Molecular diagnosis and genotyping of Dienlamoeba fragilis by polymerase chain reaction compared to microscopy and culture

ABSTRACT

The diagnosis of D. fragilis by microscopic identification of the parasite in stool is time consuming and relatively insensitive. To evaluate microscopy, culture and PCR for detection of D. fragilis in stool samples and to identify the genotypes of D. fragilis isolates among the study population. A total of 82 fresh human fecal samples were examined by microscopy using Wheatley's trichrome permanent staining, culture using the modified Boeck and Drbohlav medium and PCR targeting the small subunit [SSU] rRNA gene. Additionally, the existence of genetic variation among D. fragilis isolates [proved positive by PCR] was examined by restriction fragment length polymorphism analysis. PCR detected 25 isolates [30.5%], MBD culture detected 24 isolates [29.3%], while microscopy detected 8 isolates [9.8%]. Sensitivities of PCR, culture and microscopy were 92.6%, 88.9% and 29.6%, respectively. The agreement between PCR and culture results was substantial [KA=0.86]. PCR followed by RFLP analysis revealed the existence of two genetic variants among 25 D. fragilis isolates. Genotype I predominated in 23 [92%] samples, while the remaining two isolates corresponded to genotype II. It is recommended to use culture for routine diagnosis of D. fragilis in suspected gastrointestinal cases. Two genetic variants of D. fragilis existed in the Egyptian isolates