Identical digeneans in coral reef fishes from French Polynesia and the Great Barrier Reef (Australia) demonstrated by morphology and molecules

Three coral reef fish species, Zanclus cornutus, Chaetodon vagabundus and Naso lituratus, were collected in French Polynesia and on the Great Barrier Reef, Queensland. These fish species were each infected by one morphologically similar digenean species in both localities; Schistorchis Zancli Hanson, 1953 was found in Zanclus cornutus. Preptetos laguncula Bray and Cribb, 1996 in Naso lituratus and Neohypocreadium dorsoporum Machida and Uchida, 1987 in Chaetodon vagabundus. In addition, on the Great Barrier Reef P. laguncula was also found in Naso unicornis and N. dorsoporum in Chaetodon ephippium and Chaetodon flavirostris. Morphometric differences between the species from the two sites were only slight. Sequences from the second internal transcribed spacer of the ribosomal DNA of each worm revealed total homology or negligible divergence between samples from hosts caught in French Polynesia and on the Great Barrier Reef. These results show that across more than 6000 km these digeneans are similar in morphology and genotype. Some species of fishes and molluscs a-re considered to have distributions that encompass the entire tropical Indo-West Pacific. These findings suggest that at least some of their parasites have similarly broad distributions. (C) 2001 Australian Society for Parasitology Inc. Published by Elsevier Science Ltd. All rights reserved.

The use and implications of ribosomal DNA sequencing for the discrimination of digenean species

In just over a decade, the use of molecular approaches for the recognition of parasites has become commonplace. For trematodes, the internal transcribed spacer region of ribosomal DNA (ITS rDNA) has become the default region of choice. Here, we review the findings of 63 studies that report ITS rDNA sequence data for about 155 digenean species from 19 families, and then review the levels of variation that have been reported and how the variation has been interpreted. Overall, complete ITS sequences (or ITS1 or ITS2 regions alone) usually distinguish trematode species clearly, including combinations for which morphology gives ambiguous results. Closely related species may have few base differences and in at least one convincing case the ITS2 sequences of two good species are identical. In some cases, the ITS1 region gives greater resolution than the ITS2 because of the presence of variable repeat units that are generally lacking in the ITS2. Intraspecific variation is usually low and frequently apparently absent. Information on geographical variation of digeneans is limited but at least some of the reported variation probably reflects the presence of multiple species. Despite the accepted dogma that concerted evolution makes the individual representative of the entire species, a significant number of studies have reported at least some intraspecific variation. The significance of such variation is difficult to assess a posteriori, but it seems likely that identification and sequencing errors account for some of it and failure to recognise separate species may also be significant. Some reported variation clearly requires further analysis. The use of a yardstick to determine when separate species should be recognised is flawed. Instead, we argue that consistent genetic differences that are associated with consistent morphological or biological traits should be considered the marker for separate species. We propose a generalised approach to the use of rDNA to distinguish trematode species.

Spermiogenesis and ultrastructure of the spermatozoon of Wardula capitellata (Digenea, Mesometridae), an intestinal parasite of the sparid teleost Sarpa salpa in Senegal

The spermiogenesis process in Wardula capitellata begins with the formation of a differentiation zone containing two centrioles associated with striated rootlets and an intercentriolar body. Each centriole develops into a free flagellum orthogonal to a median cytoplasmic process. Later these flagella rotate and become parallel to the median cytoplasmic process, which already exhibits two electron-dense areas and spinelike bodies before its proximodistal fusion with the flagella. The final stage of the spermiogenesis is characterized by the constriction of the ring of arched membranes, giving rise to the young spermatozoon, which detaches from the residual cytoplasm. The mature spermatozoon of W. capitellata presents most of the classical characters reported in digenean spermatozoa such as two axonemes of different lengths of the 9 + '1' trepaxonematan pattern, nucleus, mitochondrion, two bundles of parallel cortical microtubules and granules of glycogen. However, some peculiarities such as two lateral expansions accompanied by external ornamentation of the plasma membrane and spinelike bodies characterize the mature sperm. Moreover, a new spermatological character is described for the first time, the so-called cytoplasmic ornamented buttons.

