A community based cross-sectional study was carried out in rural areas of West Bengal with the aim to assess the prevalence of vitamin A deficiency (VAD) among rural preschool children. Clinical examination was carried out on 9,228 children for the signs and symptoms of VAD and a sub-sample of 590 children were covered for the estimation of blood vitamin A levels using dried blood spot (DBS) method. The prevalence of Bitot’s spots was 0.6% (95% CI=0.44, 0.76), which is more than the public health significance, and it increased with increase in age. The prevalence was significantly higher (P<0.001) among boys (0.8%) as compared to girls (0.4%). The proportion of children with subclinical vitamin A deficiency (blood vitamin A < 20μg/dL) was 61% (95% CI: 52.3–65.1), and it was significantly (P<0.01) higher among the children of lower socioeconomic communities.