A Multiplex PCR Approach for Detecting Dual Infections and Recombinants Involving Major HIV Variants

Abstract

The cocirculation of different HIV types and groups can lead to dual infections and recombinants, which hinder diagnosis and therapeutic management. We designed two multiplex PCRs (mPCRs) coupled with capillary electrophoresis to facilitate the detection of such infections. The first, MMO2, targets three variants (HIV-1/M, HIV-1/O, and HIV-2), and the second, MMO, targets HIV-1/M and HIV-1/O. These mPCRs were validated on DNA and RNA extracts from 19 HIV-1/M, 12 HIV-1/O, and 13 HIV-2 cultures and from mixtures simulating dual infections. They were then assessed with DNA and RNA extracts from samples of 47 clinical monoinfections and HIV-1/M+O dual infections or infections with HIV-1/MO recombinants. Both mPCRs had excellent specificity. Sensitivities ranged from 80 to 100% for in vitro samples and from 58 to 100% for clinical samples, with the results obtained depending on the material used and the region of the genome concerned. Sensitivity was generally lower for DNA than for RNA and for amplifications of the integrase and matrix regions. In terms of global detection (at least one target gene for each strain), both mPCRs yielded a detection rate of 100% for in vitro samples. MMO2 detected 100% of the clinical strains from DNA and 97% from RNA, whereas MMO detected 100% of the strains from both materials. Thus, for in vitro and clinical samples, MMO2 was a useful tool for detecting dual infections with HIV-1 and HIV-2 (referred to as HIV-1+HIV-2) and HIV-1/M+O, and MMO was useful for detecting both MO dual infections and MO mosaic patterns.

FIG 1

(Left) mPCR-CE results for clinical HIV-M+O dual infections and/or infections with an HIV-MO recombinant. (a) DNA forms; (b) RNA forms. (Right) Partial or whole-genome recombination patterns associated with each recombinant. The HIV-1 genome is given above the recombinants, with the genomic targets of the mPCRs (MA, PR, IN, gp120, and gp41) indicated. a, abbreviations are as follows: M+O, HIV-1/M+O dual infection; MO, HIV-1/MO recombinant; M+O+MO, HIV-1/M+O dual infection plus HIV-1/MO recombinant; O+MO, HIV-1/O plus HIV-1/MO recombinant. b, detection of HIV-1/M and HIV-1/O; c, detection of HIV-1/O only; d, detection of HIV-1/M only.