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To evaluate an algorithm using two enzyme immunoassays (EIA) for anti-HIV-1 antibodies in a rural African population and to assess alternative simplified algorithms.Sera obtained from 7895 individuals in a rural population survey were tested using an algorithm based on two different EIA systems: Recombigen HIV-1 EIA and Wellcozyme HIV-1 Recombinant. Alternative algorithms were assessed using negative or confirmed positive sera.None of the 227 sera classified as unequivocably negative by the two assays were positive by Western blot. Of 192 sera unequivocably positive by both assays, four were seronegative by Western blot. The possibility of technical error cannot be ruled out in three of these. One of the alternative algorithms assessed classified all borderline or discordant assay results as negative had a specificity of 100% and a sensitivity of 98.4%. The cost of this algorithm is one-third that of the conventional algorithm.Our evaluation suggests that high specificity and sensitivity can be obtained without using Western blot and at a considerable reduction in cost.Although the Western blot test is widely used to confirm HIV-1 serostatus, concerns over its additional cost have prompted review of the need for supplementary testing and the evaluation of alternative test algorithms. Serostatus tends to be confirmed with this additional test especially when tested individuals will be informed of their serostatus or when results will be used for research purposes. The confirmation procedure has been adopted as a means of securing suitably high levels of specificity and sensitivity. With the goal of exploring potential alternatives to Western blot confirmation, the authors describe the use of parallel testing with a competitive and an indirect enzyme immunoassay with and without supplementary Western blots. Sera were obtained from 7895 people in the rural population survey and tested with an algorithm based on the Recombigen HIV-1 EIA and Wellcozyme HIV-1 Recombinant; alternative algorithms were assessed on negative or confirmed positive sera. None of the 227 sera classified as negative by the 2 assays were positive by Western blot. Of the 192 identified ass positive by both assays, 4 were found to be seronegative with Western blot. The possibility of technical error does, however, exist for 3 of these latter cases. One of the alternative algorithms assessed classified all borderline or discordant assay results as negative with 100% specificity and 98.4% sensitivity. This particular algorithm costs only one-third the price of the conventional algorithm. These results therefore suggest that high specificity and sensitivity may be obtained without using Western blot and at a considerable reduction in cost.

The molecular diversity and phylogenetic relationship of 22 HIV-1 group O strains were examined on the basis of the protease gene and the N-terminal region of gp41env. Analysis of the newly characterized protease sequences with 12 reference sequences revealed no specific clustering patterns, despite the distinct geographic locations of the specimens. In contrast, analysis of the newly sequenced gp41 sequences with 34 published sequences revealed two distinct clusters, each represented by one full-length sequence (MVP5180 and ANT-70). Further, four of the specimens classified as group O in the protease region clustered with group M in the gp41 region (three subtype A and one subtype G, respectively), suggesting dual and/or recombinant infections with HIV-1 groups M and O. The presence of two distinct clusters in the gp41 region indicates at least two possible subtypes within group O viruses, and this may provide useful information regarding molecular epidemiological studies of group O infections.

Strategies to discriminate group O from group M infections need to be improved. We have developed and evaluated an HIV-1 group O V3 peptide-based enzyme immunoassay (PEIA) for specific HIV-1 group O antibody detection among HIV-1-infected patients. Synthetic peptides, derived from the amino acid sequences of the V3 loop of 15 different group O strains and 7 group O consensus sequences, were evaluated in a PEIA against a panel of genetically confirmed group O (n=33), group M (n=90), and HIV-1 antibody-negative sera (n=17). The best-performing PEIA(s) were then used to screen 134 sera of European and 336 sera of Cameroonian origin for the presence of anti-HIV-1 group O antibodies. The reactivity of reference ("gold standard") sera to individual peptides in the PEIA resulted in the selection of five different peptides with sensitivities (sens), specificities (spec), and test efficiencies (TEs) in the range of 90 to 100%. Improvement of the PEIA was obtained with simultaneous reactivity of at least two different peptides in separate wells of an ELISA plate, together with stringent criteria for positivity. We were able to select seven peptide combinations each with a sens, spec, and TE of 96.9, 100, and 99.2%, respectively. None of the 134 European and 4 (1.2%) of the 336 Cameroonian samples sera were group O positive in the optimized HIV-1 group O PEIA ; this was confirmed by the repeated presence of reactives, in agreement with the present knowledge of group O infection distribution. Finally, we were able to develop a strategy with a higher TE (99.2%) than the previously used ANT-70 (98.5%) and ANT-70/MVP5180 (95.7%). Our results show that optimal specificity rather than optimal sensitivity makes the V3 PEIA a sufficiently accurate epidemiological tool to be useful in estimating specifically group O infection among HIV-1-infected patients. (Résumé d'auteur)

HIV-1-infected individuals from which syncytium-inducing (SI) viruses are isolated most often progress more rapidly to AIDS than individuals carrying only non-syncytium-inducing (NSI) viruses. The syncytium-inducing capacity of virus isolates is commonly determined in conjunction to replication in MT-2 cells. Comparison of HIV-1 env sequences and a site-directed mutagenesis study have indicated that the presence of a positively charged amino acid at position 11 or 25 in the V3 loop is minimally required for the SI capacity of HIV-1 subtype B viruses. Studies have also shown a similar correlation between positively charged signature amino acids in the V3 loop and syncytium formation in MT-2 cells for HIV-1 subtypes A, D, and E. In the present study virus phenotype was determined and compared to the V3 loop sequence of seven HIV-1 group O isolates. Three of the HIV-1 group O isolates showed the NSI/non-MT-2 tropic phenotype and two showed the SI/MT-2 tropic phenotype, whereas two isolates presented an uncommon NSI/MT-2 tropic phenotype. The V3 loop of the two SI/MT-2 tropic isolates had a high net positive charge and contained a positively charged amino acid at position 11 or 25. The V3 loop of the two NSI/MT-2 tropic isolates had a low net positive charge and contained a single positively charged amino acid at position 37

Objectives: To examine the genetic variation of HIV-1 isolates in Abidjan, Cote d'Ivoire, and to determine the extent to which phylogenetic trees based on sequence information of part of the env gene containing the principal neutralizing domain are representative for documenting genetic variability. Design: Phylogenetic comparison of 13 HIV-1 strains isolated from patients in Abidjan with previously documented HIV-1 strains of different geographic origin. Methods: To sequence a 900 base-pair fragment of the env gene containing V3, V4, V5 and the beginning of gp41 of three to four clones per isolate. Phylogenetic tree analysis was performed with the software package TREECON. Results: Eleven HIV-1 isolates of Abidjan were classified as genotype A, while two were classed as genotypes B and D. Intra-genotype A distances at the nucleotide level were a maximum of 14.1%. Inter-genotype distances between genotype A and genotypes B, C, and D varied from 16.0 to 22.6%. Phylogenetic trees, based on sequence data of a 300 base-pair fragment containing the V3 loop, showed significant differences in tree topology and statistical confidence with phylogenetic trees based on sequence data of the 900 base-pair env fragment. Conclusions: Genotype A Cote d'Ivoire HIV-1 strains, which comprise 11 out of 13 isolates, predominate in Abidjan, which may indicate a local burst of particular variants. Phylogenetic trees should be interpreted with caution when based on a more limited number of nucleotides, such as the V3 region.