Dynamic transcriptome landscape of Paragonimus proliferus developmental stages in the rat lungs.

Paragonimus proliferus, a lung fluke of the genus Paragonimus, was first reported in Yunnan province, China. P. proliferus can infect Sprague-Dawley (SD) rats and cause lung damage, but there is still no direct evidence of human infection. Until now, there has been a lack of studies on P. proliferus parasitism and development in mammalian lung tissue. The aim of this study was to perform transcriptomic profiling of P. proliferus at different developmental stages. SD rats were infected with P. proliferus metacercariae obtained from crabs; worms isolated from the lungs at different time points as well as metacercariae were subjected to whole transcriptome sequencing. Overall, 34,403 transcripts with the total length of 33,223,828 bp, average length of 965 bp, and N50 of 1833 bp were assembled. Comparative analysis indicated that P. proliferus, similar to other Paragonimus spp., expressed genes related to catabolism, whereas P. proliferus-specific transcripts were related to the maintenance of cellular redox homeostasis, sensitivity to bacteria, and immune response. Transcriptional dynamics analysis revealed that genes involved in the regulation of catabolism and apoptosis had stable expression over the P. proliferus life cycle, whereas those involved in development and immune response showed time-dependent changes. High expression of genes associated with immune response corresponded to that of genes regulating the sensitivity to bacteria and immune protection. We constructed a P. proliferus developmental model, including the development of the body, suckers, blood cells, reproductive and tracheal systems, lymph, skin, cartilage, and other tissues and organs, and an immune response model, which mainly involved T cells and macrophages. Our study provides a foundation for further research into the molecular biology and infection mechanism of P. proliferus.

Echinostoma mekongi: Discovery of Its Metacercarial Stage in Snails, Filopaludina martensi cambodjensis, in Pursat Province, Cambodia.

Echinostoma mekongi was reported as a new species in 2020 based on specimens collected from humans in Kratie and Takeo Province, Cambodia. In the present study, its metacercarial stage has been discovered in Filopaludina martensi cambodjensis snails purchased from a local market nearby the Tonle Sap Lake, Pursat Province, Cambodia. The metacercariae were fed orally to an experimental hamster, and adult flukes were recovered at day 20 post-infection. They were morphologically examined using light and scanning electron microscopes and molecularly analyzed by sequencing of their mitochondrial cox1 and nad1 genes. A total of 115 metacercariae (1-8 per snail) were detected in 60 (60.0%) out of 100 Filopaludina snails examined. The metacercariae were round, 174 µm in average diameter (163-190 µm in range), having a thin cyst wall, a head collar armed with 37 collar spines, and characteristic excretory granules. The adult flukes were elongated, ventrally curved, 7.3 (6.4-8.2)×1.4 (1.1-1.7) mm in size, and equipped with 37 collar spines on the head collar (dorsal spines in 2 alternating rows), being consistent with E. mekongi. In phylogenetic analyses, the adult flukes showed 99.0-100% homology based on cox1 sequences and 98.9-99.7% homology based on nad1 sequences with E. mekongi. The results evidenced that F. martensi cambodjensis snails act as the second intermediate host of E. mekongi, and hamsters can be used as a suitable experimental definitive host. As local people favor to eat undercooked snails, these snails seem to be an important source of human infection with E. mekongi in Cambodia.

How costly are metacercarial infections in a bivalve host? Effects of two trematode species on biochemical performance of cockles.

Bivalve stocks have been decreasing in the last decades largely due to emergent diseases and consequent mass mortality episodes. Cerastoderma edule (the edible cockle) is one of the most exploited bivalves in Europe and is among the most common hosts for trematodes, the most prevalent macroparasites in coastal waters but yet poorly studied. Therefore, in the present study, this bivalve species was used as host model to determine if trematode infection exerts a negative effect on bivalve energy metabolism and balance and if the tissues targeted by different trematodes influence the metabolic cost, with physiological and biochemical consequences. Cockles were experimentally infected with two trematode species, Himasthla elongata and Renicola roscovitus, that infect the foot and palps, respectively. Trematode infection exerted a negative effect on the metabolism of C. edule, the second intermediate host, by reduction of oxygen consumption. A different host biochemical response was found depending on trematode species, especially in regard to the level of oxygen consumption decrease and the preferential accumulation of lipids and glycogen. This study represents a step towards the understanding of host-trematode relationships that can be used to better predict potential conservation threats to bivalve populations and to maximize the success of stock and disease management.

