HIV-1C in-House RNA-Based Genotyping Assay for Detection of Drug Resistance Mutations in Samples with Low-Level Viral Loads

Ontlametse T Bareng,1,2 Wonderful T Choga,1,2 Segomotso T Maphorisa,3 Sekgabo Seselamarumo,1 Kaelo K Seatla,1 Patrick T Mokgethi,1,4 Dorcas Maruapula,1,4 Mompati L Mogwele,1 Doreen Ditshwanelo,1,5 Natasha O Moraka,1 Irene Gobe,2 Modisa S Motswaledi,2 Joseph M Makhema,1,6 Rosemary Musonda,1 Roger Shapiro,1,6 Max Essex,1,6 Vlad Novitsky,1 Sikhulile Moyo,1,2,6 Simani Gaseitsiwe1,6 1Botswana Harvard AIDS Institute Partnership, Gaborone, Botswana; 2School of Allied Health Professions, Faculty of Health Sciences, University of Botswana, Gaborone, Botswana; 3Ministry of Health and Wellness, Republic of Botswana, Gaborone, Botswana; 4Department of Biological Sciences, Faculty of Science, University of Botswana, Gaborone, Botswana; 5Department of Biological Science and Biotechnology, Botswana International University of Science and Technology, Palapye, Botswana; 6Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, MA, USACorrespondence: Simani Gaseitsiwe, Botswana Harvard AIDS Institute Partnership, Private Bag BO320, Bontleng, Gaborone, Botswana, Tel +267 390 2671, Fax +267 390 1284, Email sgaseitsiwe@bhp.org.bwPurpose: Monitoring HIV-1 drug resistance mutations (DRM) in treated patients on combination antiretroviral therapy (cART) with a detectable HIV-1 viral load (VL) is important for the selection of appropriate cART. Currently, there is limited data on HIV DRM at low-level viremia (LLV) (VL 401– 999 copies/mL) due to the use of a threshold of VL ≥ 1000 copies/mL for HIV DRM testing. We here assess the performance of an in-house HIV drug resistance genotyping assay using plasma for the detection of DRM at LLV.Methods: We used a total of 96 HIV plasma samples from the population-based Botswana Combination Prevention Project (BCPP). The samples were stratified by VL groups: 50 samples had LLV, defined as 401– 999 copies/mL, and 46 had ≥ 1000 copies/mL. HIV pol (PR and RT) region was amplified and sequenced using an