Spermiogenesis and ultrastructure of the spermatozoon of Wardula capitellata (Digenea, Mesometridae), an intestinal parasite of the sparid teleost Sarpa salpa in Senegal

The spermiogenesis process in Wardula capitellata begins with the formation of a differentiation zone containing two centrioles associated with striated rootlets and an intercentriolar body. Each centriole develops into a free flagellum orthogonal to a median cytoplasmic process. Later these flagella rotate and become parallel to the median cytoplasmic process, which already exhibits two electron-dense areas and spinelike bodies before its proximodistal fusion with the flagella. The final stage of the spermiogenesis is characterized by the constriction of the ring of arched membranes, giving rise to the young spermatozoon, which detaches from the residual cytoplasm. The mature spermatozoon of W. capitellata presents most of the classical characters reported in digenean spermatozoa such as two axonemes of different lengths of the 9 + '1' trepaxonematan pattern, nucleus, mitochondrion, two bundles of parallel cortical microtubules and granules of glycogen. However, some peculiarities such as two lateral expansions accompanied by external ornamentation of the plasma membrane and spinelike bodies characterize the mature sperm. Moreover, a new spermatological character is described for the first time, the so-called cytoplasmic ornamented buttons.

Molecules and morphology in phylogenetic studies of the Hemiuroidea (Digenea : Trematoda : Platyhelminthes)

Phylogenies of trematodes based on characters derived from morphology and life cycles have been controversial. Here, we add molecular data to the phylogenetic study of a group of trematodes, members of the superfamily Hemiuroidea Looss, 1899. DNA sequences from the V4 domain of the nuclear small subunit (18S) rRNA gene and a matrix of morphological characters modified from a previous study were used. There was no significant incongruence between the molecular and the morphological data. However, this was probably due largely to the limited resolving power of the morphological data. Analyses support a monophyletic Hemiuroidea containing at least the families Accacoeliidae, Derogenidae, Didymozoidae, Hirudinellidae, Sclerodistomidae, Syncoeliidae, Isoparorchiidae, Lecithasteridae, and Hemiuridae. These families fall into two principal clades. One contains the first six families and the other the Hemiuridae and lecithasterine lecithasterids. The positions of the hysterolecithine lecithasterids and the Isoparorchiidae were poorly resolved. The Ptychogonimidae may be the sister group of the remaining Hemiuroidea, but there was no support from the molecular data for the placement of the Azygiidae within the superfamily. (C) 1998 Academic Press.

A review of the Apocreadiidae Skrjabin, 1942 (Trematoda : Digenea) and description of Australian species

