Background: Acute diarrhea is the most common gastrointestinal disease among children. Identification of factors involved in the pathogenesis of acute bacterial diarrhea is highly critical. Objectives: This study was conducted to investigate the correlation between serum 25-hydroxyvitamin D (25 (OH) Vit. D) and acute bacterial diarrhea in children. Methods: The present study compared 60 children with acute bacterial diarrhea (the case group) and 60 healthy children (the control group) for serum 25 (OH) Vit. D concentrations. The children age ranged from two months to 12 years. The serum 25hydroxyvitamin D level was measured using ELISA method. Results: Among 60 children with acute bacterial diarrhea, 38 (63.3%) were males and 22 (36.7%) females. In the control group, 31 (51.6%) were males and 29 (48.4%) females (P = 0.26). The median (IQR) of age in case and control groups were 16 (29) and 16 (24) months, respectively (P = 0.75). The mean ± SD of serum 25 (OH) Vit. D levels in case and control groups were 19.3 ± 7.8 and 22.4 ± 7.3 ng/mL, respectively. There was a significant difference between groups regarding the serum 25 (OH) Vit. D concentrations (P = 0.02). Conclusions: This study revealed a significant correlation between serum 25 (OH) Vit. D level and acute bacterial diarrhea. It is likely that vitamin D plays a role in the pathogenesis of diarrhea.