BackgroundThis study was conducted to determine the etiology of diarrhoea in a hospital setting in Kolkata. Active surveillance was conducted for 2 years on two random days per week by enrolling every fifth diarrhoeal patient admitted to the Infectious Diseases and Beliaghata General Hospital in Kolkata.ResultsMost of the patients (76.1%) had acute watery diarrhoea in association with vomiting (77.7%) and some dehydration (92%). Vibrio cholerae O1, Rotavirus and Giardia lamblia were the important causes of diarrhoea. Among Shigella spp, S. flexneri 2a and 3a serotypes were most predominantly isolated. Enteric viruses, EPEC and EAEC were common in children <5 year age group. Atypical EPEC was comparatively higher than the typical EPEC. Multidrug resistance was common among V. cholerae O1 and Shigella spp including tetracycline and ciprofloxacin. Polymicrobial infections were common in all age groups and 27.9% of the diarrhoea patients had no potential pathogen.ConclusionsIncrease in V. cholerae O1 infection among <2 years age group, resistance of V. cholerae O1 to tetracycline, rise of untypable S. flexnerii, higher proportion of atypical EPEC and G. lamblia and polymicrobial etiology are some of the emerging trends observed in this diarrhoeal disease surveillance.

Three rare human G12 strains were detected from diarrheic clinical samples of children (<8 months of age) in Calcutta during a routine surveillance study of rotaviral diarrhea in India. The VP7 genes of G12 strains and their products showed maximum homology (97 to 99% at the nucleotide level and 98% at the amino acid level, respectively) with those of two recently reported G12 strains (from the United States and Thailand) but lesser homology with those of prototype G12 strain L26.Rotaviruses are the major cause of acute gastroenteritis in infants and young of a wide variety of mammalian and avian species (11). In developing countries, approximately 130 million infants are infected with rotaviruses and there are 800,000 annual deaths (5, 12). In group A rotaviruses, 15 VP7 G serotypes and 21 VP4 P genotypes have been reported for humans, animals, and birds (17). Though 10 G types and 10 P types from humans have been reported (15), clinically and epidemiologically the most important strains belong to the G1 to G4 serotypes with P[8] and P[4] genotypes (6,14). On the other hand, some unusual types (G5, G8, and G9) and rare combinations of G and P types from different countries have also been reported (1,2,3,4,13,16,20). The human rotavirus G12 strains (L26 and L27) were first detected from Philippines in 1990 (19). After more than a decade, in 2002, human G12 strains Se585 (9) and T152 (15) from the United States and Thailand, respectively, were reported. In this study, we report the detection of three rare G12 strains of human rotaviruses in India.As part of a routine surveillance study for diarrheal diseases, stool samples were collected from children below 4 years of age from B. C. Roy Children's Hospital and also from patients of all age groups admitted to the Infectious Diseases Hospital, Calcutta, India, in 2001. Stool samples were also collected from children with diarrhea from Assam Medical College, Dibrugarh, Assam, India; Capital Hospital and Municipality Hospital, Bhubaneshwar, Orissa, eastern India; and Post-Graduate Institute of Medical Education and Research, Chandigarh, northern India. A total of 454 samples were screened for rotaviruses by RNA electrophoresis as described earlier by Herring et al. (10). The fecal specimens were processed for extraction of rotavirus double-stranded RNA suitable for reverse transcription and amplification of the VP4 and VP7 genes as described previously (4). G and P typing of positive samples was carried out by nested and multiplex PCR with consensus and type-specific primers as described previously (4,7,8,18,21). The amplified products were purified with either the QIAquick gel extraction kit or the PCR purification kit (Qiagen, GmBH) in accordance with the manufacturer's instructions. Direct sequencing was carried out by using ABI PRISM BigDye terminator cycle sequencing kits (Applied Biosystems) with an automated DNA sequencer, the ABI PRISM 310 genetic analyzer (Applied Biosystems). The sequencing of the VP7 genes of all three isolates was repeated three times to...

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(1.25%), were also detected during this study period. Twenty two percent of specimens showed mixed infections, 38 (24%) of the total samples remained untypeable for either VP7 or VP4, while only 4 (2.5%) of the samples were untypeable for both genes. Eleven specimens collected from Manipur were also genotyped and revealed a very high degree of genomic reassortment.Group A rotaviruses are the major cause of severe dehydrating gastroenteritis among human infants and young of a wide variety of mammalian and avian species (17), and it has been estimated that nearly a million deaths occur every year (9, 20), predominantly in developing countries.Fifteen G serotypes of rotaviruses (27) are recognized, depend on the molecular characterization of VP7 (glycosylated outer capsid protein). However, G1 to G4 are the most predominant genotypes in humans. Moreover, a number of unusual genotypes, G5, G8, and G9, have also been reported recently from various countries (8). Thirteen P serotypes (22) and 21 P genotypes are recognized on the basis of VP4 (protease-sensitive outer capsid protein), but the most common P serotypes infecting humans are P1A and P1B, corresponding to the P[8] and P[4] genotypes, respectively. Although the role of VP4 protein in protective immunity is not very well established, information on G and P typing is important for identifying unusual or new virus strains circulating in different populations (19).The first human rotavirus vaccine (a human-animal tetravalent vaccine) was licensed in the United States in 1998; however, the same vaccine has not been tried extensively in other countries. Due to strain diversity in different parts of the world, knowledge of molecular epidemiology and antigenic diversity of rotaviruses in circulation is imperative for the development of a suitable, efficacious vaccine to combat rotaviral diarrhea. Therefore, in this study we report the characterization of different G and P types of rotavirus strains circulating in a particular area of India.

Long electropherotype with Subgroup I specificity is a common feature of animal rotaviruses. In an epidemic of infantile gastroenteritis in Manipur, India, long but SG I strains predominated in the outbreak in the year 1987-88. One such strain isolated from that region, following the outbreak had G9P [19] specificity. As this is a rare combination, the gene sequences encoding VP4, VP6, VP7, NSP1, NSP2, NSP3, NSP4 and NSP5 of this strain were analyzed. All these genes except VP7 were closely related to porcine rotaviruses (95-99% identity at amino acid level) and clustered with the porcine strains in phylogenetic analysis. In addition, it had subgroup I nature and belonged to NSP4 genotype B which is characteristic of animal rotaviruses. This is the first report of a rotavirus with VP6 and NSP4, two crucial proteins thought to be involved in host range restriction and pathogenicity, were of porcine origin and caused diarrhoea in a human host. Among the genes of this strain sequenced so far, only VP7 had highest identity to human strains at amino acid level. This study suggests reassortment may be occurring between human and other animal strains and some of the reassortant viruses may be virulent to humans.

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