Revision of 13 Genera of Haploporidae (Trematoda)

The Haploporidae is a family of digeneans united by the combination of the possession of a hermaphroditic sac and a single testis or, rarely, two tandem testes. The major divisions in the Haploporidae have been based on the organization, development, and nature of the male and female reproductive systems. Overstreet and Curran (2005) has been the only attempt to organize the genera of the Haploporidae in a subfamilial framework. In the present work the validity of the subfamily Waretrematinae by Overstreet and Curran (2005) is assessed by morphological and molecular methods, based on original descriptions, type and vouchered, specimens and newly collected material for morphology and rDNA sequence data analyses. These analyses conflicted with hypotheses and framework by Overstreet and Curran (2005) in that: (1) Megasoleninae is the basal subfamily within the Haploporidae; (2) Waretrematinae and Haploporinae are not sister groups; (3) species of Spiritestis and Capitimitta, both previously considered members of Waretrema, are not closely related; (4) Waretrematinae was not monophyletic when Unisaccus and New Genus 1, new species are included; (5) species of Unisaccoides and Unisaccus species are closely related; and (6) species of Intromugil are allocated to Chalcinotrematinae rather than Waretrematinae. Reports of Waretrema were determined to comprise members of three different genera. The morphology of Intromugil was accessed by the redescription of I. mugilicolus from newly collected material and a new species is described. Species from the Indo-Pacific region possessing spirally arranged pads in the hermaphroditic duct and a caecum were accessed and required changes to the organization and membership of several genera plus changes to the intergeneric relations within the family based on phylogenetic analysis. Members of the genera Platydidymus and Carassotrema were assessed and a new species was described. Two new genera of haploporid were diagnosed, a new species was described for each, and phylogenetic relations are estimated. A new genus and new species of Megaperidae are described, and molecular data were provided for three other species. Previously, megaperid species were members of the Apocreadiidae rather than the Haploporidae. Phylogenetic hypotheses based on Bayesian Inference analysis of an alignment of partial 28S gene sequences of haploporids provide a framework for the evaluation of the interrelationships within the Haploporidae

Review of Haploporid (Trematoda) Genera with Ornate Muscularization in the Region of the Oral Sucker, Including Four New Species and a New Genus

Species of the Haploporidae Nicoll, 1914 with elaborate muscularization of the oral sucker belong in three trematode genera, including three new species and a new genus from the intestine of fishes in Australian waters. Spiritestis Nagaty, 1948 is resurrected and S. herveyensis n. sp. is described from the mullet Moolgarda seheli (Forsskål) collected in Hervey Bay, Queensland, Australia; the latter differs from S. arabii Nagaty, 1948 in that the position of the genital pore is pharyngeal rather than post-pharyngeal and the geographical range is off Australia rather than the Red Sea. A new genus is proposed for two new species, with a uniquely ornamented oral sucker, which infect Australian scatophagids. Members of Capitimitta n. g. are distinguished from Waretrema Srivastava, 1937, species of which have a simple oral sucker with six radially arranged anterior muscular lobes, in that their oral sucker is V-shaped with six embedded muscular finger-like structures in the anteroventral portion. The relatively small C. darwinensis n. sp., collected from Selenotoca multifasciata (Richardson) at Darwin, Northern Territory, Australia, is distinguished from C. costata n. sp., collected from Scatophagus argus (Linnaeus) in the same locality and S. multifasciata off Brisbane, Australia, and by having smaller eggs, a vitellarium commencing at a level close to the ventral sucker rather than at greater than one ovarian length posterior to the ventral sucker, and shorter tegumental body spines. Sequence data of a c. 2,500 bp region of the 3′ end of 18S, the entire ITS region and the 5′ end of the 28S revealed that Spiritestis and Capitimitta are not as closely related as some morphological features would suggest and are probably not the closest relative of each other. What has been reported as Waretrema piscicolum Srivastava, 1937 probably consists of several species, some in different genera, and one, based on material collected by Dr. Masaaki Machida, is proposed as Spiritestis machidai n. sp. from Crenimugil crenilabis (Forsskål) off Japan. Phylogenetic hypotheses, based on analysis of an alignment of partial 28S sequences with other haploporids, provide a framework for the evaluation of interrelationships within the Haploporidae. These analyses show that: (1) Spiritestis and Capitimitta are supported within the Haploporidae; (2) branches to Forticulcita Overstreet, 1982, Saccocoelioides Szidat, 1954, Spiritestis and Capitimitta create a clade that is sister to haploporines from the Mediterranean Sea; (3) the branch to Saccocoelioides, Spiritestis, and Capitimitta create a polytomy; and (4) the two new species of Capitimitta, plus an immature specimen of an unnamed species, form a monophyletic clade

