Alteration of predominant gastrointestinal flora and oxidative damage of large intestine under simulated hypobaric hypoxia.

Hypobaric hypoxia is an immediate and crucial starting mechanism of acute mountain sickness included with some non-specific gastrointestinal (GI) complications. To study the effect of hypoxia on GI microflora and its upshot to this system, male albino rats were exposed to 55 kPa (air pressure ~ 4872.9 m altitude) consecutively 30 days for 8 hours/day. The different indicator group of large intestinal microbial populations were enumerated and correlated with the levels of antioxidant indicators like catalase (CAT), superoxide dismutase (SOD), malondialdehyde (MDA), reduced glutathione (GSH) and oxidized glutathione (GSSG) of large intestinal epithelial cells. In addition, the histological study was performed by haematoxylin eosin (HE), periodic acid schiff staining (PAS) and scanning electron microscopy (SEM). It was observed that the density of total aerobes (104 folds) significantly (p < 0.05) decreased but the population of total anaerobes (209 folds) and Escherichia coli (125 folds) elevated after 30 days of hypoxic stress. The strict anaerobes like Bifidobacterium spp. (3 folds), Bacteroides spp. (134 folds), Lactobacillus spp. (7 folds) and other selected obligate anaerobes like Clostridium perfringens (40 folds), Peptostreptococcus spp. (21 folds) increased in respect to their control population. The growth direction index (GDI) of anaerobic populations was positive and correlated with gas formation aptitude. The activities of CAT and SOD in the large intestinal epithelia decreased significantly (p < 0.05) and GSH/GSSG pool turned into oxidized state with higher MDA (p < 0.05) formation. Histological study revealed the necrotized epithelial layer with higher lymphocytes infiltration in lamina propia accompanied by reduction of acidic mucins secreting goblet cells. From this experiment, it can be hypothesized that high altitude induced hypoxia manipulated the bacterial imprint and damaged the epithelial barrier of the large intestine which may cause systemic infection.

Evaluation of anti-infective potential of a tribal folklore Odina wodier Roxb against some selected microbes and herpes simplex virus associated with skin infection.

AIM: To evaluate the in vitro antimicrobial activity of aqueous and methanol extracts of Odina wodier bark (OWB), a folk medicine, against representative bacteria, fungi and herpes simplex virus (HSV) associated with skin infections. METHODS AND RESULTS: The OWB extract(s) was found to inhibit the isolates of Staphylococcus aureus, Bacillus subtilis, Pseudomonas aeruginosa, Klebsiella pneumonia, Escherichia coli at an MIC of 256-5000 µg ml(-1) and Candida albicans at and above 4000 µg ml(-1) by agar and broth dilution assays. The growth curve of Staph. aureus revealed the highest activity within 2-6 h of methanol extract (ME) exposure. Interestingly, the MTT and plaque reduction assay showed that the extracts can inhibit HSV-1 and HSV-2 at EC50 of 22·4 and 28·8 µg ml(-1) , with Selectivity index of 11·7-15. While the time kinetic and binding assays demonstrated that the ME at 50 µg ml(-1) prevents viral attachment into Vero cells. Phytochemical and HPLC analysis of ME revealed the presence of flavonoids, phytosterols, saponins and tannins including the pseudotannin chlorogenic acid. CONCLUSION: The traditional use of OWB for the management of skin infections has scientific basis. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrated the antimicrobial potential of OWB on selected isolates of bacteria, fungi and HSV, associated with skin infections.

Purification and characterization of an endoxylanase from the culture broth of Bacillus cereus BSA1.

An extracellular xylanase from the fermented broth of Bacillus cereus BSA1 was purified and characterized. The enzyme was purified to 3.43 fold through ammonium sulphate precipitation, DEAE-cellulose chromatography and followed by gel filtration through Sephadex G-100 column. The molecular mass of the purified xylanse was about 33 kDa. The enzyme was an endoxylanase as it initially degraded xylan to xylooligomers. The purified enzyme showed optimum activity at 55 degrees C and at pH 7.0 and remained reasonably stable in a wide range ofpH (5.0-8.0) and temperature (40-65 degrees C). The Km and Vmax values were found to be 8.2 mg/ml and 181.8 micromol/(min mg), respectively. The enzyme had no apparent requirement ofcofactors, and its activity was strongly inhibited by Cu++, Hg++. It was also a salt tolerant enzyme and stable upto 2.5 M of NaCl and retained its 85% activity at 3.0 M. For stability and substrate binding, the enzyme needed hydrophobic interaction that revealed when most surfactants inhihited xylanase activity. Since the enzyme was active over wide range ofpH, temperature and remained active in higher salt concentration, it could find potential uses in biobleaching process in paper industries.

