Synchronous Identification of Entamoeba histolytica, Giardia intestinalis, and Cryptosporidium spp. in Stool Samples Using a Multiplex PCR Assay

Background: Diarrheal disease annually causes 760000 deaths in children, and 1700 million new cases are reported each year worldwide. Among the parasites, Entamoeba histolytica, Giardia intestinalis, and Cryptosporidium spp. are the most important infectious agents leading to diarrhea. Clinical presentations due to these parasites are more or less similar, and microscopy is not as much as sensitive for the detection. The aim of this study was to set up and evaluate a Multiplex PCR Assay for Synchronous Identification of Entamoeba histolytica, Giardia intestinalis, and Cryptosporidium spp. in Stool Samples Methods: Samples were obtained from different sources such as culture media and patient stool samples. Primer pairs were designed using primer-BLAST, and for the extraction of DNA, the QIAamp DNA stool mini kit was used. The study was conducted in Tehran, Iran and completed in 2016. Results: The current multiplex PCR assay for the detection of E. histolytica achieved sensitivity and specificity of 86.36% (95% CI: 65.09% to 97.09) and 95.74 % (95% CI: 85.46% to 99.48%), respectively. Sensitivity and specificity of the test for G. intestinalis was 90.91% (95% CI: 70.84% to 98.88%) and 95.74% (95%CI: 85.46% to 99.48%), respectively, and for the detection of Cryptosporidium, multiplex PCR showed a sensitivity of 90.91% (95% CI: 70.84% to 98.88%) and specificity of 95.74% (95%CI: 85.46% to 99.48%). Conclusion: Multiplex PCR in this study showed admissible sensitivity and specificity for the detection of E. histolytica, G. intestinalis, and Cryptosporidium spp. in fecal samples.