Strains of Entamoeba histolytica isolated in Brazil were characterized using the Low-Stringency Single Specific Primer PCR (LSSP–PCR), that detects single or multiple mutations in gene size DNA fragments. Using this technique, a 482-bp genomic DNA fragment from a structural gene in 8 strains and 2 clones of E. histolytica, isolated from symptomatic and asymptomatic patients in Brazil, including pathogenic and non-pathogenic zymodemes were studied. The results obtained indicate that LSSP–PCR is a valuable method for differentiating strains of E. histolytica. Moreover, the results are consistent with the concept that pathogenic and non-pathogenic strains of E. histolytica may represent distinct species or subspecies and are in accord with phenotypically characteristic isoenzyme patterns.