Molecular epidemiology and genotype diversity of Dientamoeba fragilis in a Czech hospital-based cohort: higher prevalence in paediatric patients and evidence of mixed G1/G2 infection in a single patient

Aim This study examined the prevalence, epidemiological characteristics, and genotype diversity of Dientamoeba fragilis in a Czech hospital-based cohort. Methods Patients undergoing routine parasitological diagnostics at the Regional Hospital Liberec (Liberec) between 2022 and 2023 formed the primary epidemiological cohort. Additional PCR-positive samples from the University Hospital Bulovka (Prague) collected between 2022 and 2024 were included for molecular characterization. Detection was performed using multiplex real-time PCR (Allplex, Seegene) and routine microscopy. Genotypes were identified by Sanger sequencing of a 674 bp fragment of the SSU rRNA gene, with cloning used to resolve the presence of multiple genotypes within individual samples. Results Dientamoeba fragilis was detected by PCR in 25.3% of individuals in the epidemiological cohort. Positivity was significantly associated with male sex and younger age, with the highest prevalence observed among children aged 5–9 years. Co-detection with other intestinal parasites was common, most frequently with Blastocystis spp. Abdominal pain was initially associated with D. fragilis detection, but this relationship weakened after adjustment for Blastocystis spp. co-detection. Molecular characterisation of 87 isolates revealed a strong predominance of genotype G1. In one patient, cloning confirmed the simultaneous presence of genotypes G1 and G2 within the same host. Conclusion Dientamoeba fragilis was commonly detected in this Central European clinical population, particularly among school-aged children. Molecular analysis demonstrated intra-host coexistence of genotypes G1 and G2 in a single patient, documenting mixed genotype infection and genetic diversity in human D. fragilis isolates.