Research Journal of Biotechnology

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Research Journal of Biotechnology

Biodiversity of VAM Fungi in Queensland Arrowroot (Canna Edulis Ker-Gawler) in some Forest Soils of Kerala

Jaya Kumari T.R.

Abstract: Queensland Arrowroot (Canna edulis Ker-Gawler) is a promising tropical root crop that yields the high quality ‘canna starch’ from its rhizomatous rootstocks. In the present study, assessment of the vesicular arbuscular mycorrhizal (VAM) fungal status of this crop in the forest soils in Thiruvananthapuram District of Kerala has been undertaken. Rhizosphere soil and root samples, collected from the plants growing naturally under field conditions in different localities were analysed for VAM fungal colonization. Physicochemical characteristics of the soil like pH, moisture, organic carbon and available phosphorus were also studied. Analysis of soil samples indicated significant levels of VAM fungal colonization in the rhizosphere of the crop. Total spore density/100g rhizosphere soil ranged from 211-910. All the collected plants showed distribution of VAM fungal structures like hyphae, vesicles and arbuscules. Root colonization percentage ranged from 51-100%. Five morphotypes of VAM fungi belonging to two genera viz. Glomus and Gigaspora were characterised and identified. Glomus was the most frequent genus (80%) with three identified species - G. aggregatum, G. caledonium and G. constrictum. The most frequent species recovered was G. aggregatum.

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Isolation and Characterization of Plant Growth promoting Bacillus Thuringiensis from Agricultural Soil of West Bengal

Bandopadhyay Sandip, Pal Subrata and Gangopadhyay Swati R. Pugalenthi M. Doss A. and Vadivel V.

Abstract: Plant Growth Promoting Rhizobacteria (PGPR) from different agricultural fields of North 24 parganas, West Bengal, India was isolated on the basis of their phosphate solubilizing and phytohor­mone producing properties. Among 30 different bacte­rial isolates, the strain (A5), that exhibited maximum rate of phosphate solubilization was ident­ified as Bacillus Thuringiensis by 16s rDNA analy­sis. Extra-cellular secretion of L-malic acid seems to be the major cause of microbial phosphate solubilization as evident from High Performance Liquid Chromato­graphy (HPLC) analysis. Application of this strain in pot, stimulated the plant growth. Beside this, the particular strain also showed multiple antibiotic resistant and heavy metal resistant characteristics.

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Purification and Optimum Characterization of extracellular Amylolytic Enzyme from Aspergillus Species

Haq Imdadul, Sani Wirakarnain, Philip Koshy , Rafat Arash, Hossain A.B.M. Shariff and Taha Rosna Mat Pugalenthi M. Doss A. and Vadivel V.

Abstract: In the present study, the amylase enzyme producing potential of three different Aspergillus spec­ies was analyzed. The extracted amylase enzyme was purified by DEAE cellulose and Sephadex G-50 column chromatography and the enzyme activity was measured by using synthetic substrate starch. The partially purified enzyme exhibits maximum activity at the optimum pH (6), temperature (50-600C) and substrate concentration (1.5-2.0%) under standard as­s­ay conditions. Among the three different Asper­gillus species examined, A.fumigatus registers the highest production of amylase enzyme with maximum enzyme activity. The characteristics of the partially purified enzyme such as optimum pH and temperature are also favourable for the industrial applications.

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Biosurfactant Production from Pseudomonas sp.G6 by solid Substrate Fermentation using coconut oil cake as substrate

Thomas Minu, Philip Suja and Mathew Jyothis

Abstract: With the objective of isolating a potent biosurfactant producer, bacteria isolated from soil samples from garage sites were screened for biosurfactant production by blue agar test, blood agar test and oil spreading test. Emulsification indices of the strains were also determined. A bacterium identified as Pseudomonas sp. G6 was found to be producing biosurfactant of rhamnolipid type. FT- IR spectrum of the sample gave satisfactory match with that of standard rhamnolipid. Coconut oil cake, a cheap and easily available agricultural byproduct was found to be a suitable substrate for the production of biosurfactant. The optimum temperature, pH and incubation period for production were 370C, 6 and 72 hours respectively. NaNO3 and NaCl had enhancing effect on biosurfactant production.

