HIV DNA Testing Showed Perfect Accuracy in First-Time Testers

Key Takeaways

• Among adults receiving their first HIV test at Beijing Youan Hospital, quantitative HIV DNA correctly classified those diagnosed as HIV-positive or HIV-negative.
• Qualitative HIV RNA produced more false positives and had lower specificity than the other evaluated assays.
• An ELISA–DNA sequence performed well in simulation but was not prospectively evaluated as a testing pathway.
• Both quantitative nucleic acid assays correctly classified every WB-indeterminate specimen in a limited subgroup.

An indeterminate Western blot (WB) result or undisclosed antiviral exposure can complicate initial human immunodeficiency virus (HIV) testing. Antigen/antibody enzyme-linked immunosorbent assay (ELISA) and qualitative HIV ribonucleic acid (RNA) results may require further interpretation in these circumstances. Quantitative HIV deoxyribonucleic acid (DNA) and RNA assays were compared with those tests against a clinical diagnosis.

Investigators prospectively enrolled 215 adults receiving their first HIV test at Beijing Youan Hospital, China, from June 2024 through May 2025 in Yuan and colleagues' HIV diagnostic accuracy study in the British Journal of Biomedical Science. The single-center, cross-sectional comparison required consent and self-reported no previous antiretroviral therapy (ART) or pre- or postexposure prophylaxis. Each participant underwent lymphocyte-enriched HIV-1 DNA quantification, plasma HIV-1 RNA quantification, plasma qualitative HIV-1 RNA testing, and antigen/antibody ELISA. The reference diagnosis integrated laboratory findings, clinical manifestations, and epidemiologic exposure history; participants classified as negative had at least three months of follow-up. Serial testing strategies were calculated from the cohort's concurrent assay results rather than prospectively deployed.

Quantitative DNA had 100% sensitivity (102/102) and 100% specificity (113/113) against the clinical diagnosis. Qualitative RNA had 99% sensitivity (101/102) and 75% specificity (85/113), with substantially more false positives than the other assays. Quantitative and qualitative RNA missed the same HIV-positive participant, who later disclosed a month of ART before sampling; DNA was positive in that case. This single observation does not establish DNA accuracy during antiviral treatment.

Quantitative RNA had full observed specificity but missed an infection. ELISA missed a different infection and returned false positives, although its specificity remained higher than that of qualitative RNA. A separately simulated ELISA-followed-by-DNA sequence had 99% net sensitivity and 100% net specificity. Among 18 WB-indeterminate specimens, both quantitative assays classified all specimens correctly; qualitative RNA was less accurate, and ELISA was least accurate. Detection was broadly consistent across CD4-defined groups, although some positive participants lacked CD4 data. Adjusted comparisons of receiver operating characteristic curves favored the quantitative assays and ELISA over qualitative RNA.

Perfect observed DNA classification came from one male-predominant Beijing cohort, not from diverse populations. The composite reference diagnosis incorporated laboratory findings, and its independence from the evaluated assays was not established. The serial sequence was calculated within the cohort rather than validated as a deployed pathway. Detection timeliness, operational complexity, and formal cost-effectiveness were not evaluated.

According to the investigators, quantitative DNA had the highest observed single-assay accuracy, while ELISA followed by DNA performed well in simulation. Prospective evaluation would be needed before treating that sequence as a validated testing pathway.

Clinician Questions

Were people taking antiretroviral therapy or PrEP included in the HIV DNA diagnostic accuracy comparison?

Known antiviral recipients and people with known acute-phase infection were excluded, although eligibility relied on self-reported treatment and prophylaxis history. A participant's later disclosure of ART exposure does not establish DNA test accuracy in people receiving antiviral treatment; the findings also do not establish accuracy in acute infection.

What counted as a positive qualitative HIV RNA result, and why were there false positives?

The qualitative HIV-1 RNA assay counted an individual sample as positive at a cycle-threshold value of 37 or less. Investigators noted that the kit was designed for pooled blood-bank screening and proposed that its use for individual testing contributed to false positives; they did not experimentally establish that explanation.