Tegumental topography and molecular characterisation of two trematodes (Platyhelminthes: Digenea) from Clarias gariepinus (Burchell, 1822) in Kenya.

The discoveries of new taxonomic features of digenean species through the application of contemporary techniques, such as scanning electron microscopy (SEM) and molecular analysis are still growing. Two species of trematodes Glossidium pedatum and Tylodelphys mashonensis from the intestine and vitreous humour of Clarias gariepinus were recovered from Lake Ol'Bolossat, Kenya. The two endo-helminths were prepared for morphological examination using SEM and molecular characterisation. Additional morphological features were observed for G. pedatum such as domed papillae in the anterior extremity and a protruding cirrus which was unarmed, laterally folded and with a blunt tip as the first such observation for the genus and led to additional characteristics of the diagnosis of the genus. Tylodelphys mashonensis was characterised by a round oral sucker and tribocytic organ rounded with rows of papillae symmetrically arranged. The molecular analyses using ribosomal marker 28S large subunit (LSU) rDNA and mitochondrial (mtDNA) cytochrome c oxidase subunit 1 (cox1) for both G. pedatum and T. mashonensis confirmed the identity of the species and their phylogenetic relationship within the subclass Digenea. This study provides the first mitochondrial (mt)DNA sequence for G. pedatum and also extends the geographical record of two parasites to Kenya.

RECRUITMENT AND SEASONAL OCCURRENCE OF PARASITES IN JUVENILE INVASIVE ROUND GOBIES (NEOGOBIUS MELANOSTOMUS) IN THE ST. LAWRENCE RIVER, QUEBEC, CANADA.

The round goby (Neogobius melanostomus) is an invasive species that has become one of the most abundant fish in the St. Lawrence River, Quebec, Canada over the past 15 yr. Since its introduction, the round goby has acquired a number of native parasites, yet little is known about the dynamics of parasite recruitment. To examine this question, young-of-the-year and juvenile round gobies were collected monthly from 2 localities in the river (Îles de la Paix, Île Dorval) from June through November 2012. At Îles de la Paix, round gobies (n = 180) were infected with 3 species of parasites, all larval stages (Diplostomum spp., Tylodelphys scheuringi, Neoechinorhynchus tenellus). Prevalence of the digenean Diplostomum spp. varied from 3.3 to 13.3%, and mean abundance from 0.03 to 0.53 from June through September, with a maximum in August. The digenean T. scheuringi was seen only in August, at a prevalence of 10.0% and a mean abundance of 0.53. The acanthocephalan N. tenellus was observed in June, August, and September, prevalence ranging from 3.3 to 10.0% and mean abundance from 0.03 to 0.27. Maximum infection for all 3 species occurred in August. All infected fish were ≥44 mm in total length (TL). Fish infected with more than 1 parasite species were >60 mm TL. No round goby (n = 178) was infected at Île Dorval. This study demonstrated that the invasive round goby starts to acquire parasite infections in the St. Lawrence River in the first year of life and may contribute to the transmission of some parasites within this ecosystem.

Molecular phylogeny of Diplostomum, Tylodelphys, Austrodiplostomum and Paralaria (Digenea: Diplostomidae) necessitates systematic changes and reveals a history of evolutionary host switching events.

The Diplostomidae Poirier, 1886 is a large, globally distributed family of digeneans parasitic in intestines of their definitive hosts. Diplostomum and Tylodelphys spp. are broadly distributed, commonly reported, and the most often sequenced diplostomid genera. The majority of published DNA sequences from these genera originated from larval stages only, which typically cannot be identified to the species level based on morphology alone. We generated partial large ribosomal subunit (28S) rRNA and cytochrome c oxidase subunit 1 (cox1) mtDNA gene sequences from 14 species/species-level lineages of Diplostomum, six species/species-level lineages of Tylodelphys, two species/species-level lineages of Austrodiplostomum, one species previously assigned to Paralaria, two species/species-level lineages of Dolichorchis and one unknown diplostomid. Our DNA sequences of 11 species/species-level lineages of Diplostomum (all identified to species), four species/species-level lineages of Tylodelphys (all identified to species), Austrodiplostomum compactum, Paralaria alarioides and Dolichorchis lacombeensis originated from adult specimens. 28S sequences were used for phylogenetic inference to demonstrate the position of Paralaria alarioides and Dolichorchis spp. within the Diplostomoidea and study the interrelationships of Diplostomum, Tylodelphys and Austrodiplostomum. Our results demonstrate that two diplostomids from the North American river otter (P. alarioides and a likely undescribed taxon) belong within Diplostomum. Further, our results demonstrate the non-monophyly of Tylodelphys due to the position of Austrodiplostomum spp., based on our phylogenetic analyses and morphology. Furthermore, the results of phylogenetic analysis of 28S confirmed the status of Dolichorchis as a separate genus. The phylogenies suggest multiple definitive host-switching events (birds to otters and among major avian groups) and a New World origin of Diplostomum and Tylodelphys spp. Our DNA sequences from adult digeneans revealed identities of 10 previously published lineages of Diplostomum and Tylodelphys, which were previously identified to genus only. The novel DNA data from this work provide opportunities for future comparisons of larval diplostomines collected in ecological studies.

