Insights from new cestodes of the crocodile shark, Pseudocarcharias kamoharai (Lamniformes: Pseudocarchariidae), prompt expansion of Scyphyophyllidum and formal synonymization of seven phyllobothriidean genera - at last!

Three new cestode species are described from the crocodile shark (Pseudocarcharias kamoharai) in Ecuador. All three were examined with light and scanning electron microscopy. The unique combination of morphological features in one of the new species prompted formal investigation of the non-monophyly of Paraorygmatobothrium relative to the morphologically similar genera Doliobothrium, Guidus, Marsupiobothrium, Nandocestus, Orectolobicestus, Ruhnkecestus and Scyphophyllidium. Sequence data generated for part of the 28S rDNA gene were subjected to maximum likelihood (ML) analysis. The resulting tree led to the synonymization of six of these seven genera with Scyphophyllidium, and transfer of their species to the latter genus. With the new species, the number of described members of Scyphophyllidium is now 45. The diagnosis of Scyphophyllidium is revised to accommodate these species. In addition, to expedite future descriptions, eight categories of Scyphophyllidium species are circumscribed, based largely on bothridial features. Scyphophyllidium timvickiorum n. sp. is a category 1 species. Beyond being the smallest category 1 species, it bears, rather than lacks, apical suckers and lacks, rather than bears, strobilar scutes. The two other new species are members of Clistobothrium. Clistobothrium amyae n. sp. differs from its congeners in bothridial shape, elongate cephalic peduncle and tiny size. Clistobothrium gabywalterorum n. sp. differs from the two of its congeners that also possess foliose bothridia in overall size and testis number. Despite their substantial morphological differences, the ML tree indicates they are sister taxa. Both are unique among their congeners in possessing cephalic peduncle spinitriches. The diagnosis of Clistobothrium is revised accordingly.

Four New Species of Paraorygmatobothrium (Eucestoda: Phyllobothriidea) From Sharks of the Gulf of Mexico and the Atlantic Ocean, with Comments on Their Host Specificity.

Four new species of Paraorygmatobothrium are described from carcharhinid shark species from the Gulf of Mexico and the U.S. coast of the northwestern Atlantic Ocean. Paraorygmatobothrium bullardi n. sp. is described from Carcharhinus brevipinna as its type host, and from Carcharhinus acronotus and Carcharhinus limbatus. Paraorygmatobothrium campbelli n. sp. is described from Rhizoprionodon terraenovae. Paraorygmatobothrium deburonae n. sp. is described from Carcharhinus isodon as its type host, as well as from C. brevipinna, C. limbatus, and R. terraenovae. Paraorygmatobothrium mattisi n. sp. is described from R. terraenovae as its type host, and from C. brevipinna and C. limbatus. These 4 species differ from other species of Paraorygmatobothrium and from each other in bothridial microthrix shape, and in various combinations of features such as the total length of the worm, apical sucker size, bothridial musculature, proglottid number, testes number, and genital pore position. Difficulty in differentiating these 4 species from one another, and from other species of Paraorygmatobothrium, points to limitations in the use of morphology alone to recognize species diversity in this genus. A phylogenetic analysis of Paraorygmatobothrium and related genera, based on partial (D1-D3) 28S rDNA sequence data, confirms that the genus is not monophyletic. Genetic distances of COI revealed that sequence divergence between species of Paraorygmatobothrium is generally at least an order of magnitude larger than COI divergence within a species. Patterns of host use for 3 of the new species are consistent with previous findings that point to a lack of host specificity in Paraorygmatobothrium.

Two new species and a new phyllobothriid cestode genus from sharks of the genus Negaprion Whitley (Carcharhiniformes).

