Molecular and Microscopic Detection of Enterocytozoon bieneusi Among People Living With HIV in Northwest Iran: A Cross‐Sectional Study

Microsporidian infections are increasingly recognized in humans and animals, with Enterocytozoon bieneusi having a broad host range and potential for zoonotic transmission. Data on its occurrence and molecular characteristics among immunocompromised populations in Northwest Iran remain limited. We investigated microsporidian infection among 161 people living with HIV (PLWH) in Kurdistan Province, Northwest Iran, using microscopic examination and nested PCR targeting the small subunit ribosomal RNA (SSU rRNA) gene. Microsporidia were detected in 10/161 participants (6.2%) by PCR and 8/161 (4.9%) by microscopy. All PCR‐positive specimens were identified as E . bieneusi , while no Encephalitozoon spp. were detected. In a paired analysis of the same specimens, PCR and microscopy showed 97.5% agreement (Cohen ^′ s kappa = 0.765), and the difference in positivity rates was not statistically significant (McNemar′s exact test, p = 0.625). One PCR‐positive amplicon was successfully sequenced and showed 98%–100% nucleotide identity with sequences reported from human, animal, and environmental sources. Phylogenetic analysis placed this sequence among sequences from different host and environmental sources, indicating genetic similarity but not establishing a zoonotic transmission pathway. No significant associations were observed between infection and the demographic variables examined. The detection of E . bieneusi in this population, together with its genetic similarity to sequences from animal and environmental sources, supports further investigation of potential shared transmission sources. However, the molecular characterization remains preliminary because only a single SSU rRNA sequence was obtained. Human, animal, and environmental sampling combined with more discriminatory genetic markers would help clarify the epidemiology and transmission of E . bieneusi in Northwest Iran.