Phylogenetic relationships of 9 species of ticks were inferred from nucleotide sequences of the D1 domain of large subunit ribosomal DNA (rDNA), the V4 region of small subunit rDNA and sequences immediately adjacent to these regions. Six of the 7 subfamilies in the Argasidae and Ixodidae were represented: Ornithodorinae and Argasinae from the Argasidae (soft ticks); and Ixodinae, Amblyomminae, Haemaphysalinae and Rhipicephalinae from the Ixodidae (hard ticks). A mite, Phytoseiulus persimilis, from the Mesostigmata, the putative sister group of the Ixodida, was used for out-group reference. Our sequence alignment, which contained the D1, V4 and adjacent sequences, comprised 503 sites; 209 of these varied among the species studied and 77 were phylogenetically informative, i.e. at least 2 different nucleotides were represented at least twice. This alignment was subjected to maximum parsimony, genetic distance and maximum likelihood analyses. We found strong support in bootstrap resampling experiments for the following relationships: (i) monophyly of the 7 species of Ixodidae (98-100% support); (ii) monophyly of the 2 species of Amblyomma (93-99%); (iii) monophyly of the 6 species of Metastriata (96-99%); (iv) a sister-group relationship between Ixodes pilosus and the 6 species of Metastriata (98-100%); and (v) a sister-group relationship between Ornithodoros capensis (Ornithodorinae) and the 7 species of Ixodidae (94-99% support). The last result may be preliminary evidence that the Argasidae family (soft ticks) is paraphyletic since the 2 argasid species did not form a monophyletic group. The paraphyletic status of the Argasidae has been previously suggested by Black & Piesman (1994; Proceedings of the National Academy of Sciences U.S.A. 91: 10034). Our study indicates that the D1, V4 and adjacent sequences may be used to answer long-standing questions about the phylogeny of the Ixodida.