AEGiS-14IAC: Rapid, efficient detection of HIV-1 subtypes, recombinants, and dual infections in East Africa by a multi-region hybridization assay.

14th International AIDS Conference

Barcelona, Spain - July 7-12, 2002

Rapid, efficient detection of HIV-1 subtypes, recombinants, and dual infections in East Africa by a multi-region hybridization assay.

Int Conf AIDS 2002 Jul 7-12; 14:(abstract no. TuOrC1192)

Hoelscher M, Sanders-Buell E, Dowling WE, Carr JK, Harris M, Thomschke A, Robb ML, Birx DL, McCutchan FE
Department of Infectious DIseases and Tropical Medicine, Munich, Germany

Background and

OBJECTIVE: The "gold standard" method of HIV-1 genotyping, full-genome sequencing, is costly and low-throughput, but all other assays provide incomplete discrimination of subtypes and recombinant forms. Here we describe the development, evaluation and application of the Multi-region Hybridization Assay (MHA) for the efficient determination of HIV-1 subtypes A, C, and D, recombinants, and dual infections in support of vaccine development.

METHODS: Five, well distributed genome regions (gag, pol, vpu, env and gp41) containing clustered mutations distinguishing subtypes A, C, and D were identified and used to design subtype-specific probes. DNA from primary peripheral blood mononuclear cells (PBMC) was template for real-time PCR using the fluorescent, subtype-specific probes in separate reactions.

RESULTS: A panel of 45 clinical samples from Uganda, Kenya, and Tanzania, previously characterized by full-genome sequencing and including 26 pure subtypes and 19 different recombinant strains, was evaluated by 225 probe hybridization assays. The MHA provided 90% sensitivity and 98% specificity for the three subtypes, efficiently discriminated subtypes from recombinant forms, and detected several dual infections. The MHA was also applied to a panel of 56 HIV+ samples from a high-risk cohort in Mbeya, Tanzania.

CONCLUSIONS: Accurate and efficient genotyping of HIV-1 strains in East Africa, ascertainment of the frequency and circumstances of dual infection, and elucidation of the genesis of recombinant forms in individuals will be facilitated by the application of MHA. Similar MHAs are under development for other geographic regions, each targeting 2 or 3 HIV-1 subtypes.

Keywords: AEGIS, HIV-1, Tanzania, HIV Infections, Hybridization, Genetic, Polymerase Chain Reaction, Africa, Uganda, Kenya, genetics

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