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Periodic evaluation of the impact of viral diversity on diagnostic tests is critical to ensure current technologies are keeping pace with viral evolution. To determine whether HIV diversity impacts the ARCHITECT HIV Combo Ag/Ab (HIV Combo) or RealTime HIV-1 (RT) assays, a set of N = 199 HIV clinical specimens from Cameroon, Senegal, Saudi Arabia, and Thailand were sequenced and tested in both assays. The panel included historical groups N and P specimens and a newly identified group N specimen. These and specimens classified as H, U (unclassified)/URF (unique recombinant form), CRF (circulating recombinant form) 01, 02, 06, 09, 11, 13, 18, 22, 37, and 43 were detected by both the RT assay (1.75-6.84 log copies/ml) and the HIV Combo assay (3.26-1121.96 sample to cutoff ratios). Sequence alignment identified 3 or fewer mismatches to the RT assay oligos in 82.4% of samples. Altogether, these data demonstrate the HIV Combo and RT assays detect diverse strains of HIV in clinical specimens.

HIV-1 strain diversity was examined in a study population that consisted of hospital and clinic patients from seven cities and villages located in the northwestern regions of Cameroon. Specimens were screened using a serological algorithm designed to identify HIV-1 group M, N, and O, and SIVcpz-like infections followed by RT-PCR amplification to characterize the infecting virus. The results show that the HIV epidemic in northwest Cameroon is dominated by HIV-1 group M CRF02_AG infections (57%). Additional group M subtypes present include A, D, F2, G, and CRF01_AE. Based on discordant subtype classification between gag and env sequences, a high percentage (23%) of viral strains appear to be unique intersubtype recombinants with the majority (88%) involving recombination with CRF02_AG. Group O prevalence is low accounting for only 0.4% of HIV infections. However, group O strain diversity is high; isolates from clades I, IV, and V, as well as unclassified and recombinant strains, were found. Three dual infections by HIV-1 group M and group O were identified and characterized. In two specimens, both group M and O sequences were amplified in gag, pol, and env suggesting the presence of both viruses. Analysis of the third specimen shows the presence of a group O virus and an intergroup M/O recombinant virus. Finally, no infections due to HIV-1 group N or SIVcpz-like strains were found in the study population.

HIV-1 group O strains have a level of genetic diversity similar to that of strains in group M; however, group O has not been readily classified into genetic subtypes. Phylogenetic classification of group O has been hindered by the limited sequence information available. To facilitate phylogenetic analysis, we sequenced the gag p24 (693 nt), pol p32 (864 nt), and env gp160 (approximately 2700 nt) genes from 39 group O-infected specimens. These specimens include 32 plasma samples collected in Cameroon between 1996 and 1999, 2 specimens collected in the United States, and 5 infections previously isolated in Equatorial Guinea. Phylogenetic analysis of HIV-1 group O sequences resulted in the identification of five clusters that are maintained across gag, pol, and env, generally supported by high bootstrap values, and approximately equidistant from each other. In addition to the group O clusters, several isolates branch independently and are equidistant from the other group O isolates. Cluster I comprises greater than 50% of the group O isolates and is a diverse set of isolates that is subdivided into subclusters. The average intra-, sub-, and intercluster distances for group O are similar to the corresponding distances for group M subtypes. The five group O clusters have characteristics similar to those of group M subtypes. Thus the data presented may form the basis for classification of group O into subtypes. However, full-length genomes representing each group O cluster will be required to formalize a group O subtype classification.