RNA Migration Patterns Research Articles

Rotavirus cases among Rotarix vaccinated children below five years during the 2018 outbreak in the Kingdom of Eswatini

Background: A large outbreak of diarrhoeal disease in children under 5 years was reported in four surveillance sentinel sites Mbabane Government Hospital, Raleigh Fitkin Memorial Hospital, Hlathikhulu Government Hospital and Good Shepherd Hospital. Rapid increase of number diarrhoeal cases, were reported from week 34 to 41 (August to October 2018). Approximately 1,535 children presented with acute diarrhea, vomiting and dehydration. Methods and materials: Diarrhoeal stool samples were routinely collected for surveillance from children less than 5 years of age to determine the etiology of the diarrhoea. A total of 139 stool samples were tested for rotavirus using enzyme immunoassay kit, Molecular characterization using polyacrylamide gel electrophoresis, semi nested reverse-transcription polymerase chain reaction, sequencing and phylogenetic analysis. Rotavirus negative stool samples were further tested for bacteria and parasites using molecular diagnostic assays. Results: The rotavirus positive samples were positive for group A rotavirus with a prevalence of (117/211) 53% with the age groups mostly affected being;<12 months = (68/117)58.1%, 3-24 months = 23.9% (28/117) and >24 months = 16.2%(19/117). The PAGE patterns observed were long RNA migration pattern. The G3P[8] 59/75(78.6%) rotavirus strain were mainly associated with the outbreak. Phylogenetic analysis of the partial length VP4 P[8] associated G3 clustered together with other P[8] strains detected in Africa and distantly related to vaccine strains (Rotarix™ and RotaTeq™). Pathogens that were observed in Eswatini were Shigella/Enteroinvasive E. coli 18% (11/61) was frequently detected followed by Campylobacter jejuni/coli 13% (8/61) and Giardia lamblia 13% (8/61). Conclusion: Majority of the children affected by the rotavirus received vaccination and the rotavirus genotype G3P[8] was the major cause of these outbreak. It also important to note that the G3P[8] that was circulating was the wild type stains not related to the vaccine strains. As evident from the Maximum likelihood phylogenetic tree generated, G3P[8] study strains clustered in different lineages as compared to the vaccine strains (Rotarix™ and RotaTeq™). Further studies on whole genome sequence analysis are required to determine the evolutionary relationship among the strains and surveillance of rotavirus among the infants should be continued while increasing the rotavirus vaccine coverange and add a booster dose to increase protection.

Epidemiology and genetic diversity of group A rotavirus in acute diarrhea patients in pre-vaccination era in Himachal Pradesh, India

Acute gastroenteritis due to Group A rotaviruses remains the leading cause of mortality and morbidity in children in developing countries. India introduced its indigenous rotavirus vaccine Rotavac® in 2016 and Himachal Pradesh (HP) the first state to launch it. The present study aimed to evaluate rotavirus strain diversity associated with AGE prior to vaccine introduction in HP. A total of 331 fecal specimens collected from diarrheic children hospitalized at RPGMC Tanda, HP between July-2014 and June-2016 were screened for RVA by EIA. Rotavirus RNA was extracted by TRIZOL method and analyzed by RNA-PAGE. G/P typing was performed using semi-nested multiplex reverse transcriptase PCR. Rotavirus was detected in 45% (n = 149/331) of diarrheic children, with highest rate observed in the 6–11 months age group (47%). Vomiting was found more frequently associated with RV-infection. Among G-types, G12 was found most prevalent (33.1%) followed by G1 (28.4%), G9 (12.2%), G2 (9.5%), G3 (3.4%) and G10 (2.7%). G4 (0.7%) strains were rarely detected. Among P-types, P[6] was the most prevalent (40.5%) followed by P[8] (29.1%) and P[4] (14.2%). Of note, genotypes G3 and P[11] were detected for the first time in HP. Among G/P combinations, G12P[6] was most prevalent (30.4%) followed by G1P[8] (20.3%), G2P[4] (4.7%), G1P[6] (3.4%) and G3P[8] (2.7%). Interestingly, our study observed high percentage of unusual strains (14.2%) namely G9P[4], G2P[6], G2P[8], G12P[4] and G1P[11]. The regionally common strains G3P[6], G4P[6], G9P[6], G9P[8], G10P[6], G10P[8] and G12P[8] strains were very rarely detected. Of interest, RNA migration pattern of G1P[8] was DS-1 like and genomic heterogeneity was observed within G12P[4] strains with both long and short electropherotypes. Our study highlights rich genetic diversity with emergence of rare rotavirus strains circulating in HP and provides baseline data prior to Rotavac® introduction that will help to gauge the impact of the Rotavac® vaccine in HP.

