Antibody maturation and viral diversification in HIV-infected women.

INTRODUCTION: The Post-exposure Prophylaxis in Infants (PEPI)-Malawi trial evaluated infant antiretroviral regimens for prevention of post-natal HIV transmission. A multi-assay algorithm (MAA) that includes the BED capture immunoassay, an avidity assay, CD4 cell count, and viral load was used to identify women who were vs. were not recently infected at the time of enrollment (MAA recent, N = 73; MAA non-recent, N = 2,488); a subset of the women in the MAA non-recent group known to have been HIV infected for at least 2 years before enrollment (known non-recent, N = 54). Antibody maturation and viral diversification were examined in these women. METHODS: Samples collected at enrollment (N = 2,561) and 12-24 months later (N = 1,306) were available for serologic analysis using the BED and avidity assays. A subset of those samples was used for analysis of viral diversity, which was performed using a high resolution melting (HRM) diversity assay. Viral diversity analysis was performed using all available samples from women in the MAA recent group (61 enrollment samples, 38 follow-up samples) and the known non-recent group (43 enrollment samples, 22 follow-up samples). Diversity data from PEPI-Malawi were also compared to similar data from 169 adults in the United States (US) with known recent infection (N = 102) and known non-recent infection (N = 67). RESULTS: In PEPI-Malawi, results from the BED and avidity assays increased over time in the MAA recent group, but did not change significantly in the MAA non-recent group. At enrollment, HIV diversity was lower in the MAA recent group than in the known non-recent group. HRM diversity assay results from women in PEPI-Malawi were similar to those from adults in the US with known duration of HIV infection. CONCLUSIONS: Antibody maturation and HIV diversification patterns in African women provide additional support for use of the MAA to identify populations with recent HIV infection.

Use of a high resolution melting assay to analyze HIV diversity in HIV-infected Ugandan children.

BACKGROUND: We used a novel high resolution melting (HRM) diversity assay to analyze HIV diversity in Ugandan children (age 0.6-12.4 years) who were enrolled in an observational study of antiretroviral treatment (ART). Children were maintained on ART if they were clinically and immunologically stable. METHODS: HIV diversity was measured before ART (baseline) in 76 children and after 48 or 96 weeks of ART in 14 children who were not virally suppressed. HIV diversity (expressed as HRM scores) was measured in 6 regions of the HIV genome (2 in gag, 1 in pol, 3 in env). RESULTS: Higher baseline HRM scores were significantly associated with older age (≥2 years, P ≤ 0.001 for all 6 regions). HRM scores from different regions were weakly correlated. Higher baseline HRM scores in 3 regions (1 in gag, 2 in env) were associated with ART failure. HIV diversity was lower in 4 regions (2 in gag, 1 in pol, 1 in env) after 48-96 weeks of nonsuppressive ART compared with baseline. CONCLUSIONS: Higher levels of HIV diversity were observed in older children before ART, and higher levels of diversity in some regions of the HIV genome were associated with ART failure. Prolonged exposure to nonsuppressive ART was associated with a significant decrease in viral diversity in selected regions of the HIV genome.

Passive (amyloid-ß) immunotherapy attenuates monoaminergic axonal degeneration in the AßPPswe/PS1dE9 mice.

The role of amyloid-ß (Aß in the neurodegeneration of Alzheimer's disease remains controversial, to a large extent because of the lack of robust neurodegeneration in mouse models of AD. To address this question, we examined the effects of Aß antibodies in the recently described monoaminergic (MAergic) axonal degeneration in AßPPswe/PS1dE9 mice. To determine if Aß accumulation is directly involved in degeneration of MAergic axons, we examined the effects of passive anti-Aß antibody (7B6) administration on Aß pathology and MAergic degeneration in AßPPswe/PS1dE9 mice. Injections of monoclonal antibody (mAb) 7B6 into mice (6 to 9 months of age) resulted in a modest reduction of Aß load in the brains of AßPPswe/PS1dE9 mice. In addition, 7B6 treated AßPPswe/PS1dE9 mice had significantly higher densities of MAergic axons in both cortex and in hippocampus as compared to untreated mutant mice. For example, 7B6 treated mice showed almost 2-fold greater densities of serotonergic (5-HT) axons in the cortex compared to saline treated mice. Similar findings were observed in the catecholaminergic (TH) axons. Our results demonstrate that lowering of Aß levels via passive Aß immunotherapy ameliorates ongoing degenerative processes, supporting a causal link between Aß and neurodegeneration.

Association of HIV diversity and survival in HIV-infected Ugandan infants.

