins. PAC from Escherichia coli strain ATCC 11105, Bacillus megaterium strain ATCC 14945 and mutants of these two strains is currently used in industry. Genes encoding for PAC from various bacterial sources have been cloned and overexpressed with significant improvements in transcription, translation and post-translational processing. Recent developments in enzyme engineering have shown that PAC can be modified to gain conformational stability and desired functionality. This review provides an overview of recent advances in the production, stabilization and application of PAC, highlighting the recent biotechnological approaches for the improved catalysis of PAC.