Molecular characterization of rotavirus strains among children under five years presenting with gastroenteritis at Busia county referral Hospital, Kenya

Abstract

Rotavirus has been found to be the leading cause of severe diarrhea in young children in developing countries, especially in Africa, contributing to 121,000 out of the total childhood deaths. Rotavirus has nine serogroups (A–I), but only groups A, B, and C infect humans. Group A is the most common and is responsible for most severe rotavirus gastroenteritis cases in young children. The introduction of two new live oral rotavirus vaccines, namely Rotarix® and RotaTeq®, was expected to reduce child morbidity and mortality due to rotavirus gastroenteritis. In Kenya, a previous study showed that the prevalence of rotavirus infection is 16%. The vaccine currently administered in Busia County is Rotarix®, which targets the most common strain, G1P[8]. Emerging genotypes could potentially reduce the vaccine's effectiveness. There is no documented data on the prevalence of rotavirus and circulating genotypes in Busia County. The broad objective was the molecular characterization of rotavirus strains circulating in children under five presenting with gastroenteritis at the Busia County Referral Hospital. The specific objectives were to determine the prevalence of circulating genotypes, assess vaccination status against rotavirus, and analyze the phylogenetic relationships among the identified strains. This study was a hospital based cross-sectional descriptive study on 116 patients randomly selected among those presenting with gastroenteritis. A questionnaire was used to determine their clinical and demographic characteristics and vaccination status. ELISA-based kits were used to detect Group A rotavirus antigens. Rotavirus dsRNA was extracted, separated by polyacrylamide gel electrophoresis and visualized by silver staining. Positive specimens were selected, and dsRNA extracted was reverse-transcribed and amplified using semi-nested RT-PCR in the presence of original consensus primers and genotyped using a mixture of serotype specific primers for the rotavirus genes specifying G (gene 9) and P (gene 4) classification. Phylogenetic analysis was conducted using the neighbor-joining method with MEGA software, and bootstrap analysis was performed with 1000 replications, following sample collection, RNA extraction, and sequence alignment. The prevalence of rotavirus in the study was 50.9%. The most affected age group was 7 – 12 months constituting 33.9% of the positive cases. There were more males (52.5%) than females (47.5%) infected with rotavirus. The vaccination rate was 31%. The G1P[8] genotype was the most predominant (52.5%), followed by G3P[6](20.3%), G2P[4] at (15.3%), G9P[8] (5.1%), G1,3P[6] (3.4%) and G1P[4] (3.4%) respectively. G1P[8] was common in both vaccinated and unvaccinated patients but was more prevalent in vaccinated individuals and appeared mostly in the lower severity scores indicating a less severe disease. The G3P[6] genotype accounted for 20.3% of the total infections and was higher in unvaccinated individuals, suggesting that this it may be less affected by the current vaccination regime. The G3P[6] genotype had more patients with higher severity scores (7 and 8), indicating more serious disease. Phylogenetic analysis of the circulating strains revealed that there is a high sequence similarity among many of the identified strains. The presence of multiple genotypes and slight genetic variations means that vaccines need to account for this genetic diversity, possibly due to ongoing viral evolution and adaptation. The study revealed the prevalence of rotavirus, the presence of various strains and the low vaccination coverage. These findings underscore the importance of widespread rotavirus vaccination that covers all strains. The study recommends that intervention be taken by implementing community-based programs to improve sanitation and hygiene practices, which can help reduce rotavirus transmission, as well as evaluating the effectiveness of the current vaccine, Rotarix®, against the various genotypes identified. More robust efforts also need to be put in place to increase vaccination coverage against rotavirus in the county, focusing particularly on the most affected age group, 0-12 months. Additionally, comparative genomic studies should be conducted to understand the genetic variations and evolutionary relationships among the rotavirus strains.