The Apocreadiidae is reviewed and is considered to include genera recognised previously within the families Apocreadiidae, Homalometridae, Schistorchiidae, Sphincterostomatidae and Trematobrienidae. Key features of the family are extensive vitelline follicles, eye-spot pigment dispersed in forebody, I-shaped excretory vesicle, no cirrus-sac and genital pore opening immediately anterior to the ventral sucker (usually) or immediately posterior to it (Postporus Manter, 1949). Three subfamilies and 18 genera are recognised within the Apocreadiidae. The Apocreadiinae comprises Homalometron Stafford, 1904 (new syn. Barbulostomum Ramsey, 1965), Callohelmis n. g., Choanodera Manter, 1940, Crassicutis Manter, 1936, Dactylotrema Bravo-Hollis & Manter, 1957, Marsupioacetabulum Yamaguti, 1952, Microcreadium Simer, 1929, Myzotus Manter, 1940, Neoapocreadium Siddiqi & Cable, 1960, Neomegasolena Siddiqi & Cable, 1960, Pancreadium Manter, 1954, Procaudotestis Szidat, 1954 and Trematobrien Dollfus, 1950. The Schistorchiinae comprises Schistorchis Luhe, 1906, Sphincterostoma Yamaguti, 1937, Sphincteristomum Oshmarin, Mamaev & Parukhin, 1961 and Megacreadium Nagaty, 1956. The Postporinae comprises only Postporus. A key to subfamilies and genera of the Apocreadiidae is provided. It is argued that there is no convincing basis for the recognition of the genus Apocreadium Manter, 1937 and all its constituent species are combined with Homalometron. The following new combinations are proposed for species previously recognised within Apocreadium: Homalometron balistis (Manter, 1947), H. caballeroi (Bravo-Hollis, 1953), H. cryptum (Overstreet, 1969), H. longisinosum (Manter, 1937), H. manteri (Overstreet, 1970), H. mexicanum (Manter, 1937) and H. vinodae (Ahmad, 1985). Apocreadium uroproctoferum Sogandares-Bernal, 1959 is found to lack a uroproct and is made a synonym of H. mexicanum. Homalometron verrunculi nom. nov. is proposed to replace the secondarily pre-occupied H. caballeroi Lamothe-Argumedo, 1965. Barbulostomum is made a synonym of Homalometron and H. cupuloris (Ramsey, 1965) n. comb. is proposed. Neochoanodera is made a synonym of Choanodera and Choanodera ghanensis (Fischthal & Thomas, 1970) n. comb. is proposed. Species within the Apocreadiinae and Postporinae are reviewed and the following are recorded or described from Australian fishes: Homalometron wrightae n. sp. from Achlyopa nigra (Macleay), H. synagris (Yamaguti, 1953) n. comb. from Scolopsis monogramma (Cuvier), H. stradbrokensis n. sp. from Gerres subfasciatus Cuvier, Marsupioacetabulum opallioderma n. sp. from G. subfasciatus, Neoapocreadium karwarensis (Hafeezullah, 1970) n. comb. from G. subfasciatus, N. splendens n. sp. from S. monogramma and Callohelmis pichelinae n. g., n. sp. from Hemigymnus melapterus (Bloch), H. fasciatus (Bloch), Stethojulis bandanensis (Bleeker) andChoerodon venustus (De Vis). Callohelmis is recognised by the combination of absence of tegumental spines, caeca terminating midway between the testes and posterior end of body, ventral sucker enclosed in a tegumental pouch, prominent muscles radiating through the body from the ventral sucker, vitelline follicles not extending into the forebody, and a very short excretory vesicle that opens ventrally. New combinations for species previously recognised within Crassicutis are proposed as follows: Neoapocreadium caranxi (Bilqees, 1976) n. comb., N. gerridis (Nahhas & Cable, 1964) n. comb., N. imtiazi (Ahmad, 1984) n. comb. and N. marina (Manter, 1947) n. comb. The host-specificity and zoogeography of the Apocreadiinae are considered.

A new species, Pretestis laticaecum, (Trematoda : Cladorchiidae), from Emydura krefftii Gray, 1871 (Pleurodira : Chelidae) from Central Queensland, Australia

Pretestis laticaecum is described from the small intestine of the freshwater turtle Emydura krefftil. The new species can be distinguished from its congener P. australianus by the following characters; significantly smaller ovary, main lymph vessels reach anterior to posterior testis, genital atrium in mid-oesophageal region, small vitelline follicles clumped around the ovary and significantly larger caeca overlapping. The, position of this species and related genera in fish, the life cycle of P. australianus and the presence of P. laticaecum in turtles suggest that it is a relatively recent host capture.

The nature and evolution of the association among digeneans, molluscs and fishes

Patterns of association of digenean families and their mollusc and vertebrate hosts are assessed by way of a new database containing information on over 1000 species of digeneans for lift-cycles and over 5000 species from fishes. Analysis of the distribution of digenean families in molluscs suggests that the group was associated primitively with gastropods and that infection of polychaetes, bivalves and scaphopods are all the results of host-switching. For the vertebrates. infections of agnathans and chondrichthyans are apparently the result of host-switching from teleosts. For digenean families the ratio of orders of fishes infected to superfamilies of molluscs infected ranges from 0.5 (Mesometridae) to 16 (Bivesiculidae) and has a mean of 5.6. Individual patterns of host association of 13 dipenean families and superfamilies are reviewed. Two, Bucephalidae and Sanguinicolidae. are exceptional in infecting a range of first intermediate hosts qualitatively as broad as their range of definitive hosts. No well-studied taxon shows narrower association with vertebrate than with mollusc clades. The range of definitive hosts of digeneans is characteristically defined by eco-physiological similarity rather than phylogenetic relationship. The range of associations of digenean families with mollusc taxa is generally much narrower. These data are considered in the light of ideas about the significance of different forms of host association. If Manter's Second Rule (the longer the association with a host group, the mure pronounced the specificity exhibited by the parasite group) is invoked, then the data may suggest that the Digenea first parasitised molluscs before adopting vertebrate hosts. This interpretation is consistent with most previous ideas about the evolution of the Digenea but contrary to current interpretations based on the monophyly of the Neodermata. The basis of Manter's Second Rule is. however, considered too flimsy for this interpretation to be robust. Problems of the inference of the evolution of patterns of parasitism in the Neodermata al-e discussed and considered so intractable that the truth may be presently unknowable. (C) 2001 Australian Society for Parasitology Inc. Published by Elsevier Science Ltd. All rights reserved.

