Using a combination of the cytochrome oxidase subunit I gene (COI) and internal transcribed spacer (ITS) sequence information, cross-breeding, and morphological criteria, the taxonomic status of Tetranychus takafujii was reassessed. Described from Japan and very close to Tetranychus evansi, only the male empodium II separated the two species. Six T. evansi samples collected in Brazil, France, Kenya, Spain (including the Canary Islands), Taiwan, and two samples from Japan previously species-identified as T. takafujii, were examined. DNA sequences consistently divide the samples into two groups: group I held the mites from Brazil and France and group II contained the remaining samples of T. evansi together with the two mites collected in Japan. Likewise, cross-breeding experiments detected the same two groups that were reproductively partially incompatible. However, the Japanese samples were completely compatible with the rest of the group II T. evansi samples. The empodium II shape failed to distinguish consistently between the T. evansi and T. takafujii samples, but rather highlighted its variability, questioning its utility for separating these taxa. Congruent results support the co-specificity of the two taxa. It is concluded that T. takafuji is a junior synonym of T. evansi.

The spider mite sub-family Tetranychinae includes many agricultural pests. The internal transcribed spacer (ITS) region of nuclear ribosomal RNA genes and the cytochrome c oxidase subunit I (COI) gene of mitochondrial DNA have been used for species identification and phylogenetic reconstruction within the sub-family Tetranychinae, although they have not always been successful. The 18S and 28S rRNA genes should be more suitable for resolving higher levels of phylogeny, such as tribes or genera of Tetranychinae because these genes evolve more slowly and are made up of conserved regions and divergent domains. Therefore, we used both the 18S (1,825–1,901 bp) and 28S (the 5′ end of 646–743 bp) rRNA genes to infer phylogenetic relationships within the sub-family Tetranychinae with a focus on the tribe Tetranychini. Then, we compared the phylogenetic tree of the 18S and 28S genes with that of the mitochondrial COI gene (618 bp). As observed in previous studies, our phylogeny based on the COI gene was not resolved because of the low bootstrap values for most nodes of the tree. On the other hand, our phylogenetic tree of the 18S and 28S genes revealed several well-supported clades within the sub-family Tetranychinae. The 18S and 28S phylogenetic trees suggest that the tribes Bryobiini, Petrobiini and Eurytetranychini are monophyletic and that the tribe Tetranychini is polyphyletic. At the genus level, six genera for which more than two species were sampled appear to be monophyletic, while four genera (Oligonychus, Tetranychus, Schizotetranychus and Eotetranychus) appear to be polyphyletic. The topology presented here does not fully agree with the current morphology-based taxonomy, so that the diagnostic morphological characters of Tetranychinae need to be reconsidered.