Comparison of microscopy, two xenic culture techniques, conventional and real-time PCR for the detection of Dientamoeba fragilis in clinical stool samples

Dientamoeba fragilis is a pathogenic protozoan parasite that is notoriously difficult to diagnose. The aim of this study was to determine the gold standard for laboratory detection of D. fragilis. A total of 650 human faecal samples were included in the study. All specimens underwent the following: microscopy using a permanent stain (modified iron-haematoxylin), culture using a modified Boeck and Drbohlav's medium (MBD) and TYGM-9, a conventional polymerase chain reaction (PCR) and a real-time PCR (RT-PCR). The overall prevalence of D. fragilis in the study population was 5.4% (35/650). RT-PCR detected 35 isolates, conventional PCR detected 15 isolates, MBD culture detected 14 isolates, TYGM-9 detected ten isolates, while microscopy detected 12 isolates. RT-PCR detected an additional 15 positive samples compared to the other diagnostic methods, all of which were confirmed by sequencing. When all methods were compared to each other, RT-PCR showed a sensitivity and specificity of 100 and 100%, conventional PCR 42.9 and 100%, MBD culture 40 and 100%, TYGM-9 culture 28.6 and 100%, and microscopy 34.3 and 99%, respectively. These results show that RT-PCR is the diagnostic method of choice for the detection of D. fragilis in clinical samples and, as such, should be considered as the gold standard for diagnosis.