Research Journal of Biotechnology

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Vol. 13(7) July 2018

“Doi”, a naturally fermented milk product of Assam contains indigenous Lactobacillus spp. with probiotic attributes

Fatima Jaiba, Goswami Gunajit, Bora Sudipta Sankar, Pathak Sushanta Sarmah, Boro Robin Chandra and Barooah Madhumita*

Page No: 1-15

Abstract: In the present study, 20 lactic acid bacteria (LAB) were isolated from ‘Doi’ samples collected from 8 different districts of Assam. The LAB isolates characterized both phenotypically and biochemically were screened for potential probiotic properties based on conditions simulating human gastrointestinal tract (GI). The isolates were able to survive at pH 2.5 which is similar to the pH of the human GI tract and is able to grow at a temperature ranging from 15-40ºC. Four isolates viz. ABT-Z2, ABT-Z3, ABT-Z4 and ABT-Z22 had antimicrobial properties and were able to inhibit the growth of six pathogenic bacteria (six pathogenic bacteria viz. Escherichia coli, Pseudomonas putida, Staphylococcus aureus, Staphylococcus epidermidis, Salmonella enterica and Bacillus cereus).

The isolates produced maximum exopolysaccharides (EPS) at pH-6. Characterization of the 6 best isolates with probiotic properties through 16S rRNA gene sequencing identified them as Lactobacillus plantarum (4) and L. brevis (2). Lactobacillus plantarum was the predominant species of LAB occurring in Doi and also had the best probiotic attributes among the tested isolates. In future, this strain of bacteria can be used not only for developing efficient starter culture for Doi but also can be applied to produce probiotic milk based fermented product.

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Production of pyridoxal 5՛-phosphate and pyridoxine by Lactobacillus pentosus L47I-A

Sukprasong Rattanaporn, Tongpim Saowanit and Trongpanich Yanee*

Page No: 16-23

Abstract: Vitamin B6 is known as an essential micronutrient and a potent antioxidant. One group of bacteria, lactic acid bacteria (LAB), is useful for the fermentation of food products. LAB are also probiotic which means the bacteria are beneficial for health. The aim of this study was to screen and identify LAB producing vitamin B6 from fish gut and fermented foods from the Northeastern area of Thailand. Among 216 isolates, isolate 86 L47I-A exhibited the highest production of extracellular vitamin B6 with 239.64 µg/L in synthetic medium. Total extracellular vitamin B6 was determined by agar diffusion assay which was applied from microbiological assay. The extracellular vitamin B6 was separated into two fractions (F1 and F2) by semi-preparative HPLC. Reversed phase HPLC analysis showed that the forms of vitamin B6 produced in culture media were pyridoxal 5´-phosphate (F1) and pyridoxine (F2).

Through morphology, biochemical assay and analysis of 16S rRNA gene sequence, the isolate L47I-A was designated as Lactobacillus pentosus L47I-A. The results indicate that further research to develop vitamin B6 production and utilize vitamin B6 producing LAB in foods and beverages is warranted.

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Entomopathogen and parasitic medley from pest collinear ecosystem

Shanthakumar S.P.* and Mary Ivan Roberts

Page No: 24-40

Abstract: A total of 475 rhizosphere bacteria isolated from a chick pea coplanar were screened for their control efficacy against the corresponding pest Helicoverpa armigera out of which eight potential strains were identified. Bacillus thuringiensis MSSRF S1, Bacillus amyloliquefaciens MSSRF S2, Bacillus cereus MSSRF S4, Bacillus thuringiensis MSSRF S5, Enterobacter cloacae MSSRF S8, Exiguobacterium mexicanum MSSRF S9, Proteus hauseri MSSRF S11 and Pseudomonas aeruginosa MSSRF S20 exhibited pest control attributes.

