PCR-RFLP analysis of the ITS2 region to identify Schistosoma haematobium and S. bovis from Kenya

Abstract

Schistosoma haematobium, primarily a human parasite, and the closely related Schistosoma bovis from ruminants, are sympatric in many African countries such as Kenya. Because these two species 1) can inhabit the same Bulinus snails, 2) may be found in the same freshwater habitat, and 3) have morphologically similar cercariae, better means are needed to tell them apart. The second internal transcribed spacer (ITS2) region of the ribosomal gene complex (rDNA) of recent Kenyan isolates of both species was sequenced and found to be a 98% match. The S. bovis sequences were nearly identical (99%) to conspecific sequences from Niger; the S. haematobium sequences were nearly identical (99%) to conspecific sequences from Egypt, Mali, and Niger. Restriction fragment length polymorphism (RFLP) analysis of a 480 base pair (bp) PCR product containing the ITS2 region using two restriction enzymes, Taq1 and Sau3A1, yielded species-specific fragment patterns that allowed successful identification of a single S. haematobium cercaria. The protocol outlined here is useful in providing a rapid, one-day identification of S. haematobium (and likely S. bovis) cercariae (the infective larval stage) and/or other life cycle stages in a basic molecular biology laboratory. By helping to determine whether schistosome-infected bulinid snails in a particular body of water are transmitting a human or an animal schistosome, or both, this analysis will aid in disease control and in ongoing epidemiological studies.