Draft Genome Sequence of Pasteurella multocida subsp. multocida B:2 Strain VTCCBAA264 Isolated from Bubalus bubalis in North India.

The Pasteurella multocida subsp. multocida B:2 serotype causes hemorrhagic septicemia in bubalines with high morbidity and mortality in the Indian subcontinent. We report the draft genome sequence of Pasteurella multocida strain VTCCBAA264 isolated from the small-intestine of a buffalo calf that died of high fever.

Prevalence of Serum Antibodies to TORCH Infection in and Around Varanasi, Northern India.

BACKGROUND: The acute infections which are caused by Toxoplasma gondii, Rubella virus, Cytomegalovirus (CMV) and the Herpes Simplex Virus (HSV-2) during pregnancy are often associated with adverse foetal outcomes and reproductive failures. In the Indian context, the exact seroprevalence of these infections is not known due to unavailability of baseline data. AIMS: The present study was undertaken to determine the serological evidence of the acute TORCH infections in women who were in the first trimesters of their pregnancies in and around Varanasi, north India. SETTINGS AND DESIGN: This study was carried out in the Sir Sunderlal Hospital, Varanasi and in the Department of Microbiology, Institute of Medical Sciences, BHU, Varanasi, UP, India. The study population involved pregnant women with bad obstetric histories, who were in the first trimester of their pregnancy. METHODS AND MATERIALS: Sera were collected from the women with Bon and they were tested for the presence of specific IgM antibodies against the TORCH infections by ELISA. STATISTICAL ANALYSIS: A 95% confidence interval was calculated for the positive cases in each of the TORCH components. RESULTS: The specific IgM antibodies were found to be positive in 74(19.4%) cases for toxoplasmosis, in 126 (30.4%) cases for the Rubella virus, in 130 (34.7%) cases for CMV and in 151 samples (33.5%) for the HSV-2 infections. CONCLUSIONS: The study showed a high prevalence of the infections which were caused by the TORCH complex amongst pregnant women with bad obstetric histories. Therefore, all the antenatal cases should be routinely screened for the TORCH infections, for carrying out early interventions to prevent foetal loss.

Molecular characterization of Vibrio cholerae O139 bengal isolated from water and the aquatic plant Eichhornia crassipes in the River Ganga, Varanasi, India.

A collection of ten strains of Vibrio cholerae O139, comprising six isolates from Eichhornia crassipes, two from water of the River Ganga, and one each from a well and a hand pump, were characterized. All the strains carried the CTX genetic element (ctxA, zot, and ace) except for the st gene and carried structural and regulatory genes for toxin-coregulated pilus (tcpA, tcpI, and toxR), adherence factor (ompU), and accessory colonization factor (acfB); all produced cholera toxin (CT). These strains were resistant to trimethoprim, sulfamethoxazole, streptomycin, and to the vibriostatic agent pteridine. Results obtained by ribotyping and enterobacterial repetitive intergenic consensus sequence-PCR fingerprint analysis indicate that multiple clones of toxigenic-pathogenic V. cholerae O139 were present in the aquatic environment.

Molecular analysis of Vibrio cholerae O1, O139, non-O1, and non-O139 strains: clonal relationships between clinical and environmental isolates.

