HIV populations are large and accumulate high genetic diversity in a nonlinear fashion.

HIV infection is characterized by rapid and error-prone viral replication resulting in genetically diverse virus populations. The rate of accumulation of diversity and the mechanisms involved are under intense study to provide useful information to understand immune evasion and the development of drug resistance. To characterize the development of viral diversity after infection, we carried out an in-depth analysis of single genome sequences of HIV pro-pol to assess diversity and divergence and to estimate replicating population sizes in a group of treatment-naive HIV-infected individuals sampled at single (n = 22) or multiple, longitudinal (n = 11) time points. Analysis of single genome sequences revealed nonlinear accumulation of sequence diversity during the course of infection. Diversity accumulated in recently infected individuals at rates 30-fold higher than in patients with chronic infection. Accumulation of synonymous changes accounted for most of the diversity during chronic infection. Accumulation of diversity resulted in population shifts, but the rates of change were low relative to estimated replication cycle times, consistent with relatively large population sizes. Analysis of changes in allele frequencies revealed effective population sizes that are substantially higher than previous estimates of approximately 1,000 infectious particles/infected individual. Taken together, these observations indicate that HIV populations are large, diverse, and slow to change in chronic infection and that the emergence of new mutations, including drug resistance mutations, is governed by both selection forces and drift.

ART suppresses plasma HIV-1 RNA to a stable set point predicted by pretherapy viremia.

Current antiretroviral therapy is effective in suppressing but not eliminating HIV-1 infection. Understanding the source of viral persistence is essential for developing strategies to eradicate HIV-1 infection. We therefore investigated the level of plasma HIV-1 RNA in patients with viremia suppressed to less than 50-75 copies/ml on standard protease inhibitor- or non-nucleoside reverse transcriptase inhibitor-containing antiretroviral therapy using a new, real-time PCR-based assay for HIV-1 RNA with a limit of detection of one copy of HIV-1 RNA. Single copy assay results revealed that >80% of patients on initial antiretroviral therapy for 60 wk had persistent viremia of one copy/ml or more with an overall median of 3.1 copies/ml. The level of viremia correlated with pretherapy plasma HIV-1 RNA but not with the specific treatment regimen. Longitudinal studies revealed no significant decline in the level of viremia between 60 and 110 wk of suppressive antiretroviral therapy. These data suggest that the persistent viremia on current antiretroviral therapy is derived, at least in part, from long-lived cells that are infected prior to initiation of therapy.

Frequent polymorphism at drug resistance sites in HIV-1 protease and reverse transcriptase.

BACKGROUND: Failure of antiretroviral therapy may result from the selection of pre-existing, drug-resistant HIV-1 variants, but the frequency and type of such variants have not been defined. OBJECTIVE: We used single genome sequencing (SGS) to characterize the frequency of polymorphism at drug resistance sites in protease (PR) and reverse transcriptase (RT) in plasma samples from antiretroviral naive individuals. METHODS: A total of 2229 pro-pol sequences in 79 plasma samples from 30 patients were analyzed by SGS. A mean of 28 single genome sequences was obtained from each sample. The frequency of mutations at all PR and RT sites was compared to those associated with drug resistance. RESULTS: We detected polymorphism at one or more drug resistance sites in 27 of 30 (90%) patients. Polymorphism at positions 179 and 215 of RT was most common, both occurring in 23% of patients. Most (68%) of other drug resistance sites were polymorphic with an average of 3.2% of genomes per sample containing at least one variant from wild type. Seven drug resistance sites were polymorphic in more than 1% of genomes: PR position 33; RT positions 69, 98, 118, 179, 210, and 215. Although frequencies of synonymous polymorphism were similar at resistance and nonresistance sites, nonsynonymous polymorphism were significantly less common at drug resistance sites, implying stronger purifying selection at these positions. CONCLUSIONS: HIV-1 variants that are polymorphic at drug resistance sites pre-exist frequently as minor species in antiretroviral naive individuals. Standard genotype techniques have grossly underestimated their frequency.

Plasma pharmacokinetics of sulfadiazine administered twice daily versus four times daily are similar in human immunodeficiency virus-infected patients.

