Clement Likhovole Poster 2025

Clement Likhovole

 Dr Clement Likhovole

Mount Kenya University, Kenya

Novel Diagnostic Molecular Technique based on Identical Multi Repeat Sequences on the Mycobacterium tuberculosis genome for the Detection of Tuberculosis

 

Poster Abstract

Introduction: Globally in 2020, approximately 1.3 million deaths occurred among HIV-negative people because of Tuberculosis (TB) infection. The COVID-19 pandemic has reversed years of progress in reducing TB cases. To effectively manage TB cases, rapid, affordable and accurate diagnostic approaches are required. Nucleic acid amplification methods such as PCR offer high specificity and sensitivity for smear-positive TB but poorer sensitivity and specificity for smear-negative TB. These assays require specialized laboratory infrastructure and expertise. While being open systems, they are also at risk of contamination particularly in set-ups with sub-optimal laboratory infrastructure. Therefore, uptake of these assays in high-burden settings with resource scarcity is limited.

Methods: We used genome-mining approaches to identify identical multi repeat sequences (IMRS) distributed throughout the Mycobacterium tuberculosis (MTB) genome to design a primer pair that target 32 repeat sequences. H37Rv genomic DNA was diluted 10-folds from a starting concentration of 2.0×10^7 and dilutions used as template for MTB IMRS PCR and Isothermal assays. The 16S rRNA PCR assay was used as a reference method. Probit analysis was used to calculate the lower limit of detection (LLOD) of the MTB-IMRS PCR and the conventional 16S rRNA PCR. The MTB-IMRS PCR assay was also validated using 29 freshly collected sputum samples.

Results: The IMRS primers could amplify MTB genomic DNA from a concentration less than 0.1 fg/L as compared to the gold standard 16S rRNA primers (10 fg/L). the LLOD of the

MTB-IMRS was 0.1259 ng/μl and the conventional 16S rRNA PCR method was 25.1189 ng/μl. Isothermal amplification products were observed up to a concentration of 0.363pg/l using the MTB-IMRS primers.

Conclusion: De novo genome mining of MTB IMRS as amplification primers can serve as a platform for developing ultrasensitive diagnostics for TB and potentially a wide range of infectious pathogens.

 

Biography

Dr. Clement Shiluli is currently a postdoctoral research scientist at the Centre for Research in Infectious Diseases (CRID) of the Directorate of Research, College of Graduate Studies and Research of the Mount Kenya University (MKU). Since joining the CRID in 2021, Dr. Shiluli has pioneered research on point of care diagnostics for various infectious diseases. These include curable sexually transmitted infections, Schistosomiasis, Malaria, Leishmaniasis, Tuberculosis and HIV. Findings from his research at the CRID has positively impacted the design and development of point of care diagnostics in resource limited settings. In addition, he has played a critical role in providing mentorship and supervision to more than 10 postgraduate and undergraduate scholars at the CRID who are undertaking various research projects. Before joining the CRID, Dr. Shiluli was the Head of the Microbiology Department of the School of Medicine of UZIMA University. Dr. Shiluli has also worked with the Kenya Medical Research Institute Malaria and Tuberculosis research laboratories where he has contributed significantly to efforts towards eradicating infectious diseases in Kenya.