Alexandercestus n. g. (Cestoda: Tetraphyllidea) is erected for two cestode species found parasitising the two known species of lemon sharks (Carcharhiniformes: Negaprion spp.). This new genus differs from all other phyllobothriid genera except for Hemipristicola Cutmore, Theiss, Bennett & Cribb, 2011, Marsupiobothrium Yamaguti, 1952, Nandocestus Reyda, 2008, Orectolobicestus Ruhnke, Caira & Carpenter 2006, Orygmatobothrium Diesing, 1863, Paraorygmatobothrium Ruhnke, 1994 and Phyllobothrium van Beneden, 1849 in possessing uniloculate bothridia with an apical sucker and neck scutes. Alexandercestus differs from Orectolobicestus and Nandocestus in lacking marginal loculi on the bothridia, from Paraorygmatobothrium in possessing uninterrupted vitelline follicles at the level of the ovary and from Phyllobothrium in being euapolytic as opposed to anapolytic and in lacking posteriorly bifid bothridia. The new genus lacks the central accessory bothridial organ seen in specimens of Orygmatobothrium, and lacks the central bothridial accessory sucker of specimens of Marsupiobothrium. Alexandercestus spp. compare most favourably with specimens of Hemipristicola, especially with respect to aspects of proglottid morphology, but differ in possessing aristate gladiate spinitriches rather than serrate gladiate spinitriches on the proximal bothridial surface. In addition, the bothridia of Alexandercestus spp. are comparatively more fleshy and foliose than those in specimens of Hemipristicola. Two new species of Alexandercestus n. g. are described, Alexandercestus gibsoni n. sp. from Negaprion acutidens, collected from off northern Australia and the Marshall Islands, and Alexandercestus manteri n. sp. from N. brevirostris, collected off the islands of Bimini and the Florida Keys. The two new species differ in total length and vitelline follicle distribution. Bayesian inference and parsimony analysis of the D1-D3 region of the large nuclear ribosomal DNA of 17 published and seven novel sequences placed A. gibsoni as the sister taxon to a clade containing Hemipristicola gunterae Cutmore, Theiss, Bennett & Cribb, 2011 and species of Paraorygmatobothrium. This result supports the erection of Alexandercestus as a genus separate from Hemipristicola and Paraorygmatobothrium. At the present time, species of Alexandercestus are known only from hosts of the carcharhinid genus Negaprion Whitley; examination of extensive survey data suggests this may be the extent of the host distribution of this genus.

Two new species of Anthobothrium van Beneden, 1850 (tetraphyllidea: phyllobothriidae) from carcharhinid sharks, with a redescription of Anthobothrium laciniatum Linton, 1890.

Anthobothrium laciniatum Linton, 1890 is redescribed based on specimens taken from the dusky shark Carcharhinus obscurus (Lesueur) collected from the Northwestern Atlantic Ocean, and a neotype is designated. A. laciniatum differs from A. cornucopia van Beneden, 1850, A. altavelae Euzet & Ben Hassine, 2002, A. lesteri Williams, Burt & Caira, 2004 and A. spinosum Subhapradha, 1955 in total length. It further differs from A. cornucopia, A. altavelae and A. spinosum in proglottid number, and differs from A. galeorhini Suriano, 2002, A. cornucopia, and A. spinosum in testis number. A. lyndoni n. sp. is described from the sandbar shark C. plumbeus (Nardo). This new species differs from A. laciniatum in ovarian width and from A. cornucopia, A. altavelae, A. galeorhini and A. spinosum in the total number of proglottids. It further differs from A. cornucopia, A. galeorhini, and A. spinosum in total length, and from A. cornucopia and A. galeorhini in the number of testes. A. lyndoni n. sp. differs from A. lesteri in bothridial muscle and ovarian morphology. Anthobothrium caseyi n. sp. is described from Prionace glauca (Linnaeus). This new species differs conspicuously from the other six species of Anthobothrium van Beneden, 1850 (sensu stricto) in the shape of its proglottid laciniations. The taxonomic status of 43 species that have been associated with Anthobothrium is addressed. Taxonomic actions regarding Anthobothrium during the past century have resulted in a polyphyletic taxon.

Three new species of Anthocephalum Linton, 1890 (Cestoda: Tetraphyllidea) from dasyatid stingrays of the Gulf of California.

