Molecular Epidemiology and Characterization of Rotavirus strains isolated from dogs' stool

Abstract

Background: Rotavirus is one significant viral cause of acute gastroenteritis in humans and animals. It is estimated to cause approximately 218 000 deaths annually in children of less than 5 years of age globally, predominantly in developing countries. Dogs are considered as reservoirs of viral, bacterial, or parasitic zoonosis for persons of all ages due to the close relationship of dogs and humans. Contamination of the stools with rotavirus of infected dogs embodies a solemn hygienic problem in many capacities and this may be of epidemiological importance and therefore the need to conduct rotavirus prevalence studies amongst dogs especially in rural areas where animal health facilities are a scarcity. Methods: Two hundred and forty five (245) stool samples (diarrhoeal and none diarrhoeal) were randomly collected from dogs in Mafikeng and its immediate proximity, in the North West Province, between 2014 and 2016. The samples were screened for the presence of Rotavirus using a VIKIA Rota/Adenovirus Kit to determine antigen presence. Samples positive for rotavirus antigens were further analysed by Polyacrylamide Gel Electrophoresis (PAGE) to determine electrophoretic patterns and the morphology of the virus was studied using Transmission Electron Microscopy (TEM). Only five (5) PAGE Positive samples were confirmed using Real Time RT-PCR and from the five real time positive samples only two were positive by conventional RT-PCR. The resulting PCR products were sent to lnqaba for sequencing and BLASTING was completed through the National Center of Biotechnology Information (NCBI). The relatedness of the strains was analysed through a phylogenetic tree. Results: The overall prevalence based on antigen detection of rotavirus from this study was found to be 9.76% (24/245). Comparing the 3 years of data collection, the prevalence of rotavirus was shown to be higher in 2015 (16.84%) than 2014 (10.84%) and 2016 (2.94%) and when the winter and the summer season collection were compared, rotavirus prevalence was found to be higher during the winter season at 15.46%, than during the summer season which was 6.08% in line with most human rotavirus reports. The age distribution of the dogs studied was estimated to be less than 3 months and older and rotavirus prevalence considering age, was found to be 15.9% in dogs less than 3 months and 8.46% in older dogs. The prevalence between diarrhoeal and non-diarrhoeal animals was found to be 7.84% and 8.46% respectively. Two samples visualised by TEM for structural morphology revealed viral like particles of about 50nm. From PAGE analysis, only 5 samples showed rotavirus migration patterns. All (5), PAGE positive samples were amplified successfully with Real Time RT-PCR and only two samples were successfully amplified by conventional RT-PCR. Discussion: Currently there are no rotavirus prevalent studies published concerning dogs in the country. This study was considered, therefore, as a baseline study. The prevalence results obtained need to be interpreted with discretion as we do not have any other data in the country to compare with except studies from abroad. Therefore the antigen/antibody of 9.46% indicates an exposure of some dogs to the virus. The confirmation tests also revealed a low molecular prevalence of 2.04%. The two phylogenetic trees constructed indicated that the isolated rotavirus strains are not of canine origin but of human origin. The strains were also placed in Groups A and C which are common in humans. We may therefore conclude, with caution, that rotavirus infection among dogs in the area of study is currently not a problem corresponding with some reports from abroad. Though the study will contribute to the missing body of scientific knowledge further cross-sectional studies with higher numbers are recommended in order to be able to make informed conclusions about the molecular epidemiology of rotavirus infection among dogs, not only from the rural areas of Mafikeng but also from other rural areas in the country.