Use of a mutation-specific genotyping method to assess for HIV-1 drug resistance in antiretroviral-naïve HIV-1 Subtype C-infected patients in Botswana.

Background: HIV-1 drug resistance poses a major threat to the success of antiretroviral therapy. The high costs of available HIV drug resistance assays prohibit their routine usage in resource-limited settings. Pan-degenerate amplification and adaptation (PANDAA), a focused genotyping approach based on quantitative PCR (qPCR), promises a fast and cost-effective way to detect HIV drug resistance mutations (HIVDRMs).  Given the high cost of current genotyping methods, we sought to use PANDAA for screening key HIVDRMs in antiretroviral-naïve individuals at codons 103, 106 and 184 of the HIV-1 reverse transcriptase gene. Mutations selected at these positions have been shown to be the most common driver mutations in treatment failure.  Methods: A total of 103 samples from antiretroviral-naïve individuals previously genotyped by Sanger population sequencing were used to assess and verify the performance of PANDAA. PANDAA samples were run on the ABI 7500 Sequence Detection System to genotype the K103N, V106M and M184V HIVDRMs. In addition, the cost per sample and reaction times were compared. Results: Sanger population sequencing and PANDAA detected K103N mutation in three (2.9%) out of 103 participants.  There was no evidence of baseline V106M and M184V mutations observed in our study. To genotype the six HIVDRMs it costs approximately 40 USD using PANDAA, while the reagents cost per test for Sanger population sequencing is approximately 100 USD per sample. PANDAA was performed quicker compared to Sanger sequencing, 2 hours for PANDAA versus 15 hours for Sanger sequencing. Conclusion: The performance of PANDAA and Sanger population sequencing demonstrated complete concordance. PANDAA could improve patient management by providing quick and relatively cheap access to drug-resistance information.