Histopathological studies of insect treated with B. thuringiensis MSSRF S1, B. amyloliquefaciens MSSRF S2, E. cloacae MSSRF S8, B. thuringiensis MSSRF S5, B. thuringiensis MSSRF S4 and E. mexicanum MSSRF S9 showed tissue fragmentation, columnar cell disintegration on the larval midgut epithelium and P. aeruginosa MSSRF S20 exhibited relatively higher insect pathogenic activity over all the other isolates. Dipping and diet incorporation methods confirmed the control potential of the selected bacterium and its toxic metabolite secretion.

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Exploring hypoglycemic potential of Vicia faba crude seed extract in Saccharomyces cerevisiae 3187

Choudhary Dhiraj Kumar, Mishra Abha* and Mittal Aishwarya

Page No: 41-48

Abstract: The aim of this investigation was to examine the hypoglycaemic potential of Vicia faba seed extract by various in vitro methods. Vicia faba seed extracts were exploited for their effects on glucose adsorption capacity, in vitro glucose diffusion, in vitro amylolysis kinetics and glucose transport inside the yeast cells. It was observed that seed extracts adsorbed glucose remarkably with an increase in glucose concentration. In vitro glucose inhibition method and in vitro amylolysis, kinetic experimental model showed the rate of glucose diffusion was increased with time from 30 to 180 min.

Ethanol and methanol seed extracts had major inhibitory effects on movement of glucose into external solution across dialysis membrane compared to control. Seed extract also promoted glucose uptake in yeast cells on temperature and different sugar molecules. Percent increase in glucose uptake was dependent on both the sample, temperature, glucose concentration and sugars. FTIR and HPLC analysis confirmed the polyphenols present in seed extract. The output of this study confirmed the hypoglycaemic activity of the extract of Vicia faba seed. Hypoglycemic effect may due to synergistic effect of all the constituents present in seed extract or acting separately.

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Effect of pre-harvest potassium foliar spray and postharvest storage methods on quality and shelf life of cherry tomatoes

Islam Mohammad Zahirul, Lee Young-Tack, Mele Mahmuda Akter and Kang Ho-Min*

Page No: 49-54

Abstract: Potassium (K) foliar sprays as a pre-harvest treatment and storage methods as a postharvest treatment were applied to get the effects on quality and shelf life of cherry tomatoes. The supplied nutrient solutions were regulated to EC 2.5 dS m-1 and pH 5.8-6.2. The control (distilled water), 10.23 mM potash of potassium chloride (KCl), 10.23 mM potash of potassium nitrate (KNO3) and 36.74 mM potash of monopotassium phosphate (KH2PO4) were sprayed for 5 weeks. Harvested tomato fruits were stored at 50C in boxes (commercial size) and in oxygen transmission rate film as a modified atmosphere packaging. Ca, K, P and Cu contents in KH2PO4-treated tomatoes were higher compared with other treatments.

KH2PO4-treated tomatoes had the lowest fresh weight loss, respiration rates, ethylene emission which prolonged the shelf life. The shelf life KH2PO4-treated tomatoes were 25 days for MAP storage and 20 days for box storage. Moreover, the firmness, soluble solids and glucose content were increased by the KH2PO4 treatment and they were retained after storage. KH2PO4-treated tomatoes influenced the color, lycopene and titratable acidity during the storage period. Therefore, K foliar sprays from KH2PO4 used for both preharvest treatment and postharvest storage methods maintain the quality and shelf life of cherry tomatoes.

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Establishing the frequency of IL-10Rα polymorphisms rs2508445, rs4252270, rs149491038 in Indian Tamils

Joshi Suraj kumar, Devaraju Panneer and Jayaraman Megala*

Page No: 55-58

Abstract: IL-10, an anti-inflammatory cytokine exerts its immune regulatory effect by binding to its receptor IL-10R, which is a hetero tetrameric protein belonging to type-II cytokine family, expressed in all hematopoietic cells. The incidence of autoimmune and inflammatory disorders (AIIDs) is on the rise in Indian population. Individual genetics plays a major role in determining the susceptibility, clinical phenotype and response to therapy for these diseases. The higher incidence of AIIDs and a lack of genetic data on South Indian Tamils served as a basis for this study geared to establish the normotype frequency of three genetic variants of IL-10RA gene (rs2508445, rs4252270, rs149491038) in this population.

