Bibhuti Bhusan Pal

Hyperglycemia-induced oxidative stress induces apoptosis by inhibiting PI3-kinase/Akt and ERK1/2 MAPK mediated signaling pathway causing downregulation of 8-oxoG-DNA glycosylase levels in glial cells.

Glial cells are very important for normal brain function and alterations in their activity due to hyperglycemia, could contribute to diabetes-related cognitive dysfunction. Oxidative insults often cause rapid changes in almost all cells including glial cells. However, pathophysiologic mechanisms that lead to diabetic complications are not completely elucidated. Therefore, we examined whether elevated glucose levels directly or indirectly disrupt antioxidant defense mechanisms causing alterations in signaling pathways, cell cycle dysregulation, and reactive oxygen/nitrogen species-mediated apoptosis in glial cells. Findings of this study demonstrated that exposure of glial cells to high glucose markedly induces cellular and molecular injuries, as evidenced by elevated levels of reactive oxygen/nitrogen species, biomolecules damage, cell cycle dysregulation, decrease in antioxidant enzymes, and decrease in cell viability. Pretreatment of cells with N-acetyl-L-cysteine reduced high glucose-induced cytotoxicity by increasing the levels of antioxidant enzymes, and decreasing the number of apoptotic cells. Further, at molecular level high glucose treatment resulted in a significant increase in phosphorylation of Akt, MAPKs, tuberin, down regulation of 8-oxoG-DNA glycosylase and increase in 8-hydroxydeoxyguanosine accumulations. Pretreatment of cells with N-acetyl-L-cysteine, phosphatidylinositol3-kinase/Akt and ERK1/2 inhibitors completely abolished the apoptotic effects of high glucose. Moreover, N-acetyl-L-cysteine significantly inhibited reactive oxygen/nitrogen species generation, elevated antioxidants levels, inhibited Akt, ERK1/2, tuberin phosphorylation, decreased 8-hydroxydeoxyguanosine accumulation and upregulated 8-oxoG-DNA glycosylase expression. Our results demonstrate that high glucose induces apoptosis and inhibits proliferation of glial cells, which may be mediated by the phosphorylation of tuberin, down regulation of 8-oxoG-DNA glycosylase and 8-hydroxydeoxyguanosine accumulation via activation of Akt and ERK1/2MAPK pathways.

Epidemics of severe cholera caused by El Tor Vibrio cholerae O1 Ogawa possessing the ctxB gene of the classical biotype in Orissa, India

BACKGROUND We investigated the epidemic of cholera that occurred in Kashipur and Dasmantpur blocks of Orissa, reported during July-September 2007. METHODS Sixty-two rectal swabs and 28 water samples collected from diarrhea patients at different hospitals and villages were bacteriologically analyzed for the identification, antibiogram, and detection of toxic genes of Vibrio cholerae. RESULTS The cholera outbreaks were caused by V. cholerae O1 Ogawa biotype El Tor in both Kashipur and Dasmantpur blocks. All the V. cholerae isolates from the clinical and environmental samples were sensitive to tetracycline, gentamicin, azithromycin, and chloramphenicol, but were resistant to ampicillin, ciprofloxacin, norfloxacin, co-trimoxazole, nalidixic acid, neomycin, and furazolidone, except the water isolates, which were sensitive to ciprofloxacin and norfloxacin. The multiplex PCR assay revealed that all the clinical and environmental V. cholerae isolates were positive for the ctxA and tcpA genes, showing biotype El Tor. Interestingly, 88% of the clinical and environmental isolates of V. cholerae were El Tor biotype with mutation at the ctxB gene of the classical strain, as confirmed by mismatch amplification of mutation (MAMA)-PCR assay. CONCLUSIONS This is the first report of the El Tor variant of V. cholerae O1 Ogawa having the ctxB gene of the classical strain with altered antibiogram causing epidemics of cholera in Orissa, India.

Quadruplex PCR for Simultaneous Detection of Serotype, Biotype, Toxigenic Potential, and Central Regulating Factor of Vibrio cholerae

ABSTRACT A quadruplex PCR was developed for the simultaneous detection of genes specific for Vibrio cholerae O1 and/or O139 serogroup (wbe and/or wbf), cholera toxin A subunit (ctxA), toxin-coregulated pilus (tcpA), and central regulating protein ToxR (toxR) in a single tube reaction. This is a simple, rapid, and accurate approach for the detection of toxigenic V. cholerae O1 and/or O139 and can prevent the rapid spread of the disease by early detection.

Emergence and Spread of Tetracycline resistant Vibrio cholerae O1 El Tor variant during 2010 cholera epidemic in the tribal areas of Odisha, India

OBJECTIVES The epidemics of cholera were reported in the Kashipur, K.singhpur, B cuttack blocks of Rayagada district and Mohana block of Gajapati district of Odisha during 2010. The present study was carried out to isolate the bacterial pathogen, its drug sensitivity pattern and to describe the spread of the disease in those areas. METHODS A total of 68 rectal swabs collected from patients with severe diarrhea, admitted to different health centers and diarrhea affected villages were bacteriologically analyzed. Similarly 22 water samples collected from different villages from nala, chua, etc were tested for the presence of V cholerae. RESULTS Out of 68 rectal swabs tested 35 (51.5%) were V cholerae O1 Ogawa and 14(20.6%) were E coli; which might be commensals. All water samples were negative for V cholerae. The V cholerae strains were sensitive to gentamicin, norfloxacin, ciprofloxacin, azithromycin and ofloxacin; but were resistant to ampicillin, tetracycline, nalidixic acid, furazolidone, streptomycin, erythromycin, co-trimoxazole, neomycin and chloramphenicol. All V cholerae strains were 100% resistant to tetracycline and they were El Tor variants harboring ctxB gene of classical strain. CONCLUSIONS The present study indicated the emergence and spread of tetracycline resistant V cholerae O1 El Tor variant in the tribal areas which needs close monitoring.