Three new species of Anthocephalum Linton, 1890 are described from dasyatid stingrays collected in the Gulf of California. Anthocephalum michaeli n. sp. is described from Dasyatis longus (Garman). This species most closely resembles A. alicae Ruhnke, 1994, but differs from this species in proglottid number. A. lukei n. sp. is also described from D. longus. This new species is most similar to A. cairae Ruhnke, 1994, but differs from that species in marginal loculi number and number of proglottids. The third new species, A. currani n. sp., is described from D. brevis (Garman). This species is most similar to A. centrurum (Southwell, 1925) Ruhnke, 1994, but differs from that species in marginal loculi number, number of testes and ovarian length. Phyllobothrium kingae Schmidt, 1978 is also consistent in morphology with species of Anthocephalum and is transferred to this genus, forming the new combination Anthocephalum kingae n. comb. This species most closely resembles A. michaeli n. sp., but differs in testicular shape. This brings the total number of species of Anthocephalum to nine. The transfer of the species Phyllobothrium arctowskii Wojciechowska, 1991, P. georgiense Wojciechowska, 1991, P. rakusai Wojciechowska, 1991 and P. siedleckii Wojciechowska, 1991 to Anthocephalum is not warranted, as these four species lack a posteriorly recurved cirrus-sac and a sinuous vagina, and have vitelline follicles uninterrupted by the ovary. Of the nine known species, all are parasitic in batoid fishes, and six are found in species of Dasyatis Garman. The phylogenetic status of Anthocephalum species in relationship to Rhinebothroides Mayes, Brooks & Thorson, 1981, Pararhineothroides Zamparo, Brooks & Barriga, 1999 and other rhinebothriin taxa is discussed.

Two new species of Paraorygmatobothrium Ruhnke, 1994 (Tetraphyllidea: Phyllobothriidae) from the smooth-hound Mustelus mustelus (L.) and the gummy shark M. antarcticus Günther (Carcharhiniformes: Triakidae).

Two new tetraphyllidean species, Paraorygmatobothrium bai n. sp. and P. rodmani n. sp., are described from the smooth-hound shark Mustelus mustelus (Linnaeus) and the gummy shark M.s antarcticus Günther, respectively. The two species are generally consistent in morphology with the nine existing species of Paraorygmatobothrium Ruhnke, 1994. P. bai n. sp. and P. rodmani n. sp. differ from the existing species of Paraorygmatobothrium in exhibiting gravid proglottids on the strobila. P. bai n. sp. differs from P. rodmani n. sp. in testicular shape and number, in addition to significant differences in body length, and terminal and subterminal proglottis length to width ratios. Description of these two new species increases the known number of species of Paraorygmatobothrium to 11.

Orectolobicestus n. g. (Cestoda: Tetraphyllidea), with the description of five new species and the transfer of Phyllobothrium chiloscyllii to the new genus.

ORECTOLOBICESTUS N. G. (CESTODA: Tetraphyllidea) is erected for six cestode species parasitising bamboo sharks (Orectolobiformes: Chiloscyllium). Members of this genus differ from all other phyllobothriid genera in possessing modified maisiform spinitriches on their distal bothridial surfaces. In addition, they are easily recognised in that they share the following unique combination of characters: their vitelline fields are interrupted by the ovary, their necks are scutellate, and their bothridia bear an apical sucker and marginal loculi. Five new species of Orectolobicestus are described, including O. tyleri n. sp. from Chiloscyllium punctatum off Borneo, O. lorettae n. sp. from C. cf. punctatum off Australia, O. mukahensis n. sp. and O. kelleyae n. sp. from C. indicum off Borneo, and O. randyi n.sp. from C. hasselti also from off Borneo. In addition, Phyllobothrium chiloscyllii Subhapradha, 1955 is transferred to the new genus. O. chiloscyllii (Subhapradha, 1955) n. comb. is readily distinguished from all five new species in its greater total length. In addition to a number of proglottid features, O. kelleyae n. sp. and O. randyi n. sp. clearly differ from the other three new species in their possession of trifid, rather than fully serrate, spinitriches on their proximal bothridial surfaces. The latter two species conspicuously differ from one another in total number of proglottids (11-21 vs 27-38). O. tyleri n. sp. generally has fewer proglottids than O. lorrettae n. sp. (7-17 vs 13-23) and, like O. mukahensis n. sp., possesses scutes that are spathate rather than elongate. O. tyleri n. sp. is readily distinguished from O. mukahensis n. sp. in its possession of fewer proglottids (7-17 vs. 19-29). The five new species of Orectolobicestus share derived bothridial microthrix features with Phyllobothrium squali Yamaguti, 1952, Thysanocephalum sp., Orygmatobothrium sp., Ruhnkecestus Caira & Durkin, 2006 and species of Paraorygmatobothrium Ruhnke, 1994. Among these taxa, Orectolobicestus most closely resembles Paraorygmatobothrium and Ruhnkecestus in its vitelline fields being interrupted by the ovary and the possession of a scutellate neck.

