Case Report: The First Direct Evidence of Gnathostoma spinigerum Migration through Human Lung.

Gnathostomiasis is a helminthic infection caused by the third-stage larvae of nematodes of the genus Gnathostoma. The life cycle in humans starts with an enteric phase, with the worm perforating the gastric or intestinal mucosa to reach the peritoneal cavity and migrating through the human body. Subsequent penetration through the diaphragm may produce pleuropulmonary symptoms. We herein present a previously healthy 56-year-old Thai man from Southern Thailand who was an ex-smoker presented with chronic dry cough progressing to hemoptysis after consuming grilled swamp eels and freshwater fish. Chest computed tomography showed consolidation at the lingular segment, and the differential diagnosis was primary lung cancer and pulmonary tuberculosis. The lung tissue biopsied during bronchoscopy displayed segments of organisms with the phenotypic characteristics of Gnathostoma spp., and abundant eosinophils were seen in the alveolar tissue. Gnathostoma spinigerum infection was confirmed by a Western blot assay for G. spinigerum-specific 24-kDa reactive band. The patient received albendazole, and a follow-up chest radiograph revealed improvement in the consolidation in the lung and reduction in hemoptysis. We report the first direct evidence including pathology and immunohistochemistry of Gnathostoma invasion via the human lung, with clinical and radiographic presentations mimicking either malignancy or chronic infection.

Entamoeba chiangraiensis n. sp. (Amoebozoa: Entamoebidae) isolated from the gut of Asian swamp eel (Monopterus albus) in northern Thailand.

The genus Entamoeba comprises mostly gut parasites and commensals of invertebrate and vertebrate animals including humans. Herein, we report a new species of Entamoeba isolated from the gut of Asian swamp eels (Monopterus albus) in northern Thailand. Morphologically, the trophozoite is elongated and has a single prominent pseudopodium with no clear uroid. The trophozoite is actively motile, 30-50 µm in length and 9-13 µm in width. Observed cysts were uninucleate, ranging in size from 10 to 17.5 µm in diameter. Chromatin forms a fine, even lining along the inner nuclear membrane. Fine radial spokes join the karyosome to peripheral chromatin. Size, host and nucleus morphology set our organism apart from other members of the genus reported from fish. The SSU rRNA gene sequences of the new isolates are the first molecular data of an Entamoeba species from fish. Phylogenetic analysis places the new organism as sister to Entamoeba invadens. Based on the distinct morphology and SSU rRNA gene sequence we describe it as a new species, Entamoeba chiangraiensis.

Fish Parasite Dinoflagellates Haidadinium ichthyophilum and Piscinoodinium Share a Recent Common Ancestor.

The dinoflagellate Haidadinium ichthyophilum Buckland-Nicks, Reimchen and Garbary 1997 is an ectoparasite of the spine-deficient, three-spine stickleback Gasterosteus aculeatus L. Reimchen 1984, a fish endemic to Rouge Lake, Haida Gwaii. Haidadinium ichthyophilum proved difficult to assign taxonomically because its morphology and complex life cycle exhibited defining characteristics of both autotrophic and heterotrophic dinoflagellates, and was tentatively assigned to the Phytodiniales. Here, we characterized a 492 bp fragment of the small subunit ribosomal RNA (SSU rRNA) from preserved H. ichthyophilum cysts. In SSU phylogeny, H. ichthyophilum branches with the fish parasites, Piscinoodinium sp., strongly supporting the inclusion of H. ichthyophilum within the Suessiales.

Moderate pathogenic effect of Ligophorus uruguayense (Monogenoidea, Ancyrocephalidae) in juvenile mullet Mugil liza (Actinopterygii, Mugilidae) from Brazil

ABSTRACT Monogenoidea pathogenic activity can elicit various histological responses in fish. Species of Ligophorus are specific parasites of mullets, and its relationship with host fish may result in a moderate pathogenic action. In order to ascertain this relationship, estuarine mullets (Mugil liza) were collected in an estuary, reared in laboratory, for three weeks, and forwarded for histological and parasitological analyses. Ligophorus uruguayense (Monogenoidea) infestation in the gills of the mullets was identified. The severe infestation by only one species of Monogenoidea may result from the specificity of these parasites to mullets. Mullets submitted to histological analysis exhibited respiratory epithelium detachment; mild, moderate and severe hyperplasia of the respiratory epithelium; atrophy; and telangiectasia of the gills. This is the first study reporting that mullets highly infested by Monogenoidea can show mild (100%) to severe (20%) gill changes with a distinct frequency of occurrence. Because of the high prevalence of mild alterations observed, it is possible to accept that L. uruguayense is moderately pathogenic to M. liza, even during high prevalence and intensity of infestation, as a result of its specificity.