A total of 180 healthy South Indian Tamils had a genotype run for IL-10RA variants by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). It was observed that the mutant allele frequency of rs2508445, rs4252270 was 23% and 12.3% respectively. The Arg262Cys rs149491038 was completely absent in our population. Screening the polymorphisms in IL-10R might be helpful in terms of predicting the risk of susceptibility and to determine the outcome of therapeutic modalities for AIIDs.

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Assessment of the genetic diversity of Ashwagandha (Withania somnifera L. Dunal) using agro-morphological, phytochemical and molecular markers

Kumar Vinay*, Gajbhiye N.A. and Manivel P.

Page No: 59-68

Abstract: Ashwagandha (Withania somnifera Dunal) is a crucial medicinal plant known as Indian ginseng; the dried roots of which are used in traditional systems of medicine. In the present investigation, genetic diversity was estimated among 24 morphologically distinct ashwagandha accessions using molecular, phytochemical and morphological markers. Out of 60 randomly amplified polymorphic DNA (RAPD) markers used to estimate the genetic relationships, 27 primers produced better polymorphic and reproducibility pattern. An average of 9 DNA fragments was amplified by each primer within the range of 200 to 2000 bp in size. The genetic diversity among the accessions measured by Jaccard’s similarity coefficient varied from (0.440 to 0.831). The highest (0.831) and lowest (0.440) was measured between DWS-95 and DWS-141 and between DWS-327 and DWS-133 respectively.

Cluster generated through RAPD data grouped 24 accessions into two major clusters; the first cluster was solitary having only a single accession DWS-327 (a dwarf line with less than 30 cm height). Based on morphological traits, the 24 accessions were grouped in to six major clusters, of which four clusters were solitary containing only single accessions. Cluster III contained the highest value for the studied traits. Similarly, three phytochemical compounds namely withanolide-A, 12- deoxy withastramonolide and withaferin-A of pharmacological importance revealed the presence of highest withanolide-A content ≥1 mg/g in DWS-327 line. The quantitative and qualitative status of active constituents along with estimation of genetic diversity using morpho-molecular traits in ashwagandha would be useful to devise conservation strategies and selection of right parent/sample for maximum yield.

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Seroepidemiology of infectious bovine rhinotracheitis in Chhattisgarh, India

Jain Lata*, Kumar Vinay, Chaturvedi Sameer, Roy Goutam and Barbuddhe Sukhadeo B.

Page No: 69-72

Abstract: Infectious bovine rhinotracheitis (IBR) is important infection of bovines causing respiratory and reproductive problems and abortions leading to huge economic loss to the dairy industry. This disease is known to be transmitted through natural service and artificial insemination causing severe implication on international trade of livestock and their products. In the present study, seroepidemiology and risk factors infectious bovine rhinotracheitis were investigated in cattle and buffaloes of Chhattisgarh, India. A total of 374 serum samples from bovines were collected randomly from 94 villages of six districts of Chhattisgarh state with different attributes like districts, species, breeds, sex, age, farming sectors and clinical condition of animal.

Samples were subjected to i-ELISA for detection of antibodies for IBR. The overall apparent seroprevalence for IBR was observed to be 63.9%. Statistical analyses indicated a significant difference in seroprevalence among districts (P<0.05) for all the three diseases. IBR was significantly more prevalent in Indian cattle age above 4 years and clinically ailing animals as compared to cross bred cattle, buffaloes and apparently healthy ones. Further, positivity for IBR in rearing sector and sex of animal was non-significant. The finding of the study represented a major impact on animal health and productivity.

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Chemoprotective role of garlic extract on GST-Pi expression in the liver of Cyclophosphamide administered rats

Devi A.*, Harishankar M.K. and Aruljothi K.N.