Identification of group B rotavirus as an etiological agent in the gastroenteritis outbreak in Maharashtra, India.

Acute gastroenteritis outbreak occurred at Pargaon, Maharashtra, India in 1789 cases with an attack rate of 32.5% between November to December 2015. The stool specimens (n = 32) were investigated for different enteric viral agents using conventional methods. Transmission electron microscopy and RNA polyacrylamide gel electrophoresis respectively identified morphologically distinct rotavirus particles in 28% and RNA migration pattern of Group B Rotavirus (GBR) in 72% of the specimens. Reverse transcription polymerase chain reaction and nucleotide sequencing confirmed presence of GBR in 97% of the samples analyzed. The predominance of GBR infections and absence or insignificant presence of other agents confirmed GBR as an etiological agent of the gastroenteritis outbreak occurred in Maharashtra, India.

Evolution of DS-1-like G1P[8] double-gene reassortant rotavirus A strains causing gastroenteritis in children in Vietnam in 2012/2013

Rotavirus A (RVA) strains, a leading cause of severe gastroenteritis in children worldwide, commonly possess the Wa or DS-1 genotype constellations. During a hospital-based study conducted in Hanoi, Vietnam, in the 2012-2013 rotavirus season, G1P[8] strains with a virtually identical short RNA migration pattern were detected in 20 (14%) of 141 rotavirus-positive samples. Two representatives of these strains were shown by whole-genome sequencing to be double-gene reassortants possessing the genotype constellation of G1-P[8]-I2-R2-C2-M2-A2-N2-T2-E2-H2. Sequencing and a database search revealed that these Vietnamese G1P[8] double-gene reassortant strains shared an immediate ancestor with a locally circulating G2P[4] strain in all of the inner-capsid and non-structural protein genes, whereas they were more closely related in the VP7 and VP4 genes to a Chinese G1P[8] strain and a Chinese G3P[8] strain, respectively, than to locally circulating G1P[8] strains. Despite the marked similarity between Japanese and Thai G1P[8] double-gene reassortant strains, phylogenetic analysis suggested that the Vietnamese and Japanese/Thai G1P[8] double-gene reassortant strains originated from independent reassortment events. Clinically, children infected with Vietnamese G1P[8] double-gene reassortant strains experienced severe diarrhoea, but it was not more severe than that in children infected with ordinary G1P[8] strains. In conclusion, Vietnamese G1P[8] double-gene reassortant strains originated from a locally circulating G2P[4] strain and caused severe diarrhoea, but there was no evidence of increased virulence.

Detection of a novel equine-like G3 rotavirus associated with acute gastroenteritis in Brazil.

Genotype G3P[8] of rotavirus A (RVA) is detected worldwide, usually associated with Wa-like constellation and exhibiting a long RNA migration pattern. More recently, a novel inter-genogroup, G3P[8] reassortant variant with a short electropherotype, has emerged in Asia, Oceania and Europe, denoting an overall potential of unusual rotavirus strains. During a RVA surveillance in Brazil, G3P[8] strains were found displaying a short electropherotype pattern, which had not been detected before in this region. This study aims to characterize the complete genome of 10 G3P[8] strains detected in the northern region of Brazil. All G3P[8] samples were subjected to partial sequencing, and the whole-genome phylogenetic analysis demonstrated that all strains possessed I2-R2-C2-M2-A2-N1-T2-E2-H2 genotype background, representing reassortants with an equine-like G3 VP7 and amino acid changes in VP4 and VP7 antigenic regions as compared to vaccine strains. Phylogenetic analysis demonstrated high nucleotide identity in almost all RNA segments of G3P[8] DS-1 samples detected in Asia, Oceania and Europe as well as G3P[4] strains in Japan. This study reports a novel, equine-like G3P[8] strain circulating in Brazil and isolated from children hospitalized for severe gastroenteritis, and highlights the complex dynamics of RVA molecular epidemiology. Our findings point to a novel RVA strain emerging in this region, and studies should be done to detect whether this may represent a challenge to current vaccine strategies.

