TANGONG CALEB NKEMTA1*
1Faculty of medicine and biomedical sciences, the university of Yaounde1
CaHReF 2018, Yaoundé Congres hall, 08 – 11 January 2019 , OAEI013
Background: The widespread use of antiretrovirals in resource-limited settings has drastically reduced mortality and morbidity as regarding HIV-1 infection. The occurrence of resistant viruses due to mutations have led to serious concerns about the reliability of our drug regimens. This therefore necessitates a cheap and simple method to identify these mutations.
Objectif: To develop and use an amplification refractory mutation system-PCR assay to determine mutation profile of HIV-1 infected patients using plasma and dried blood spots (DBS)
Methodology: A cross-sectional study in sixty HIV-1 infected adult patients on first-line ART attending the infirmary of CIRCB was conducted. Venous blood of 10ml was collected from each patient, viral loads measured, plasma and DBS prepared and stored at -20°C. RNA was extracted from samples. A 2-step PCR was conducted and the products were visualized under UV light after agarose gel electrophoresis. The mutation profiles were both detected by the ARMS-PCR and Sanger sequencing methods. Resistance mutations were interpreted and phylogenetic analysis performed. The performance of ARMS-PCR was evaluated.
Results: Amongst the 60 patients recruited, 30 samples amplified by PCR. RNA from DBS had an average concentration of 21.7ng/ul. Viral load threshold for amplification of DBS was 4.1log(12987 copies/ml). ARMS-PCR had a good performance to Sanger sequencing in detecting various major mutations(K65R, K103N, Y181C, M184V and T215Y/F) tested. The overall sensitivity, specificity, negative predictive value(NPV) and positive predictive value(PPV) were 54.5%, 75.0%, 72.0%, 57.4%, respectively; accuracy of the test being 64.75%. The p-value 0.8028 (α=0.05) showed no statistical significance between the two methods. ARMS-PCR was able to detect the above mutations in DBS with similar values in plasma.
Conclusion/Recommandation: Conclusion: We concluded that (ARMS)-PCR is a reliable method to detect major HIV drug resistance mutations using plasma and dried blood spots (DBS) with high precision.
Recommendations:
- To evaluate the use of ARMS-PCR, as a point-of-care for the detection of major drug resistance mutations against our current ART regimens.
- To carry out larger epidemiological studies on the different drug resistance mutations that can affect the current ART regimens.
Key Words: Amplification Refractory Mutation System, Drug resistance