Bifidobacteria and its rice fermented products on diet induced obese mice: analysis of physical status, serum profile and gene expressions.

Obesity is highly correlated with the dysbiosis of intestinal microbiota, and bifidobacteria are one of the soft targets of this metabolic syndrome. The aim of this study is to evaluate the efficacy of Bifidobacterium sp. MKK4 and rice-based fermented foods on physical, haematological, gut microbiota and lypogenic-lypolytic marker genes in diet-induced obese mice. Adult male mice (21±0.7 g) were randomly divided into four groups (n=10) according to the type of diet: normal diet (ND), high fat diet (HFD), HFD supplemented with Bifidobacterium sp. MKK4 and HFD supplemented with MKK4 associated rice-fermented food. 8 weeks of bacterial therapy in the obese mice resulted in significant reduction of body and organ weights, improved serum levels of glucose, triglyceride and cholesterol, the histological structure of the liver (steatosis), and re-establishment of gut Lactobacillus, Bifidobacterium and Bacteroides species. The bacterial therapy led to up-regulation of lipolytic transcription factors, such as peroxisome proliferator-activated receptor (PPAR)-α, PPAR-δ, and their regulated gene products in fatty acid metabolism and glucose uptake, such as acyl-CoA oxidase, carnitine palmitoyl-transferase-1, uncoupling protein-3 and glucose transporter-4. Concomitantly, both adipocytogenesis and fatty acid synthesis were arrested as reflected by the down-regulation of sterol regulatory element binding protein-1c, acetyl-CoA carboxylase, fatty acid synthase and tumour necrosis factor alpha genes. The effectiveness of the fermented product was more profound than the single bacterium. These data provide experimental support with regard to the use of Bifidobacterium sp. MKK4 as a natural therapeutic agent to control obesity.

A rapid colony screening method for the detection of arsenate-reducing bacteria.

A rapid and simple method has been developed for the detection of arsenate reducing bacteria based on the presence of arsenite [As (III)], the end product of anaerobic arsenate [As (V)] respiration. Confirmation of As (III) product is made by the reduction of starch-iodine complex. The method can be used over a large pH range (5.5-9.0) and can easily be determined at arsenite concentration as low as 0.025 mM. Major advantages of this technique are that a large number of samples can be analyzed easily at a time.

Epidemiology of Japanese encephalitis.

Seven hundred and sixty-two cases of Japanese Encephalitis (JE) were studied during the last 5 years (1985-1989) in relation to age, sex, religion, nutritional status, living habits, exposure to domestic animals and mosquitos, clinical profile, seasonal variation and mortality pattern. The maximum occurrence was in 1987-1988 and it showed a preponderance in males (51-82%). The disease is progressively decreasing in Muslims (3-7%) and gradually increasing in tribes (25-60%). Children in the age-group of 6-7 years (19-25%) were maximally affected and the disease was rare in infancy. The common features were coma, convulsions, neck rigidity and fever (88-97%). Gastrointestinal manifestations were rare (3.6%) but were associated with the highest mortality. About 80-95% had exposure to domestic animals directly or indirectly and 95% of the patients were not using mosquito nets. The CSF protein and sugar content were normal, with or without slight leucocytosis while the lymphocyte count was variable. The CSF and blood picture had no significant relation with clinical presentation and prognosis.

Production of tannase by the immobilized cells of Bacillus licheniformis KBR6 in Ca-alginate beads.