An unusual blood sequestering tapeworm (Sanguilevator yearsleyi n. gen., n. sp.) from Borneo with description of Cathetocephalus resendezi n. sp. from Mexico and molecular support for the recognition of the order Cathetocephalidea (Platyhelminthes: Eucestoda).

Sanguilevator yearsleyi n. gen., n. sp. and Cathetocephalus resendezi n. sp. are described from the Broadfin shark, Lamiopsis temmincki, in Malaysian Borneo and Carcharhinus leucas in Mexico, respectively. The new genus is unusual in its possession of internal chambers and channels in its scolex that appear to house extensive quantities of host white and red blood cells, respectively. Histology reveals an extremely intimate association between host tissue and the surface of the apical pad of the scolex. Positive staining with periodic acid-Schiff of the surface of the pad of the scolex and the linings of the chambers and channels suggests that an adhesive substance may be produced in these regions. However, explanations for how and why host blood cells come to reside within the scolex remain elusive. Cathetocephalus resendezi n. sp. is distinctive in the form of the papillae in the papillate band of the scolex and also in the inconspicuous nature of the rugose base of the scolex. Scanning electron microscopy of both new taxa as well as Cathetocephalus thatcheri, Cathetocephalus australis and an undescribed species of Cathetocephalus collected from Carcharhinus amboinensis in Australia, suggests that the papillae surrounding the pad of the scolex are of significant taxonomic utility in distinguishing among species in these groups. Parsimony and Bayesian analyses of sequence data (766 bases of 18S rDNA and 405 bases of 28S rDNA) generated from ethanol preserved specimens of C. thatcheri and S. yearsleyi, when compared with equivalent data available for 40 cestode species in GenBank, resulted in trees that support previous propositions that Cathetocephalus should be placed in the order Cathetocephalidea. The results suggest that Sanguilevator should also be considered to belong to this order.

Two new species of Duplicibothrium Williams & Campbell, 1978 (Tetraphyllidea: Serendipidae) from the Pacific cownose ray Rhinoptera steindachneri.

Two new tetraphyllidean species, Duplicihothrium caircae n. sp. and D. paulum n. sp., are described from Pacific cownose rays Rhinoptera steindachneri collected from the Gulf of California. D. cairae n. sp. differs from the only other known species in this genus, D. minutum, in its possession of two posteriorly bifurcating longitudinal septa on each bothridium and having a greater number of loculi per bothridium, wider bothridia and a greater number of segments per worm. D. paulum n. sp. differs from the above two species in being shorter, having two continuous longitudinal septa on each bothridium, rather than posteriorly bifurcating or absent longitudinal septa, having a greater number of loculi per bothridium and having fewer segments per worm. The generic diagnosis of Duplicibothrium is amended to reflect the inclusion of the two new species. Species of Duplicibothrium appear to be restricted to host species of the genus Rhinoptera. Systematic information gained from the study of these two new species supports the monophyly of the tetraphyllidean family Serendipidae and suggests that Duplicibothrium shares a phylogenetic heritage with species in the genera Glyphobothrium and Serendip, taxa that are also parasitic in cownose rays. Evidence for a phylogenetic relationship between serendipid species and species of Dioecotaenia is discussed, as are potential molluscan hosts for cestode species in Duplicibothrium and Dioecotaenia.

Evidence for host-specific clades of tetraphyllidean tapeworms (Platyhelminthes: Eucestoda) revealed by analysis of 18S ssrDNA.

