Differential detection of Entamoeba histolytica, Entamoeba dispar, and Entamoeba moshkovskii by a single-round PCR assay.
A single-round PCR assay was developed for detection and differential diagnosis of the three Entamoeba species found in humans, Entamoeba moshkovskii, Entamoeba histolytica, and Entamoeba dispar, that are morphologically identical as both cysts and trophozoites. A conserved forward primer was derived from the middle of the small-subunit rRNA gene, and reverse primers were designed from signature sequences specific to each of these three Entamoeba species. PCR generates a 166-bp product with E. histolytica DNA, a 752-bp product with E. dispar DNA, and a 580-bp product with E. moshkovskii DNA. Thirty clinical specimens were examined, and the species present were successfully detected and differentiated using this assay. It was possible to detect as little as 10 pg of E. moshkovskii and E. histolytica DNA, while for E. dispar the sensitivity was about 20 pg of DNA. Testing with DNA from different pathogens, including bacteria and other protozoa, confirmed the high specificity of the assay. We propose the use of this PCR assay as an accurate, rapid, and effective diagnostic method for the detection and discrimination of these three morphologically indistinguishable Entamoeba species in both routine diagnosis of amoebiasis and epidemiological surveys.
Hamzah
Zulhainan
Z
Department of Protozoology, Faculty of Tropical Medicine, Mahidol University, 420/6 Rajvithi Road, Bangkok 10400, Thailand.
Petmitr
Songsak
S
Mungthin
Mathirut
M
Leelayoova
Saovanee
S
Chavalitshewinkoon-Petmitr
Porntip
P
D000818:Animals; D017931:DNA Primers; D016054:DNA, Protozoan / Q000032:analysis; D003937:Diagnosis, Differential; D004747:Entamoeba / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D004748:Entamoeba histolytica / Q000145:classification / Q000235:genetics / Q000302:isolation & purification; D004749:Entamoebiasis / Q000175:diagnosis* / Q000469:parasitology*; D006801:Humans; D008101:Liver Abscess, Amebic / Q000175:diagnosis / Q000469:parasitology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013785:Thailand
D017931:DNA Primers; D016054:DNA, Protozoan