Page No: 73-79

Abstract:Cyclophosphamide (CY) remains as one of the most widely utilized alkylating agent in cancer chemotherapy. However, the innumerable side effects associated with CY treatment were considered to be the major concern for oncologists. Glutathione S-transferase-Pi (GST-Pi), an isoform of GST is an important Phase II detoxification enzyme and garlic extracts play an important chemoprotective role against number of carcinogenic compounds. Infinite number of studies on plant extracts and antioxidant compounds has shown to scavenge reactive oxygen species and possess immunomodulating effects in CY administered animals. The current study was undertaken to determine the potential effects of garlic extract in abrogating the deleterious effects induced by CY via monitoring GST-Pi expression.

The chemoprotective effects of garlic against CY were investigated in male Wistar rats, randomized into control (garlic extract treated alone) and experimental (CY + garlic extract treated) groups. The experiment was carried out for 10 days and during the course of the study liver was isolated and fixed in 10% formalin. The effect of garlic extract on CY was assessed by analyzing the expression pattern of GST-Pi in experimental and control groups after 2,4,6,10 days (post treatment) by immunoblot and RT-PCR analysis. Our results clearly indicate that in experimental group animals (CY + garlic extract treated), only single cells positive for GST-Pi expression was detected and no foci expression was detected. This could be due to the chemoprotective effect of garlic extract which was used as a supplement to counteract the cytotoxic effects of CY.

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Multivariate Analysis of Plant Morphological Characters for Aluminium Tolerance in Cowpea Genotypes under Hydroponics

Kushwaha Jitendra Kumar and Manivannan S.*

Page No: 80-90

Abstract: Aluminium toxicity is a major problem in the acid soil which affects nutrient uptake behaviour of the plants and in turn the morphological characters. In order to ascertain the effect of aluminium toxicity on plant morphological characteristics, multivariate analysis was carried out on fifteen cowpea genotypes in hydroponics having four aluminium levels i.e. 0, 25, 50, 100 µM in a factorial complete randomized design.

Plant heights at weekly interval up to vegetative stage and root length, total biomass, root dry matter and shoot dry matter at the end of the experiment were observed for studying their individual and interaction effect of genotype and aluminium levels. Principal component analysis and cluster analysis were carried out to ascertain the most tolerant and susceptible genotype for aluminium toxicity and also to group the cowpea genotypes based on their tolerance level to aluminium. Based on the ANOVA, principal component analysis and cluster analysis genotypes were classified into four groups viz. highly susceptible, susceptible, tolerant and highly tolerant.

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Molecular cloning and activity analysis of a novel vascular-preferential promoter from poplar

Wang Like, Yang Liheng, Xu Chen, Zhang Le, Ma Guojia and Zhuge Qiang*

Page No: 91-107

Abstract: Thaumatin-like proteins (TLPs) belong to the pathogenesis-related protein 5 family (PR-5) and constitute a large, multifunctional gene family involved in host defense against pathogenic infections and environmental stress. A 2078 bp upstream sequence of a TLP gene, designated pPeTLP, was isolated by homologous polymerase chain reaction (PCR) from poplar (Populus deltoides × P. euramericana cv. ‘Nanlin895’). Using PlantCARE and PLACE prediction, we found that the putative cis-regulated elements of pPeTLP include 12 light-responsive elements, 1 low-temperature responsive element, 1 drought-responsive element, 2 gibberellin-responsive elements and 1 salicylic acid-responsive element. Promoter activity analyses showed that the pPeTLP::GFP construct is localized in the cytoplasm, nucleus and periphery of the plasma membrane in onion epidermal cells.

Transgenic analyses of the pPeTLP::GUS construct indicated that the GUS protein is expressed in the vascular tissues of leaves, stems and roots of Arabidopsis thaliana. Expression vectors of four chimeric pPeTLP were constructed and transferred into poplar. GUS activities declined in the shorter promoter fragments. These results suggest that pPeTLP is a vascular-preferential promoter and can be induced by drought, salicylic acid, gibberellin, light and low-temperature treatment.

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