Evidence of hepatitis A virus infection in the patients with acute encephalitis syndrome in Gorakhpur region, North India.

The etiological agent remained unidentified in a large number of patients hospitalized for acute encephalitis syndrome (AES) in 2008-2009 in Uttar Pradesh and Bihar, north India. All patients were found to present with fever and altered sensorium, while 28%, 19% and 13% showed hepatomegaly, splenomegaly and meningeal signs, respectively. Involvement mostly of children with abnormal hepatic features prompted us to undertake an exploratory study on viral hepatitis A to determine its association, if any, with hepatic derangements. AES patients (n = 2515) and healthy children (n = 167) were investigated for the presence of serum anti-hepatitis A virus (anti-HAV) IgM and anti-Japanese encephalitis (anti-JE) virus IgM by ELISA. Cerebrospinal fluids (CSFs, n = 595) and rectal swabs (n = 182) were examined for anti-HAV IgM and/or HAV RNA. Anti-HAV IgM was detected in the sera of 14.6% patients as against 6.6% of healthy children (p = 0.0042). Anti-JE virus IgM positivity was Keywords: acute encephalitis syndrome; cerebrospinal fluid; hepatitis A virus; anti-HAV IgM; non-Japanese encephalitis.

Group C rotavirus infection in patients with acute gastroenteritis in outbreaks in western India between 2006 and 2014.

Faecal specimens collected from outbreak (n = 253) and sporadic (n = 147) cases of acute gastroenteritis that occurred in western India between 2006 and 2014 were tested for group C rotavirus (GCR) using partial VP6 gene-based RT-PCR. All specimens were tested previously for the presence of other viral and bacterial aetiological agents by conventional methods. The rate of GCR detection was 8·6% and 0·7% in outbreak and sporadic cases, respectively. GCR infections prevailed in outbreaks reported from rural areas (10·9%) compared to urban areas (1·6%). Clinical severity score of the patients with GCR infection (n = 23) indicated severe disease in the majority (70%) of cases. The age distribution analysis indicated 52·1% of GCR infections in children aged <10 years. The male:female ratio in GCR-positive patients was 2·3:1. Of the 23 GCR-positive cases, 17 (73·9%) had a sole GCR infection and six had mixed infections with other viral and/or bacterial agents. Phylogenetic analysis of nucleotide sequences classified GCR strains of the study in to I2 genotype of the VP6 gene. This is the first study to show the occurrence of GCR in gastroenteritis outbreaks in India.

Cellular antioxidant and pro-oxidant actions of nitric oxide.

We describe a biphasic action of nitric oxide (NO) in its effects on oxidative killing of isolated cells: low concentrations protect against oxidative killing, while higher doses enhance killing, and these two effects occur by distinct mechanisms. While low doses of NO (from (Z)-1-[N-(3-ammonio propyl)-N-(n-propyl)-amino]-diazen-1-ium-1,2(2) diolate [PAPA/NO] or S-nitroso-N-acetyl-L-penicillamine [SNAP] prevent killing of rat hepatocytes by t-butylhydroperoxide (tBH), further increasing doses result in increased killing. Similar effects occur with rat hepatoma cells treated with PAPA/NO and tBH or H2O2. Increased killing with higher concentrations of NO donor is due to both NO and tBH, because NO donor alone is without effect. Glutathione (GSH) is not involved in either of these actions. Based on measurements of thiobarbituric acid-reactive substances (TBARS) and effects of lipid radical scavenger (DPPD) and deferoxamine, the protective effect, but not the enhancing effect, involves peroxidative chemistry. Fructose has no effect on tBH killing alone but provides substantial protection against killing by higher concentrations of NO plus tBH, suggesting that the enhancing effect involves mitochondrial dysfunction. Hepatocytes, when stimulated to produce NO endogenously, become resistant to tBH killing, indicative of the presence of an NO-triggered antioxidant defensive mechanism. The finding that the protective effects of low concentrations of NO and the harmful effects of high concentrations of NO are fundamentally different in nature suggest that therapeutic interventions could be designed, which selectively prevent its pro-oxidant activity at high concentrations, thus converting NO from a "Janus-faced" modulator of oxidant injury into a "pure" protectant.

Isolation, cell culture, and characterization of oviduct epithelial cells of the cow.

