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.

Kaelo Seatla,Dorcas Maruapula,Rosemary Musonda,Max Essex,Christopher F Rowley,Kesaobaka Molebatsi,Sikhulile Moyo,Iain J Macleod,Simani Gaseitsiwe,Melvin Leteane

doi:10.12688/aasopenres.13107.2

Abstract

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.

Highlights

HIV remains a major global health problem; currently, 37.9 million adults and children are estimated to be living with HIV with sub-Saharan Africa being the most severely affected region[1]
Here, we show that the HIV drug resistance mutations results of Pan-degenerate amplification and adaptation (PANDAA) are comparable to those produced by Sanger population sequencing
Our findings proved the potential use of PANDAA assay for testing drug resistance mutations in resource-limited settings

Summary

Introduction

HIV remains a major global health problem; currently, 37.9 million adults and children are estimated to be living with HIV with sub-Saharan Africa being the most severely affected region[1]. Major nonnucleoside reverse transcriptase inhibitor (NNRTI) mutations, such as K103N and V106M, are selected when HIV is exposed to nevirapine (NVP) and efavirenz, which is still used in both low and high resource settings as part of patient management. 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 HIV1 reverse transcriptase gene. Mutations selected at these positions have been shown to be the most common driver mutations in treatment failure. Reviewer’s comments; The combination of mutations evaluated in this study is suboptimal in the context of the current ISTI based regimens. It is most likely that the list of important mutations to be evaluated will always change and more mutations added to the list

Methods

Results

Discussion

Conclusion