Vibrio cholerae O1 Ogawa Strains Carrying the ctxB7 Allele Caused a Large Cholera Outbreak during 2014 in the Tribal Areas of Odisha, India

The large outbreak of cholera reported during July to September 2014 in the Narla block of Kalahandi district, India, was investigated to determine the causative organism. Rectal swabs collected from patients with diarrhea and environmental water samples were cultured following standard techniques. The causative organism was identified as Vibrio cholerae O1 Ogawa biotype El Tor, and analysis by double mismatch mutation assay PCR confirmed that all strains were the ctxB7 variant of Haitian V. cholerae O1. The environmental water samples were negative for V. cholerae. The V. cholerae O1 strains were sensitive to tetracycline, ciprofloxacin, norfloxacin, ofloxacin, doxycycline, and azithromycin, but were resistant to erythromycin, gentamicin, chloramphenicol, furazolidone, neomycin, cotrimoxazole, nalidixic acid, and ampicillin. In the 2014 cholera outbreak, the early reporting of the pathogen enabled the government authorities to implement adequate control measures in time to curtail the spread of the disease. That was the second large cholera outbreak due to Haitian variants of V. cholerae O1 after the 2010 Haiti cholera outbreak reported from Odisha, India, and other locations globally. Active surveillance is required to track the spread of this strain in the Odisha region.

Prevalence of Staphylococcus aureus associated with Skin and Soft Tissue Infection (SSTI) among septic patients from Bhubaneswar

Staphylococcus aureus is a major gram positive bacterial pathogen that causes a wide spectrum of clinical infections, ranging from localized soft-tissue infections to life-threatening bacteremia and endocarditis. S. aureus can infect tissues when the skin or mucosal barriers have been breached. This can lead to many different types of infections, including boils, carbuncles (a collection of boils) and abscesses. Deeply penetrating S. aureus infections can be severe. The incidence of methicillin resistant S. aureus (MRSA) in India ranges from 30-70%. The present study investigates the detection of S. aureus from pus swabs of hospitalized patients (Capital Hospital, Bhubaneswar) having skin infections and abscesses and its’ susceptibility pattern to different antibiotics. Out of 230 samples collected 204 (88.9%) were culture positive for different bacterial pathogens from which S. aureus was 54 (23%). The incidence rate of S. aureus among male and female group studied was 56.3% and 43.7%, respectively. The isolated S. aureus was found to be resistant to most of the antibiotics such as azithromycin, doxycycline, ciprofloxacin, tetracycline, gentamycin, ofloxacin, chloramphenicol, ampicillin and oxacillin. Among the various antibiotics, the isolated S. aureus strains revealed resistant to methicillin (MRSA) and vancomycin (VRSA) were 90.7% and 14.8, respectively. The MRSA strains were confirmed genotypically by amplification of methicillin resistant (mec A) gene. S. aureus identification and its antibiogram profile are highly essential for implementation of treatment and control of the infection in Odisha. Citation: Mohanty, A. and Pal, B.B. Prevalence of Staphylococcus aureus associated with Skin and Soft Tissue Infection (SSTI) among septic patients from Bhubaneswar [Abstract]. In: Abstracts of the NGBT conference; Oct 02-04, 2017; Bhubaneswar, Odisha, India: Can J biotech, Volume 1, Special Issue, Page 127. https://doi.org/10.24870/cjb.2017-a113

Genomic profile of antibiotic resistant, classical ctxB positive Vibrio cholerae O1 biotype El Tor isolated in 2003 and 2005 from Puri, India: A retrospective study

Objectives: To examine eight strains of Vibrio cholerae O1 isolated in 2003 and 2005 from Puri, India, for antibiotic susceptibility, presence of virulence and regulatory genes, cholera toxin (CT) production, CTX arrangement and genomic profiles. Materials and Methods: Bacterial strains were tested for antibiotic susceptibility using disc diffusion assay. Polymerase chain reaction determined the presence of antibiotic resistance, virulence and regulatory genes. To determine the type of cholera toxin subunit B (ctxB), nucleotide sequencing was performed. Southern hybridisation determined the number and arrangement of CTXΦ. Ribotyping and pulsed-field gel electrophoresis (PFGE) were used to determine the genomic profile of isolates. Results: All the eight strains, except one strain, showed resistant to nalidixic acid, sulphamethoxazole, streptomycin and trimethoprim and possessed the sullI, strB, dfrA1 and int SXT genes. All the strains carried the toxin-co-regulated pilus pathogenicity island, the CTX genetic element, the repeat in toxin and produced CT. Restriction fragment length polymorphism (RFLP) analysis showed that V. cholerae O1 possess a single copy of the CTX element flanked by tandemly arranged RS element. Nucleotide sequencing of the ctxB gene showed the presence of classical ctxB. RFLP analysis of conserved rRNA gene showed two ribotype patterns. PFGE analysis also showed at least three PFGE patterns, irrespective of year of isolations, indicating the genomic relatedness among them. Conclusion: Overall, these data suggest that classical ctxB-positive V. cholerae O1 El Tor strains that appeared in 2003 continue to cause infection in 2005 in Puri, India, and belong to identical ribotype(s) and/or pulsotype(s). There is need to continuous monitor the emergence of variant of El Tor because it will improve our understanding of the evolution of new clones of variant of V. cholerae.