DISTRIBUTION AND DIVERSITY OF DIPLOSTOMIDS IN NEW ZEALAND.

Parasitism is one of the most common consumer strategies and contributes a large portion to biological diversity. Trematodes in the family Diplostomidae are common in freshwater ecosystems worldwide, often residing in the eyes or brain of fish and then infecting fish-eating birds as adults. As a result, some species have broad geographic distributions due to the bird host's motility. In contrast to the cosmopolitan nature of diplostomids, only a single species, Tylodelphys darbyi, has been identified in New Zealand to date, and only from the South Island. Tylodelphys darbyi has a 3-host life cycle consisting of an unidentified snail, a freshwater fish (Gobiomorphus cotidianus), and the Australasian crested grebe (Podiceps cristatus australis). To date, T. darbyi has been found in 2 locations, Lake Hayes, in the eyes of G. cotidianus, and Lake Wanaka, adults recovered from grebes. Considering the near ubiquity of the fish host in New Zealand, it is likely the bird, listed as nationally vulnerable, is the limiting factor in the range of T. darbyi. Up to 10 G. cotidianus were sampled from 10 mountain lakes known to have populations of grebe in the Otago and Canterbury regions of New Zealand's South Island. The eyes of all fish were examined and any metacercariae present were set aside for genetic analysis. In addition to expanding the known range of T. darbyi to at least 4 water bodies across the South Island, 2 new taxa of diplostomid were identified. A lens-infecting metacercariae clustered with Diplostomum spathaceum, while the metacercariae from the humor clustered with Diplostomum baeri.

Eye fluke (Tylodelphys clavata) infection impairs visual ability and hampers foraging success in European perch.

Visual performance and environmental conditions can influence both behavioral patterns and predator-prey interactions of fish. Eye parasites can impair their host's sensory performance with important consequences for the detection of prey, predators, and conspecifics. We used European perch (Perca fluviatilis) experimentally infected with the eye fluke Tylodelphys clavata and evaluated their feeding behavior and competitive ability under competition with non-infected conspecifics, in groups of four individuals, for two different prey species (Asellus aquaticus and Daphnia magna). To test whether the effect of T. clavata infection differs at different light conditions, we performed the experiments at two light intensities (600 and 6 lx). Foraging efficiency of perch was significantly affected by infection but not by light intensity. The distance at which infected fish attacked both prey species was significantly shorter in comparison to non-infected conspecifics. Additionally, infected fish more often unsuccessfully attacked A. aquaticus. Although the outcome of competition depended on prey species, there was a general tendency that non-infected fish consumed more of the available prey under both light intensities. Even though individual prey preferences for either A. aquaticus or D. magna were observed, we could not detect that infected fish change their prey preference to compensate for a reduced competitive foraging ability. As infection of T. clavata impairs foraging efficiency and competitive ability, infected fish would need to spend more time foraging to attain similar food intake as non-infected conspecifics; this presumably increases predation risk and potentially enhances transmission success to the final host.

Description of Tylodelphys darbyi n. sp. (Trematoda: Diplostomidae) from the threatened Australasian crested grebe (Podiceps cristatus australis, Gould 1844) and linking of its life-cycle stages.