Complete DNA sequence and gene organization of the mitochondrial genome of the liverfluke, Fasciola hepatica L. (Platyhelminthes; Trematoda)

The complete nucleotide sequence of the mitochondrial (mt) DNA molecule of the liverfluke, Fasciola hepatica (phylum Platyhelminthes, class Trematoda, family Fasciolidae), was determined, It comprises 14462 bp, contains 12 protein-encoding, 2 ribosomal and 22 transfer RNA genes, and is the second complete flatworm (and the first trematode) mitochondrial sequence to be described in detail. All of the genes are transcribed from the same strand. Of the genes typically found in mitochondrial genomes of eumetazoans, only atp8 is absent. The nad4L and nad4 genes overlap by 40 nt. Most intergenic sequences are very short. Two larger non-coding regions are present. The longer one (817 nt) is located between trnG and cox3 and consists of 8 identical tandem repeats of 85 nt, rich in G and C, followed by 1 imperfect repeat. The shorter non-coding region (187 nt) exhibits no special features and is separated from the longer region by trnG. The gene arrangement resembles that of some other trematodes including the eastern Asian Schistosoma species (and cyclophyllidean cestode species) but it is strikingly different from that of the African schistosomes, represented by Schistosoma mansoni. The genetic code is as inferred previously for flatworms. Transfer RNA genes range in length from 58 to 70 nt, their products producing characteristic 'clover leaf' structures, except for tRNA(S-VON) and tRNA(S-AGN) lacking the DHU arm.

Phylogeny and classification of the Digenea (Platyhelminthes : Trematoda)

Complete small subunit ribosomal RNA gene (ssrDNA) and partial (D1-D3) large subunit ribosomal RNA gene (lsrDNA) sequences were used to estimate the phylogeny of the Digenea via maximum parsimony and Bayesian inference. Here we contribute 80 new ssrDNA and 124 new lsrDNA sequences. Fully complementary data sets of the two genes were assembled from newly generated and previously published sequences and comprised 163 digenean taxa representing 77 nominal families and seven aspidogastrean outgroup taxa representing three families. Analyses were conducted on the genes independently as well as combined and separate analyses including only the higher plagiorchiidan taxa were performed using a reduced-taxon alignment including additional characters that could not be otherwise unambiguously aligned. The combined data analyses yielded the most strongly supported results and differences between the two methods of analysis were primarily in their degree of resolution. The Bayesian analysis including all taxa and characters, and incorporating a model of nucleotide substitution (general-time-reversible with among-site rate heterogeneity), was considered the best estimate of the phylogeny and was used to evaluate their classification and evolution. In broad terms, the Digenea forms a dichotomy that is split between a lineage leading to the Brachylaimoidea, Diplostomoidea and Schistosomatoidea (collectively the Diplostomida nomen novum (nom. nov.)) and the remainder of the Digenea (the Plagiorchiida), in which the Bivesiculata nom. nov. and Transversotremata nom. nov. form the two most basal lineages, followed by the Hemiurata. The remainder of the Plagiorchiida forms a large number of independent lineages leading to the crown clade Xiphidiata nom. nov. that comprises the Allocreadioidea, Gorgoderoidea, Microphalloidea and Plagiorchioidea, which are united by the presence of a penetrating stylet in their cercariae. Although a majority of families and to a lesser degree, superfamilies are supported as currently defined, the traditional divisions of the Echinostomida, Plagiorchiida and Strigeida were found to comprise non-natural assemblages. Therefore, the membership of established higher taxa are emended, new taxa erected and a revised, phylogenetically based classification proposed and discussed in light of ontogeny, morphology and taxonomic history. (C) 2003 Australian Society for Parasitology Inc. Published by Elsevier Science Ltd. All rights reserved.

