Cryptosporidium spp. Enterocytozoon bieneusi and Encephalitozoon intestinalis are opportunistic pathogens responsible for gastrointestinal diseases. We evaluated the ParaGENIE Crypto-Micro Real-Time PCR kit (Ademtech, France), the first CE-IVD compliant PCR assay available for these pathogens. This study was conducted blindly against a reference panel of 115 stool specimens including positive samples for Cryptosporidium spp. (n = 48) and E. bieneusi (n = 38) as well as negative or positive samples for other parasites to test for cross-reactivity. An additional set of samples corresponding to 8 rare Cryptosporidium species was also included. Discrepancies were evaluated with external in-house PCR tests. The ParaGENIE Crypto-Micro PCR assay displayed a sensitivity/specificity of 91.7%/100% and 97.3%/98.7% for Cryptosporidium spp. and E. bieneusi, respectively, and was able to detect all 12 Cryptosporidium species of the reference panel, including rare species. This new CE-IVD assay will facilitate the diagnosis of intestinal cryptosporidiosis and microsporidiosis, a major concern in immunocompromised patients and travelers.