Description of Three Species of \u3ci\u3eIsorchis\u3c/i\u3e (Digenea: Atractotrematidae) from Australia

Three species of Isorchis Durio and Manter, 1969 are described from Australian waters. Isorchis megas sp. nov. is described from the spotbanded scat, Selenotoca multifasciata (Richardson), off Western Australia (WA) and Northern Territory (NT); Isorchis currani sp. nov. is described from S. multifasciata off NT; and Isorchis anomalus sp. nov. is described from the milkfish, Chanos chanos Forsskål, off WA. Isorchis megas sp. nov. can be differentiated from the other species of Isorchis by possessing a single, large egg that is greater than 20% of the body length; having a shorter body (the largest specimen is less than 500 μm); and utilizing a scatophagid rather than a chanid host. Isorchis currani sp. nov. can be differentiated from all other species of Isorchis in possessing an irregular-shaped genital pore rather than one that is circular to oblong. A Bayesian inference analysis of partial 28S rDNA sequences of the three new species of Isorchis and 30 other haploporoids revealed (1) the monophyly of the Atractotrematidae Yamaguti, 1939, (2) the two species of Isorchis infecting S. multifasciata were each other\u27s closest relative, and (3) that Isorchis was most closely related to Pseudomegasolena Machida and and Komiya, 1976 rather than Atractotrema Goto and Ozaki, 1929 although sequence data are not yet available for a member of Pseudisorchis Ahmad, 1985

Erection of the Haploporid Genus \u3ci\u3eLitosaccus\u3c/i\u3e n. g. and Its Phylogenetic Relationship within the Haploporidae Nicoll, 1914

Litosaccus n. g. is erected for Paralecithobotrys brisbanensis Martin, 1974 n. comb. for which an amended description is given. The new genus is morphologically similar to the haploporine Lecithobotrys Looss, 1902 but with a more elongate and cylindrical body; an infundibuliform oral sucker; a thin-walled hermaphroditic sac; a shallow genital atrium; and unequal, cylindrical, and elongated caeca. It also resembles Pseudolecithobotrys Blasco-Costa, Gibson, Balbuena, Raga & Kostadinova, 2009, but the only member of that genus has a hermaphroditic sac that is twice the length of the ventral sucker, a hermaphroditic duct with intensely staining cuboidal cells, an elongate testis, and single or paired caeca. A Bayesian inference analysis of partial 28S rDNA sequences of L. brisbanensis and 24 other haploporoids revealed that L. brisbanensis grouped with other haploporines and placed Intromugil Overstreet & Curran, 2005 in a clade with the chalcinotrematine Saccocoelioides Szidat, 1954 rather than the other seven tested waretrematine species. This analysis represents the first phylogenetic study of the Haploporidae Nicoll, 1914 that incorporates a haploporine from outside of the Mediterranean Sea

Large Scale Screening of Digeneans for \u3cem\u3eNeorickettsia\u3c/em\u3e Endosymbionts Using Real-Time PCR Reveals New \u3cem\u3eNeorickettsia\u3c/em\u3e Genotypes, Host Associations and Geographic Records

Digeneans are endoparasitic flatworms with complex life cycles including one or two intermediate hosts (first of which is always a mollusk) and a vertebrate definitive host. Digeneans may harbor intracellular endosymbiotic bacteria belonging to the genus Neorickettsia (order Rickettsiales, family Anaplasmataceae). Some Neorickettsia are able to invade cells of the digenean\u27s vertebrate host and are known to cause diseases of wildlife and humans. In this study we report the results of screening 771 digenean samples for Neorickettsia collected from various vertebrates in terrestrial, freshwater, brackish, and marine habitats in the United States, China and Australia. Neorickettsia were detected using a newly designed real-time PCR protocol targeting a 152 bp fragment of the heat shock protein coding gene, GroEL, and verified with nested PCR and sequencing of a 1371 bp long region of 16S rRNA. Eight isolates of Neorickettsia have been obtained. Sequence comparison and phylogenetic analysis demonstrated that 7 of these isolates, provisionally named Neorickettsia sp. 1–7 (obtained from allocreadiid Crepidostomum affine, haploporids Saccocoelioides beauforti and Saccocoelioides lizae, faustulid Bacciger sprenti, deropegid Deropegus aspina, a lecithodendriid, and a pleurogenid) represent new genotypes and one (obtained from Metagonimoides oregonensis) was identical to a published sequence of Neorickettsia known as SF agent. All digenean species reported in this study represent new host records. Three of the 6 digenean families (Haploporidae, Pleurogenidae, and Faustulidae) are also reported for the first time as hosts of Neorickettsia. We have detected Neorickettsia in digeneans from China and Australia for the first time based on PCR and sequencing evidence. Our findings suggest that further surveys from broader geographic regions and wider selection of digenean taxa are likely to reveal new Neorickettsia lineages as well as new digenean host associations

