A point-of-care (POC) HIV drug resistance (HIV-DR) test is needed for low- and middle-income countries (LMICs). Oligonucleotide Ligation Assay (OLA)-Simple, designed as a near-POC HIV-DR test, was assessed for its overall usability in Kenya by technicians with and without molecular laboratory PCR experience. Further, its diagnostic accuracy was evaluated by PCR-experienced technicians utilizing 147 plasma samples with known Sanger sequence genotypes—based on seven major HIV-DR mutations of nucleotide and non-nucleoside reverse transcriptase inhibitors. Thirteen laboratory technicians were recruited, including five with prior PCR experience. Twelve technicians completed the training and attained OLA-Simple testing competency, ten of whom were able to perform the OLA-Simple test within 6 h. Technicians’ survey feedback indicated the user-friendliness of OLA-Simple, citing straightforward reagent reconstitution, concise instructions in prompts, and a shorter sample-to-result test time compared to standard genotyping assays. Of the 147 archived plasma samples tested, 132 (90%) yielded interpretable results. OLA-Simple assay demonstrated a sensitivity of 97.3% (95% CI 94.5, 98.9), a specificity of 97.2% (95% CI 95.5, 98.3), and a percent agreement of 97.1% (95% CI 95.9, 98.2) compared to Sanger sequencing. This evaluation found that OLA-Simple was user-friendly among intended end-users and performed well. LMIC HIV programs would benefit from strategizing on case-use scenarios for such near-POC HIV-DR assays to improve HIV outcomes.

HIV-1 genetic diversity may influence the selection of antiretroviral drug resistance mutations (DRMs); however, subtype-specific resistance patterns remain incompletely characterized in East Africa, where subtype A1 predominates. We examined the distribution of genotypic DRMs across HIV-1 subtypes among patients with advanced HIV disease failing NNRTI-based first-line therapy in western Kenya. In this cross-sectional molecular sub-analysis, 65 hospitalized individuals (≥15 years) with virological failure (HIV-1 RNA ≥1000 copies/mL) had a 1,084-bp pol gene fragment sequenced. Subtypes were assigned using REGA. DRMs were identified using the Stanford HIV Drug Resistance Database. Associations between subtype and specific mutations were evaluated using Fisher’s exact test. Subtype A1 dominated (70.8%), followed by D (13.8%), recombinants (9.2%), and A2 (6.2%). We detected dual-class resistance in 73.8% of participants. The most frequent mutations were M184V (64.6%) and K103N/S (60.0%). Subtype A1 sequences were less frequently observed to have K101E/H (8.7%, p=0.011) and Y181C/I/V (17.4%, p=0.027) than non-A1 subtypes. No significant associations were observed between subtype and NRTI mutations. These findings suggest possible subtype-associated differences in NNRTI resistance patterns that warrant validation in larger studies, underscoring the importance of continued molecular surveillance to guide resistance monitoring and optimize long-term treatment strategies in the era of integrase inhibitor–based therapy.