Background: Cryptosporidium is one of the most important causes of gastroenteritis in humans. Objectives: This study aimed to investigate the distribution of Cryptosporidium species in children with acute diarrheal disease in Zahedan, Iran. Methods: Stool specimens collected from 764 children aged < 10 years with diarrheal disease admitted to Aliasghar Pediatric Hospital and the Reference Laboratory in Zahedan were assessed for the presence of Cryptosporidium oocysts using the conventional microscopic examination and acid-fast staining. To characterize the oocysts at the molecular level, the 18S rRNA was nested-PCR amplified and sequenced. PCR products were also digested using Vsp1 restriction enzyme for genotype differentiation. Results: Of the 764 stool specimens, 7 (0.91%) were positive for Cryptosporidium oocysts using the microscopic examination. Restriction endonuclease digestion pattern successfully revealed the presence of Cryptosporidium parvum in two microscopic positive samples that were confirmed by DNA sequencing. Conclusions: Cryptosporidium parvum is responsible for cryptosporidiosis in children under 10 years of age in the study region. Although C. parvum may be an important pathogen associated with diarrhea, it is the cause of only a small proportion of diarrheal episodes.