Molecular Characterization of Two Opecoelid Trematodes From Fishes in the Gulf of Mexico, With a Desceiption of a New Species of \u3ci\u3eHelicometra\u3c/i\u3e

The plagioporine opecoelids Helicometra fasciata (Rudolphi, 1819) Odhner, 1902, and Macvicaria crassigula (Linton, 1910) Bartoli, Bray, and Gibson, 1989 have been reported from fishes in expansive geographic regions, disjointed from their type localities. New material of M. crassigula was collected from near its type locality as well as specimens resembling Helicometra fasciata sensu lato from three triglids in the Gulf of Mexico. Comparisons of the ribosomal DNA (rDNA) sequences, comprising the partial 18S rDNA, internal transcribed spacer region (= ITS1, 5.8S, and ITS2), and partial 28S rDNA gene, from M. crassigula and Helicometra fasciata sensu lato in the Gulf of Mexico were made with sequences deposited in GenBank from those species from the Mediterranean Sea. Results reveal that M. crassigula sensu stricto from the Gulf of Mexico is distinct from the two cryptic species of M. crassigula sensu lato from the Mediterranean Sea and Helicometra fasciata sensu lato in this study differs from H. fasciata sequences from the Mediterranean Sea, thus Helicometra manteri sp. nov. is described

Review of Haploporid (Trematoda) Genera with Ornate Muscularization in the Region of the Oral Sucker, Including Four New Species and a New Genus

Species of the Haploporidae Nicoll, 1914 with elaborate muscularization of the oral sucker belong in three trematode genera, including three new species and a new genus from the intestine of fishes in Australian waters. Spiritestis Nagaty, 1948 is resurrected and S. herveyensis n. sp. is described from the mullet Moolgarda seheli (Forsskål) collected in Hervey Bay, Queensland, Australia; the latter differs from S. arabii Nagaty, 1948 in that the position of the genital pore is pharyngeal rather than post-pharyngeal and the geographical range is off Australia rather than the Red Sea. A new genus is proposed for two new species, with a uniquely ornamented oral sucker, which infect Australian scatophagids. Members of Capitimitta n. g. are distinguished from Waretrema Srivastava, 1937, species of which have a simple oral sucker with six radially arranged anterior muscular lobes, in that their oral sucker is V-shaped with six embedded muscular finger-like structures in the anteroventral portion. The relatively small C. darwinensis n. sp., collected from Selenotoca multifasciata (Richardson) at Darwin, Northern Territory, Australia, is distinguished from C. costata n. sp., collected from Scatophagus argus (Linnaeus) in the same locality and S. multifasciata off Brisbane, Australia, and by having smaller eggs, a vitellarium commencing at a level close to the ventral sucker rather than at greater than one ovarian length posterior to the ventral sucker, and shorter tegumental body spines. Sequence data of a c. 2,500 bp region of the 3′ end of 18S, the entire ITS region and the 5′ end of the 28S revealed that Spiritestis and Capitimitta are not as closely related as some morphological features would suggest and are probably not the closest relative of each other. What has been reported as Waretrema piscicolum Srivastava, 1937 probably consists of several species, some in different genera, and one, based on material collected by Dr. Masaaki Machida, is proposed as Spiritestis machidai n. sp. from Crenimugil crenilabis (Forsskål) off Japan. Phylogenetic hypotheses, based on analysis of an alignment of partial 28S sequences with other haploporids, provide a framework for the evaluation of interrelationships within the Haploporidae. These analyses show that: (1) Spiritestis and Capitimitta are supported within the Haploporidae; (2) branches to Forticulcita Overstreet, 1982, Saccocoelioides Szidat, 1954, Spiritestis and Capitimitta create a clade that is sister to haploporines from the Mediterranean Sea; (3) the branch to Saccocoelioides, Spiritestis, and Capitimitta create a polytomy; and (4) the two new species of Capitimitta, plus an immature specimen of an unnamed species, form a monophyletic clade