Molecular identification of a G2 rotavirus that provided a G1P[4] mono-reassortant with a DS-1-like genotype constellation.

The segmented nature of the rotavirus genome provides an opportunity for the virus to reassort upon co-infection of more than one rotavirus strain. Previously, two G1P[4] strains isolated from children with diarrhoea, AU64, and AU67, were shown by RNA-RNA hybridization to be a VP7 mono-reassortant possessing a DS-1 genogroup background. However, the origin of the parental G2 strain was not sought for at that time. The aim of this study, therefore, was to identify the G2 strain that provided AU64/AU67 with the DS-1-like genotype constellation. By carefully comparing the genomic RNA migration patterns on polyacrylamide gels of strains circulating at the same and nearby geographic locations around the time of the mono-reassortant detection (1989), a G2 strain designated 88H449 was shown to possess 10 genome segments co-migrating with those of AU64. Sequencing of all genome segments of AU64 and 88H449 showed that those two strains were 99.6-100% identical at the nucleotide level in all except the VP7 gene, indicating that the parental G2 strain that provided AU64 with the 10 genome segments was a sibling of 88H449. Sequencing of the VP7 genes of 36 G1P[8] strains circulating locally between 1981 and 1990 revealed the presence of strains bearing diverse VP7 sequences among which the closest nucleotide identity to AU64 was 98.6% and the closest amino acid identity in the major antigenic regions was 100%. This unusual mono-reassortant was concluded to be the result of a contemporary reassosrtment event occurring between locally co-circulating strains.

Electropherotypes and G-Types of Group A Rotaviruses Detected in Children with Diarrhea in Lagos, Nigeria

Approximately over 500,000 children die annually due to severe dehydrating diarrhea caused by rotaviruses. This work investigated rotavirus infection among children less than 5 years with diarrhea in Lagos and determined the circulating electropherotypes and genotypes of the virus isolates. Three hundred and two (n=302) stool samples from children below 60 months were collected from different hospitals and health care centers in Lagos and subjected to enzyme immunoassay (EIA) to determine the presence of Group A rotavirus, RT-PCR to determine the G-types, and polyacrylamide gel electrophoresis (PAGE) to determine the electropherotypes. The results show that 60.3% of the samples showed distinct rotavirus RNA migration pattern, having long electropherotypes (55.3%) of seven variations dominating over the short electropherotypes (44.5%). Six different G-types were detected (G1, G2, G3, G4, G9, and G12). Serotypes G1 and G12 showed long electropherotypic pattern while G2, G3, and G9 exhibited either short or long electropherotype. All G4 detected show short electropherotypic pattern. In conclusion, information on the genomic diversity and RNA electropherotypes of rotaviruses detected in children with diarrhea in Lagos is reported in this study.

Circulation of group A rotaviruses among neonates of human, cow and pig: study from Assam, a north eastern state of India

Rotavirus (RV) infections are worldwide in distribution causing high morbidity and mortality in human and animal neonates. Human settlements in close proximity of animals aids for genetic re-assortment of the virus by interspecies transmission and consequent emergence of new viral antigenic strain. Therefore, the present study was designed to explore RV incidence in a single approach from human and animal neonates sharing similar environment. Altogether, 200 diarrheal samples from children (50), piglets (80) and calves (70) were collected during the year of 2010-2012 from various locality, farms and hospitals, initially screened through monoclonal antibody based enzyme immunoassay followed by RNA-PAGE and VP7 gene amplification by Reverse transcription PCR. The overall prevalence of rotavirus was found to be 41.5% (83/200) where maximum numbers of positive cases were found in piglets (46.3%) followed by human (40%) and cow (37.1%). Majority of samples demonstrated characteristic group A rotavirus (RVA) electropherotype of 4:2:3:2 pattern. Moreover, RNA profiles of seven samples from piglets and calves revealed variation in the migration pattern of class II, III and class IV segments. The study, for the first time from the valley, detected 43.7% of neonatal RVA positive cases from human and animal sharing similar setting. The variation in RNA migration pattern in seven cases signifies tentative cases of gene re-assortment that warrant further evaluation.

Molecular characterization of rotavirus strains detected during a clinical trial of a human rotavirus vaccine in Blantyre, Malawi.

