Showing papers by "I. S. Bright Singh published in 2017"

Production and characterization of polyhydroxybutyrate from Vibrio harveyi MCCB 284 utilizing glycerol as carbon source.

Abstract: Aim Production and characterization of polyhydroxybutyrate (PHB) from moderately halophilic bacterium Vibrio harveyi MCCB 284 isolated from tunicate Phallusia nigra. Methods and Results Twenty five bacterial isolates were obtained from tunicate samples and three among them exhibited an orange fluorescence in Nile red staining indicating the presence of PHB. One of the isolates, MCCB 284, which showed rapid growth and good polymer yield, was identified as V. harveyi. The optimum conditions of the isolate for the PHB production were pH 8.0, sodium chloride concentration 20 g l−1, inoculum size 0.5% (v/v), glycerol 20 g l−1 and 72 h of incubation at 30°C. Cell dry weight of 3.2 g l−1, PHB content of 2.3g l−1 and final PHB yield of 1.2 g l−1 were achieved. The extracted PHB was characterized by FTIR, NMR and DSC-TGA techniques. Conclusion An isolate of Vibrio harveyi that could effectively utilize glycerol for growth and PHB accumulation was obtained from tunicate Phallusia nigra. PHB produced was up to 72% based on cell dry weight. Significance and impact of the study This is the first report of an isolate of Vibrio harveyi which utilizes glycerol as the sole carbon source for PHB production with high biomass yield. This isolate could be of use as candidate species for commercial PHB production using glycerol as the feed stock or as source of genes for recombinant PHB production or for synthetic biology. This article is protected by copyright. All rights reserved.

Streptomyces artemisiae MCCB 248 isolated from Arctic fjord sediments has unique PKS and NRPS biosynthetic genes and produces potential new anticancer natural products.

Abstract: After screening marine actinomycetes isolated from sediment samples collected from the Arctic fjord Kongsfjorden for potential anticancer activity, an isolate identified as Streptomyces artemisiae MCCB 248 exhibited promising results against the NCI-H460 human lung cancer cell line. H460 cells treated with the ethyl acetate extract of strain MCCB 248 and stained with Hoechst 33342 showed clear signs of apoptosis, including shrinkage of the cell nucleus, DNA fragmentation and chromatin condensation. Further to this treated cells showed indications of early apoptotic cell death, including a significant proportion of Annexin V positive staining and evidence of DNA damage as observed in the TUNEL assay. Amplified PKS 1 and NRPS genes involved in secondary metabolite production showed only 82% similarity to known biosynthetic genes of Streptomyces, indicating the likely production of a novel secondary metabolite in this extract. Additionally, chemical dereplication efforts using LC-MS/MS molecular networking suggested the presence of a series of undescribed tetraene polyols. Taken together, these results revealed that this Arctic S. artemisiae strain MCCB 248 is a promising candidate for natural products drug discovery and genome mining for potential anticancer agents.

Regulating gonad inhibition and vitellogenin/vitellin induction in Penaeus monodon using mature GIH fusion protein and polyclonal antisera.

Abstract: Gonad inhibiting hormone (GIH), type II class of the CHH family neuropeptides, is released by the neurohaemal XO-SG complex of the eyestalk. The inhibitory function of GIH has a pivotal role in gonad development and reproduction. In this study, we report the expression and production of a thioredoxin-fused mature GIH protein (mf-PmGIH) of Penaeus monodon in a bacterial system and its use as antigen to raise polyclonal antiserum (anti-mf-PmGIH). The mature GIH gene of 237bp that codes for 79 amino acids, was cloned into the Escherichia coli thioredoxin gene fusion expression system. The expression vector construct (mf-PmGIH+pEt32a+) upon induction produced 32.16kDa mature GIH fusion protein (mf-PmGIH)·The purified fusion protein was used as exogenous GIH and as antigen to raise polyclonal antisera. The fusion protein when injected into juvenile shrimp significantly reduced vitellogenin/vitellin levels by 31.55% within 72h in comparison to the controls showing the gonad inhibiting property. Vitellogenin/vitellin levels were significantly induced by 74.10% within 6h when polyclonal antiserum (anti-mf-PmGIH - 1:500) was injected in P. monodon. Anti-mf-PmGIH immunolocalized GIH producing neurosecretory cells in the eyestalk of P. monodon. The present manuscript reports an innovative means of gonad inhibition and vitellogenin/vitellin induction with thioredoxin fused GIH and antisera developed.

Molecular Identification and Comparative Evaluation of Tropical Marine Microalgae for Biodiesel Production

Abstract: Marine microalgae have emerged as important feedstock for liquid biofuel production. The identification of lipid-rich native microalgal species with high growth rate and optimal fatty acid profile and biodiesel properties is the most challenging step in microalgae-based biodiesel production. In this study, attempts have been made to bio-prospect the biodiesel production potential of marine and brackish water microalgal isolates from the west coast of India. A total of 14 microalgal species were isolated, identified using specific molecular markers and based on the lipid content; seven species with total lipid content above 20% of dry cell weight were selected for assessing biodiesel production potential in terms of lipid and biomass productivities, nile red fluorescence, fatty acid profile and biodiesel properties. On comparative analysis, the diatoms were proven to be promising based on the overall desirable properties for biodiesel production. The most potential strain Navicula phyllepta MACC8 with a total lipid content of 26.54 % of dry weight of biomass, the highest growth rate (0.58 day-1) and lipid and biomass productivities of 114 and 431 mgL-1 day-1, respectively, was rich in fatty acids mainly of C16:0, C16:1 and C18:0 in the neutral lipid fraction, the most favoured fatty acids for ideal biodiesel properties. The biodiesel properties met the requirements of fuel quality standards based on empirical estimation. The marine diatoms hold a great promise as feedstock for large-scale biodiesel production along with valuable by-products in a biorefinery perspective, after augmenting lipid and biomass production through biochemical and genetic engineering approaches.

