The results of this study suggest that different gene amplification methods can be used to arrive at comparable determinations of HIV-1 RNA copy number in the female genital tract. In previous studies of blood plasma (3
), we observed differences in estimated RNA copy number between the OTC NASBA-QT assay and the Roche AMPLICOR HIV-1 MONITOR assay; these differences could be eliminated by adjusting to a common set of external standards. In the current study, we confirmed our previous findings concerning the NASBA-QT and AMPLICOR MONITOR assays but found no significant differences between HIV-1 RNA copy number estimates obtained with the NucliSens and AMPLICOR MONITOR assays.
Importantly, there were no significant differences among the assays in terms of HIV-1 RNA quantification for the female genital tract. Thus, the use of a common set of external standards would not appear to be necessary when making comparisons between assays using genital tract specimens. The absence of a difference between assays using peripheral blood specimens and those using genital tract specimens could not be explained by a decrease in the dynamic range. The adjusted and unadjusted dynamic ranges for peripheral blood and genital tract HIV-1 RNA were equivalent. Similarly, although the assays use different processing technologies, each uses the same processing technology for peripheral blood and genital tract specimens.
In contrast to previous findings (3
), however, this study found a higher SD for the OTC assays than for the Roche assay. Thus, the lack of differences between the estimated RNA copy numbers obtained with these assays may be confounded by the higher assay SD. Nonetheless, as previously reported (3
), we observed similar differences when the NASBA-QT and AMPLICOR MONITOR assays were used to test peripheral blood, in spite of the higher SD.
The effect of VQA standard adjustment was greatest for the NASBA-QT assay and was more pronounced at higher and lower HIV-1 RNA concentrations. For both peripheral blood and genital tract specimens and for both amplification technologies, the copy number estimates obtained with the VQA standards were higher than the kit-based estimates at the high end and lower than the kit-based estimates at the low end.
An issue of concern is the high level of discordance in HIV-1 detection between the OTC and Roche gene amplification systems. This discordance was not confined to either compartment or to any group of women. Many of these differences were not at the level of the limit of detection for the assays. Sequence variation among the highly discordant isolates may have resulted in differences in primer pair efficiencies, which could explain the discordance.
In summary, the OTC NucliSens and Roche AMPLICOR HIV-1 MONITOR assays appear to be equivalent for determining HIV-1 RNA levels in either the female genital tract or peripheral blood, although the SD for the NucliSens assay was high. These equivalent determinations can be made without using an external set of common standards. However, the two amplification technologies do produce some discordant positive and negative results.