The human, G1P[8] rotavirus vaccine (Rotarix™) significantly reduced severe rotavirus gastroenteritis episodes in a clinical trial in South Africa and Malawi, but vaccine efficacy was lower in Malawi (49.5%) than reported in South Africa (76.9%) and elsewhere. The aim of this study was to examine the molecular relationships of circulating wild-type rotaviruses detected during the clinical trial in Malawi to RIX4414 (the strain contained in Rotarix™) and to common human rotavirus strains. Of 88 rotavirus-positive, diarrhoeal stool specimens, 43 rotaviruses exhibited identifiable RNA migration patterns when examined by polyacrylamide gel electrophoresis. The genes encoding VP7, VP4, VP6 and NSP4 of 5 representative strains possessing genotypes G12P[6], G1P[8], G9P[8], and G8P[4] were sequenced. While their VP7 (G) and VP4 (P) genotype designations were confirmed, the VP6 (I) and NSP4 (E) genotypes were either I1E1 or I2E2, indicating that they were of human rotavirus origin. RNA-RNA hybridization using 21 culture-adapted strains showed that Malawian rotaviruses had a genomic RNA constellation common to either the Wa-like or the DS-1 like human rotaviruses. Overall, the Malawi strains appear similar in their genetic make-up to rotaviruses described in countries where vaccine efficacy is greater, suggesting that the lower efficacy in Malawi is unlikely to be explained by the diversity of circulating strains.

Clinical presentation and molecular characterization of group B rotaviruses in diarrhoea patients in Bangladesh

A total of 1106 stool samples collected from diarrhoea patients admitted to Dhaka hospital of the International Centre for Diarrhoeal Disease Research, Bangladesh, during January-December 2008 were analysed for the presence of rotavirus-specific RNA by PAGE. The group B-specific RNA migration pattern was detected in 26 patients (2.4%) and group A-specific pattern in 259 patients (23.4%). Clinical data from group A and group B rotavirus-infected patients indicated that episodes did not differ much in the prevalence of diarrhoea, number of stools, outcome or differences in gender. However, abdominal pain was more common in group B rotavirus infections (36 vs 15%, P=0.02) and the virus was responsible for more severe dehydration compared with group A-infected patients (12 vs 3%, P=0.04). Sequence analyses of VP4, VP7 and NSP2 indicated that an Indian-Bangladeshi lineage of the virus, which is different from both the prototype (Chinese) lineage and from the animal group B rotaviruses, has been circulating in Bangladesh. Continuous monitoring of group B rotaviruses both in hospitals and in the community will be helpful to determine the true burden of group B rotaviruses.

Genetic diversity and classification of the outer capsid glycoprotein VP7 of porcine group B rotaviruses

We determined the nucleotide sequences of the outer capsid glycoprotein (VP7) genes of 38 porcine group B rotaviruses (GBRs) from feces of pigs at 27 farms in Japan between 2000 and 2007. Substantial diversity among porcine GBR VP7 genes was observed, with up to 42.4% difference in nucleotides and 49.8% in amino acids. On comparison of VP7 genes, porcine GBRs were clearly distinct from the published corresponding genes from human, bovine and murine GBRs (53.7-70.8% identity in nucleotides and 45.8-73.4% identity in amino acids). Phylogenetic analysis showed that the VP7s of GBRs could be divided into five genotypes: the murine strain was genotype 1, human strains were genotype 2, bovine and some porcine strains were genotype 3, and other porcine strains belonged to genotype 4 or 5. In addition, GBR VP7s in genotypes 3 and 5 were further divided into four and five clusters, respectively. No relationship between VP7 genotype and double-stranded RNA migration patterns of porcine GBRs in polyacrylamide gel electrophoresis were observed. However, an antigen enzyme-linked immunosorbent assay using antiserum to recombinant bovine GBR VP6 did not react with fecal samples containing one cluster of genotype 5 of porcine GBRs. The abundant divergence of porcine GBR VP7 genes suggests that porcine species might be an original natural host of GBR infection and that different serotypes might exist among porcine GBRs. To our knowledge, this is the first report to describe the gene sequences and typing of porcine GBR VP7s.

Detection of human rotavirus in hospitalized diarrheic children in central India.