Evaluation of pyocyanin toxicity in various life stages of Penaeus monodon and in nitrifying bacterial consortia for their safe application in recirculating aquaculture systems (RAS) to abrogate pathogenic vibrios

Abstract: Pyocyanin has recently been recognized as antimicrobial agent in aquaculture industry, an alternative to antibiotics, as it accorded higher level inhibitory effect on aquaculture pathogens. As pyocyanin-induced toxicity is a major concern, this study has been undertaken to evaluate the toxicity of purified pyocyanin and to determine the LC50 value for its safe application in aquaculture. The dose-dependent pyocyanin-induced toxicity was tested on various life stages of P. monodon such as nauplius, zoea, mysis and post-larva as well as on nitrifying bacterial consortia used in aquaculture in order to fix the concentration limits to the animal rearing systems. The study revealed that pyocyanin was a potential inhibitory compound to V. harveyi, bacteriostatic at 5 mg L−1 and bactericidal at 10 mg L−1. The LC50 values of pyocyanin at various life stages of P. monodon were higher than the concentration required to abrogate pathogenic V. harveyi. Moreover, pyocyanin did not inhibit nitrification, specifically of nitrifying bacterial consortia used for activating nitrifying bioreactors, at different salinities, suggesting its potential application in recirculating aquaculture systems to control pathogenic vibrios.

Molecular Characterisation of a Novel Isoform of Hepatic Antimicrobial Peptide, Hepcidin (Le-Hepc), from Leiognathus equulus and Analysis of Its Functional Properties In Silico

Abstract: Hepcidin represents a family of cysteine-rich antimicrobial peptides that are mainly expressed in the liver of living organisms. In this study, we have identified and characterised a novel isoform of hepcidin from the common pony fish, Leiognathus equulus (Le-Hepc). A 261-bp fragment cDNA coding for 86 amino acids was obtained. Homologous analysis showed that Le-Hepc belongs to the hepcidin super family and shares sequence identity with other known fish pre-propeptide hepcidin sequences. The ORF encodes for a 24-amino acid (aa) signal peptide coupled to a 36-aa prodomain followed by a 26-aa mature peptide. The mature peptide region has a calculated molecular weight of 2.73 kDa, a net positive charge of +2 and a theoretical pI of 8.23. Phylogenetic analysis of Le-Hepc showed a strong relationship with other fish hepcidin sequences and clustered into HAMP2 group hepcidins. Secondary structural analysis indicated that Le-Hepc mature peptide contains two antiparallel β-sheets strengthened by four disulphide bonds formed by eight conserved cysteine residues. The physicochemical properties of the peptide and its structural parameters are in agreement with characteristic features of an antimicrobial peptide. This is the first report of an antimicrobial peptide from the common pony fish, L. equulus.

Purification and characterization of highly active LasB protease from Pseudomonas aeruginosa MCCB 123

Abstract: Proteolytic enzymes constitute one of the most important groups of industrial enzymes and have received worldwide attention due to their wide range of biotechnological applications and their sustainability in reducing environmental degradation. In this study, a LasB protease with dual substrate specificities was purified from Pseudomonas aeruginosa MCCB 123 with 6.85 fold increase in protease specific activity and 9.05 fold increase in elastase-specific activity. The enzyme revealed the presence of a 33 kDa protein. Optimum temperature and pH of protease activity were 60°C and 9.0, and that of elastase activity were 40°C and 8.0, respectively. Based on the cytotoxicity assay on Hep2 cells, the enzyme has an IC50 (inhibitory concentration) value of 47.28 μg mL . According to the deduced amino acid sequence, the MCCB 123 LasB protease is a zinc dependent metalloendopeptidase belonging to M4 neutral protease GluZincin superfamily. Appreciable protease activity of the enzyme at high temperature and alkaline pH proposes its potential application in detergency. MCCB 123 LasB protease is 3.96 fold more active when compared to commercial protease Savinase suggesting its industrial relevance.

Crustacean hyperglycemic hormone family gene silencing in Penaeus monodon mediated through dsRNA synthesized in vitro from genomic and cDNA

Abstract: S Vrinda1, C Reshmi1, Seena Jose1, P Reynold2, K K Vijayan 2, Rosamma Philip3 and I S Bright Singh1* 1National Centre for Aquatic Animal Health, Cochin University of Science and Technology, Cochin 682 016, India 2Central Marine Fisheries Research Institute, Cochin 682 018, India 3Department of Marine Biology, Microbiology and Biochemistry, Cochin University of Science and Technology Cochin 682 016, India