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Abstract
HIV-proviral DNA and plasma-RNA sequencing for detection of drug resistance mutations in antiretroviral naive patients
Parisi S.G.1, Mazzi R.2, Zazzi M.3, Manfrin V.4, Nicolini G.5, Carolo G.6, Boldrin C.1, Franchin E.1, Dal Bello F.1, Gatti F.2, Palù G.1
1University Hospital, Padua, Italy, 2Hospital, Verona, Italy, 3University Hospital, Siena, Italy, 4Hospital, Vicenza, Italy, 5Hospital, Schio, Italy, 6Hospital, Legnago, Italy
Introduction: Genotypic analisys is commonly used in drug naive HIV-patients (pts), and plasma-RNA (PL) is taken as representative of viral population. However, in the absence of drug pressure, resistant variants may revert to or be outcompeted by more fit wild-type viruses. We tested if DNA obtained from peripheral blood mononuclear cells (PBMC) provides complementary informations to PL about drug resistance primary mutations (DRPM) in naive pts.
Methods: PL and PBMCs from 60 pts diagnosed in 2004 as HIV-infected were analyzed. The protease (PR) and the reverse transcriptase (RT) were sequenced by in-house method, and analyzed for the presence of DRPMs and for drug susceptibility profile.
Results: In 45 pts both genotype and subtype were concordant (Copt); several DRPMs were detected, conferring resistance to NRTI (3 pts), to NRTI+NNRTI (1), to PI (2). 15 pts showed substancial differences between PL-virus and PBMCs-virus (Dipt). 5 Dipts showed RT-DRPMs and resistance to NRTI (3) and NNRTI (2) in PL; in their PBMCs-DNA also PR-DRPMs were found, conferring PI-resistance. 6 Dipts revealed in PBMCs both RT and PR-DRPMs, and NRTI and PI-resistance, with drug sensitive viruses in PL. 3 out of these 6 Dipts had CRF02_AG subtype in PBMC, and B-subtype in PL. 4 Dipt showed RT and PR-DRPMs in PL, and wild-type viruses in PBMCs.
Conclusions: We found drug resistance in 21 out of 60 naive-pts (35%), but only in 6 Copts and 9 Dipts (25%) this resistance was at least partially evident with conventional PL-analysis. A substantially different virus was found in PBMCs of 10 Dipts, revealing a dual infection, a recent superinfection or a reversion of an originally infecting resistant virus, with the storage of the less fit variant, still detectable as transient prevalent population in the PBMC-archive. We suggest that PBMCs analysis is useful in naive pts to detect stored resistant variants that may compromise future therapy options.
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