This report describes an easy method of isolation and cell culture of the epithelial cells of cow oviduct. Incubation of cow oviduct with 0.1 mg/ml collagenase in the lumen for 90 minutes helped to dislodge large numbers of ciliated and secretory epithelial cells. The isolated cells, when seeded on plastic, proliferated very quickly and became confluent in 8-10 days in 35 mm Petri dishes. The isolated ciliated cells which attached to the plastic dish lost their cilia after 4-5 days in culture. The cultured epithelial cells were keretin positive. The isolated bovine oviduct epithelial cells, when cultured on plastic precoated with 10 mg/ml matrigel, organized themselves into hollow tubes or spheres with microvilli directed towards the lumen. The epithelial cells seeded on 2 mg/ml matrigel became subconfluent in 15-20 days after seeding. The histoarchitecture of the secretory cells growing in vitro on matrigel resembled that of intact oviduct secretory epithelial cells. Occasional ciliated cells containing large number of mitochondria were observed in the monolayer cultured on 2 mg/ml matrigel substratum but possessed few cilia. The oviduct epithelial cells cultured on 2 mg/ml matrigel incorporated 35S-methionine linearly into protein up to 8 hours in the presence of estradiol or progesterone. The fluorograph of the newly synthesized proteins indicated the presence of an additional 60 kd protein in the cell extract of epithelial cells incubated with estradiol.

Preparation and short-term culture of enriched populations of guinea pig spermatocytes and spermatids.

In order to study acrosome biogenesis in vitro, the authors developed techniques for the isolation and culture of enriched populations of guinea pig spermatogenic cells using a modification of the technique of Romrell et al (1976). The modifications include changes in the medium, enzyme concentrations, cell loads for gradient separations, and sedimentation times. Three major cell populations were pooled: Pachytene spermatocytes (PS: 2.0 x 10(7), 80-85% pure), round spermatids, (RS: 1.2 x 10(8), 80-85% pure), and condensing spermatids (CS: 2.0 x 10(8), 50-60% pure, contaminated with residual bodies). The ultrastructural properties of the isolated cells appeared similar to those of cells in situ. PS were 3-4 times more active than RS and 10 times more active than CS in synthesizing proteins. These experiments demonstrate that highly enriched populations of guinea pig spermatogenic cells can be isolate, and that these cells can be cultured in the presence of radioactive precursors for studies of protein synthesis during spermatogenesis.

Fecal shedding of hepatitis A virus in Indian patients with hepatitis A and in experimentally infected Rhesus monkey.

Hepatitis A is highly endemic in India. The surveillance reports for the disease from this region are primarily based on the demonstration of hepatitis A virus (HAV) specific serum IgM and IgG antibodies. The present study was conducted to assess the presence and duration of fecal shedding of HAV in patients with hepatitis A and in an experimentally infected rhesus monkey. Nested reverse transcription polymerase chain reaction (RT-PCR) was applied to fecal specimens from 67 sporadic cases of hepatitis A. Recent infection with HAV in these cases was evidenced by the presence of serum anti-HAV IgM. Fecal HAV RNA positivity was observed in nearly 40% patients. The proportion of HAV RNA positivity in fecal specimens obtained within the first week (36.58%) was not different from those collected in 2-12 weeks post onset (42.42%) (P>0.05). A significant number of HAV RNA positive stool specimens showed presence of full virus particles by immune electron microscopy (IEM). Extended fecal shedding of HAV could be a major contributory factor for high circulation of virus thereby maintaining hyperendemicity of the disease. One of the IEM positive samples was inoculated into an anti-HAV negative rhesus monkey. Serum alanine amino transferase levels of the monkey remained within the normal limits. However, HAV RNA positivity in the feces was noted from 3 to 50 days post inoculation. The monkey seroconverted to anti-HAV IgM on day 31. This study records prolonged excretion of HAV in humans as well as in experimentally infected rhesus monkey.

Primary rhabdomyosarcoma of the seminal vesicle.

Sarcomas of the seminal vesicle are very rare and poorly documented; as it is not always possible to pinpoint a truly vesicular origin of the pelvic mass due to local spread at the time of presentation. The purpose of the article is to document and characterize a rhabdomyosarcoma of the seminal vesicle of which to the knowledge of the authors there has been no previous report in the English literature.

Prevalence of hepatitis a antibodies in western Indian population: changing pattern.