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Extraction and Separation Studies of U(VI) from Salicylate Media using Neutral Organophosphorous Extractant, Cyanex-923 in Toluene

Ghag S. M. and Pawar S. D.*

The neutral extractant, Cyanex-923 has been used for the extraction and separation of U(VI) from sodium salicylate media. The metal ion was found to be quantitatively extracted with Cyanex-923 in toluene at pH 5.0 and from the organic phase it can be stripped with 4.0M H2SO4 solution. The effect of pH, sodium salicylate concentration, reagent concentration, equilibration period, diluents, diverse ions and stripping agent on the extraction of U(VI) has been studied. The stoichiometry of the extracted species of this metal ion was determined on the basis of the slope analysis method. The reaction proceeded by solvation and the probable extracted species found were UO2(HSal)2. 2 Cyanex-923.

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Isolation and Identification of 1, 3-Propanediol producing Strain of K. Pneumoniae 141B from Soil and Optimization of process Parameters

Jalasutram Vanajakshi * and Jetty Annapurna

Research on 1,3-propanediol (1.3-PDO) has markedly increased due to its potential applications in polyester preparations, cosmetics, foods, lubricants and medicines. The present study was aimed at isolation and characterization of 1,3-PDO producing strain from soil. More than 40 isolates of glycerol fermenting micro­organisms were isolated from IICT campus, Hydera­bad, India and screened for 1,3-PDO production. Among these isolates, 141B strain had shown the maximum 1,3-PDO production of 7.41 and 5.94 g/l under aerobic and anaerobic conditions respectively. Based on morphol­ogical, physiological, biochemical and phylogenetic analysis, the strain had been confirmed as a novel strain of Klebsiella pneumoniae and hence it was deposited in IMTECH, Chandigarh, India, with the accession number MTCC 9751 and the EMBL accession number of this strain is FN820293. The effect of process parameters on 1,3-PDO production and on enzyme production of glycerol dehydratase (GDHt) and 1,3-propanediol oxidoreductase (PDOR) production was evaluated. GDHt and PDOR were the two key intracellular enzymes involved in the production of 1,3-PDO. The optimal process parameters were found to be fermentation time, 8h; temperature, 37ºC; pH, 6; aeration, semi aerobic. The yield of 1,3-PDO had improved from 7.41 g/l with the basal medium to 11.17 g/l with the optimized medium resulting in an increase of 50 %. The specific activities of GDHt and PDOR have been increased to 35.1 % and 29.3 % with the optimized medium. The results demonstrated that K. pneumoniae 141B is an efficient strain for 1,3-PDO production as it produced a molar yield of 0.67 mol/mol of glycerol.

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Identification of Chilling-inducible Genes in Sweet potato by Suppression Subtractive Hybridization

Lin K. H.,1 Chiang C. M., Lai Y.C., You S. H.1 and Lo H.F.*

Abstract The objectives of this study were to identify chilling stress-response genes of a chilling-tolerant sweet potato (Ipomoea batatas (L.) Lam) variety, ‘Tainung 71’ and to measure the regulation of gene expression after exposure to 7°C for 1 day. RNAs from plants subjected to chilling periods for 6 h as well as non-chilled plants (control) were used as the tester and driver samples respectively. After suppression subtractive hybridization (SSH), polymerase chain reaction (PCR) products were cloned into a vector and transformed into Escherichia coli. The unique cDNA inserts were screened for differential expression by dot blotting and the identified clones were then picked for DNA sequencing. Using BLAST and Gene Ontology database searches to annotate the putative functions of the genes, 8 of these unigenes were found to be involved in metabolism, stress, regulation and development categories, whereas 2 genes with unknown functions were also identified. Two differentially expressed gene transcript with known function were selected for real-time PCR and found to be upregulated across chilling times in contrast to the actin gene (control). Our results indicate that chilling-induced genes are related to metabolic pathways and genetic regulation of stress and development and can serve as a foundation for future studies to elucidate chilling stress mechanisms of sweet potato.