BACKGROUND: The level of viral diversity in an HIV-infected individual can change during the course of HIV infection, reflecting mutagenesis during viral replication and selection of viral variants by immune and other selective pressures. Differences in the level of viral diversity in HIV-infected infants may reflect differences in viral dynamics, immune responses, or other factors that may also influence HIV disease progression. We used a novel high resolution melting (HRM) assay to measure HIV diversity in Ugandan infants and examined the relationship between diversity and survival through 5 years of age. METHODS: Plasma samples were obtained from 31 HIV-infected infants (HIVNET 012 trial). The HRM assay was used to measure diversity in two regions in the gag gene (Gag1 and Gag2) and one region in the pol gene (Pol). RESULTS: HRM scores in all three regions increased with age from 6-8 weeks to 12-18 months (for Gag1: P = 0.005; for Gag2: P = 0.006; for Pol: P = 0.016). Higher HRM scores at 6-8 weeks of age (scores above the 75(th) percentile) were associated with an increased risk of death by 5 years of age (for Pol: P = 0.005; for Gag1/Gag2 (mean of two scores): P = 0.003; for Gag1/Gag2/Pol (mean of three scores): P = 0.002). We did not find an association between HRM scores and other clinical and laboratory variables. CONCLUSIONS: Genetic diversity in HIV gag and pol measured using the HRM assay was typically low near birth and increased over time. Higher HIV diversity in these regions at 6-8 weeks of age was associated with a significantly increased risk of death by 5 years of age.

Association of recent HIV infection and in-utero HIV-1 transmission.

OBJECTIVE: We previously developed a multiassay algorithm (MAA) to identify recent HIV infection that includes the BED-capture enzyme immunoassay, an avidity assay based on the Genetic Systems HIV-1/HIV-2 + O enzyme immunoassay, CD4 cell count, and HIV viral load. We used this MAA to evaluate the association between recent maternal HIV infection and in-utero transmission of HIV. METHODS: Plasma samples were collected at delivery from 2561 HIV-infected women in the postexposure prophylaxis of infants-Malawi trial. The MAA described above was used to identify women with recent HIV infection. Logistic regression models assessed association between recent HIV infection and in-utero HIV transmission (defined as a positive infant HIV DNA test at birth). RESULTS: Seventy-three women were identified as recently infected using the MAA. Those women were younger and had lower parity than women who were identified as not recently infected using the MAA (P < 0.0001 for age and parity). The frequency of in-utero HIV transmission was 17.8% among women identified as recently infected, compared with 6.7% among women identified as not recently infected (13/73 vs. 166/2488, P = 0.001). In a multivariate model, three factors were independently associated with in-utero HIV transmission: recent infection [adjusted odds ratio (AOR): 2.49, 95% confidence interval (CI): 1.30-4.78, P = 0.006], log(10) HIV viral load at delivery (AOR: 2.01, 95% CI: 1.60-2.51, P < 0.0001), and younger age (per 10 year increase, AOR: 0.66, 95% CI: 0.43-0.93, P = 0.02). CONCLUSION: Results obtained using a MAA suggest that recent maternal HIV acquisition is strongly associated with in-utero HIV transmission, independent of HIV viral load at delivery.

Analysis of HIV diversity using a high-resolution melting assay.

HIV viruses are usually genetically homogeneous shortly after infection, and become more heterogeneous over time. We developed a high-resolution melting (HRM) assay to analyze HIV diversity without sequencing. Plasma samples from the HIVNET 012 trial were obtained from nine Ugandan mother-infant pairs. DNA amplified from the HIV gag region was analyzed to determine the number of degrees over which the DNA melted (HRM score). HRM gag DNA was also cloned and sequenced (50 clones/mother; 20 clones/infant). The median HRM score for infants (4.3, range 4.2-5.3) was higher than that for control plasmids (3.4, range 3.2-3.8, p < 0.001) and lower than that for mothers (5.7, range 4.4-7.7, p = 0.005, exact Wilcoxon rank sum test). The intraclass correlation coefficient reflecting assay reproducibility was 94% (95% CI: 89-98%). HRM scores were also compared to sequenced-based measures of HIV diversity; higher HRM scores were associated with higher genetic diversity (p < 0.001), complexity (p = 0.009), and Shannon entropy (p = 0.022), but not with length variation (p = 0.111). The HRM assay provides a novel, rapid method for assessing HIV diversity without sequencing. This assay could be applied to any region of the HIV genome or to other genetic systems that exhibit DNA diversity.