This report pertains to a retrospective study conducted between 1983 and 1995 at three time points to evaluate the prevalence of hepatitis A virus (HAV) infection in the population of Bhor Taluk, situated in western India. Serum samples from children and adults were tested for anti-HAV antibodies using blocking ELISA test. There was a significant decrease in anti-HAV prevalence among children aged 5-10 years in 1995 (87.36%) as compared to that of 1983 (97.58%) and 1987 (96.48%). All individuals >11 years of age were seropositive for anti-HAV antibodies. Anti-HAV prevalence was similar in the users of well water, but was significantly reduced in individuals supplied with piped water in 1995 (88.61%) compared with that in 1983 (98.77%). A significant decrease in anti-HAV positivity was noted in children from Bhor Taluk as compared to children from Pune bled in 1992. These results underline the need for periodic surveillance of seroepidemiology of hepatitis A to determine the measures for prevention and control of the disease.

Portal hypertension in north Indian children.

Etiological factors associated with portal hypertension in children influence the decision about therapy and the prognosis. This cross-sectional observational study was performed at a tertiary care centre in northern India from January, 1990 to December, 1994. Children below the age of 14 years with suspected portal hypertension were prospectively assembled into a cohort to determine the etiology and clinical profile of portal hypertension. Of the 115 patients with portal hypertension, 76.5% had extrahepatic portal hypertension (EHPH). Remaining 23.5% of the cases had intrahepatic and post-hepatic causes of portal hypertension. Children with EHPH had a significantly earlier onset of symptoms as compared to those with intrahepatic portal hypertension (p = 0.002) and bled significantly more frequently (p = 0.00). Forty per cent of patients with chronic liver disease (CLD) never had jaundice. History suggestive of potential etiological factors could be elicited in only 7% of EHPH patients. The commonest site of block in splenoportal axis was at the formation of the portal vein. An inverse relation of bleeding rates with duration of illness was seen in EHPH. Of the 10 CLD patients in whom liver biopsy could be done, cirrhosis was present in 6 patients. Understanding the natural history of EHPH and portal hypertension due to other etiologies may have significant implications in choosing the appropriate intervention and predicting the outcome.

Electron microscopy of buffalo green monkey kidney cells persistently infected with hepatitis A virus and immunolocalization of HAV antigens.

Studies were carried out to analyse the ultrastructural changes and the distribution of hepatitis A virus (HAV)/antigens at subcellular level in buffalo green monkey kidney (BGMK) cells persistently infected with HM-175 strain of HAV. HAV infected BGMK cells showed distinct abnormalities in the endoplasmic reticulum and cytoplasmic membrane as compared to uninfected cells. The abnormalities were characterized by wavy arrays, structures like myelin, annulate lamellae, cytoplasmic inclusion bodies and vesicles. The wavy arrays within the cytoplasm of the host cells appeared to represent degenerating membranes. A complex myelin like body was found in close association with a group of virus like particles. Annulate lamellae like structures involving single paired membrane were detected infrequently whereas the cytoplasmic vesicles were numerous in these cells. An indirect immunogold technique was utilized to localize the HAV antigenin infected cells. A high density immunogold label for HIV like particles was predominantly detected in cytoplasmic vesicles. These results suggest a strong association of membrane substructure in vesicle forms with the compartmentalized replication of HAV within persistently infected host cells.

Cytogenetic alterations in buccal mucosa cells of chewers of areca nut and tobacco.

OBJECTIVE: the rationale of the study was to evaluate the cytological alterations especially micronucleus (MN) and other nuclear anomalies in buccal mucosa cells of chewers to understand the genotoxic and clastogenic potential of chewing mixture (containing areca nut and tobacco as main ingredients). METHODS: the buccal cytome assay involves the examination of epithelial smear to determine micronucleated cell and other nuclear anomalies after the Feulgen plus light green staining. The assay was applied to exfoliated buccal mucosa cells of 262 subjects [non-chewers - 161 and chewers - 101 (includes 20 subjects with OSMF)] and 1000 cells per individual were examined microscopically. Nuclear anomalies were compared among chewers, non-chewers and OSMF subjects and correlated with consumption of quids per day and duration of chewing in years. RESULTS: MN cells were found significantly (p<0.0001) higher among chewers and OSMF subjects as compared to non-chewers. Further analysis indicated that MN was significantly higher in OSMF subjects with respect to even chewers. Nuclear buds were significantly higher (p<0.0001) in OSMF subjects as compared to chewers as well as non-chewers. Nuclear anomalies viz. binucleated, karyorrhexis and karyolysis were also considerably higher in OSMF subjects as compared to non-chewers. CONCLUSION: the MN and other nuclear anomalies reflected genetic damage and cytotoxicity, associated with tobacco and areca nut consumption. Further, these data reveal a risk for development of OSMF among chewers of mixture containing areca nut and/or tobacco, as all the OSMF subjects were chewers.