Procedures

Testing followed the national algorithm¹⁴: first, BBT by Determine (Alere Determine HIV-1/2 Ag/Ab Combo; Abbott, Chicago, IL) (fourth generation), followed by First Response (First Response HIV-1-2; Premier Medical Corporation Ltd, Kachigam, India) (third generation), if Determine was reactive. In the case of 2 reactive BBTs, the same 2 BBTs were performed by a different provider to confirm a positive diagnosis. In the case of discordant BBTs, both tests were repeated. If discordance persisted, a third test, CHEMBIO was performed (CHEMBIO HIV 1/2 STAT-PAK Assay; CHEMBIO Diagnostic Systems, Inc., New York, NY). If this third test was positive, the result was reported as inconclusive and a retest conducted in 14 days. OMT was conducted by clinic staff blinded to BBT results.

The national algorithm mirrored that in Zimbabwe with the following exceptions: the Determine HIV test was third instead of fourth generation and DNA PCR from dry blood spot specimens was the third test and was considered conclusive.^14–16 In addition BBT was conducted by research staff for those enrolled in the FIT trial and non-research staff for those enrolled from routine testing points. Research staff performed OMT and were not blinded to BBT results.

The reference standard used for our study was the HIV status as per the national algorithim of each country.

In Zimbabwe and Kenya, OMT sample collection and processing was performed bedside by qualified HIV testing lay providers who are typically lower than nurse level providers and are responsible for HIV testing in both countries. The qualification for these providers is a standard national training for HIV services conducted over 2 weeks. Testing was conducted according to manufacturer details (OraQuick ADVANCE Rapid HIV-1/2 Antibody Test; OraSure Technologies, Inc., Bethlehem, PA), whereby the research staff collected an oral fluid sample from the participants by running the test device between the lips and outer gums of the client once on top and once at the bottom and then place the test device pad directly into the reaction fluid immediately after collection.¹⁷ OMT results were read once between 20 and 40 minutes in Zimbabwe, and twice in Kenya at both 20 and 40 minutes to assess test performance at the lower and upper recommended times. OMT results were not shared with caregivers, because the test was undergoing validation.

Data were analyzed using STATA 14 (StataCorp, College Station, TX). Sensitivity was calculated by dividing the number of OMT and BBT-positive children by the number of BBT-positive children. Specificity was calculated by dividing the number of OMT and BBT-negative children by the number of BBT-negative children. Positive predictive value (PPV) and negative predictive value (NPV) were calculated in the Zimbabwean cohort by dividing the number with both positive OMT and BBT by all the positive OMT tests (PPV) and by dividing the number with both negative OMT and BBT results by the total negative by OMT (NPV). PPV and NPV were not calculated in the Kenyan cohort, because positive children were oversampled. Ninety-five percent (95%) or 97.5% (when the estimate was 100%) confidence intervals (CIs) were calculated using a binomial distribution. Stability of the test results using result interpretation pictures from the manufacturer was described in Kenya.

Adolescents ≥16 gave independent written informed consent without parental/guardian consent. Parents/guardians of children aged 18 months–15 years provided written consent; adolescents 13–15 years signed a paragraph within the parental consent form to give their assent, whereas children 7–12 years signed a separate assent document, which was optional in Kenya. B-GAP received approval from the Biomedical Research and Training Institute, the Medical Research Council of Zimbabwe and institutional review boards at Duke University and the London School of Hygiene and Tropical Medicine. The Kenyan study received approval from the Kenyatta National Hospital Ethics and Research Committee and the University of Washington Institutional Review Board.