Genetic Diversity of Clinical Isolates of Mycobacterium Tuberculosis Circulating Within Western Kenya Between 2013 And 2014

Abstract

Tuberculosis (TB) is a pulmonary disease caused by Mycobacterium tuberculosis complex. It is a major global public health problem with an estimated 8 million incidences and 3 million mortalities annually. In Kenya, the prevalence rate of TB was 233 per 100, 000 persons in the year 2017. Majority of the counties in Western Kenya, have been noted to have a higher than national prevalence rate of tuberculosis infections. Previous studies have identified various species and strains of Mycobacteria. However, in many clinical setups in Kenya and other developing countries, it is assumed that the most identified species is M. tuberculosis complex (MTBC). This increases the chance of misdiagnosis when other species of Mycobacteria causes TB infection in humans. Human migration has been associated with spatial dissemination of foreign strains from their local evolutionary regions to different geographical regions and this increases the risk of transfer of strains associated with drug resistance to different regions. Knowledge on the genotypes and genetic diversity is needed in ensuring adequate prevention, control and treatment strategies. This study aimed at determining the genetic diversity of the clinical isolates of Mycobacterium tuberculosis in Western Kenya. Specifically, it identified the MTBC species and strains and their proportions in Western Kenya. Furthermore, it determined the genetic diversity of M. tuberculosis complex in Western Kenya. This was a cross-sectional study on 40 archived clinical isolates(between 2013 to 2014) from patients with confirmed tuberculosis following Lowenstein-Jensen (LJ) medium cultures. The study was conducted at Moi Teaching and Referral Hospital (a national referral hospital serving Western Kenya Counties), in Eldoret –Kenya. Mycobacterial deoxyribonucleic acids (DNA) was extracted using molecular grade water in an ultrasonicator bath before 12-loci MIRU-VNTR genotyping was performed to determine the species and strains of MTBC. Oligonucleotide primer sequences for amplification used were on MIRU 02, 04, 10, 16, 20, 23, 24, 26, 27, 31, 39 and 40 loci. Descriptive statistics techniques were used to describe the study population. Genetic data analysis was conducted on the MIRU-VNTR plus web server to display NJ Tree Dendrogram (for cluster analysis) and conduct similarity searches to identify the species. Allelic diversity was calculated using Hunters-Gaston diversity index (HGDI). The species identified in this population were M. tuberculosis, M. africanum and M. bovis and the strains identified in the M. tuberculosis species were: Beijing, LAM, URAL, Uganda 1 and EAI strains. However, some isolates had a mixture of more than one species, while others had unknown species (new/unassigned). The strains were grouped into five clusters of: Beijing (n=10), LAM (n=5), Uganda 1 (n=3), EAI (n=1) and Ural (n=1) with clustering rate of 56.4%. The study revealed that there is more than a single species and strains of Mycobacterium that causes tuberculosis in Western Kenya. These strains were highly diverse as seen in the high allelic diversity index of 0.808. There is need for more awareness among healthcare and other stakeholders on the existence of foreign species and strains in Western Kenya. Policy makers should adopt molecular detection techniques of the strains circulating in Western Kenya to boost prevention, control and treatment strategies. Further studies using 24 loci MIRU-VNTR should be conducted to build on findings