Variable relationship between proviral DNA load and infectious virus titre in the peripheral blood mononuclear cells of HIV-1-infected individuals.

OBJECTIVES: To determine the relationship between infectious virus titre and proviral copy number in peripheral blood mononuclear cells (PBMC) of infected subjects and to ascertain which, if either, is most closely related to CD4+ cell loss and disease progression. DESIGN AND METHODS: Cellular HIV-1 viraemia was quantified in 45 infected subjects who had not received antiretroviral therapy using limiting dilution tissue culture infective dose (PBMC TCID) and quantitative polymerase chain reaction (PCR) techniques. RESULTS: Proviral DNA was detected in 44 (98%) and infectious virus in 38 (82%) of the 45 subjects. Viraemia as measured by both culture and PCR was inversely correlated with patient CD4+ cell count and associated with disease status. Measurement using both techniques correlated with each other (Spearman's rank correlation rho = 0.52; P = 0.0006). The ratio of proviral copies to PBMC TCID ranged from 1:1 1000:1. to > 1000:1. CONCLUSIONS: The ratio of provirus:PBMC TCID was highest when the PBMC TCID was low, and approached unity when PBMC TCID was high. This ratio could be influenced by a variety of factors but did not correlate significantly with patient disease status or CD4+ cell count.

Comparison of culture- and non-culture-based methods for quantification of viral load and resistance to antiretroviral drugs in patients given zidovudine monotherapy.

Virological assays for human immunodeficiency virus type 1 load and drug resistance can broadly be divided into culture-based and molecular biology-based methods. Culture-based methods give a direct measure of infectious virus load and phenotypic drug resistance, whereas molecular biology-based methods are indirect, assaying nucleic acid levels to determine virus load and point mutations associated with drug resistance. We have compared culture-based and non-culture-based methods for patients enrolled in a placebo-controlled trial of zidovudine (the Concorde Trial). Virus loads were assayed by culture of peripheral blood mononuclear cells (PBMCs) or quantitative PCR, and drug resistance was assayed in culture or in a quantitative, PCR-based point mutation assay. The rates of detection of viremia and drug resistance were higher by PCR than by culture for this population of subjects. Comparison of the virus loads by the two measures showed a good correlation for virus loads in PBMCs but a poor correlation for virus loads in plasma. The latter result probably reflected the inaccuracies of culture in assaying plasma with the low infectious virus titers seen in the study population. The concordance of phenotypic and genotypic drug resistance methods was high, with all phenotypically resistant isolates having at least one resistance-associated mutation and with no mutations being found in a drug-sensitive isolate. Genomic resistance scores (weighted sums of levels of resistance mutations) showed good correlations with the levels of phenotypic resistance, and both resistance measures were observed to increase as the duration of exposure to drug increased. Overall, non-culture-based methods were shown to correlate well with culture-based methods and offer a low-cost, high-throughput alternative. However, culture-based methods remain the final arbiters of infectious virus load and phenotypic drug resistance and are unlikely to be superseded entirely.