The active form of vitamin D (VD) exerts hormonal effects by regulating the expression of genes involved in T‐cell activity, cell differentiation, and proliferation. Human T‐cell leukemia virus type 1 (HTLV‐1) is a causative agent of life‐threatening diseases, adult T‐cell leukemia (ATL) and HTLV‐1‐associated myelopathy (HAM). Among ATL patients, hypercalcemia is one of the most serious complications due to bone resorption. In this study, wild‐type mice administered UV‐irradiated HTLV‐1‐infected cells showed up to 47% decrease of plasma VD level compared with untreated mice. To clarify the effect of HTLV‐1 on plasma VD level, 315 samples registered in nationwide cohort study on ATL onset were measured. The VD level in HAM (14.98 ± 8.5 ng/mL) was significantly lower than those in asymptomatic carriers and ATL (p < 0.05). Upon comparing the VD levels in ATL stratified by disease subtypes, acute ATL showed a lower level (15.81 ± 12.0 ng/mL) than chronic and smoldering types (p < 0.05). In the longitudinal observation, VD levels were significantly higher in untreated spontaneous remission cases than in ATL progression cases, in which the VD levels decreased approximately 40% after onset. In cases of relapse after transplantation, the plasma VD level dropped to 38.7% of the pre‐relapse level, while in cases of complete remission, the VD level increased with improvement of the performance status. Taken together, these results suggest that plasma VD level is a potential indicator for the onset and relapse of HTLV‐1‐associated diseases.