HIV Type 2 Proviral Load Measured by Quantitative Polymerase Chain Reaction Correlates with CD4⁺Lymphopenia in HIV Type 2-Infected Individuals

Abstract

The efficiency of detection of 2 sets of primer pairs from putatively conserved regions of the human immunodeficiency virus type 2 (HIV-2) genome were assessed in 86 seropositive individuals from The Gambia by nested polymerase chain reaction (PCR). HIV-2 long terminal repeat (LTR) target sequences were detected in DNA extracted from peripheral blood mononuclear cells (PBMCs) in 84 of 86 (97%) individuals whereas HIV-2 integrase (pol) gene sequences were detected in 39 of 41 (95%) individuals. The use of LTR target sequences and recombinant Pfu DNA polymerase, rather than Taq polymerase, in a modified secondary amplification reaction mediated the incorporation of 35S-labeled nucleotides in a quantitative radiometric assay. This sensitive assay was used to quantify HIV-2 proviral DNA in clinical samples and compared well with estimations by limiting end-point dilution of target molecules. A linear response between counts and the number of copies amplified from serial dilutions of pROD10 plasmid DNA (3-2000 copies) yielded a standard curve to allow extrapolation to clinical data. Increased levels of HIV-2 proviral DNA, expressed as copies per 10(5) CD4-positive lymphocytes, were associated with declining CD4 count in 63 adult patients (Spearman rank correlation, r = -0.71, n = 63, p < 0.001) and with the occurrence of HIV-related clinical disease. Kruskall-Wallis analysis of variance analysis showed the mean proviral copy number (log10) to be significantly different between groups (p < 0.001) where CD4 counts were grouped as < 200/mm3 (3.4 +/- 1.05 copies), 200-500/mm3 (2.84 +/- 0.93 copies), and > 500/mm3 (1.88 +/- 0.43 copies).(ABSTRACT TRUNCATED AT 250 WORDS)

Polyfunctional T cell responses are a hallmark of HIV‐2 infection

“…Most HIV-2-infected individuals have low to undetectable viral loads [4], yet levels of proviral DNA in peripheral blood CD4 + T cells is equivalent to that found in HIV-1-infected individuals at the same stage of disease [5][6][7]. These observations are consistent with control of HIV-2 viral replication by host immune responses leading to the lack of subsequent CD4 + T cell decline in typical HIV-2 infection [8].…”

New Approach to mRNA Quantification: Additive RT-PCR

“…These fittings showed similar results, but the logistic one was more accurate when applied to a wider range of concentrations (Figure 2). This behavior of TaqDNA polymerase has also been shown by other authors who have studied activity vs. increasing cycles and template concentrations (1).…”

Cell-Associated Viral Burden Provides Evidence of Ongoing Viral Replication in Aviremic HIV-2-Infected Patients

“…Notwithstanding the two-log difference in viremia levels that characterize HIV-2 and HIV-1 infection, similar levels of cellassociated viral DNA have been reported, suggesting a comparable number of infected cells (6,24,45,52). This apparent paradox highlights the potential contribution of quiescent latent virus to proviral load and the relevance of quantifying the ongoing viral replication.…”