Molecular-Genotyping Detection of Entamoeba histolytica in Diarrheic Patients

Amebiasis is caused by Entamoeba histolytica, a protozoan that is found worldwide. The degree of pathogenesis of clinical isolates varies greatly. This study was aimed to molecular identification of E. histolytica in children using the nested polymerase chain reaction (nPCR), and then, a genotyping of positive E. histolytica isolates using the quantitative PCR (qPCR) assay through targeting serine-rich E. histolytica protein (SREHP) gene. For this purpose, a total of 50 bloody diarrheic stool samples were collected from the children attended to the Al-Zahraa' Teaching Hospital and Alkut Hospital for Gynecology, Obstetric and Pediatrics (Alkut, Wasit, Iraq) were subjected to the present study from September to December 2021. Firstly, the extracted DNAs that amplified using specific primers through targeting 18S rRNA gene and tested using nPCR assay were revealed an overall 48% (24/50) positive samples for E. histolytica. For genotyping, our results were detected an existence of four different genotypes (I, II, III and IV) with a significant prevalence of Genotype-II (54.17%) when compared to Genotype-I (20.83%), Genotype-III (12.5%) and Genotype- IV (12.5%). In addition, results of melting temperature of targeted genotypes were 84ºC, 83 - 83.5ºC, 82.5ºC and 81ºC for Genotype-I, II, III and IV, respectively. In conclusion, molecular amplification of 18S rRNA gene was revealed the large prevalence of E. histolytica among bloody diarrheic children of study areas; while, amplification of SREHP gene was reflected the widespread phenotypic variation of the Genotype-II suggesting the high ability of this genotype to spread infection in children. In different endemic areas as Iraq, the utilization of high-resolution genotyping methods showed the extremely polymorphic genetic structure of this parasite.