During the present study, group A human rotaviruses were detected among diarrheic children using polyacrylamide gel electrophoresis (PAGE) technique, with a typical RNA migration pattern of 4:2:3:2, suggestive of group A rotavirus. During the study, a total of 46 fecal samples collected from hospitalized children with acute diarrhea as well as children inhabiting nearby animal farms with history of presence of animal rotaviruses on the farms were processed for detection of human rotavirus. Out of 33 diarrheic children, 12 showed presence of rotavirus infection (36.36%), however, none of the children from animal farm areas showed presence of rotavirus. Female children were more susceptible to rotavirus infection (46.15%) than males (30%). Majority of the cases of rotavirus gastroenteritis belonged up to one year of the age, with an incidence of 40.91%. RNA profile of rotaviruses suggested circulation of 5 different electropherotypes in this geographical locale of the country, indicating existence of genomic diversity among human rotaviruses. Majority of the isolates were of long pattern (66.67%), whereas short pattern was detected only in one third of the viruses. This preliminary study emphasizes for further detailed studies on the molecular characterization of rotaviruses circulating in this part of country and their relationship with other human rotavirus strains and animal strains in the country.

Detection of human rotavirus G9P[8] strains circulating in Argentina: Phylogenetic analysis of VP7 and NSP4 genes

During the surveillance of rotavirus strains that were circulating in Argentinean children from 2000 to 2004, seven rotaviruses were detected bearing the genotype combination G9P[8]. The molecular characterization of the VP7 and NSP4 genes and the RNA migration patterns support the hypothesis that rotaviruses G9 could have been reintroduced into Argentina as a novel G9P[8] strain, rather than represent VP7 gene reassortants from G9P[6] strains that had been circulating previously in this country.

Group B rotaviruses similar to strain CAL-1, have been circulating in Western India since 1993.

Generally, group A rotaviruses are the most common cause of paediatric diarrhoea. However, group B rotavirus, adult diarrhoea rotavirus (ADRV), was found to be involved in epidemics of severe gastroenteritis in several areas of China during 1982-1983 and had resulted in more than one million cases among adults as well as older children. Human group B rotavirus has been rarely reported outside China, but has been detected first from five adults with diarrhoea in Kolkata, India during 1997-1998 (strain CAL-1). During epidemiological studies at the National Institute of Virology (NIV) on hospitalized diarrhoea patients at Pune, India, faecal specimens from patients of >5 years age, which were negative for group A rotavirus by ELISA were tested by polyacrylamide gel electrophoresis (PAGE). We detected rotavirus RNA migration patterns similar to that of group B rotavirus in three faecal specimens from adults, two from the specimens collected in 1993 and one in 1998 from sporadic diarrhoea cases. RT-PCR was carried out using primers derived from gene 8 which codes for the NS2 protein, followed by nested PCR, which confirmed the presence of group B rotavirus in all three specimens. The sequences of the PCR products of NIV specimens were compared with that of CAL-1, ADRV and IDIR (infectious diarrhoea of infant rat) belonging to group B rotaviruses. The sequence analysis of the PCR products showed the highest identity with CAL-1, which was reported from Kolkata, India during 1997--1998. The finding suggests that human group B rotaviruses have been circulating in Pune. India, since 1993. This emerging virus may lead to more severe disease among adults in India. There is a need for surveillance of group B rotavirus infections, especially in adult diarrhoea cases and seroepidemiological studies on group B rotavirus are required among humans and animals of Western Maharashtra, India.

Genomic diversity and prevalence of Rotavirus in cow and buffalo calves in northern India.

Faecal samples were collected from seventy-eight diarrhoeic cow and buffalo calves between November 1998 and February 1999 to study the genomic diversity and prevalence of Rotavirus infection by ribonucleic acid polyacrylamide gel electrophoresis (RNA-PAGE) and enzyme-linked immunosorbent assay (ELISA). In the organised dairy farm (where daily production and health records were maintained), the overall prevalence of infection with Rotavirus, recorded by RNA-PAGE and ELISA, was 27.02% (10/37) in both cow and buffalo calves. In unorganised dairy herds (where no production or health records were maintained), RNA-PAGE and ELISA detected infection with Rotavirus in 26.8% (11/41) of cow and 19.5% (8/41) of buffalo calves. Five distinct electropherotypes were found to circulate in cow and buffalo calves. All were short electropherotypes except the single long electropherotype observed in a buffalo calf in an unorganised dairy herd. Some differences in RNA migration pattern were observed when these electropherotypes were compared with the neonatal calf diarrhoea virus strain of Rotavirus. Some electropherotypes were restricted to one farm while others were found in both organised and unorganised dairy herds and in both cow and buffalo calves.

