Background: Objective measures of antiretroviral therapy adherence, such as tenofovir (TFV) concentrations, may allow for targeted interventions to prevent future HIV viremia. We evaluated 3 TFV adherence measures and their association with current and 24-week viral load outcomes. Methods: In a South Africa-based trial, we measured urine TFV using a point-of-care (POC) antibody-based assay and 2 metrics assessed through liquid-chromatography-mass-spectrometry: quantitative urine TFV and TFV-diphosphate (TFV-DP) concentrations in dried blood spots (DBS). Logistic regression assessed the association between current and subsequent 24-week viral load outcomes and these 3 measures. We also compared the baseline characteristics of individuals with detectable vs. undetectable POC urine TFV results at enrollment. Results: Of 124 participants, 54.8% female, median age 39 years, 100 (81.5%) had detectable POC urine TFV and 23 (18.5%) did not. Higher TFV-DP concentrations in DBS were negatively associated with viremia at 24 weeks [odds ratio (OR) 0.83, 95% confidence interval (CI): 0.725–0.928, P = 0.003], while a detectable POC urine TFV (OR 0.62, 95% CI: 0.22–1.82, P = 0.380) and quantitative urine TFV (OR 0.98, 95% CI: 0.95–1.00, P = 0.153) showed no significant association. Compared with those with detectable POC urine TFV, those with undetectable POC urine TFV were more likely to be viremic at enrollment (78.3% vs. 25.7%, P < 0.001) and have a CD4 count <200 cells/uL (34.8% vs. 12.9%, P = 0.001). Conclusions: DBS TFV-DP was associated with viremia at 24 weeks and could help predict future viremia. Undetectable POC urine TFV was associated with recent antiretroviral therapy initiation, current viremia, and lower CD4 count. Trial Registration: Pan African Clinical Trials Registry (PACTR202001785886049) and the South African Clinical Trials Registry (DOH-27-072020-6890).