Species of the genus Tylodelphys (Diplostomidae) have a cosmopolitan distribution. Metacercariae of these species infect the eye, brain, pericardial sac or body cavity of fish second intermediate hosts, and the adults are found in piscivorous birds of many orders. An unnamed species of Tylodelphys from the eyes of bullies (Gobiomorphus cotidianus) was characterized molecularly and morphologically as a metacercaria in a previous study, in which it was predicted that the adult of this species would be found in the Australasian crested grebe. Two specimens of this bird became available and specimens of the unnamed Tylodelphys species were, indeed, found in them, confirmed by identity of genetic sequence data. Found to differ morphologically from its congeners, the new species is here described as Tylodelphys darbyi n. sp. Three species are closest to the new species in morphology: Tylodelphys glossoides, T. immer and T. podicipina robrauschi. Compared with T. darbyi n. sp. these three species are slightly larger and possess longer eggs. Tylodelphys glossoides also differs in having a wider oral sucker and T. podicipina robrauschi in having comma- or kidney-shaped pseudosuckers and an ovary that reaches a larger size, along with higher upper limits for body width, hind body and sucker width, holdfast and oesophagus length, and pharynx, pseudosucker and testes length and width. Tylodelphys immer also differs from T. darbyi n. sp. in having a shorter ventral sucker and the largest pseudosuckers of any Tylodelphys species.

Exploring the genetic diversity of Tylodelphys (Diesing, 1850) metacercariae in the cranial and body cavities of Mexican freshwater fishes using nuclear and mitochondrial DNA sequences, with the description of a new species.

Members of the genus Tylodelphys Diesing, 1850 are endoparasites of fish-eating birds, particularly ciconiids, anhingids, and podicipedids across the globe. Metacercariae of Tylodelphys spp. were collected from the cranial and body cavities of freshwater fishes in central and northern Mexico; adults were recovered from the intestine of two species of freshwater diving birds of the family Podicipedidae, commonly known as grebes, in two locations of central Mexico. Specimens were sequenced for two molecular markers, the internal transcribed spacers (ITS1 and ITS2) plus 5.8S gene of the nuclear ribosomal DNA and of the cytochrome c oxidase subunit 1 from mitochondrial DNA. The genetic divergence among the 25 samples (16 metacercariae and 9 adults) and between the newly sequenced specimens and those deposited in the GenBank were estimated. Maximum likelihood and Bayesian inference analyses inferred with each data set revealed the existence of five genetic lineages. Eight metacercariae analyzed in this study were nested in two divergent lineages previously recognized as Tylodelphys sp. 5 and Tylodelphys sp. 6 (sensu Locke et al., Int J Parasitol, 45:841-855, 2015). Five adult specimens recovered from the intestine of the least grebe (Tachybaptus dominicus Linnaeus, 1766) in Tecocomulco Lake, Hidalgo State, nested in a single clade with other sequences identified previously as Tylodelphys aztecae, expanding its distribution range in other areas of central Mexico. The isolates of the metacercariae found in the cranial cavity of the shortfin silverside, Chirostoma humboldtianum Valenciennes, 1835 from Zacapu Lake in central Mexico formed a monophyletic lineage and were recognized as an undescribed species of Tylodelphys. The lack of adult specimens of this lineage in our samples prevented a formal description. However, the metacercariae collected in the cranial cavity of the silverside, Chirostoma jordani Woolman, 1894 and the adult specimens recovered from the intestine of the western grebe, Aechmophorus occidentalis (Lawrence, 1858) from Cuitzeo Lake formed a monophyletic clade, allowing us to link both stages of the life cycle and to describe this as a new species, Tylodelphys kuerepus n. sp. The new species represents the eighth species of the genus described in the Americas and the fourth in the Nearctic region. We briefly discuss the ecological associations between the metacercariae and their second intermediate hosts in relation to the genetic diversity patterns uncovered in our study.

Morphological and molecular analyses of Tylodelphys spp. metacercaria (Trematoda: Diplostomidae) from the vitreous humour of two freshwater fish species, Channa gachua (Ham.) and Puntius sophore (Ham.).

Trematodes of family Diplostomatidae consists of a large and diverse group of parasites which have larval stages that are very important pathogens of wild and cultured freshwater fishes worldwide, can cause serious impacts. Our understanding of the diplostomids diversity, though, remains deficient and limited especially in India. In the present study, the morphology and molecular characterization of Tylodelphys spp. from the vitreous humour of eye, collected from two economically important food fish, dwarf snakehead Channa gachua (Perciformes: Channidae) and pool barb Puntius sophore (Cypriniformes: Cyprinidae) were described from Bijnor and Meerut, Uttar Pradesh (U.P.), India respectively. Tylodelphys spp. were characterized morphologically by light microscopy and SEM observations. Partial sequences of the ribosomal ITS1-5.8S-ITS2 gene cluster were also used for molecular identification of the species. Combined morphological and molecular analyses revealed the presence of three species of Tylodelphys: Tylodelphys sp. CG from C. gachua and Tylodelphys sp. PS 1, Tylodelphys sp. PS 2 from P. sophore respectively. This study demonstrated that molecular diversity of Tylodelphys spp. in Meerut, UP, India that may contribute to our knowledge of the diagnosis and taxonomy of diplostomids in fish.

