The cestode order Rhinebothriidea no longer family-less: A molecular phylogenetic investigation with erection of two new families and description of eight new species of Anthocephalum.

The spiral intestines of a total of 30 specimens of 14 species of batoids from around the world were examined for rhinebothriideans. These consisted of Taeniura grabata, Dasyatis margaritella, and Dasyatis sp. from Senegal, Dasyatis americana from Florida, Dasyatis dipterura and Dasyatis longa from México, Himantura jenkinsii, Himantura leoparda, Himantura uarnak 2, Urogymnus asperrimus 1, and Neotrygon kuhlii 4 from Australia, in addition to Himantura uarnacoides and Neotrygon kuhlii 1 from Borneo. Each of these hosted one or more species of Anthocephalum. Eleven of the cestode species were new to science; four represented described species. In addition, Urotrygon aspidura from Costa Rica hosted a species of Escherbothrium. Sufficient material was available for formal description of the following eight species of Anthocephalum: A. decrisantisorum n. sp., A. healyae n. sp., A. jensenae n. sp., A. mattisi n. sp., A. meadowsi n. sp., A. odonnellae n. sp., A. papefayi n. sp., and A. philruschi n. sp. These species differ from their nine described congeners in overall size, number of proglottids and marginal loculi, number and arrangement of testes, apical sucker size, arrangement and distribution of vitelline follicles particularly with respect in the post-poral field, and muscularity of the genital pore. The diagnosis of Anthocephalum is emended slightly to accommodate these new species. Material of four previously described Anthocephalum species, seven of the novel Anthocephalum species described here, 3 undescribed Anthocephalum species, and the species of Escherbothrium was preserved in 95% ethanol and partial 28S rDNA (D1-D3) and complete 18S rDNA sequence data were generated de novo. These data were combined with data from GenBank for Anthocephalum cf. centrurum (recognized as A. mattisi n. sp. below) and 29 species representing 12 other putative rhinebothriidean genera. Phylogenetic analyses using Bayesian Inference and Maximum Likelihood methods were conducted using a total of five representatives of the Lecanicephalidea, Cathetocephalidea and "Tetraphyllidea" as outgroups. The analyses yielded trees that were largely congruent and that supported the existence of four major subgroups of rhinebothriideans. Family designations were established for each of these clades. Echeneibothriidae was elevated from subfamily to family level to accommodate the group consisting of Echeneibothrium and Pseudanthobothrium; this family is unique in retaining the apical organ (as a myzorhynchus) into adulthood. Rhinebothriidae was elevated from subfamily to family level to accommodate the group consisting of Rhabdotobothrium, Rhinebothrium, Rhinebothroides, Rhodobothrium, Scalithrium and Spongiobothrium. This family is distinctive in its lack of apical suckers and also of a definitive anterior/posterior orientation to the bothridia. Anthocephaliidae n. fam. was established to house Anthocephalum and taxa identified as New Genus 1, New Genus 2, and New Genus 4 by previous authors. The bothridia of its members exhibit a conspicuous anterior/posterior orientation signaled by the presence of an apical sucker. In addition, its members bear marginal loculi or one or more rows of facial loculi and vitelline follicles that are usually interrupted by the ovary. Escherbothriidae n. fam. was established to house Escherbothrium and the taxon identified as New Genus 3 by previous authors. It most closely resembles Anthocephaliidae but the facial loculi are arranged in columns anteriorly and rows posteriorly, rather than arranged in multiple rows or entirely lacking. A key to the families is also provided.