The nucleotide sequences of the first and second internal transcribed spacers of nuclear ribosomal DNA were determined for adults of Cylicostephanus minutus from different geographical origins. The lengths of first and second internal transcribed spacer sequences ranged from 370 to 372bp and 215 to 216bp, respectively. Pairwise sequence comparisons revealed that some individuals of C. minutus had identical first and second internal transcribed spacer sequences, whereas others differed by 3.0% and 7.4% in their first and second transcribed spacers, respectively. Some individuals with sequence differences originated from the same host. The levels of difference within C. minutus were higher than that between the morphologically distinct species, Cylicostephanus goldi and Cylicostephanus longibursatus (0.8% for the first internal transcribed spacer and 3.8% for the second internal transcribed spacer). The data provide support for the proposal that C. minutus represents a complex of at least two species. In order to study the population genetic structure of C. minutus, a PCR-linked single-strand conformation polymorphism technique was also established.