Parasitological Analysis and Gill Histopathology of Pearl Dace (Semotilus Margarita) and Brook Stickleback (Culaea Inconstans) Collected from the Athabasca Oil Sands Area (Canada).

Pearl dace (Semotilus margarita) and brook stickleback (Culaea inconstans) were collected from tributaries of the Athabasca River (Alberta, Canada), upstream (reference site) and downstream of oil sands deposits where fish were expected to be exposed to naturally occurring oil sands constituents. The objective was to determine if fish collected from these sites exhibited differences in the prevalence or intensity of infection by parasites or in gill histology. Dace did not display significant differences in these parameters. Alternately, upstream stickleback were predominantly infected by complex life history parasites, while downstream fish were primarily infected by parasites with simpler life histories. Moreover, downstream stickleback exhibited significantly more clubbing and aneurysms in secondary gill lamellae relative to upstream fish. This suggested a difference in habitat quality between upstream and downstream sites. However, based on basic body condition parameters of the fish, it would appear that any impacts upon the health of the fish due to the presence of naturally occurring oil sands associated chemical constituents would have been minor.

Parasites contribute to ecologically dependent postmating isolation in the adaptive radiation of three-spined stickleback.

Spatial variation in parasitic infections is common, and has the potential to drive population divergence and the reproductive isolation of hosts. However, despite support from theory and model laboratory systems, little strong evidence has been forthcoming from the wild. Here, we show that parasites are likely to cause reproductive isolation in the adaptive radiation of three-spined stickleback. Adjacent wild populations on the Scottish island of North Uist differ greatly and consistently in the occurrence of different parasites that have substantial effects on fitness. Laboratory-reared fish are more resistant to experimental infection by parasite species from their own population. Furthermore, hybrid backcrosses between the host populations are more resistant to parasites from the parental population to which they are more closely related. These patterns provide strong evidence that parasites can cause ecological speciation, by contributing to selection against migrants and ecologically dependent postmating isolation.

Molecular characterization and morphological aspects of Myxobolus parvus (Myxozoa) from Liza saliens (Mugilidae) off the Turkish Black Sea coasts.

Members of the phylum Myxozoa are among the most cosmopolitan parasites of fish, and the genus Myxobolus is the largest within the class Myxosporea in freshwater and marine environment from all over the world. Myxobolus parvus has been reported from mainly mugilid fishes from different localities including the Black Sea. Leaping mullet Liza saliens collected by fishermen off Sinop coasts of the Black Sea in the period between September 2014 and December 2015 was investigated for myxosporean parasites using conventional methods. M. parvus Schulman, 1962 was the only Myxobolus species identified and it was found in the gills, gall bladder, kidney tubules, and inside several cysts located on the lower jaw of examined fish in the present study. Morphometric data and all details of parasitic infection were determined and presented in tables and illustrated in figures. Molecular characterization was also conducted to reveal its position within Myxosporea. Two M. parvus samples from gills and kidney showed the same haplotype and appeared in the same sub-lineage with marine Myxobolus species. The closest haplotypes to M. parvus were Triactinomyxon sp. and Endocapsa sp. which are the actinosporeans forms of unknown myxozoan species. These data are the first in molecular characterization of M. parvus, its occurrence in lower jaw of leaping mullet along with its infection prevalence values off the Turkish Black Sea coasts.

Oestrogenic pollutants promote the growth of a parasite in male sticklebacks.

