The Genetic Diversity of Ndumu Virus Strains Isolated from Mosquitoes from Garissa, Ijara, Busia and Baringo during Routine Arbovirus Surveillance in Kenya between 2007 and 2009

Abstract

ABSTRACT Members of the genus Alphavirus, family Togaviridae are a diverse group of principally mosquito-borne RNA viruses that cause a variety of diseases worldwide. There are at least 29 species and many more subtypes of alphaviruses. Most of them infect humans for example Chikungunya and O’nyong-nyong viruses and cause public health problems whereas some are considered potential bioweapons, an example being Venezuelan equine encephalitis virus. Being RNA viruses, they are prone to genetic mutation like other RNA viruses, a good example being Chikungunya virus which has recently shown this phenomenon. During routine arbovirus surveillance study in selected study sites in Kenya, carried out in Viral hemorrhagic fever laboratory Centre for Virus research in Kenya Medical Research Institute, alphaviruses, identified as Ndumu viruses were isolated. They were from pools of mosquitoes that were collected from four different counties in Kenya, namely: Garissa, Ijara, Busia and Baringo. In this study, Ndumu virus isolates from the four districts in Kenya were sequenced to determine their genetic diversity. This was achieved by first culturing the virus isolates in Vero cells.The infected cells were harvested and the cell culture supernatant was used to isolate total RNA after which Reverse transcriptase polymerase chain reaction was performed using primers specific for the envelope gene (E1) to confirm the identity of the virus. This was followed by pyro-sequencing of the isolates using 454 sequencer. A mapping assembly of the sequence reads from the 454 sequencer was done against the only available partial genome sequence of Ndumu virus using GS Runmapper (Roche). Nucleotide and amino acid sequence alignments were done using the software xvi Muscle. Molecular Evolutionary Genetics Analysis software was used for phylogenetic analysis. The Ndumu virus isolates from Baringo, Busia and Garissa counties showed minimal genetic variation, only one isolate from Ijara was distinct. This study has facilitated the identification of single nucleotide polymorphisms (SNPs) within Ndumu virus genome. These SNPs however, have not changed the charge or hydrophobicity of proteins coded for. Nevertheless, this does not rule out that additional SNPs may change the protein coding sequence and affect the virus’ virulence and/or host susceptibility. The information generated from this study may be used in the design of a control strategy for alphavirus infections.