Background: The combination of automated sample preparation and real-time RT-PCR for measurement of HIV-1 viral load has the potential to significantly enhance throughput, reduce operator-associated error, and improve assay sensitivity and dynamic range. Due to the high rate of evolution and ever-changing distribution of HIV-1, reliable patient monitoring requires that viral load assays detect and accurately quantify genetically diverse strains of HIV-1. The aim of this study was to evaluate performance of the Abbott RealTime^TM HIV-1 assay with the m2000sp automated sample preparation instrument (Abbott Molecular, Des Plaines, IL; not available in the U.S.) on a panel of genetically diverse specimens.

Methods: The specimen panel consisted of 91 HIV-1 seropositive plasmas collected from blood donors in Brazil. Subtype was determined by sequence/phylogenetic analysis of three independent genomic regions: gag p24, pol integrase, and env gp41. Specimens for which subtype assignments differed between the regions analyzed were categorized as mosaics. Viral loads were measured in the Abbott RealTime HIV-1 assay (m2000sp sample preparation and m2000rt amplification and detection instruments) and LCx^® HIV RNA Quantitative assay (LCx HIV; Abbott Laboratories, Abbott Park, IL; not available in the U.S.).

Results: Group/subtype was determined for 89 of 91 specimens. The panel included 69 subtype B, 1 C, 2 F, and 17 mosaic strains. Eighty-nine specimens were quantified by the RealTime HIV-1 assay, 87 by the LCx HIV assay. The observed correlation for 86 specimens within the dynamic range of both assays was 0.908 with a slope of 0.843 and intercept of 0.823. Two specimens below the LLD of the LCx HIV assay (50 copies/ml) were quantified in the RealTime HIV-1 assay. Notably, two specimens below the LLD of both assays were also PCR-negative for all three regions used for subtype analysis. Good agreement was observed between assays with 91% of values within 0.5 log₁₀ copies/ml.

Conclusions: In the present study, the automated RealTime HIV-1 assay and sample preparation system accurately quantified this genetically diverse panel. Viral load determinations were highly correlated with the LCx HIV assay. The automated RealTime HIV-1 assay offers the advantages of increased throughput and reduced labor while providing reliable quantification of diverse HIV strains.