Background: Tenofovir disoproxil fumarate (TDF) increases serum parathyroid hormone (PTH) and 1,25 dihydroxy vitamin D (1,25-(OH) 2 D), and decreases bone mineral density (BMD). Optimal treatment of TDF-associated BMD loss requires an understanding of the primary cause of these abnormalities. Methods: Secondary review of data from two studies of TDF use in youth, comparing the relationship of PTH, 25-hydroxy vitamin D (25-OHD) and 1,25-(OH) 2 D in three groups with varying exposures to TDF: youth without HIV enrolled in a trial of TDF/emtricitabine (FTC) for HIV preexposure prophylaxis (PrEP) at baseline (no TDF exposure) and after 12 weeks of TDF (short-term TDF exposure); and youth with HIV treated with TDF-containing combination antiretroviral therapy (cART) for at least 6 months at study entry (long-term TDF exposure). Relationships were evaluated by correlation analyses. Results: Participants ranged in age from 17 to 24 years and >50% were Black/African American. In persons not treated with TDF, PTH had the physiologically appropriate negative correlation with 25-OHD (r=-0.3504, P=0.004). Correlations between PTH and 25-OHD in groups treated with TDF were weaker or absent. With longer term TDF treatment in persons with HIV, 25-OHD and 1,25-(OH) 2 D had the positive correlation similar to that found in vitamin D deficiency. Conclusions: TDF changes the relationship of 25-OHD to PTH, suggesting that in persons using TDF for PrEP or cART, a higher than usual target for serum 25-OHD concentration might be needed to reduce PTH and optimize bone health.