Aquatic environments are especially susceptible to anthropogenic chemical pollution. Yet although knowledge on the biological effects of pollutants on aquatic organisms is increasing, far less is known about how ecologically-important interspecific interactions are affected by chemicals. In particular, the consequences of anthropogenic pollution for the interaction of hosts and parasites are poorly understood. Here, we examine how exposure to 17ß-oestradiol (E2)-a natural oestrogen and a model endocrine disrupting chemical (EDC) -affects infection susceptibility and emergent infection phenotypes in an experimental host-parasite system; three spined sticklebacks (Gasterosteus aculeatus) infected with the common, debilitating cestode Schistocephalus solidus. We exposed individual sticklebacks to a 0ngl(-1) (control), 10ngl(-1) or 100ngl(-1) E2 treatment before feeding them infective stages of S. solidus. E2 exposure significantly elevated vitellogenin (VTG) levels-a biomarker of exposure to xenoestrogens-in both female and male fish, and reduced their body condition. Susceptibility to parasite infection was unaffected by EDC exposure; however, E2 treatment and fish sex interacted significantly to determine the growth rate of parasites, which grew quickest in male hosts held under the higher (100ngl(-1)) E2 treatment. Tissue VTG levels and parasite mass correlated positively across the whole sample of experimentally infected fish, but separate regressions run on the male and female datasets demonstrated a significant relationship only among male fish. Hence, among males-but not females-elevated VTG levels elicited by E2 exposure led to more rapid parasite growth. We outline plausible physiological mechanisms that could explain these results. Our results demonstrate that oestrogenic pollutants can alter host-parasite interactions by promoting parasite growth, and that male hosts may be disproportionately affected. Because ecologically-relevant effects of infection on host antipredator responses, growth, energetics and reproductive development all depend on parasite mass in this host-parasite system, our results indicate that EDCs can mediate the ecological consequences of infections. We therefore consider the implications of our results for the ecology of hosts and parasites in polluted environments.

Three-spined stickleback Gasterosteus aculeatus, as a possible paratenic host for salmonid nematodes in a subarctic lake.

In Takvatn, a subarctic lake in northern Norway, 35 of 162 three-spined sticklebacks examined were infected with 106 specimens of third-stage larvae of Philonema oncorhynchi. The prevalence and mean intensity of P. oncorhynchi were 10 % and 2.0 in 2013 and 24 % and 3.0 in 2014, respectively. A single specimen of Cystidicola farionis was found in an additional sample. While the latter is considered an accidental infection, three-spined sticklebacks may function as paratenic hosts of P. oncorhynchi, potentially enhancing its transmission to salmonids due to their central role in the lacustrine food web of this subarctic lake.

Diseases of captive yellow seahorse Hippocampus kuda Bleeker, pot-bellied seahorse Hippocampus abdominalis Lesson and weedy seadragon Phyllopteryx taeniolatus (Lacépède).

Seahorses, pipefish and seadragons are fish of the Family Syngnathidae. From 1998 to 2010, 172 syngnathid cases from the Toronto Zoo were submitted for post-mortem diagnostics and retrospectively examined. Among the submitted species were yellow seahorses Hippocampus kuda Bleeker (n=133), pot-bellied seahorses Hippocampus abdominalis Lesson (n=35) and weedy seadragons Phyllopteryx taeniolatus (Lacépède; n=4). The three most common causes of morbidity and mortality in this population were bacterial dermatitis, bilaterally symmetrical myopathy and mycobacteriosis, accounting for 24%, 17% and 15% of cases, respectively. Inflammatory processes were the most common diagnoses, present in 117 cases. Seven neoplasms were diagnosed, environmental aetiologies were identified in 46 cases, and two congenital defects were identified.

Study of Myxosporea (Myxozoa), infecting worldwide mullets with description of a new species.

Existing data on Myxozoa parasites infecting mullets were reviewed. The validity of nine species names was updated. Sixteen species were registered during analysis of original material collected in the Mediterranean, Black, Azov, and Japan Seas in 2004-2005. A new bivalvulid myxozoan parasite, Myxobolus adeli n. sp., was described from the inner organs of the golden grey mullet Liza aurata (Risso, 1810) collected in the Mediterranean (Ebro Delta, Spain), Black Sea (Kerch Strait, Ukraine), and Azov Sea (Genichesk, Ukraine) coastal waters. It is characterized by the presence of elongated, spindle-like cysts 0.5-1.3 mm in size, filled with wide transverse-oval spores about 6.2 × 7.2 × 4.6 µm in size, with two equal polar capsules measuring about 3.0 × 1.8 µm and short polar filament, turned into four coils. The obtained data show that this species differs from all previously described Myxobolus spp. with equal polar capsules. Comparative study of Myxobolus spp. recorded in worldwide mullets indicates a close relationship with M. adeli n. sp. and Myxobolus improvisus Isjumova, 1964 registered in mullets. Probably, the last species includes representatives of some different species, infecting freshwater and marine hosts.