The eye fluke Tylodelphys clavata affects prey detection and intraspecific competition of European perch (Perca fluviatilis).

Parasites that occupy the eyes of fish have the potential to affect visual perception and consequently alter the host's behaviour, as these organs provide information about their surroundings. In an experimental study, the feeding behaviour of European perch (Perca fluviatilis) infested with the eye fluke Tylodelphys clavata was examined. The results showed that an individual's ability to identify and approach food items was negatively affected by the infestation intensity of T. clavata. Additionally, the foraging success of an individual was reduced in competition with another, less heavily infested conspecific, when the same food resource was exploited. These alterations in the ability to locate food may have important consequences on the feeding strategy of the fish. Furthermore, the impaired feeding capability caused by T. clavata may also increase the predation risk as heavily infested fish need to spend more time foraging to attain a rate of food intake equivalent to less infested conspecifics.

First report on molecular evidence of Tylodelphys cerebralis (Diplostomulum cerebralis) Chakrabarti, 1968 (Digenea: Diplostomidae) from snakehead fish Channa punctata.

Freshwater snakehead fish, Channa punctata (Perciformes: Channidae) from Meerut (U.P.), India were screened for infection with metacercaria from the eye (vitreous humor) and brain (cranial cavity) and were analyzed by molecular methods using PCR and sequencing of the complete internal transcribed spacers (ITS1-5.8S-ITS2) of the ribosomal RNA gene. Recovered metacercariae from sites, vitreous humor and cranial cavity were previously diagnosed as Diplostomulum cerebralis Chakrabarti, 1968 on the basis of morphological characteristics. A combination of molecular methods in this study depicts that this is a species of genus Tylodelphys Diesing, 1850 which was misinterpreted as Diplostomulum. Hence, in this study, we validated the status of T. cerebralis (=D. cerebralis) after 49 years from its original description. The present work might contribute to expand our knowledge for identification, biodiversity and taxonomy of diplostomids in Indian fishes.

Digenea in the surubim Pseudoplatystoma corruscans (Spix and Agassiz, 1829) (Siluriformes: Pimelodidae) of the upper São Francisco River, State of Minas Gerais, Brazil

The specimens of surubim, Pseudoplatystoma corruscans (Spix and Agassiz, 1829), an important food fish with high commercial value in the Upper São Francisco River, were collected downstream from the Três Marias Dam, in the municipality of Três Marias, State of Minas Gerais, Brazil. Thirty-six individuals were examined and among the parasites, four species of Digenea were found: Witenbergia witenbergi Vaz, 1932 and Tylodelphys sp. (metacercarie), both with prevalence above 10 percent, followed by Acanthostomum gnerii (Szidat, 1958) and Acanthostomum sp.. The total length of the hosts did not influence the parasite prevalence. Witenbergia witenbergi occurred only in female hosts and the abundance of Tylodelphys sp. was higher in the males. These Digenea were recorded for the first time in P. corruscans, and their known geographic distribution has been expanded to the São Francisco Basin in Brazil.

Espécimes de surubim, Pseudoplatystoma corruscans (Spix e Agassiz, 1829), importante peixe na pesca profissional e de maior valor comercial, do Alto Rio São Francisco, foram coletados à jusante da barragem de Três Marias, município de Três Marias, Minas Gerais. Foram examinados 36 indivíduos e dentre outros parasitos, foram encontradas quatro espécies de Digenea: Witenbergia witenbergi Vaz, 1932 e Tylodelphys sp. (metacercárias) ambas com prevalência superior a 10 por cento, seguidas por Acanthostomum gnerii (Szidat, 1958) e Acanthostomum sp.. O comprimento total dos hospedeiros não influenciou a prevalência e a abundância parasitária. Witenbergia witenbergi ocorreu apenas em hospedeiros fêmeas e a abundância de Tylodelphys sp. foi mais elevada nos hospedeiros machos. Esses Digenea foram registrados pela primeira vez em P. corruscans e tiveram sua distribuição geográfica conhecida ampliada para a bacia do São Francisco, Brasil.