AIMS: The present study was aimed at finding the optimal conditions for immobilization of Bacillus licheniformis KBR6 cells in calcium-alginate (Ca-alginate) beads and determining the operational stability during the production of tannin-acyl-hydrolase (tannase) under semicontinous cultivation. METHODS AND RESULTS: The active cells of B. licheniformis KBR6 were immobilized in Ca-alginate and used for the production of tannase. The influence of alginate concentration (5, 10, 20 and 30 g l(-1)) and initial cell loading on enzyme production were studied. The production of tannase increased significantly with increasing alginate concentration and reached a maximum enzyme yield of 0.56 +/- 0.03 U ml(-1) at 20 g l(-1). This was about 1.70-fold higher than that obtained by free cells. The immobilized cells produced tannase consistently over 13 repeated cycles and reached a maximum level at the third cycle. Scanning electron microscope study indicated that the cells in Ca-alginate beads remain in normal shape. CONCLUSIONS: The Ca-alginate entrapment is a promising immobilization method of B. licheniformis KBR6 for repeated tannase production. Tannase production by immobilized cells is superior to that of free cells because it leads to higher volumetric activities within the same period of fermentation. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report of tannase production from immobilized bacterial cells. The bacterium under study can produce higher amounts of tannase with respect to other fungal strains within a short cultivation period.

Tannase production by Aspergillus aculeatus DBF9 through solid-state fermentation.

Tannase an industrially important enzyme was produced by Aspergillus aculeatus DBF9 through a solid-state fermentation (SSF). The organism produced good amount of enzyme and gallic acid in wheat bran among the solid substrate used in SSF. Maximum enzyme and gallic acid production occurred in 5% tannic acid after 72 h. Eighty percent initial substrate moisture and 30 degrees C temperature was found suitable for tannase production.

Studies on the extracellular tannase from newly isolated Bacillus licheniformis KBR 6.

A tannase producing bacterial strain KBR 6 has been isolated from lateritic soil and identified as Bacillus licheniformis. It is capable of producing tannase in the medium containing only tannic acid. The rapid degradation of tannic acid and production of extracellular tannase was observed in three different media containing tannic acid (M1), tannic acid + basal salt (M2) and tannic acid + basal salt + glucose (M3). Maximum enzyme production and growth of the organism was obtained at 18-21 h and 30-36 h, respectively. The increased order of enzyme production in relation to different media is as per the following sequence, M3 > M2 > M1. The maximum growth and enzyme production was observed at pH 5.0. The pH and temperature optima of the enzyme activity were found to be at 5.75 and 60 degrees C respectively. Paper chromatographic analysis indicates that gallic acid is the enzymatic degradative product of tannic acid.

Production and characterization of extracellular and intracellular tannase from newly isolated Aspergillus aculeatus DBF 9.

A comparative study on the simultaneous production of extra and intracellular tannase was made from newly isolated fungal strain Aspergillus aculeatus DBF 9. This strain produced five times more intracellular enzyme within 24 h in liquid culture than the extracellular form. Maximum tannase production occurred in the culture broth containing 1-2% (w/v) tannic acid and 0.05-0.1% (w/v) glucose. The pH and temperature optima of both the enzymes were found at 5.0 and 50-60 degrees C, respectively. Extra and intracellular tannase showed good stability at higher temperature, pH values and salt (NaCl) concentration. These properties make the enzyme suitable for pollution control and bioprocess industry.

Colorimetric assay method for determination of the tannin acyl hydrolase (EC 3.1.1.20) activity.

A new colorimetric method of tannase (tannin acyl hydrolase, EC 3.1.1.20) assay has been developed using its specific substrate tannic acid. It is based on the changes in optical density of substrate tannic acid after enzymatic reaction at 530 nm. The residual tannic acid was measured by a modified BSA precipitation method. This assay is very simple, reproducible, and very convenient, and with it tannase activity can be measured in relation to the growth of the organism.

Distribution of tannic acid degrading microorganisms in the soil and comparative study of tannase from two fungal strains.

A quantitative survey on microbial population including tannase producing organisms have been made from different soil samples. Most of the samples harbour negligible number of tannase producers in comparison to total microbial flora. Among the tannase producers, fungal members are more frequent than bacteria. Tannase production and tannic acid degradation have been studied in two newly isolated potent fungal strains. Both the strains produce maximum tannase at their stationary phases of growth. Enzymes produced by both the strains remain active within pH 3.5-6.0 and temperature 30-60 degrees C.