Rotavirus genetic diversity, disease association, and temporal change in hospitalized rural Kenyan children

Background. The effectiveness of rotavirus vaccines will be dependent on the immunity conferred against prevalent and emergent variants causing severe diarrheal disease. Longitudinal surveillance of disease-causing strains is a prerequisite to intervention. Methods. Molecular characterization was conducted on rotavirus-positive stool samples from children admitted with diarrhea to a rural district hospital during 2002-2004. Extracted viral RNA was separated by polyacrylamide gel electrophoresis, and rotavirus VP4 (P types) and VP7 (G types) specificities were determined. Results. Among 558 investigated cases, the predominant genotype was P[8]G1 (42%), followed by P[8]G9 (15%), P[4]G8 (7%), P[6]G8 (6%), and P[8]G8 (4%), with 10% mixed strains. Overall, there were 6 different P types and 7 G types. No association was identified between genotype and child age, sex, or severity of diarrhea. The P and G genotypes and polyacrylamide gel electropherotypes showed significant temporal variation in frequency: P[8]G1 decreased from 51% (95% confidence interval [CI], 43%-58%) in 2002 to 30% (95% CI, 24%-37%) in 2004, and P[4]G8 increased from 2% (95% CI, 0%-5%) in 2002 to 13% (95% CI, 9%-19%). Quarterly data revealed seasonally endemic and emergence and/or decay patterns. Conclusions. Our study of rotavirus strains causing severe diarrhea in rural Kenyan children showed a predominance of P[8]G1 and confirms the importance of G8 and G9 strains in sub-Saharan Africa. Considerable genetic diversity of rotavirus strains was observed, including substantial mixed and unusual types, coupled with significant temporal strain variation and emergence. These results warn of variable vaccine efficacy and the need for long-term surveillance of circulating rotavirus genotypes

Recommendations for the introduction of metagenomic high-throughput sequencing in clinical virology, part I:Wet lab procedure

Metagenomic high-throughput sequencing (mHTS) is a hypothesis-free, universal pathogen detection technique for determination of the DNA/RNA sequences in a variety of sample types and infectious syndromes. mHTS is still in its early stages of translating into clinical application. To support the development, implementation and standardization of mHTS procedures for virus diagnostics, the European Society for Clinical Virology (ESCV) Network on Next-Generation Sequencing (ENNGS) has been established. The aim of ENNGS is to bring together professionals involved in mHTS for viral diagnostics to share methodologies and experiences, and to develop application recommendations. This manuscript aims to provide practical recommendations for the wet lab procedures necessary for implementation of mHTS for virus diagnostics and to give recommendations for development and validation of laboratory methods, including mHTS quality assurance, control and quality assessment protocols

Rotavirus genetic diversity, disease association, and temporal change in hospitalized rural Kenyan children

Background. The effectiveness of rotavirus vaccines will be dependent on the immunity conferred against prevalent and emergent variants causing severe diarrheal disease. Longitudinal surveillance of disease-causing strains is a prerequisite to intervention. Methods. Molecular characterization was conducted on rotavirus-positive stool samples from children admitted with diarrhea to a rural district hospital during 2002-2004. Extracted viral RNA was separated by polyacrylamide gel electrophoresis, and rotavirus VP4 (P types) and VP7 (G types) specificities were determined. Results. Among 558 investigated cases, the predominant genotype was P[8]G1 (42%), followed by P[8]G9 (15%), P[4]G8 (7%), P[6]G8 (6%), and P[8]G8 (4%), with 10% mixed strains. Overall, there were 6 different P types and 7 G types. No association was identified between genotype and child age, sex, or severity of diarrhea. The P and G genotypes and polyacrylamide gel electropherotypes showed significant temporal variation in frequency: P[8]G1 decreased from 51% (95% confidence interval [CI], 43%-58%) in 2002 to 30% (95% CI, 24%-37%) in 2004, and P[4]G8 increased from 2% (95% CI, 0%-5%) in 2002 to 13% (95% CI, 9%-19%). Quarterly data revealed seasonally endemic and emergence and/or decay patterns. Conclusions. Our study of rotavirus strains causing severe diarrhea in rural Kenyan children showed a predominance of P[8]G1 and confirms the importance of G8 and G9 strains in sub-Saharan Africa. Considerable genetic diversity of rotavirus strains was observed, including substantial mixed and unusual types, coupled with significant temporal strain variation and emergence. These results warn of variable vaccine efficacy and the need for long-term surveillance of circulating rotavirus genotypes

Prevalence and diversity of rotavirus strains in children with acute diarrhea from rural communities in the Limpopo Province South Africa, from 1998 to 2000

BACKROUND: Data regarding the prevalence and molecular epidemiology of rotavirus infection in rural areas of Africa are limited. In this study the prevalence and genetic diversity of rotaviruses in a rural South African setting were investigated. METHODS: During June 1998 to June 2000, 420 stool specimens were collected from children with acute diarrhea who visited primary health care clinics in the rural Vhembe region, Limpopo Province, South Africa. Group A rotaviruses were detected by enzyme-linked immunosorbent assay, and the G and P types were determined by reverse-transcription polymerase chain reaction. RESLUTS: Of the 420 specimens, 111 (26.4%) were positive for group A rotavirus; P[6]G1 strains predominated (32.4%), followed by P[8]G1 (13.5%), P[6]G9 (4.5%), P[4]G8 (3.6%), P[4]G1 (3.6%), P[6]G8 (3.6%), and P[6]G2 (2.7%). Dual infections, with 11 P type, were seen in 33 (37.1%) of the positive specimens. CONCLUSIONS: The unusual serotype and genotype combinations of rotavirus circulating in the rural communities of the Limpopo Province highlight the need for more studies to monitor the geographic distribution of rotavirus strains in rural African settings