HIV drug resistance testing among patients failing second line antiretroviral therapy. Comparison of in-house and commercial sequencing

Abstract

Introduction HIV genotyping is often unavailable in low and middle-income countries due to infrastructure requirements and cost. We compared genotype resistance testing in patients with virologic failure, by amplification of HIV pol gene, followed by “in-house” sequencing and commercial sequencing. Methods Remnant plasma samples from adults and children failing second-line ART were amplified and sequenced using in-house and commercial di-deoxysequencing, and analyzed in Harare, Zimbabwe and at Stanford, U.S.A, respectively. HIV drug resistance mutations were determined using the Stanford HIV drug resistance database. Results Twenty-six of 28 samples were amplified and 25 were successfully genotyped. Comparison of average percent nucleotide and amino acid identities between 23 pairs sequenced in both laboratories were 99.51 (±0.56) and 99.11 (±0.95), respectively. All pairs clustered together in phylogenetic analysis. Sequencing analysis identified 6/23 pairs with mutation discordances resulting in differences in phenotype, but these did not impact future regimens. Conclusions The results demonstrate our ability to produce good quality drug resistance data in-house. Despite discordant mutations in some sequence pairs, the phenotypic predictions were not clinically significant.

Diagnostic Accuracy of Pan-Degenerate Amplification and Adaptation Assay for HIV-1 Drug Resistance Mutation Analysis in Low- and Middle-Income Countries

“…This has been proven to be cost-effective in high-income countries (14) but practically limited and not feasible in many LMICs, as it remains complex and costly (15,16). In Zimbabwe, although a validation of an in-house genotyping method at the African Institute for Biomedical Sciences and Technology laboratory showed good-quality HIVDR results, Chimukangara et al concluded that the implementation of such techniques in Zimbabwe was too costly (17). Additionally, Phillips et al showed that the use of GRT at the time of first-line-ART failure as part of the decision whether to switch to second-line therapy was not cost-effective in LMICs (16).…”