Distribution and differential diagnosis of entamoeba histolytica from entamoeba dispar by the PCR-RFLP method in central Iran

ABSTRACT

Entamoeba histolytica and Entamoeba dispar are two morphologically indistinguishable human protozoan parasites that are genetically distinct species. The potential invasive pathogenic Entamoeba histolytica and non-invasive parasite Entamoeba dispar can be differentiated by molecular and other methods. We used polymerase chain reaction [PCR] to determine the ratio of the two species in a population in Tehran and Karaj in central Iran. Materials and Human stool samples [n=12 148] were randomly collected in Tehran and Karaj and examined for E. histolytica/E. dispar cysts with direct and formalin-ether methods. Eighty-seven [0.7%] cases were positive, of which 49 [62.8%] isolates were successfully cultured in Robinson oligonucleotide primers designed from sequence data for genomic DNA coding the 30-KD surface antigen of E. histolytica/E. dispar was used to amplify a 374 base-pair [bp] fragment. The electrophoretic pattern of the PCR product digested with Hinfl restriction enzyme was used for differentiation of the two species. The restriction fragment length polymorphism [RFLP] pattern obtained from a standard E. histolytica isolate had two fragments [219 bp and 155 bp], but the standard isolate of E. dispar showed three fragments [155, 152 and 67 bp]. Differential diagnosis of 49 isolates of E. histolytica/E. dispar from Tehran and Karaj using PCR-RFLP revealed that 46 [93.9%] were E. dispar while only 2 [4.1%] were E. histolytica. One person [2%] had a mixed infection and showed both patterns. The differential diagnosis of the potentially pathogenic parasite E. histolytica from the non-pathogenic E. dispar is of clinical and epidemiological importance. This study demonstrated that E. dispar is much more prevalent than E. histolytica among the cyst passers Iran