Abstract
INTRODUCTION: Drug resistance mutations (DRM) in viral RNA are important in defining to provide effective antiretroviral therapy (ART) in HIV-1 infected patients. Detection of DRM in peripheral blood mononuclear cell (PBMC) DNA is another source of information, although the clinical significance of DRMs in proviral DNA is less clear. MATERIALS AND METHODS: From 25 patients receiving ART at a center in Zimbabwe, 32 blood samples were collected. Dideoxy-sequencing of gag-pol identified subtype and resistance mutations from plasma viral RNA and proviral DNA. Drug resistance was estimated using the calibrated population resistance tool on www.hivdb.stanford.edu database. Numerical resistance scores were calculated for all antiretroviral drugs and for the subjects' reported regimen. Phylogenetic analysis as maximum likelihood was performed to determine the evolutionary distance between sequences. RESULTS: Of the 25 patients, 4 patients (2 of which had given 2 blood samples) were not known to be on ART (NA) and had exclusively wild-type virus, 17 had received Protease inhibitors (PI), 18, non-nucleoside reverse transcriptase inhibitors (NNRTI) and 19, two or more nucleoside reverse transcriptase inhibitors (NRTI). Of the 17 with history of PI, 10 had PI mutations, 5 had minor differences between mutations in RNA and DNA. Eighteen samples had NNRTI mutations, six of which demonstrated some discordance between DNA and RNA mutations. Although NRTI resistance mutations were frequently different between analyses, mutations resulted in very similar estimated phenotypes as measured by resistance scores. The numerical resistance scores from RNA and DNA for PIs differed between 2/10, for NNRTIs between 8/18, and for NRTIs between 17/32 pairs. When calculated resistance scores were collapsed, 3 pairs showed discordance between RNA and DNA for at least one PI, 6 were discordant for at least one NNRTI and 11 for at least one NRTI. Regarding phylogenetic evolutionary analysis, all RNA and DNA sequence pairs clustered closely in a maximum likelihood tree. CONCLUSION: PBMC DNA could be useful for testing drug resistance in conjunction with plasma RNA where the results of each yielded complementary information about drug resistance. Identification of DRM, archived in proviral DNA, could be used to provide for sustainable public health surveillance among subtype C infected patients.
Highlights
Drug resistance mutations (DRM) in viral RNA are important in defining to provide effective antiretroviral therapy (ART) in human immunodefficiency virus type-1 (HIV-1) infected patients
Eighteen samples had nucleoside reverse transcriptase inhibitors (NNRTI) mutations, six of which demonstrated some discordance between DNA and RNA mutations
nucleoside reverse transcriptase inhibitors (NRTI) resistance mutations were frequently different between analyses, mutations resulted in very similar estimated phenotypes as measured by resistance scores
Summary
Drug resistance mutations (DRM) in viral RNA are important in defining to provide effective antiretroviral therapy (ART) in HIV-1 infected patients. Detection of DRM in peripheral blood mononuclear cell (PBMC) DNA is another source of information, the clinical significance of DRMs in proviral DNA is less clear. Drug resistance is a current concern in the provision of effective antiretroviral therapy (ART) for human immunodefficiency virus type-1 (HIV-1) infection [1,2]. Detection of drug resistance mutations (DRM) and assessment of genotypic susceptibility is dependent on isolation, reverse transcription, amplification and sequencing of Plasma viral RNA (vRNA). Retrieving HIV resistance data from PBMCs may be easier and less expensive, but the clinical significance of DRM in proviral DNA is less clear. The appearance of DRM in PBMCs may be delayed, relative to vRNA, but once drug resistance is established, mutations are retained in archival form in PBMCs for months to years even in the absence of drug pressure [9]
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