Multiple gene analyses of caligid copepods indicate that the reduction of a thoracic appendage in Pseudocaligus represents convergent evolution

Background: The Caligidae is a family of parasitic copepods containing over 30 recognised genera. They are commercially important parasites as they cause disease in numerous finfish aquaculture facilities globally. Morphological features are used to distinguish between the genera and Pseudocaligus has traditionally been differentiated from Caligus solely by the presence of a much reduced form of the fourth thoracic leg. Currently there are numerous DNA sequences available for Caligus spp. but only the type species, Pseudocaligus brevipedis, has molecular data available, so systematic studies using molecular phylogenetic analyses have been limited. Methods: Three gene regions, SSU rDNA, 16S and CO1, for Pseudocaligus fugu from puffer fish from Japan and Pseudocaligus uniartus from rabbit fish from Indonesia are sequenced and molecular phylogenetic analyses performed in order to infer phylogenetic relationships between Pseudocaligus and other caligid copepods. Results: The analysis revealed that there was no discrete grouping of Pseudocaligus spp. and that they had a polyphyletic distribution within Caligus taxa. Pseudocaligus fugu grouped with Caligus elongatus and contained a unique synapomorphy in the SSU rDNA region only seen in members of that clade. Pseudocaligus uniartus formed a well-supported group, in the SSU rDNA analyses, with a Caligus sp. that also infects rabbit fish, but was unresolved in the other analyses. Pseudocaligus brevipedis consistently and robustly grouped with Caligus curtus and C. centrodonti in all analyses. The majority of Lepeophtheirus spp. form a monophyletic sister group to the Caligus clade; however, L. natalensis is unresolved in all analyses and does not form part of the main Lepeophtheirus clade. Conclusions: These findings do not support the morphological-based distinction between Pseudocaligus and Caligus, suggesting that the reduced fourth leg is a feature that has evolved on multiple occasions throughout caligid evolution. Congruent molecular phylogenetic data support groupings based on the presence of morphological features, such as lunules, geography and host fish type rather than appendage morphology. Therefore, we support the synonymy of Pseudocaligus with Caligus.