Direct amplification and differentiation of pathogenic and nonpathogenic Entamoeba histolytica DNA from stool specimens

Discrimination of pathogenic and nonpathogenic Entamoeba
histolytica is of great clinical importance. A simple and rapid DNA
extraction method that can be used directly with stool specimens
was developed without the need for prior cultivation of the
parasites. The entire protocol can be performed at room temperature
in a 1.5-ml microcentrifuge tube format. There is no DNA
precipitation step. The subsequent nested polymerase chain reaction
consists of an initial E. histolytica-specific amplification,
followed by two separate amplifications using two primer pairs
specific for pathogenic and nonpathogenic E. histolytica,
respectively. Amplification products can be verified by restriction
endonuclease digests. There is no need for hybridizations or the
use of radionucleotides. One trophozoite per milligram of stool
sample could be detected and differentiated in a 0.1-g specimen.