A total of 26 strains of Vibrio cholerae, including members of the O1, O139, and non-O1, non-O139 serogroups from both clinical and environmental sources, were examined for the presence of genes encoding cholera toxin (ctxA), zonula occludens toxin (zot), accessory cholera enterotoxin (ace), hemolysin (hlyA), NAG-specific heat-stable toxin (st), toxin-coregulated pilus (tcpA), and outer membrane protein (ompU), for genomic organization, and for the presence of the regulatory protein genes tcpI and toxR in order to determine relationships between epidemic serotypes and sources of isolation. While 22 of the 26 strains were hemolytic on 5% sheep blood nutrient agar, all strains were PCR positive for hlyA, the hemolysin gene. When multiplex PCR was used, all serogroup O1 and O139 strains were positive for tcpA, ompU, and tcpI. All O1 and O139 strains except one O1 strain and one O139 strain were positive for the ctxA, zot, and ace genes. Also, O1 strain VO3 was negative for the zot gene. All of the non-O1, non-O139 strains were negative for the ctxA, zot, ace, tcpA, and tcpI genes, and all of the non-O1, non-O139 strains except strain VO26 were negative for ompU. All of the strains except non-O1, non-O139 strain VO22 were PCR positive for the gene encoding the central regulatory protein, toxR. All V. cholerae strains were negative for the NAG-specific st gene. Of the nine non-ctx-producing strains of V. cholerae, only one, non-O1, non-O139 strain VO24, caused fluid accumulation in the rabbit ileal loop assay. The other eight strains, including an O1 strain, an O139 strain, and six non-O1, non-O139 strains, regardless of the source of isolation, caused fluid accumulation after two to five serial passages through the rabbit gut. Culture filtrates of all non-cholera-toxigenic strains grown in AKI media also caused fluid accumulation, suggesting that a new toxin was produced in AKI medium by these strains. Studies of clonality performed by using enterobacterial repetitive intergenic consensus sequence PCR, Box element PCR, amplified fragment length polymorphism (AFLP), and pulsed-field gel electrophoresis (PFGE) collectively indicated that the V. cholerae O1 and O139 strains had a clonal origin, whereas the non-O1, non-O139 strains belonged to different clones. The clinical isolates closely resembled environmental isolates in their genomic patterns. Overall, there was an excellent correlation among the results of the PCR, AFLP, and PFGE analyses, and individual strains derived from clinical and environmental sources produced similar fingerprint patterns. From the results of this study, we concluded that the non-cholera-toxin-producing strains of V. cholerae, whether of clinical or environmental origin, possess the ability to produce a new secretogenic toxin that is entirely different from the toxin produced by toxigenic V. cholerae O1 and O139 strains. We also concluded that the aquatic environment is a reservoir for V. cholerae O1, O139, non-O1, and non-O139 serogroup strains.

Synthesis and antimicrobial activity of 1-aryl-2-amino-3-(4-arylthiazol-2-yl)/(benzothiazol-2-yl)guanidines.

The synthesis of fifteen new 1-aryl-2-amino-3-(4-arylthiazol-2-yl)/(benzothiazol-2-yl)gua nidines is described. They were screened for their antimicrobial susceptibility by the standard disc diffusion method of the World Health Organization (WHO) and the activities compared with that of standard strain of Escherichia coli NCTC 10418. The sensitive aminoguanidines were further subjected to the minimum inhibitory concentration (MIC) test.

Significance of Cryptosporidium in acute diarrhoea in North-Eastern India.

In a hospital-based study, stool samples from 2095 patients of all ages were examined for different fungal, protozoal and bacterial enteropathogens over a period of 2 years (July 1994-June 1996). Cryptosporidium was detected in 151 specimens (7.2%) and was the third commonest pathogen found. The highest prevalence of this organism was in the group aged 16-45 years and during the rainy months (July-Oct.). Diarrhoea caused by the protozoon was of mild to moderate severity and features of dysentery were absent. Amongst other enteropathogens, Candida albicans was the most frequently isolated, followed by enteropathogenic and enterotoxigenic Escherichia coli, Salmonella spp., Campylobacter jejuni, Entamoeba histolytica, Giardia duodenalis (lamblia), Shigella spp., Vibrio cholerae and Aeromonas spp.

Haemolysin produced by Vibrio cholerae non-O1 is not enterotoxic.

Of 28 isolates of Vibrio cholerae non-O1 (10 from diarrhoeal patients and 18 from environmental sources) examined for haemolytic activity and its correlation, if any, with enterotoxic activity, 24 showed haemolysis. The four non-haemolytic isolates showed haemolysis after consecutive passages through rabbit ileal loops (RILs). The titres of haemolytic activity were 4-64 HU/ml irrespective of their source. Eight (28.5%) of the non-O1 isolates caused fluid accumulation; six (25%) were haemolytic and two (50%) non-haemolytic. The remaining isolates showed enterotoxic activity after one-to-three consecutive passages through RILs irrespective of their haemolytic character and source. Environmental isolates caused significantly more fluid accumulation than the diarrhoeal isolates. All these isolates reverted to their original non-toxigenic character on repeated subculture or on storage in the laboratory, but continued to show haemolytic activity. The results of the present study indicate that V. cholerae non-O1 strains are potentially enterotoxigenic independent of their haemolytic character and source, and enterotoxin, not haemolysin, is the factor most likely to be responsible for their enterotoxic activity.

Development of an improved synthetic medium for a better production of the new cholera toxin and its immunological relationship with the toxin produced by Vibrio cholerae O139 strains.