Molecular characterization of Urosporidium tapetis sp. nov., a haplosporidian hyperparasite infecting metacercariae of Parvatrema duboisi (Dollfus 1923), a trematode parasite of Manila clam Ruditapes philippinarum on the west coast of Korea.

Recently, a putative new hyperparasitic haplosporidian in the genus Urosporidium was identified from metacercariae of the trematode Parvatrema duboisi infecting Manila clam Ruditapes philippinarum on the west coast of Korea. In this study, we applied small subunit ribosomal DNA (SSU rDNA) sequences as a marker to substantiate the phylogenetic relationship of the unidentified Urosporidium within the Order Haplosporida. In our phylogenetic analysis, the 1890 bp of SSU rDNA sequences obtained were closely related to a haplosporidian parasite forming a sister clade to Urosporidium group, although the gene sequences were only 89.22-89.70% similar to Urosporidium spp. Such molecular phylogenetic distance within the genus suggested that the unidentified Urosporidium is a new member of the genus. Accordingly, we report the unidentified haplosporidian hyperparasite as Urosporidium tapetis sp. nov.

Studies on the life cycle of Pleurogenoides wayanadensis Shinad & Prasadan, 2018 (Digenea: Pleurogenidae) from the Western Ghats, India.

The life cycle of Pleurogenoides wayanadensis Shinad & Prasadan, 2018, infecting the frogs Hoplobatrachus tigerinus and Euphlyctis cyanophlyctis, is elucidated in this study. All the life cycle stages from egg to egg-producing adults were elucidated under natural conditions and successfully established in the laboratory. The life cycle took about 58 to 65 days for completion. Miracidia were released by teasing the eggs with fine needles. Sporocysts were found in the freshwater snail, Bithynia (Digoniostoma) pulchella, collected from paddy fields at Payode, Western Ghats, Wayanad region, in the months of October and November 2019. Cercariae were of the virgulate xiphidiocercous type. Metacercariae were recovered from the eyes of the damselfly naiads of the species Ischnura sp. and Copera sp., and the thorax and abdomen of the dragonfly naiads, Orthetrum sp. The metacercariae showed progenetic development. The growth and development of the metacercariae in the naiads that were exposed to cercariae, and development of the trematode in frogs that were force-fed with encysted metacercariae, have been studied at regular intervals. The prepatent period is 14-19 days. The present life cycle study of a Pleurogenoides spp. forms the seventh report from the world, fourth report from India and the third from Kerala.

Following the infection dynamics of the tropical trematode Oligogonotylus mayae in its intermediate and definitive hosts for 13 years.

We present a time series of 13 years (2003-2016) of continuous monthly data on the prevalence and mean abundance of the trematode Oligogonotylus mayae for all the hosts involved in its life cycle. We aimed to determine whether annual (or longer than annual) environmental fluctuations affect these infection parameters of O. mayae in its intermediate snail host Pyrgophorus coronatus, and its second and definitive fish host Mayaheros urophthalmus from the Celestun tropical coastal lagoon, Yucatan, Mexico. Fourier time series analysis was used to identify infection peaks over time, and cross-correlation among environmental forcings and infection parameters. Our results suggest that the transmission of O. mayae in all its hosts was influenced by the annual patterns of temperature, salinity and rainfall. However, there was a biannual accumulation of metacercarial stages of O. mayae in M. urophthalmus, apparently associated with the temporal range of the El Niño-Southern Oscillation (five years) and the recovery of the trematode population after a devasting hurricane. Taking O. mayae as an example of what could be happening to other trematodes, it is becoming clear that environmental forcings acting at long-term temporal scales affect the population dynamics of these parasites.

DNA sequencing demonstrates the importance of jellyfish in life cycles of lepocreadiid trematodes.