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Production of a thermostable and alkaline Protease by Bacillus Subtilis MTCC 9226

Prakash Chandan, Sharma Iti and Kumar Dinesh

Abstract: A thermostable alkaline protease producing strain identified as Bacillus subtilis MTCC 9226 was isolated from cow dung compost sample. Culture conditions were optimized for protease production. Maximum enzyme production was detected in statio­nary phase with Bacillus subtilis MTCC 9226 at 450 C­, pH 9.0 with 5% inoculum after 32 h incubation. The optimum temperature and pH for the activity of this protease were 45­­­­0C and 9.0 respectively. Out of various carbon and nitrogen source tested, glucose (1%) and yeast extract (0.5%)+peptone (5%) proved to be very good source/s of carbon and nitrogen respectively for the production of protease by this organism. The enzyme was stable from pH 7.0 to 11.0 at temperature 300 C to 600 C. Activity of protease increased in presence of 4mM Mg2+ (107%) and was slightly effected by EDTA 50mM (79%) and comp­letely inhibited by SDS. The enzyme activity was stable up to in 15% of H2O2 and sodium hypochlorite and inhibited at higher concentrations. The enzyme was very stable in benzene (97%) followed by methanol (86%). Complete removal of gelatin from X-ray films was achieved in 25 min at 450C.

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Study of Penicillium expansum and Penicillium citrinum for their Antibacterial Potential and Optimizing their Activity

Arora Daljit Singh* and Kaur Harpreet

Abstract: Various soil fungi are prolific producers of antimicrobial agents. One hundred and thirteen fungi were isolated and screened for antibacterial activity by agar well diffusion assay. Their microscopic examination revealed that 41.5% of the fungal isolates belonged to the Aspergillus group, 34.5% to the Penicillium and rest 23% to different systematic groups. Thirty five percent of the total isolates showed activity against one or more tested bacteria while none of the isolate was effective against Pseudomonas aeruginosa. Two species of Penicillium (Penicillium expansum and Penicillium citrinum) were selected for further optimizing their antibacterial activity. Starch was found to be the most promising carbon source, while yeast extract and soyabean meal were good sources of nitrogen to support best antibacterial activity. Heating the culture filtrate of both the Penicillium sp. showed that the compound responsible for antibacterial activity should be quite thermostable even upto 50°C. Extraction of aqueous extract of both the fungi with different solvents revealed butanol to be the best to elute the compound responsible for antibacterial activity. Butanolic extract of Penicillium expansum inhibited all the bacteria tested including Pseudomonas aeruginosa and gave better activity than aqueous extract (which was active against only three bacteria) and standard antibiotics (Penicillin G and Ampicillin).

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Evaluation of the Effect of Autochthonous Probiotics and Lactobacillus divergens on Rotifers Brachinous plicatilis and Blue-Fin Porgy Sparidentex hasta Larvae Important Cucurbit, Citrullus Colosynthis (L.) Schrad

Al Marzouk A. *, Azad I. S., Rezq T. Abu, Al Abdul Elah K. and Al Gharabally H.

