| Summary: | <i>Encephalitozoon intestinalis</i> and <i>Enterocytozoon bieneusi</i> can cause diarrhea in humans, especially severe diarrhea in immunocompromised patients. However, there have been few studies on <i>Enc. intestinalis</i> and <i>Ent. bieneusi</i> in patients with acute diarrhea in the Republic of Korea (ROK). In this study, fecal samples were collected from 1241 patients with acute diarrhea in 2020. Among these, 24 cases of <i>Enc. intestinalis</i> and one case of <i>Ent. bieneusi</i> were detected via PCR amplification of small subunit ribosomal RNA. Genotyping of the internal transcribed spacer region sequence revealed that the detected <i>Ent. bieneusi</i> genotype was in Group 1. This study provides the first evidence that <i>Ent. bieneusi</i> exists in humans in addition to animals in the ROK. To identify the causative agent, continuous monitoring of <i>Enc. intestinalis</i> and <i>Ent. bieneusi</i> is necessary for patients with acute diarrhea in the ROK.
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