Sequence data were combined with morphological analyses to identify two lepocreadiid trematode species from jellyfishes and fishes. Three species of jellyfish were captured within Port Phillip Bay, Australia, and three species of fish that feed on jellyfish were obtained from Moreton Bay (Queensland) and Port Phillip Bay and Portland (Victoria). The digeneans were distributed throughout most parts of the jellyfish. Opechona cf. kahawai Bray & Cribb, 2003 parasitized the scyphozoan jellyfish Aequorea eurodina and the scombrid fish Scomber australasicus. Cephalolepidapedon warehou Bray & Cribb, 2003 parasitized the scyphozoans Pseudorhiza haeckeli and Cyanea annaskala, and the centrolophid fishes Seriolella brama and Seriolella punctata. Intensities ranged from four to 96 in the jellyfish, and one to 30 in the fish. For both trematode species, internal transcribed spacer 2 of ribosomal DNA sequences from mature adults in the fishes matched those from metacercariae from the jellyfish. This is the first record of larval stages of C. warehou and O. cf. kahawai, and the first use of DNA sequencing to identify digenean trematode metacercariae from jellyfish. Three new host records are reported for C. warehou and two for O. cf. kahawai.

Phylogenetic relationships, expanded diversity and distribution of Crassiphiala spp. (Digenea, Diplostomidae), agents of black spot disease in fish.

Crassiphiala is a monotypic genus of diplostomid digeneans and is the type genus of the subfamily Crassiphialinae. The type species Crassiphiala bulboglossa parasitizes kingfishers in the Nearctic and has a Neascus-type metacercaria that encysts on fish intermediate hosts, often causing black spot disease. While recent molecular phylogenetic studies included some members of the Crassiphialinae, no DNA sequence data of Crassiphiala is currently available. Our molecular and morphological study of adult and larval crassiphialines from the Americas revealed the presence of at least three lineages of Crassiphiala from the Nearctic and two lineages from the Neotropics. This is the first record of Crassiphiala from the Neotropics. Herein, we provide the first molecular phylogeny of the Diplostomoidea that includes Crassiphiala. Our data revealed 0.2-2.4% divergence among 28S sequences and 11-19.8% among CO1 sequences of lineages of Crassiphiala. The results of our analyses did not support the monophyly of Crassiphialinae. Our results clearly demonstrated that the diversity of Crassiphiala has been underestimated.

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.

Survey of Gymnophalloides seoi Metacercariae in Natural and Cultured Oysters from Several Western Coastal Areas, Korea.

Gymnophalloides seoi (Digenea: Gymnophallidae) is a human intestinal trematode contracted by eating raw oysters (Crassostrea gigas) in the Republic of Korea (=Korea). It has been known to be highly endemic in Aphae Island, Shinan-gun, Jeollanam-do (Province). However, recent epidemiological status of G. seoi has not been reported since the 1990s. In this study, we investigated the prevalence of G. seoi metacercariae in natural and cultured oysters collected from 3 islands and 2 coastal areas in western parts of Korea. The oysters were examined using the artificial digestion method followed by stereomicroscopy. The overall positive rate of G. seoi metacercariae in natural oysters was 66.0% (99/150), and the oysters collected from Yubu Island showed the highest infection rate (74.0%). However, the metacercarial density per oyster was relatively low (1.5-2.4 per oyster). By contrast, no metacercaria was found in cultured oysters purchased from 2 coastal areas in Chungcheongnam-do. Thus, we could confirm that natural oysters produced from 3 western coastal islands are infected with G. seoi metacercariae, whereas cultured oysters purchased from 2 coastal areas were free from infection.

Infections with Digenetic Trematode Metacercariae in Freshwater Fishes from Two Visiting Sites of Migratory Birds in Gyeongsangnam-do, Republic of Korea.