Infection of wild mullet (Mugil cephalus) with Myxobolus episquamalis in Korea.

Wild mullet (Mugil cephalus) with white cysts on their scales were obtained from Yeosu on the south coast of Korea in 2009. Cyst-like plasmodia consisted of a large number of mature myxosporean spores and numerous sporogonic stages. Spores were oval-shaped in their front view, tapering anteriorly to a blunt apex, and lenticular in their lateral view. They measured 7.0 µm (6.2-7.6) in length, 5.2 µm (4.0-6.2) in width, and 4.9 µm (3.8-6.0) in thickness. Polar capsules contained a polar filament with five to six turns and measured 3.5 µm (2.5-4.5) in length and 2.0 µm (1.6-2.3) in width. Nucleotide sequences of the 18S rRNA gene of the myxosporean parasites in our study showed 99.8 % identity with Myxobolus episquamalis Egusa, Maeno and Sorimachi, 1990 from mullet in Tunisia. These results suggest that the Myxobolus sp. found on the scales of wild mullet is M. episquamalis. In the histopathological examination, spores were observed not only in the plasmodia on the scales, but also in the intestine, pancreas, heart, kidney, stomach, gill, skin, spleen, and liver, suggesting the possibility of the coinfection by different Myxobolus species.

A new gyrodactylid (Monogenea) parasitizing bay pipefish (Syngnathus leptorhynchus) from the Pacific Coast of North America.

Gyrodactylus leptorhynchi n. sp. (Monogenea) is described from bay pipefish ( Syngnathus leptorhynchus ) (Syngnathidae) in coastal waters of southern California and British Columbia, and from an outbreak of gyrodactylosis at the Cabrillo Marine Aquarium in California. Gyrodactylus leptorhynchi is morphologically similar (stout hamuli, superficial bar with no anterolateral processes, and a small, triangular membrane, similarly shaped marginal hook sickles, and a male copulatory organ [MCO] with numerous small spines) to the other 6 species of Gyrodactylus known from pipefish in north and south regions of the Atlantic Ocean. It resembles most closely Gyrodactylus corleonis Paladini, Cable, Fioravanti, Faria, and Shinn, 2010 , parasitizing Syngnathus typhle L. from the French Mediterranean in having relatively large hamuli (58 µm). However, in G. leptorhynchi, the marginal hook sickle has a reduced heel and a ledged toe, while in G. corleonis, it has a noticeable heel and a toe with no distinct ledge. DNA sequence data of a partial ITS1 (700+ bp), complete 5.8S (157 bp), complete ITS2 (392 bp), and a partial 18S (441 bp) are included in the description; the data are distinct from those available for other species of Gyrodactylus. The molecular data reveal that G. leptorhynchi is a member of a basal lineage of marine species within Gyrodactylus sensu lato that is known to have radiated among coastal syngnathid, anguillid, and gobiid fishes throughout the Atlantic Ocean and some adjacent waters. Occurrence of G. leptorhynchi in the eastern Pacific supports the idea that such lineages may have global distributions. Sixty-three percent (15 of 24) of bay pipefish caught at Inner Cabrillo Beach, California, were infected with 1-3 worms, predominately located on the dorsal fin, but also on the anterior body surfaces. Intensely infected pipefish at the marine aquarium had parasites distributed all over the body surface, including the open edge of the brood pouch and, on 2 occasions, inside the brood pouch. A quarantine protocol, involving the treatment of wild pipefish with serial repeats of topical anthelmintic chemicals (formalin, Trichlorfon, and Praziquantel), that helps to diminish outbreaks of G. leptorhynchi in aquarium exhibits is described.

Gnathostoma spinigerum: immunodepression in experimental infected mice.