Description of Three Species of \u3ci\u3eIsorchis\u3c/i\u3e (Digenea: Atractotrematidae) from Australia

Three species of Isorchis Durio and Manter, 1969 are described from Australian waters. Isorchis megas sp. nov. is described from the spotbanded scat, Selenotoca multifasciata (Richardson), off Western Australia (WA) and Northern Territory (NT); Isorchis currani sp. nov. is described from S. multifasciata off NT; and Isorchis anomalus sp. nov. is described from the milkfish, Chanos chanos Forsskål, off WA. Isorchis megas sp. nov. can be differentiated from the other species of Isorchis by possessing a single, large egg that is greater than 20% of the body length; having a shorter body (the largest specimen is less than 500 μm); and utilizing a scatophagid rather than a chanid host. Isorchis currani sp. nov. can be differentiated from all other species of Isorchis in possessing an irregular-shaped genital pore rather than one that is circular to oblong. A Bayesian inference analysis of partial 28S rDNA sequences of the three new species of Isorchis and 30 other haploporoids revealed (1) the monophyly of the Atractotrematidae Yamaguti, 1939, (2) the two species of Isorchis infecting S. multifasciata were each other\u27s closest relative, and (3) that Isorchis was most closely related to Pseudomegasolena Machida and and Komiya, 1976 rather than Atractotrema Goto and Ozaki, 1929 although sequence data are not yet available for a member of Pseudisorchis Ahmad, 1985

Confirmation of \u3ci\u3ePseudolepidapedon balistis\u3c/i\u3e In the Acanthocolpidae (Digenea) Based On Phylogenetic Analysis of Ribosomal DNA

Psuedolepidapedon balistis Manter, 1940 is reported from the grey triggerfish, Balistis capriscus Gmelin, 1789, from off Destin, Florida in the northern Gulf of Mexico. A fragment of ribosomal DNA comprising a short portion of the 3\u27 end of 18S nuclear rDNA gene internal transcribed spacer (ITS) region (=ITSI + 5.85 rDNA gene + ITS2) and the 5\u27 end of the 228S rDNA gene (including variable domains D1-D3) was sequenced for the species and deposited in GenBank. The fragment of 28S rDNA gene was aligned with similar fragments from 25 other digenean species, including 14 species of acanthocolpids, and a Bayesian Inference phylogenetic analysis was conducted on the alignment. The resulting phylogram confirmed that Pseudolepidapedon Yamaguti, 1938 belongs in the family Acanthocolpidae Lühe, 1906. Neoapocreadium viguerasi (Zhukov, 1983) Ahmad, 1987 is considered a junior synonym of P. balistis

Erection of the Haploporid Genus \u3ci\u3eLitosaccus\u3c/i\u3e n. g. and Its Phylogenetic Relationship within the Haploporidae Nicoll, 1914

Litosaccus n. g. is erected for Paralecithobotrys brisbanensis Martin, 1974 n. comb. for which an amended description is given. The new genus is morphologically similar to the haploporine Lecithobotrys Looss, 1902 but with a more elongate and cylindrical body; an infundibuliform oral sucker; a thin-walled hermaphroditic sac; a shallow genital atrium; and unequal, cylindrical, and elongated caeca. It also resembles Pseudolecithobotrys Blasco-Costa, Gibson, Balbuena, Raga & Kostadinova, 2009, but the only member of that genus has a hermaphroditic sac that is twice the length of the ventral sucker, a hermaphroditic duct with intensely staining cuboidal cells, an elongate testis, and single or paired caeca. A Bayesian inference analysis of partial 28S rDNA sequences of L. brisbanensis and 24 other haploporoids revealed that L. brisbanensis grouped with other haploporines and placed Intromugil Overstreet & Curran, 2005 in a clade with the chalcinotrematine Saccocoelioides Szidat, 1954 rather than the other seven tested waretrematine species. This analysis represents the first phylogenetic study of the Haploporidae Nicoll, 1914 that incorporates a haploporine from outside of the Mediterranean Sea