An improved synthetic medium (M4) comprising syncase medium supplemented with sodium chloride (1%) and sucrose (0.5%) pH adjusted to 7.4 was developed for a better production of the new cholera toxin (NCT). The culture filtrates prepared in the M4 medium caused significantly (P > 0.05) more fluid accumulation than that in syncase medium. Crude toxin, prepared in the M4 medium with V. cholerae O1 strains (X-392 and 2740-80) caused a reaction similar to that of the same amount of NCT (32 micrograms) prepared in the syncase medium. The neutralization of the optimal loop reacting dose of the NCT prepared in the M4 medium by anti-NCT raised against syncase prepared toxin indicates the release of the same kind of toxin in both media. These observations indicate that the modified M4 medium may be used for NCT preparation and further characterization. All the strains of Vibro cholerae O139 used in this study produced a toxin antigenically similar to NCT.

Biochemical characterisation, enteropathogenicity and antimicrobial resistance plasmids of clinical and environmental Aeromonas isolates.

One hundred and eight strains of Aeromonas from clinical and environmental samples were speciated. Seven species were identified, the most prevalent of which was A. hydrophila. Experimental studies in an animal model with 36 representative strains of different species revealed that all strains could cause significant fluid accumulation in rabbit ileal loops. Of 107 strains showing single or multiple antimicrobial resistance, the highest incidence of resistance was shown for beta-lactam antibiotics other than cefotaxime. Transferable resistance plasmids, encoding resistance to ampicillin, cephalexin, cefoxitin, erythromycin and furazolidone, either alone or in combination, were detected in 35 strains. A further proportion of strains could be cured of one or more resistance markers, including resistance to nalidixic acid, and this was accompanied by the loss of plasmid DNA. The plasmids ranged in size between 85.6 and > 1 50 kb.

Attachment of non-culturable toxigenic Vibrio cholerae O1 and non-O1 and Aeromonas spp. to the aquatic arthropod Gerris spinolae and plants in the River Ganga, Varanasi.

Non-cultivable, pathogenic O1 and non-O1 Vibrio cholerae and Aeromonas spp. were resuscitated from aquatic arthropods and plant homogenate respectively, by rabbit ileal loop (RIL) assay. These organisms adhered to the aquatic arthropod Gerris spinolae and various species of phytoplankton in the River Ganga, but failed to grow after direct inoculation on artificial media except for only 10 homogenates of the arthropod. The number of non-O1 V. cholerae and Aeromonas recovered on direct inoculation of G. spinolae homogenates were in the order of 10(5)-10(6) whereas those of the Ganga water were 10(2)-10(3) ml-1. A total of 119 strains of O1 and non-O1 V. cholerae and Aeromonas spp. (69 isolates from G. spinolae and 50 from aquatic plants) were recovered from the loop contents. The results indicate that production of the enzyme chitinase by O1 and non-O1 V. cholerae and Aeromonas spp. might facilitate their adsorption and multiplication on different species of zoo- and phyto-plankton. Most of the isolates were enterotoxic, haemolytic and resistant to different antibiotics. This study suggests that species of zoo- and phyto-planktons, until now not reported to be associated with O1 and non-O1 V. cholerae, may act as reservoirs of these organisms as well as different species of Aeromonas in a fresh-water riverine ecosystem.

A community study on the aetiology of childhood diarrhoea with special reference to Campylobacter jejuni in a semiurban slum of Varanasi, India.

In a community study of 607 diarrhoeal and 529 non-diarrhoeal (control) patients less than 5 years old carried out between August 1988 and July 1989, the Campylobacter jejuni isolation rate was 4% in the diarrhoeal and 0.9% in the control group. It was the second most common bacterial enteropathogen isolated after Escherichia coli. Its incidence was more common among 1-2 year old children (4.8%) and during rainy season (July-October). Features of dysentery were absent in C. jejuni diarrhoea. Findings strongly suggest its aetiological role in childhood diarrhoea. Among other enteropathogens in diarrhoeal specimens, rotavirus was the commonest (16.4%) followed by enterotoxigenic E. coli (13.8%), G. lamblia (10.3%), enteropathogenic E. coli (7.0%), E. histolytica (5.0%), Cryptosporidium (3.8%), H. nana spp. (1.5%), NAG vibrios (0.5%), P. shigelloides (0.5%), V. mimicus and Salmonella spp. (0.3%). Approximately one quarter of the stool specimens (22.6%, 256/1136) tested were positive for the ova of A. lumbricoides.