Abstract: A standard probiotic, Lactobacillus divergens (ATCC, 35677) and three autochthonous isolates, obtained from cultured yellow-fin porgy Acanthopagrus latus (SHPB), wild silver pomfret Pampus argenteus (4SQ) and wild orange-spotted grouper Epinephelus coioides (5L8) were evaluated in the present study. They were tested for their antagonism against potential bacterial pathogens of blue-fin seabream Sparidentex hasta (sobaity). Antagonism of the probiotics against Vibrio alginolyticus, V. anguillarum, V. harveyi, V. parahaemolyticus, V. vulnificus and Streptococcus agalactiae was tested along with their ability to enhance growth of rotifers and survival of sobaity larvae. The results showed that the filter paper disk overlay antagonism method was suitable to assess the effect of probiotic bacteria on pathogenic bacteria. The autochthonous SHPB showed a significant zone of inhibition against all the pathogenic bacteria with a distinctive competitive exclusion. The other probiotics showed some effect, but they were unable to inhibit the growth of all the pathogenic bacteria tested. The addition of SHPB and L. divergens increased the growth performance in respect to cell count of the rotifer population with time, while the 4SQ and 5L8 showed lower cell proliferation response compared to the control. The effects of dietary administration of either of SHPB or L. divergens and their combination on sobaity larval survival were investigated. All the probiotics tested showed significant survival rate of larvae compared to the control. The survival in L. divergens fed larvae was the highest, 11.7%, whereas it was 9.2% in the probiotic combination-fed larvae, 8.9% in SHPB and 6.3% in the control. After the challenge against vibrio harveyi (0.1 mL of 109 cells ml-1), the mortality of the fish larvae receiving the diet supplemented with the SHPB and mixture of SHPB and L. divergens was significantly lower than that observed in the control.

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Use of Agro-residues for Protease Production and Application in Degelatinization of Waste Photographic Films

Zambare V. P., Nilegaonkar S. S. * and Kanekar P. P.

Abstract: Protease production from P. aeruginosa MCM B-327, isolated from vermicompost pit soil, was achieved by using different agro-origin substrates. Maximum protease production was observed with wheat bran- tryptone followed by bengal gram flour - soya flour. The crude enzyme (3.35U) showed effective degelatinization of waste photographic films in tap water at room temperature. The present study has potential application in waste management as well as recovery of silver from photographic films.

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Sequence Analysis of Divergent Domain 28S Gene in four Species from Family: Siluridae (Pisces)

Verma Jyoti*, Lakra W. S., Kushwaha B., Sirajuddin M., Nagpure N. S. and Kumar Ravindra

Abstract: The present study was aimed to find the nucleotide sequence variation in divergent domain 28S and further utilize these variations in establishing phylogenetic relationships among the four species, viz. Ompok pabda, O. bimaculatus, O. pabo and Wallago attu, belonging to family siluridae. The size of different domains of 28S (D1-D11) rDNA ranged from 302 to 688 with 54 to 63% GC content. The sequence alignment suggested the presence of insertions/ deletions (indels) and substitutions (transitions and transversions) in these regions. The indels were responsible for length variations. The study suggested that Ompok species has a closer relationship forming one cluster, in which O. pabo and O. pabda were closely related as compared to O. bimaculatus. The species W. attu radiated far and formed separate cluster. The aim of this study was to follow a series of investigations to understand molecular characterization.

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Antimicrobial Peptides and their Mechanism of Action

Anis Mohd.* and Gupta U. S.

Abstract: Antimicrobial peptides (AMPs) are small molecular weight proteins with broad spectrum antimicrobial activity against bacteria, viruses and fungi. These evolutionarily conserved peptides are usually positively charged and have both a hydrophobic and hydrophilic side that enables the molecule to be soluble in aqueous environments yet also enters lipid-rich membranes. Once in a target microbial membrane, the peptide kills target cells through diverse mechanisms. During the past two decades several AMPs have been isolated from a wide variety of animals, both vertebrates and invertebrates, plants as well as from bacteria and fungi. Most of these peptides are obtained from different sources like macrophages, neutrophils, epithelial cells, haemo­cytes, fat body etc. These peptides exhibit broad-spectrum activity against a wide range of microorganisms including Gram-positive and Gram-negative bacteria, protozoa, yeast, fungi and viruses. A few peptides have also been found to be cytotoxics to sperm and tumor cells. In this review we present an overview of the general mechanism of action of antimicrobial peptides and discuss some types of antimicrobial peptides.

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