Emergence of serotype G9 human rotaviruses in Australia.

Emergence of serotype G9 human rotaviruses in Australia.

Prevalence and characteristics of severe rotavirus infections in Nicaraguan children.

We analyzed the prevalence of rotavirus in 296 children age between 3 and 36 months who were hospitalized in 1994 with severe gastro-enteritis at two health centres for diarrhoea treatment in León, Nicaragua. Enteric viruses were detected in 96 (32.4%) of the children and rotaviruses were the most common pathogens detected in 84 (28%). The majority of rotavirus infections occurred in children less than 1 year old and all strains isolated belonged to subgroup II and had 'long' RNA patterns. Molecular epidemiology of 55 rotavirus strains revealed that all had the same RNA migration pattern and serotyping of 37 strains by PCR technology revealed that all isolates belonged to serotype 3. A significant observation was that only one electropherotype of rotavirus circulated. No non-group A rotaviruses were found by RNA gel electrophoresis. Adenoviruses were found by ELISA in 14 of 265 (5%) children and were most frequently detected during the 1st year of life. Of 103 faecal samples analyzed by electron microscopy, four contained small round structured viruses.

Identification of group B rotaviruses with short genome electropherotypes from adult cows with diarrhea

Two field strains (BB-RVLV and KD) of group B rotaviruses from adult dairy cows with diarrhea displayed short genome electropherotypes. Gnotobiotic calves inoculated with fecal filtrates of each group B rotavirus developed diarrhea, and only group B rotaviruses or antigens were detected in the feces by immunoelectron microscopy and in intestinal epithelial cells by immunofluorescent staining, respectively. The feces or intestinal contents of the cows and inoculated calves were negative for group A and C rotaviruses by enzyme-linked immunosorbent assay, immunoelectron microscopy, or cell culture immunofluorescence assays. Comparison of the genome electropherotypes of the calf-passaged BB-RVLV and KD strains with the original samples and reference bovine group A, B, and C rotaviruses revealed conservative of their short-genome electropherotypes and double-stranded RNA migration patterns characteristics of group B rotaviruses. To our knowledge, our previous study (L.J. Saif, K.V. Brock, D.R. Redman, and E.M. Kohler, Vet. Rec. 128:447-449, 1991) and this report are the first description of bovine group B rotaviruses (in a mixed infection with bovine coronavirus or singly in fecal contents) in adult cows with diarrhea and this is the first report of short-genome electropherotypes among group B rotaviruses.

Persistence of a single electropherotype and serotype (G6P5) of bovine rotavirus in calves on a closed dairy farm from 1990 to 1993

SUMMARY A virologic survey was conducted on calves with diarrhea associated with bovine rotavirus (brv) on a closed dairy farm. The brv was detected from 32 of 219 (14.6%) fecal specimens repeatedly collected from 56 calves born during the years 1992-1993, regardless of whether they had diarrhea. Most of the 32 strains were isolated from fecal specimens obtained from 2- to 6-week-old calves. After electrophoresis of double-stranded viral rna from the 32 strains, genomic rna migration patterns were similar to those of the predominant brv strains isolated at the same farm during the years 1990-1991. All representative strains were identified as G serotype 6 (G6) and P type 5 (P5) by results of the virus-neutralization test and polymerase chain reaction procedure. Thus, brv had no change in genomic rna electropherotypes and serologic antigenicities in a closed dairy herd over a period of several years.

Rotavirus electropherotypes in Malaysian children.

A 12-month study was carried out on the molecular epidemiology of rotavirus in urban and suburban Malaysian children. Analysis of faecal samples from 973 hospitalized diarrhoeic children by polyacrylamide gel electrophoresis detected 268 rotaviruses (28%). All isolates were group A rotaviruses, which produced 22 electropherotypes: 16 (91.5%) with long RNA migration patterns and 6 (8.5%) with short patterns. One of the long-pattern electropherotypes was the predominant strain (71.1% of the total electropherotypes) isolated during this study. Although 3 other strains were detected sporadically over the study period, 16 others were present only during the first 7 months and 2 others were confined to the last 5 months. Long- and short-pattern electropherotypes were found to co-circulate extensively. There was a significant association of short-pattern electropherotypes with infection in older children. In addition, the prevalence of vomiting and mean duration of diarrhoea were significantly associated with different electropherotypes.