Journal Article

Evaluation of a New Single-Tube Multiprobe Real-Time PCR for Diagnosis of Entamoeba histolytica and Entamoeba dispar

Liang, Shih-Yu, Chan, Yun-Hsien, Landt, Olfert, Jiang, Donald Dah-Shyong, Ji, Dar-Der, Fan, Chia-Kwung, Hsia, Kan-Tai

2010

Overview

A single-tube multiprobe real-time PCR assay for simultaneous detection of Entamoeba histolytica and Entamoeba dispar was developed. One primer pair with 2 species-specific probes was designed based on new SSU RNA regions of the ribosomal DNA-containing episome. The sensitivity is 1 parasite per milliliter of feces and thus superior to the conventional nested PCR and comparable to other published real-time PCR protocols. The applicability for clinical diagnosis was validated with 218 stool specimens from patients. A total of 51 E. histolytica and 39 E. dispar positive samples was detected by the multiprobe real-time PCR compared to 39 and 22 by routine nested PCR diagnosis. The detection rate of Entamoeba species for the multiprobe real-time PCR assays was significantly higher than the nested PCR (40.8% vs. 28.0%, P < 0.01). The test did not show cross reactivity with DNA from Entamoeba moshkovskii, Giardia lamblia, Cryptosporidium sp., Escherichia coli, or other nonpathogenic enteric parasites. The multiprobe real-time PCR assay is simple and rapid and has high specificity and sensitivity. The assay could streamline the laboratory diagnosis procedure and facilitate epidemiological investigation.

• Publisher: American Society of Parasitologists,Allen Press Inc
• Subject: Amebiasis / Cryptosporidium / DNA Primers - standards / DNA probes / DNA Probes - standards / DNA, Protozoan - analysis / E coli / Electrophoresis, Agar Gel / Entamoeba - classification / Entamoeba - genetics / Entamoeba - isolation & purification / Entamoeba dispar / Entamoeba histolytica / Entamoeba histolytica - classification / Entamoeba histolytica - genetics / Entamoeba histolytica - isolation & purification / Entamoeba moshkovskii / Entamoebiasis - diagnosis / Entamoebiasis - parasitology / Epidemiology / Escherichia coli / Feces / Feces - parasitology / Giardia lamblia / Humans / Infections / Linear Models / Microbiology / Parasites / Parasitic diseases / Parasitology / Polymerase chain reaction / Polymerase Chain Reaction - methods / Polymerase Chain Reaction - standards / Primers / Product category rules / RNA / Sensitivity and Specificity / Species Specificity / Specimens / THERAPEUTICS-DIAGNOSTICS / Trophozoites