Morphological constraint obscures richness: a mitochondrial exploration of cryptic richness in Transversotrema (Trematoda: Transversotrematidae).

Species of Transversotrema Witenberg, 1944 (Transversotrematidae) occupy a unique ecological niche for the Trematoda, living externally under the scales of their teleost hosts. Previous studies of the genus have been impeded partly by limited variation in ribosomal DNA sequence data between closely related species and partly by a lack of morphometrically informative characters. Here, we assess richness of the tropical Indo-west Pacific species through parallel phylogenetic and morphometric analyses, generating cytochrome c oxidase subunit 1 mitochondrial sequence data and morphometric data for hologenophore specimens from Australia, French Polynesia, Japan and Palau. These analyses demonstrate that molecular data provide the only reliable basis for species identification; host distribution, and to a lesser extent morphology, are useful for identifying just a few species of Transversotrema. We infer that a combination of morphological simplicity and infection site constraint has led to the group displaying exceptionally low morphological diversification. Phylogenetic analyses of the mitochondrial data broadly support previous systematic interpretations based on ribosomal data, but also demonstrate the presence of several morphologically and ecologically cryptic species. Ten new species are described, eight from the Great Barrier Reef, Australia (Transversotrema chrysallis n. sp., Transversotrema daphnidis n. sp., Transversotrema enceladi n. sp., Transversotrema hyperionis n. sp., Transversotrema iapeti n. sp., Transversotrema rheae n. sp., Transversotrema tethyos n. sp., and Transversotrema titanis n. sp.) and two from off Japan (Transversotrema methones n. sp. and Transversotrema panos n. sp.). There are now 26 Transversotrema species known from Australian marine fishes, making it the richest trematode genus for the fauna.

Rich but morphologically problematic: an integrative approach to taxonomic resolution of the genus Neospirorchis (Trematoda: Schistosomatoidea).

Neospirorchis Price, 1934 is a genus of blood flukes that infect the cardiovascular system, including vessels surrounding the nervous systems of marine turtles. Although the genus comprises just two named species, the available molecular data suggest substantial richness which has not yet been formally described. The lack of description of species of Neospirorchis is probably explained by their small, slender, elongate bodies, which allow them to infect numerous organs and vessels in their hosts, such as the heart and peripheral vessels of nervous system, endocrine organs, thymus, mesenteric vessels, and gastrointestinal submucosa. This morphology and site of infection means that collecting good quality, intact specimens is generally difficult, ultimately hampering the formal description of species. Here we supplement limited morphological samples with multi-locus genetic data to formally describe four new species of Neospirorchis infecting marine turtles from Queensland, Australia and Florida, USA; Neospirorchis goodmanorum n. sp. and Neospirorchis deburonae n. sp. are described from Chelonia mydas, Neospirorchis stacyi n. sp. is described from Caretta caretta, and Neospirorchis chapmanae n. sp. from Ch. mydas and Ca. caretta. The four new species are delineated from each other and the two known species based on the arrangement of the male and female reproductive organs, on the basis of cytochrome c oxidase subunit 1 (cox1), internal transcribed spacer 2 (ITS2), and 28S ribosomal DNA (rDNA) molecular data, site of infection, and host species. Molecular evidence for three further putative, presently undescribable, species is also reported. We propose that this integrated characterisation of species of Neospirorchis, based on careful consideration of host, molecular and key morphological data, offers a valuable solution to the slow rate of descriptions for this important genus. We provide the first known life cycle data for Neospirorchis in Australian waters, from Moreton Bay, Queensland; consistent with reports from the Atlantic, sporocysts were collected from a terebellid polychaete and genetically matched to an unnamed species of Neospirorchis infecting Ch. mydas from Queensland and Florida.

Plesiochorus irwinorum n. sp. (Trematoda: Gorgoderidae) from the urinary bladder of the hawksbill turtle, Eretmochelys imbricata (Testudines: Cheloniidae), off the east coast of Australia.

Plesiochorus Looss, 1901 is a genus of Gorgoderidae infecting the urinary bladders of marine turtles globally. Currently, just two morphologically similar species are recognised, Plesiochorus cymbiformis (Rudolphi, 1819) Looss, 1901 and Plesiochorus elongatus Pigulevsky, 1953, which have been distinguished by molecular data and subtle morphological differences. Here we describe a new species, Plesiochorus irwinorum n. sp., infecting hawksbill turtles (Eretmochelys imbricata (L.)), which is primarily distinguished from the other two species of Plesiochorus on the basis of ITS2, cox1 and 28S sequence data. Morphometric data for specimens examined during this study overlap between P. cymbiformis and P. irwinorum n. sp. for every measured feature, rendering them functionally cryptic. However, principal components analysis clearly distinguishes the two species. Additionally, we report new specimens of P. cymbiformis, and provide new sequence data for specimens from Australian loggerhead (Caretta caretta (L.)) and hawksbill turtles. There is little understanding of the host-specificity or geographical distribution of the three species of Plesiochorus, and it remains possible that some of the previously reported sequences have been attributed to the wrong species.

Vermetid gastropods as key intermediate hosts for a lineage of marine turtle blood flukes (Digenea: Spirorchiidae), with evidence of transmission at a turtle rookery.