Ocorrência no Brasil de Thiara (Melanoides) tuberculata (O.F. Muller, 1774) (Gastropoda, Prosobranchia), primeiro hospedeiro intermediário de Clonorchis sinensis (Cobbold, 1875) (Trematoda, Plathyhelmintes)

Relata-se a introdução em nosso meio de Thiara (Melanoides) tuberculata possivelmente anterior a 1967, quando pela primeira vez foi coletada em Santos. Essa espécie pode comportar-se como primeiro hospedeiro intermediário de Paragonimus westermani e de Clonorchis sinensis. Somente o último trematódeo tem alguma possibilidade de vir a parasitar o homem, em nosso meio.

Infeccion natural de Ptychophallus tristani (Crustacea: decapoda) con metacercarias de Paragonimus mexicanus (Trematoda) en Tabarcia de Mora, Costa Rica

Se determinó el grado de infección de los crustáceos Ptychophallus tristani presentes en una área endémica de Paragonimus mexicanus, encontrándose que de un total de 182 cangrejos, 161 resultaron positivos (88,5%) por metacercarias. Al comparar la infección entre machos y hembras no se encontró diferencias estadísticamente significativas y en relación a su tamaño se observó una relación directa entre tamaño y porcentaje y grado de infección. Se estableció que no existen diferencias significativas en relación a la época del año en que los cangrejos fueron capturados lo que epidemiológicamente es importante pues indica que las fuentes de infección permanecen potencialmente activas durante todo el año.

Human parasitism by Phagicola sp (Trematoda, Heterophyidae) in Cananéia, São Paulo State, Brazil

We report one case of parasitism by Phagicola sp. (Trematoda, Heterophyidae) in a 31 years-old woman who, in 1987, travelled and stayed several months in the municipality of Cananéia (SP), where she ingested, in various occasions, raw mullet (Mugil sp.). The patient refered mild intestinal pain and laboratory examinations showed eggs of Phagicola sp. in the stools and a slight increase in eosinophil blood levels (8%). After treatment with praziquantel (75 mg/kg per day for three days) all the symptoms and signs disappeared. This is, certainly, the first record of human infection by Phagicola sp. in Brazil and, perhaps, in countries other than the U.S.A. where unclear references to a few human cases were reported in the South-eastern region.

Melanoides tuberculatus (Gastropoda: Thiaridae) as intermediate host of Heterophyidae (Trematoda: Digenea) in Rio de Janeiro metropolitan area, Brazil

In the late 1960s, Melanoides tuberculatus snails were introduced in Brazil from North/East Africa and Southeast Asia. The first records of specimens infected with cercariae were registered in Rio de Janeiro State in 2001. The present study reports the occurrence of M. tuberculatus infected with larval trematodes in Rio de Janeiro City. Bottom sediment was collected with dip nets and sieved through 0.25 inch-mesh screening. Snails were transported to the laboratory in vials with stream water, then measured and individually isolated in glass vials with distilled water. They were exposed to artificial light and temperature to induce cercarial emergence. The most actively emerging cercariae were processed by differential staining and silver nitrate impregnation methods. Negative snails were subsequently dissected. Approximately 700 snails were collected. Snail total lengths ranged from 1.2 to 3.3 cm. The prevalence rate was 15.76% although 53.76% of the snails were found infected in one of the sites. Infected snails were infected with rediae and pleurolophocercous cercariae. Cercarial morphology and chaetotaxy were consistent with those of the family Heterophyidae mostly due to the presence of median dorsal and ventral fins on the tail and the absence of CI dorsal sensory receptors.

Melanoides tuberculata (Mollusca: Thiaridae) as an intermediate host of Centrocestus formosanus (Trematoda: Heterophyidae) in Brazil

Pleurolophocercous cercariae emerged from naturally infected Melanoides tuberculata from Minas Gerais State, Brazil, were used to perform experimental infection of laboratory-reared Poecilia reticulata. Mature metacercariae were obtained from the gills of fishes and force-fed to Mus musculus. The adult parasites which recovered from small intestines of mice were identified as Centrocestus formosanus. This is the first report of M. tuberculata as intermediate host of this heterophyid in Brazil.