Vitamin D has been suggested to have an effect on erythropoiesis. We sought to evaluate the prevalence of anemia in a population of individuals with vitamin D deficiency compared with those with normal levels in a population of a large integrated healthplan. A cross-sectional analysis in the period 1 January 2004 through 31 December 2006 of subjects with documented concurrent levels of 25-hydroxyvitamin D and hemoglobin were evaluated. Vitamin D deficiency was defined as <30 ng/mL and anemia was defined as a hemoglobin <11 g/dL. A total of 554 subjects were included in the analysis. Anemia was present in 49% of 25-hydroxyvitamin D-deficient subjects compared with 36% with normal 25-hydroxyvitamin D levels (p < 0.01). Odds ratio for anemia in subjects with 25-hydroxyvitamin D deficiency using logistic regressions and controlling for age, gender, and chronic kidney disease was 1.9 (95% CI 1.3-2.7). 25-hydroxyvitamin D-deficient subjects had a lower mean Hb (11.0 vs. 11.7; p = 0.12 ) and a higher prevalence of erythrocyte stimulating agent use (47% vs. 24%; p < 0.05). This study demonstrates an association of vitamin D deficiency and a greater risk of anemia, lower mean hemoglobin, and higher usage of erythrocyte-stimulating agents. Future randomized studies are warranted to examine whether vitamin D directly affects erythropoiesis.