Moniezia benedeniandM. expansaare common ruminant tapeworms of worldwide distribution, causing gastrointestinal disorders and even death in sheep and goats. In this study, a polymerase chain reaction- (PCR-) based approach for precise species identification was developed and also applied to faecal DNA diagnosis of the tapeworm infection. Since nuclear ribosomal DNA (rDNA) appears to be a useful target for species and/or strain markers, the 18S regions of the rDNA ofM. benedeniandM. expansawere amplified and sequenced. The lengths and GC contents of the regions sequenced were 2476–2487 bp and 51.9–52.1% forM. benedeniand 2473 bp and 51.9–52.0% forM. expansa, respectively. Alignment and comparison of the 18S sequences of the two species showed 92.5–93.3% homology. No matches for the 18S regions ofM. benedeniandM. expansawere found with other species by BLAST search, which made the 18S sequences appropriate markers for the design of distinctive primers for the twoMonieziaspecies. Our 18S-based PCR could detect as low levels as 0.5 pg genomic DNA or the DNA extracted from 0.2 g faecal sample collected from the rectum of anM. expansa-infected goat. The results indicate that this PCR approach is a reliable alternative for the differential diagnosis ofMonieziaspecies in faecal samples.