We investigated whether V3-binding assays might be useful to analyze human immunodeficiency virus type 1 (HIV-1) variants in different geographic regions. We showed that strong cross-reactivity between subtype-specific V3 peptides is almost inevitable in standard indirect enzyme-linked immunosorbent assays (EIA), impairing precise serological subtyping. We therefore developed a subtype-specific EIA (HIV-1 SSEIA) that uses the principle of blocking by an excess of peptide in the liquid phase. Using 231 serum samples collected from HIV-1-infected individuals in 10 different geographical areas from 4 continents, we showed that this approach detected the dominant subtype reactivity in more than 97% of the cases. Internal controls (0 and 100% blocking) were used for every sample such that comparative analysis was possible, independent of both the individual humoral response and the time of collection during the course of infection. This was validated by the excellent concordance of the serological profiles of couples and the temporal stability of the serological profile in individuals. The geographical distribution of the various subtypes in the SSEIA was in agreement with the present knowledge of the distribution of the various genotypes. Although the goal of this study was not an extensive seroepidemiological survey, our results showed that the various profiles in most of the regions were relatively homogeneous, but in central Africa there was a large diversity of serological profiles. Cluster analysis identified a limited number of V3 serogroups of serotypes within the HIV-1 group M. Five serogroups, some of them divided into subgroups, were identified and characterized by a mean serological profile. Our data confirmed that subtypes A and C, although being dissimilar genetic subtypes, present conserved antigenic properties in the V3 region, and that the D subtype is probably the most divergent within the group M (B Korber et al., J Virol 1994;68:6730). Cluster analysis showed a clear correlation between position within the dendrogram and geographical origin of the samples. This is further support for the reliability and thereof the usefulness of the SSEIA. This simple methodology may help facilitate the analysis of the distribution of various HIV-1 subtypes circulating in different populations and regions.
We observed an increasing diversity of HIV-1 strains in the population of blood donors residing in France. This stresses the necessity to broaden the surveillance of HIV-1 diversity in order to improve measures to prevent HIV-1 infections.
The Semliki Forest virus (SFV) system seems to be a useful new approach for generating effective immune responses against HIV-1 in animal models. We evaluated this system by comparing the humoral immune responses raised in mice immunized against the HIV-1 envelope with the SFV system, a DNA vaccine, and a recombinant Env glycoprotein. gp160 ELISA antibody titers (204,800) were highest in the sera from mice immunized with recombinant Semliki Forest virus particles. These sera contained antibodies to the CD4-binding site and recognized linear epitopes on gp120 and gp41 that were also recognized by a pool of sera from HIV1-infected individuals. This demonstrates that the HIV-1 envelope produced in vivo by the SFV system does not fold aberrantly. A low level of neutralizing antibodies against the HIV-1LAI strain was also detected in the serum of one mouse immunized with recombinant SFV particles, suggesting that booster injections should be given to achieve a more effective immune response. SFV recombinant particles induced the strongest humoral responses to the HIV-1 envelope of all the potential HIV env vaccines tested.
HIV-1 V3 serotyping is a classification of immunodeficiency viruses based on antibody binding to V3 peptides that allows obtaining information on circulating subtypes that could be important for population-based epidemiologic studies. Recently, several laboratories have developed V3 enzyme-immunoassays (EIAs) using V3 peptides of subtypes A to E. In the present study, the utility of including additional peptides of subtypes F to H to the EIA was evaluated on a panel of 203 well-characterized serum samples from patients with diverse geographic origins (22 countries) and known HIV-1 genotype (79 A, 61 B, 21 C, 7 D, 7 E, 21 F, 6 G, 1 H). The results indicate a high predictive value (ppv) for serotypes B (> or =0.86), D (1) and E (0.88), and confirm the difficulty of predicting genotype A or C based on serotype A or C. Results also indicate that inclusion of the F peptide in the V3 EIAs may be useful (ppv = 0.61), but introduction of peptides G and H failed to demonstrate significant sensitivity or specificity for these subtypes. Correlation between serotyping and amino-acid sequences of the V3 region from 103 samples allowed the identification of key amino-acids that appear essential for subtype-specific seroreactivity.
Hepatitis C virus (HCV) morphology and physicochemical properties remain unclear because HCV usually circulates in a complexed form in association with immunoglobulins. In the present work, we were interested in the characterization of HCV particles derived from the serum of an anti-HCV negative/HCV RNA positive agammaglobulinemic patient suffering from chronic type C hepatitis. Physicochemical properties of the virus particles were determined by serum centrifugation on a 10-60% isopycnic sucrose density gradient. HCV RNA quantified by bDNA was found in a major peak at density 1.13 g/ml and in a minor peak at densities 1.05-1.07 g/ml. By electron microscopy, 45 nm large core-like particles were found at the 1.13 g/ml density while 60 nm large virus-like particles similar to other members of the Flaviviridae family were visualized at the 1.06-1.07 g/ml densities. This confirms some studies reporting the low density of HCV as compared to other members of the Flaviviridae family.
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