Characterization of HIV-1 RNA forms in the plasma of patients undergoing successful HAART.

An assay to characterize plasma human immunodeficiency virus 1 (HIV-1) sequences for patients with low viral loads was developed by combining the selective binding of anti-CD44 MicroBeads with a nested RT-PCR targeting the env C2V4 region. Sequences were obtained from 10 of 20 HIV+ patients who had viral loads below 48 copies/ml. Sequences derived from plasma were compared to those from CD14+ CD16 +monocytes and CD4+ T cells. The plasma sequences were most closely related to those amplified from monocytes, suggesting that during successful antiretroviral therapy, the predominant plasma virus originates from myeloid cells. By characterizing HIV-1 RNA sequences from 8 ml of plasma while avoiding multiple steps, which can lead to contamination and deterioration, this method can help elucidate the viral forms in patients with therapeutically suppressed HIV-1. Understanding the source of residual viremia is crucial in developing approaches for viral eradication.

Meniscus allograft survival in patients with moderate to severe unicompartmental arthritis: a 2- to 7-year follow-up.

PURPOSE: We present meniscus allograft survival data at least 2 years from surgery for 45 patients (47 allografts) with significant arthrosis to determine if the meniscus can survive in an arthritic joint. TYPE OF STUDY: Prospective, longitudinal survival study. METHODS: Data were collected for 31 men and 14 women, mean age 48 years (range, 14 to 69 years), with preoperative evidence of significant arthrosis and an Outerbridge classification greater than II. Failure is established by previous studies as allograft removal. No patient was lost to follow-up. RESULTS: The success rate was 42 of 47 allografts (89.4%) with a mean failure time of 4.4 years as assessed by Kaplan-Meier survival analysis. Statistical power is greater than 0.9, with alpha = 0.05 and N = 47. There was significant mean improvement in preoperative versus postoperative self-reported measures of pain, activity, and functioning, with P = .001, P = .004, and P = .001, respectively, as assessed by a Wilcoxon rank-sum test with P = .05. CONCLUSIONS: Meniscus allografts can survive in a joint with arthrosis, challenging the contraindications of age and arthrosis severity. These results compare favorably with those in previous reports of meniscus allograft survival in patients without arthrosis. LEVEL OF EVIDENCE: Level IV.

Changes in the human immunodeficiency virus V3 region that correspond with disease progression: a meta-analysis.

In order to determine the changes in the human immunodeficiency virus type-1 (HIV-1) envelope that corresponds with disease progression, a meta-analysis of viral forms was performed using HIV-1 sequences obtained from GenBank. Studies were selected that included longitudinally derived V3 envelope region sequences from multiple time points along with CD4 values as a marker of disease progression. Studies with a total of 58 subjects, 327 time points, and 380,000 total amino acid residues were included in this meta-analysis. Changes at specific amino acid sites over the course of disease progression stages were analyzed. The most common specific changes were found at amino acid sites 324D to N, 306S/G to R, and 360N to R. Other sites had changes from one amino acid type to another including the appearance of a basic form at 327, a charged form at 319, and 320D/E changing to basic or neutral. The timing of these changes was contrasted to CD4 decline with changes at 324 and 327 appearing before and 306, 320, and 319 appearing after the initiation of CD4 decline.

Genetic characterization of human immunodeficiency virus type 1 Tat before and after highly active antiretroviral therapy.

Two HIV-infected individuals were followed for changes in the first exon of Tat. Plasma virus collected at 2 months after commencement of highly active antiretroviral therapy (HAART) was compared with the virus collected before initiation of HAART. This short-term therapy significantly reduced the plasma viral burden and also helped modest CD4 recovery in the blood. One of the two individuals (ACG) showed only one form of the Tat before commencement of HAART whereas five different forms (including a form identical to the pre-HAART sample) were found in the post-HAART sample. Some of the post-HAART clones showed a difference of 12.8% suggesting that the source of virus replication might be a reservoir. In the other patients multiple variants of the virus were found within each time point and also between two time points. This patient also showed a debilitating mutation (25-C/R) in three clones suggesting that some of the post-HAART viral forms were not viable in this patient. The dS/dN ratios in the patients were >2 suggesting lack of positive selection.

Influence of CD4+ T cell counts on viral evolution in HIV-infected individuals undergoing suppressive HAART.

We analyzed the viral C2-V4 envelope diversity, glycosylation patterns, and dS/dN ratios of plasma HIV-1 in an attempt to better understand the complex interaction between viral quasispecies and the host-selective pressures pre- and post-HAART. Phylogenetic analysis of the envelope gene of five patients revealed monophyletic clustering in patients with higher CD4+ T cell counts and sequence intermingling in those with lower CD4+ T cells in relation to the stage of HAART. Our analyses also showed clear shifts in N-linked glycosylation patterns in patients with higher CD4+ T cells, suggesting possible distinct immunological pressures pre- and post-HAART. The relative preponderance of synonymous/nonsynonymous changes in the envelope region suggested a positive selection in patients with higher CD4+ T cells, whereas lack of evidence for positive selection was found in the patients with lower CD4+ T cells. An exception to the last analysis occurred in the only patient who reached complete viral suppression, maybe due to drug pressure exerted over the pol gene that may obscure the immune pressure/selection at the envelope in this analysis. All these indications may suggest that even when HAART generates viral suppression, quasispecies evolve in the envelope gene probably resulting from host-selective pressure.