The infection status of digenetic trematode metacercariae (DTM) was investigated in fishes from 2 representative visiting sites of migratory birds in Gyeongsangnam-do, the Republic of Korea (Korea). A totaly 220 freshwater fishes (7 species) were collected from Junam-jeosuji (reservoir), and 127 fishes (7 species) were also collected from Woopo-neup (swamp) in June and October 2017. As the control group, total 312 fish (22 spp.) from Yangcheon in Sancheong-gun, Gyeongsangnam-do were also collected in June and October 2017. All fishes collected in 3 sites were examined with the artificial digestion method. In the fishes from Junam-jeosuji, more than 4 species, i.e., Clonorchis sinensis, Echinostoma spp., Diplostomum spp. and Cyathocotyle orientalis, of DTM were detected and their endemicy was very low, 0.70. More than 6 species, i.e., C. sinensis, Echinostoma spp., Metorchis orientalis, Clinostomum complanatum, Diplostomum spp. and C. orientalis, of DTM were found in the fishes from Woopo-neup, and their endemicy was low, 5.16. In the fishes from Yangcheon, more than 8 species, i.e., C. sinensis, Metagonimus spp., Centrocestus armatus, C. complanatum, C. orientalis, M. orientalis, Echinostoma spp., and Diplostomum spp., of DTM were detected, and their endemicity was relatively high, 95.48. The percentages of avian trematode metacercariae (ATM) were 99.6% and 94.7% in fishes from Junam-jeosuji and Woopo-neup whereas it was 74.1% in the control site, Yangcheon. The above findings suggested that migratory birds partly affect in endemicity of DTM in the fish in the 2 visiting sites in Gyeongsangnam-do, Korea.

Infection Status with Digenetic Trematode Metacercariae in Fishes from Coastal Lakes in Gangwon-do, Republic of Korea.

The infection status of digenetic trematode metacercariae (DTM) was investigated in fishes from coastal lakes in Gangwon-do, the Republic of Korea (Korea). All fishes collected in 5 lakes were examined with the artificial digestion method. More than 10 species, i.e., Metagonimus spp., Pygidiopsis summa, Centrocestus armatus, Metorchis orientalis, M. taiwanensis, Clinostomum complanatum, Echinostoma spp., Stictodora spp., Diplostomum sp. and Diplostomid No. 1. by Morita (1960), of DTM were detected in fishes from 5 coastal lakes in Gangwon-do. Metagonimus spp. metacercariae were found in 52 (41.3%) out of 126 sea rundace, Tribolodon hakonensis, from 5 lakes, and their density was 14.6 per fish infected. P. summa metacercariae were detected in 48 (84.2%) out of 57 mullets from 5 lakes, and their density was 316 per fish infected. C. armatus metacercariae were detected in 7 (14.6%) T. hakonensis and 3 (15.0%) Tridentiger brevispinis from Hyang-ho, and 5 (19.2%) Acanthogobius flavimanus from Gyeongpo-ho. Stictodora spp. metacercariae were found in 4 fish species, i.e., Tridentiger obscurus, Tridentiger trigonocephalus, Chelon haematocheilus, and Acanthogobius lactipes, from Gyeongpo-ho. Total 15 C. complanatum metacercariae were detected in 2 (9.1%) crucian carp, Carassius auratus, from Songji-ho. M. taiwanensis metacercariae were found in T. hakonensis from Hyang-ho and Gyeongpo-ho and in Pseudorasbora parva from Gyeongpo-ho. Total 11 M. orientalis metacercariae were detected in 3 (6.3%) T. hakonensis from Hyang-ho. From the above results, it was confirmed that various species of DTM are infected in fishes from coastal lakes in Gangwon-do, Korea.

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.

Patterns of Clinostomum marginatum infection in fishes and amphibians: integration of field, genetic, and experimental approaches.