Sequence data from the V4 and V7-V9 variable regions of the 18S small subunit ribosomal DNA (ssrDNA) gene were used to examine relationships among 26 tetraphyllidean and two lecanicephalidean taxa. Newly collected specimens of 21 of the tetraphyllidean species were used to generate ssrDNA sequences that were combined with sequences previously available, including those of two diphyllidean taxa used for outgroup rooting. The sequences were aligned by eye according to secondary structural motifs of the conserved core of the molecule. Of the 1520 sites in the alignment, 874 (58%) were excluded from analysis due to alignment gaps and lack of positional homology as inferred by manual inspection. Genetic variability of the ssrDNA gene regions compared was greater than would be expected, based on the present taxonomy of the ingroup species, and the genetic divergences among tetraphyllidean 'families' and genera were comparable to that among tapeworm orders. Phylogenetic hypotheses were generated by the methods of maximum parsimony and maximum likelihood (GTR + I + Gamma nucleotide substitution model). Four most parsimonious trees resulted from analysis by maximum parsimony. Strict consensus of the four trees supported the monophyly of the Tetraphyllidea, with the lecanicephalidean taxa forming a sister lineage. Among the tetraphyllidean taxa included in the analysis were three major clades: a basal clade including species of the phyllobothriid genera Anthocephalum, Echeneibothrium, Rhinebothrium, Rhodobothrium and Spongiobothrium; a clade uniting the phyllobothriids of the genus Duplicibothrium with the dioecotaeniid genus Dioecotaenia; and a larger sister clade to the Duplicibothrium + Dioecotaenia clade that included the phyllobothriid genera Caulohothrium, Ceratobothrium, Clistobothrium, Paraoryigmatobothrium and Prosobothrium, the litobothriid genus Litobothrium and the onchobothriid genera Acanthobothrium, Calliobothrium, Phoreiobothrium and Platybothrium. Maximum likelihood analysis resulted in a topology that was congruent where nodes were strongly supported by parsimony analysis, but differed in the relative positions of the well-supported clades. In addition,maximum likelihood analysis grouped the lecanicephalidean taxa among the tetraphyllidean taxa, indicating paraphyly of the order Tetraphyllidea as currently defined. Relationships suggested by both methods of analysis reflected common host associations of the taxa better than their current classification, suggesting that coevolution has had a significant role in the evolution of the group.

Systematic resolution of Crossobothrium Linton, 1889, and taxonomic information on four species allocated to that genus.

The systematic status of the tetraphyllidean genus Crossobothrium is resolved. The description of the type species Crossobothrium laciniatum is emended to include new information from light and electron microscopy. Orygmatobothrium dohrnii is transferred to Crossobothrium and its description is emended. Crossobothrium dohrnii n. comb. differs from C. laciniatum in the morphology of large microtriches on the distal bothridial surface, the presence of irregular papillose projections on the distal bothridial surface, the possession of irregular, incomplete loculi on the bothridial margin morphology, and exhibiting oblong testes. The genital pore position of C. dohrnii also differs from C. laciniatum. Species of Crossobothrium differ from species in other sampled tetraphyllidean genera in apical sucker tegumentary surface morphology. The 2 species differ from species in all tetraphyllidean genera in possessing a field of deep longitudinal muscles and a greater number of follicles in the vitelline field at the level of the ovary. Two species formerly allocated to Crossobothrium, Phyllobothrium filiforme, and Phyllobothrium triacis, are transferred to Paraorygmatobothrium, and their descriptions are emended. Three other species previously allocated to Crossobothrium, namely Orygmatobothrium angustum, Phyllobothrium squali, and Tetrabothrius longicolle, are considered incertae sedis. As a taxonomic entity, Crossobothrium has suffered from a case of mistaken identity with tetraphyllidean species consistent in morphology with the genus Paraorygmatobothrium.

A new species of Calliobothrium (Tetraphyllidea: Onchobothriidae) from the whiskery shark, Furgaleus macki, in Australia.

Calliobothrium pritchardae n. sp. is described from the whiskery shark, Furgaleus macki (Whitley, 1943), collected from Young Rocks in South Australia. This species differs from the 7 other known species of Calliobothrium in its possession of 2 rather than 3 posthook loculi. It is a further distinguished from all other Calliobothrium species except Calliobothrium evani in its possession of asymmetrical hooks. The lack of an accessory piece between the bases of the axial hooks and medial axial hook bases that are longer than the lateral axial hook bases further distinguish the new species from C. evani in which an accessory piece is present and the relative lengths of the medial and lateral axial hook bases are reversed. Scanning electron microscopy reveals spiniform microtriches on the neck and proximal bothridial surfaces, filiform microtriches on the apical bothridial surfaces, and a combination of the 2 microthrix types on the distal bothridial surfaces. The extended bases of the axial hooks are densely covered with spiniform microtriches. In addition, the number of testes within worms decreases substantially in progressively posterior (older) segments. The generic diagnosis of Calliobothrium is emended to include this species with 2 rather than 3 posthook loculi.