Blood flukes of the family Spirorchiidae Stunkard, 1921 are significant pathogens of marine turtles, both in the wild and in captivity. Despite causing considerable disease and mortality, little is known about the life cycles of marine species, with just four reports globally. No complete life cycle has been elucidated for any named species of marine spirorchiid, but the group is reported to use vermetid and fissurellid gastropods, and terebelliform polychaetes as intermediate hosts. Here we report molecular evidence that nine related spirorchiid species infect vermetid gastropods as first intermediate hosts from four localities along the coast of Queensland, Australia. ITS2 rDNA and cox1 mtDNA sequence data generated from vermetid infections provides the first definitive identifications for the intermediate hosts for the four species of Hapalotrema Looss, 1899 and Learedius learedi Price, 1934. Additionally, we provide a new locality report for larval stages of Amphiorchis sp., and evidence of three additional unidentified spirorchiid species in Australian waters. Based on the wealth of infections from vermetids during this study, we conclude that the previous preliminary report of a fissurellid limpet as the intermediate host for L. learedi was likely mistaken. The nine species found infecting vermetids during this study form a strongly supported clade exclusive of species of the other two marine spirorchiid genera for which sequence data are available; Carettacola Manter & Larson, 1950 which falls sister to the vermetid-infecting clade + a small clade of freshwater spirorchiids, and Neospirorchis Price, 1934 which is distantly related to the vermetid-infecting clade. We provide further evidence that spirorchiid transmission can occur in closed system aquaria and show that spirorchiid transmission occurs at both an important turtle rookery (Heron Island, southern Great Barrier Reef, Australia) and foraging ground (Moreton Bay, Australia). We discuss the implications of our findings for the epidemiology of the disease, control in captivity, and the evolution of vermetid exploitation by the Spirorchiidae.

Gastropod first intermediate hosts for two species of Monorchiidae Odhner, 1911 (Trematoda): I can't believe it's not bivalves!

The trematode superfamily Monorchioidea comprises three families of teleost parasites: the Monorchiidae Odhner, 1911, Lissorchiidae Magath, 1917, and Deropristidae Cable & Hunninen, 1942. All presently known lissorchiid and deropristid life cycles have gastropods as first intermediate hosts, whereas those of monorchiids involve bivalves. Here, we report an unexpected intermediate host for monorchiids; two species of Hurleytrematoides Yamaguti, 1954 use gastropods as first intermediate hosts. Sporocysts and cercariae were found infecting two species of the family Vermetidae, highly specialised sessile gastropods that form calcareous tubes, from two locations off the coast of Queensland, Australia. These intramolluscan infections broadly corresponded morphologically to those of known monorchiids in that the cercariae have a spinous tegument, oral and ventral suckers, a simple tail and distinct eye-spots. Given the simplified morphology of intramolluscan infections, genetic data provided a definitive identification. ITS2 rDNA and cox1 mtDNA sequence data from the gastropod infections were identical to two species of Hurleytrematoides, parasites of butterflyfishes (Chaetodontidae); Hurleytrematoides loi McNamara & Cribb, 2011 from Moreton Bay (south-eastern Queensland) and Heron Island (southern Great Barrier Reef) and Hurleytrematoides morandi McNamara & Cribb, 2011 from Heron Island. Notably, species of Hurleytrematoides are positioned relatively basal in the phylogeny of the Monorchiidae and are a sister lineage to that of species known to infect bivalves. Thus, the most parsimonious evolutionary hypothesis to explain infection of gastropods by these monorchiids is that basal monorchiids (in our analyses, species of Cableia Sogandares-Bernal, 1959, Helicometroides Yamaguti, 1934 and Hurleytrematoides) will all prove to infect gastropods, suggesting a single host switching event into bivalves for more derived monorchiids (17 other genera in our phylogenetic analyses). A less parsimonious hypothesis is that the infection of vermetids will prove to be restricted to species of Hurleytrematoides, as an isolated secondary recolonisation of gastropods from a bivalve-infecting lineage. Regardless of how their use arose, vermetids represent a dramatic host jump relative to the rest of the Monorchiidae, one potentially enabled by their specialised feeding biology.

A new genus of Bucephalidae Poche, 1907 (Trematoda: Digenea) for three new species infecting the yellowtail pike, Sphyraena obtusata Cuvier (Sphyraenidae), from Moreton Bay, Queensland, Australia.

Three new species of the family Bucephalidae Poche, 1907 (Trematoda: Digenea) are described from the yellowtail pike, Sphyraena obtusata Cuvier (Sphyraenidae), from Moreton Bay, Queensland, Australia. The three species are morphologically consistent with the present broad concept of the genus Bucephalus Baer, 1827, but significant phylogenetic and ecological differences relative to the type-species of Bucephalus require the proposal of a new genus. Aenigmatrema n. g. is proposed for A. undecimtentaculatum n. sp. (type-species), A. inopinatum n. sp. and A. grandiovum n. sp. In addition, based on morphological, ecological and biogeographical similarities, we recombine two existing species of Bucephalus as Aenigmatrema kaku (Yamaguti, 1970) n. comb. and Aenigmatrema sphyraenae (Yamaguti, 1952) n. comb. Although the three species described in this study are extremely morphologically similar, they can be differentiated from each other, and from A. kaku and A. sphyraenae, morphometrically on the basis of egg size, tentacle number and a combination of the caecum and vitelline field lengths. Complete ITS2 rDNA, partial 28S rDNA and partial cox1 mtDNA sequence data were generated for the three new species, which formed a well-supported clade in all 28S phylogenetic analyses. An expanded phylogenetic tree for the subfamily Bucephalinae Poche, 1907 is presented, demonstrating unresolved issues with the morphology-based taxonomy of the subfamily. The three largest genera, Bucephalus, Rhipidocotyle Diesing, 1858 and Prosorhynchoides Dollfus, 1929 remain extensively polyphyletic, indicating the need for significant further systematic revision.