The pharmacokinetics of 2,000 mg of sulfadiazine administered twice daily (BID) versus those of 1,000 mg administered four times a day were compared in eight human immunodeficiency virus-infected patients. No differences in pharmacokinetic parameters were detected between the regimens. These data provide a pharmacokinetic rationale for BID dosing of sulfadiazine for the treatment and suppression of toxoplasmosis.

Increases in CD4+ T lymphocytes occur without increases in thymic size in HIV-infected subjects receiving interleukin-2 therapy.

OBJECTIVE: To examine the potential contribution of the thymus to CD4+ T-lymphocyte increases in HIV-infected patients receiving intermittent interleukin-2 (IL-2) therapy. DESIGN: Fifteen HIV-infected patients treated with antiretroviral regimens who were enrolled in a study of intermittent IL-2 therapy and were willing to undergo serial thymic computed tomography (CT) were prospectively studied. METHODS: Thymic CT was performed before and approximately 6 and 12-17 months after intermittent IL-2 therapy was started. Scans were graded in a blinded manner. Changes in lymphocyte subpopulations were determined by flow cytometry. RESULTS: Statistically significant increases in CD4+ T lymphocytes occurred with IL-2 administration, with a preferential increase in naive relative to memory CD4+ T cells. Despite this increase in naive CD4+ T cells, overall there was a modest decrease in thymic volume observed during the study period. No correlation was found between changes in thymic volume indices and total, naive, or memory CD4+ T-lymphocyte counts. CONCLUSIONS: These findings demonstrate that the profound CD4+ T-lymphocyte increases seen with intermittent IL-2 administration are not associated with increases in thymic volume and more likely are due to peripheral expansion rather than increased thymic output.

A proof-of-concept study of short-cycle intermittent antiretroviral therapy with a once-daily regimen of didanosine, lamivudine, and efavirenz for the treatment of chronic HIV infection.

BACKGROUND: We previously demonstrated that short-cycle structured intermittent therapy (SIT; 7 days without therapy followed by 7 days with antiretroviral therapy [ART]) with a ritonavir-boosted, indinavir-based, twice-daily regimen maintained suppression of plasma HIV viremia while reducing serum levels of lipids. Adherence to such a regimen may be problematic for certain patients. METHODS: Eight patients with a history of receiving combination ART that maintained suppression of plasma HIV RNA to <50 copies/mL received a once-daily SIT regimen of didanosine, lamivudine, and efavirenz. RESULTS: For 7 patients, suppression of plasma HIV RNA to <50 copies/mL was maintained for 60-84 weeks. Four patients with adequate samples had no evidence for an increase in plasma viremia for up to 72 weeks, by use of an assay with a limit of detection of <1 copy/mL. The lack of rebound viremia may be the result of the persistence of efavirenz in plasma on day 7 of the no-therapy period, as was detected in 7 of 7 patients. There was no significant change in CD4(+) T cell counts or serum hepatic transaminase or lipid levels. CONCLUSION: A once-daily short-cycle SIT regimen maintained suppression of plasma HIV RNA while preserving CD4(+) T cell counts. Such a regimen may have importance in resource-limited settings where the monetary cost of continuous ART is prohibitive.

Long-cycle structured intermittent versus continuous highly active antiretroviral therapy for the treatment of chronic infection with human immunodeficiency virus: effects on drug toxicity and on immunologic and virologic parameters.

We evaluated the effect of long-cycle structured intermittent therapy (SIT; 4 weeks without highly active antiretroviral therapy [HAART] followed by 8 weeks with HAART) versus continuous HAART. The study was prematurely terminated to new enrollment because of the emergence of genetic mutations associated with resistance to antiretroviral drugs in 5 patients. After 48 weeks, there was no significant difference between groups in lipid, hepatic transaminase, and C-reactive protein levels in 41 patients. Although there were no differences in CD4(+) or CD8(+) T cell counts or the percentage of cells that were CD4(+)CD25(+), CD8(+)CD25(+), or CD4(+)DR(+), patients who received SIT had a significantly higher percentage of CD8(+)CD38(+) and CD8(+)DR(+) cells. There was no clear autoimmunization effect by immunologic or virologic parameters. There was no benefit to long-cycle SIT versus continuous HAART with regard to certain toxicity, immunologic, or virologic parameters.