Mice were infected with 8- or 25-infective worms of advanced third stage Gnathostoma spinigerum larvae (L3) which were obtained from natural infected eels. On day 14, 60 and 200 post infections (PI), spleen cells of infected mice were tested for lymphoproliferative responses in vitro against the mitogen and specific L3 somatic antigen in order to clarify the cellular immune status of the host upon this nematode infection. Reduced responsiveness to Con A was observed in infected mice. These depressed responses were more pronounced in chronically infected mice (day 200, PI) than in day 14 and day 60, PI. There was no significant difference of lymphoproliferative response between groups of high (25 L3) and low (8 L3)-infective dose in the chronic readily stage. Regarding to the L3 somatic Ag stimulation, the depressed response was obviously detected in high dose and chronic infection. Our results demonstrated that in this G. spinigerum-mouse system T-cell response is defective. The depression could be reversible and was associated with active infection because it was abolished by anthelmintic (ivermectin) treatment. This study shows the involvement of Th-2 response to this nematode in regulating T cell proliferation.

Schistocephalus solidus infections increase gonadotropins and gonadotropin releasing hormone (GnRH3) mRNA levels in the three-spined stickleback, Gasterosteus aculeatus.

Parasites often impair the reproduction of their hosts, one well known case being the cestode Schistocephalus solidus which is a common parasite in three-spined sticklebacks, Gasterosteus aculeatus. One of the possible ways that this could be exerted is by suppression on the brain-pituitary-gonadal (BPG) axis. In this study, mRNA levels of FSH-ß and LH-ß and of GnRH2 (cGnRH II) and GnRH3 (sGnRH) were measured via Q-PCR in infected and uninfected fish sampled from the field a few weeks before the onset of breeding. The pituitary mRNA levels of both FSH-ß and LH-ß were higher in infected males than in uninfected males. Also in females, FSH-ß mRNA levels were higher in infected individuals than in others, whereas there was no significant difference found in LH-ß expression. Brain mRNA levels of GnRH3 were higher in infected fish than in uninfected fish in both sexes, but no difference was found in GnRH2 mRNA levels. Thus, infection by S. solidus was able to alter the expressions not only of gonadotropins (GtHs), but also of GnRH which has not been observed previously. However, the effects are opposite to what should be expected if the parasite suppressed reproduction via actions on the brain-pituitary level. The gonads are perhaps more likely to be impaired by the parasites in other ways, and changed feedbacks on the BPG axis could then lead to the increases in GtHs and GnRH.

Occurrence of Ascocotyle (Phagicola) longa Ransom, 1920 (Digenea: Heterophyidae) in Mugil incilis from Cartagena Bay, Colombia.

Fishborne metacercariae have not been reported in Colombia. During an 8-month period, 250 mullet/lisa (Mugil incilis) were examined for trematode metacercariae in the liver. Average prevalence and intensity were 93.4+/-2.5% and 8476+/-1238 cysts per gram of liver, respectively. Monthly prevalence of the trematode, identified as Ascocotyle (Phagicola) longa Ransom (1920), was always high (>79%) and there was no difference between sampling periods (P>0.05). Trematode intensity in lisa had a significant negative effect on host size (R=-0.325, P<0.001), condition factor (R=-0.352, P<0.001), and liver-somatic index (R=-0.372, P<0.001); in contrast, a positive relationship was found with gill-somatic index (R=0.446, P<0.001). Histopathological analysis indicated the presence of abundant cysts together with inflammatory reactions involving macrophage aggregates and necrosis. These results suggest that trematodes infecting the liver of M. incilis affect the growth and well-being of these fish, and their consumption could represent a major risk for human infection.

Identification and description of Bucephalus minimus (Digenea: Bucephalidae) life cycle in Portugal: morphological, histopathological, and molecular data.

The cercaria of Bucephalus minimus infects the digestive gland and gonads of its first intermediate host, the edible cockle, Cerastoderma edule. Light microscopy (LM) and scanning electron microscopy (SEM) of the cercaria showed a tail formed by a central stem, with 2 long contractile arms presenting distinct morphological surfaces. The encysted metacercaria naturally infected the flathead grey mullet, Mugil cephalus. The cysts found in the heart, liver, and spleen were shown to be identical by the internal transcribed spacer (ITS 1) sequence and morphological features and were associated with encapsulation, recruitment of cell infiltrates, and presence of melanomacrophages and adipose tissue. To establish the life cycle, we compared the ITS1 sequence in an adult from the known definitive host, Dicentrarchus labrax; encysted metacercariae from the liver, heart, and spleen of M. cephalus; and a cercaria from C. edule. With this comparison, we determined that they had a 100% similarity. Therefore, the ITSI sequence data clearly indicate that these 3 parasitic stages belong to the same species, i.e., B. minimus.

