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Background/objectives: The objective of this study was to test the hypothesis that vitamin D deficiency (i.e., serum 25-hydroxyvitamin D (25(OH)D) ≤ 20 ng/mL) associates with the increased occurrence and shortened time to a knee osteoarthritis (OA) diagnosis after anterior cruciate ligament reconstruction (ACLR)., Methods: This study consisted of a retrospective, case-control design. The inclusion criteria consisted of (1) patients (≥18 y) who underwent arthroscopic ACLR with (cases; n = 28) and without (controls; n = 56) a subsequent knee OA diagnosis (≥90 d from the date of ACLR) and (2) with a documented serum 25(OH)D concentration after ACLR (and before a knee OA diagnosis for the cases). Controls were matched (2:1) to cases based on sex, age at ACLR, date of ACLR, and body mass index. After matching, patients were separated into two groups: (1) vitamin D deficient (serum 25(OH)D ≤ 20 ng/mL) or (2) non-vitamin D deficient (serum 25(OH)D > 20 ng/mL). Data were extracted from the medical records., Results: Thirty-one percent ( n = 26) of patients included were vitamin D deficient. Fifty percent ( n = 13) of the vitamin D deficient and twenty-six percent ( n = 15) of the non-vitamin D deficient patients were subsequently diagnosed with knee OA ( p = 0.03). Time from ACLR to a knee OA diagnosis was significantly ( p = 0.02) decreased in the vitamin D deficient (OA-free interval, 95% confidence interval [CI] = 7.9 to 10.9 y) compared to the non-vitamin D deficient group (OA-free interval, 95% CI = 10.5 to 12.5 y)., Conclusions: Vitamin D deficiency after ACLR may serve as a prognostic biomarker for knee OA following ACLR.