MOLECULAR ANALYSIS, DISTRIBUTION AND PREVALENCE OF ROTAVIRUSES IN NIGERIA

Abstract

Information on the epidemiology of rotavirus in any particular area is necessary for vaccine development against the disease. In the study, rotavirus strains in stool samples collected from children with diarrhoea in different parts of Nigeria were characterised by: 1.Isolation of RNA and determination of its migration pattern (electropherotypes), 2. typing the nucleic acid using the PCR technique and 3. determination of the nucleotide sequence of the viral RNA. A total of 314 stool samples were collected from children aged five years and below who presented with diarrhoea from different areas of Nigeria between December 1993 and August 1995. Samples were analysed by ELISA, PAGE, PCR, Radioactive Dot-Blot hybridization and viral RNA from three of the positive cases were sequenced by the Dideoxy Chain Termination DNA techniques. Forty-five (14.3%) of the stool specimen, from the children were positive for presence of rotavirus antigens. Electropherotype were determined in 26 (57.7%) of the 45 positive samples. PCR analysis showed 41 (91.1%) and 39 (86.7%) of the 45 positive samples to be typeable for G and P serotypes and genotypes respectively. Whereas serotype G1 and G3 were evenly distributed (34.2%) across northern and southern Nigeria during the time of the study (P>0.05), mixed infection specificities were more prevalent (61. 5%) in the northern part of the country, although the difference was statistically insignificant (P>0.05). Genotype P6 was the most frequently encountered P type of rotavirus in Nigeria, occurring in 16 (41.03%) of the 39 typed specimens followed by genotype P8 accounting for 33.3% of the typed specimens. Observation on the geographical distribution of P genotypes showed that P6 was more frequently encountered (68.75%) among infected neonates in southern Nigeria while mixed infection was more prevalent (70%) among infants in northern Nigeria. Overall, strain G1P8 predominated (22.22%) followed by G3P6 (17.8%). In addition, strain G1P8 was the most prevalent among infants aged 3-9 months while strain G3P6 was the most widespread among neonates below 3 months of age (P<0.05). While strain G1P8 was the most prevalent across the country during the time of this study, type G3P6 was the most commonly identified rotavirus strain in Southern Nigeria. The occurrence of mixed infection type-specificities indicates potential for reassortment events among different rotavirus genogroups in Nigeria. The VP7 gene9 sequence data of three strains of Nigerian rotavirus were determined. Nucleotide and deduced amino acid and phylogenetic analysis of strain HMG89, showed it to be a serotype G8. Primer sequence analysis of the primer (aAT8) used initially in the serotyping assay revealed a mutation in one of the three nucleotides at the 3'end of the primer binding site accounting for its inability to detect Nigerian strains of rotavirus bearing VP7 G8 serotype. These findings demonstrate that PCR analysis can, albeit infrequently, lead to error in typing of rotaviruses due to primer mismatch in the primer binding region. The epidemiological implications of nucleotide sequence analysis for serotyping of rotavirus strains originating from different geographical regions and for vaccine development and application are discussed.