Kudoa unicapsula n. sp. (Myxosporea: Kudoidae) a parasite of the Mediterranean mullets Liza ramada and L. aurata (Teleostei: Mugilidae).

A new multivalvulid myxozoan parasite, Kudoa unicapsula n. sp., is described from the intestinal mesentery, intestine and pyloric caeca of the thin-lipped grey mullet Liza ramada (Risso 1826) and the golden grey mullet L. aurata (Risso, 1810) from the Mediterranean coastal waters of Spain. It is characterized by the presence of elongated, rice corn-like white cysts of 0.47-0.56 x 0.18-0.38 mm, filled with tetracapsulate, slightly asymmetric spores, rectangular in apical view and tear-shaped in lateral view with four polar capsules of considerably different size and slightly unequal spore valves with rounded edges, overlapping each other on the apex of the spore. One large polar capsule includes a polar filament coiled in two to three turns, and the other three polar capsules, which are very small, posses only a rudimental filament. Both light and electron microscopy data showed that this species differs from all previously described Kudoa spp. with unequal polar capsules. The molecular analysis based on 18S and 28S ribosomal deoxyribonucleic acid DNA sequence data of K. unicapsula n. sp. indicates a close relationship and thus phylogenetic clustering together with K. trifolia, a myxozoan from the same host and the same geographical location.

A new endemic focus of Heterophyes nocens and other heterophyid infections in a coastal area of Gangjin-gun, Jeollanam-do.

To know the prevalence of heterophyid trematodes among inhabitants of a southern coastal village, i.e., Sacho-ri, Gangjin-gun, Jeollanam-do (Province), 82 stool samples were examined on helminth eggs and protozoan cysts using Kato-Katz and formalin-ether sedimentation techniques. Total 33 people (40.2%) were positive for trematodes (Heterophyes nocens 15 people, Pygidiopsis summa 3, Metagonimus sp.; 4, Clonorchis sinensis 7, Gymnophalloides seoi 6) and/or protozoa (Entamoeba coli 3). Among intestinal trematode egg positive cases, 17 were treated with praziquantel and their whole diarrheic stools were collected after purgation. Adult flukes of H. nocens (number of specimens = 1,294), P. summa (386), Stellantchasmus falcatus (5), Stictodora lari (4), and Heterophyopsis continua (1) were collected using a stereomicroscope. To know the source of human H. nocens infections in this village, metacercarial infections in mullets (10) were examined and most H. nocens metacercariae (101/105, 96.2%) were found in the trunk portion. From above results, the surveyed coastal village has been newly known as an endemic focus of human H. nocens infection and consuming raw mullets was the presumable source of human heterophyid infections.

Robinia aurata n. g., n. sp. (Digenea: Hemiuridae) from the mugilid Liza aurata with a molecular confirmation of its position within the Hemiuroidea.

Robinia aurata n. g., n. sp. is described from Liza aurata (Mugilidae), the golden grey mullet, from the Ebro Delta, Spanish Mediterranean. The new genus differs from all other hemiurid genera in the combined possession of muscular flanges and a vestigial ecsoma. Within the Bunocotylinae, which currently accommodates 2 genera, Bunocotyle and Saturnius, the new genus exhibits a unique combination of blind caeca, Juel's organ, post-ovarian bulk of the uterus in the hind-body, and tegumental papillae surrounding the oral and ventral sucker apertures. Furthermore, Robinia n. g. differs from both Bunocotyle and Saturnius in the nature of the muscular extensions around the oral sucker, with the shape of a muscular belt in the latter and numerous muscular papillae in the former. The phylogenetic hypothesis for the Bunocotylinae developed from sequence data analyses based on partial lsrDNA and complete ssrDNA combined (22 species) and V4 domain of the ssrRNA gene (37 species) supports the erection of the new genus and confirms its position within the Hemiuroidea. Both molecular analyses confirmed the monophyly of the Hemiuroidea, its division into 2 major clades and the polyphyly of the Derogenidae, as in previous studies, and suggest that the Gonocercinae (with 2 genera, Gonocerca and Hemipera), may require a distinct familial status. Finally, there was poor support for the distinct status of the Lecithasteridae and Hemiuridae, following previous suggestions based on different sequence data sets. A key to genera of the Bunocotylinae is presented.