Digenetic trematodes of the genus Clinostomum are cosmopolitan parasites infecting fishes, amphibians, reptiles, and snails as intermediate hosts. Despite the broad geographical distribution of this genus, debate about the number of species and how they vary in host use has persisted. To better understand patterns of infection among host species and across life stages, we used large-scale field surveys and molecular tools to examine five species of amphibians and seven species of fishes from 125 California ponds. Among the 12,360 examined hosts, infection was rare, with an overall prevalence of 1.7% in amphibians and 9.2% in fishes. Molecular evidence indicated that both groups were infected with Clinostomum marginatum. Using generalized linear mixed effects models, host species identity and host life stage had a strong influence on infection status, such that Lepomis cyanellus (green sunfish) (49.3%) and Taricha granulosa (rough skinned newt) (9.2%) supported the highest overall prevalence values, whereas adult amphibians tended to have a higher prevalence of infection relative to juveniles (13.3% and 2.5%, respectively). Experimentally, we tested the susceptibility of two amphibian hosts (Pseudacris regilla [Pacific chorus frog] and Anaxyrus boreas [western toad]) to varying levels of cercariae exposure and measured metacercariae growth over time. Pseudacris regilla was 1.3× more susceptible to infection, while infection success increased with cercariae exposure dose for both species. On average, metacarcariae size increased by 650% over 20 days. Our study highlights the importance of integrating field surveys, genetic tools, and experimental approaches to better understand the ecology of host-parasite interactions.

Experimental investigation of alternative transmission functions: Quantitative evidence for the importance of nonlinear transmission dynamics in host-parasite systems.

Understanding pathogen transmission is crucial for predicting and managing disease. Nonetheless, experimental comparisons of alternative functional forms of transmission remain rare, and those experiments that are conducted are often not designed to test the full range of possible forms. To differentiate among 10 candidate transmission functions, we used a novel experimental design in which we independently varied four factors-duration of exposure, numbers of parasites, numbers of hosts and parasite density-in laboratory infection experiments. We used interactions between amphibian hosts and trematode parasites as a model system and all candidate models incorporated parasite depletion. An additional manipulation involving anaesthesia addressed the effects of host behaviour on transmission form. Across all experiments, nonlinear transmission forms involving either a power law or a negative binomial function were the best-fitting models and consistently outperformed the linear density-dependent and density-independent functions. By testing previously published data for two other host-macroparasite systems, we also found support for the same nonlinear transmission forms. Although manipulations of parasite density are common in transmission studies, the comprehensive set of variables tested in our experiments revealed that variation in density alone was least likely to differentiate among competing transmission functions. Across host-pathogen systems, nonlinear functions may often more accurately represent transmission dynamics and thus provide more realistic predictions for infection.

First report of Euryhelmis parasites (Trematoda, Heterophyidae) in Africa: conservation implications for endemic amphibians.

In this study, we report, through molecular identification, the first African records of a digenean trematode parasite of the genus Euryhelmis. We recovered metacercariae encysted in an anuran, the endemic Moroccan painted frog (Discoglossus scovazzi), and a vulnerable caudate, the North African fire salamander (Salamandra algira), from four localities in North Africa (Morocco). Our records go back to the past century and have been confirmed in successive fieldwork seasons thereafter. Metacercarial stages of these parasites require amphibians as the last intermediate host, but the exact identity of the primary hosts and predators of the infected animals in Africa remain unknown. Our searches with basic local alignment search tool (BLAST) from Genbank revealed that hosts were infected by parasites of Euryhelmis costaricensis, which showed almost the same genetic identity (with only one substitution) to previous reports from Costa Rica and Japan, suggesting a recent introduction in Morocco. We proceed to discuss the likely role of introduced mustelids as the potential definitive hosts of trematode adults. Under this assumption, we conclude that the infestation of Discoglossus scovazzi and Salamandra algira might pose a risk to these threatened species.

Worm expulsion of Gymnophalloides seoi from C57BL/6 mice: role of metacercarial exosomes in upregulating TLR2 and MUC2 expression in intestinal tissues.

Gymnophalloides seoi worms were rapidly expelled from C57BL/6 mice within days 3-6 post-infection probably due to operation of mucosal innate immunity. To understand better the mucosal immunity related to worm expulsion from the host, we isolated exosomes of G. seoi metacercariae and investigated their role in induction of mRNA and protein expression of several Toll-like receptors and mucin-related factors in vitro. G. seoi-secreted exosomes were collected using differential ultracentrifugation, and cellular internalization of the exosomes into HT-29 intestinal epithelial cells was visualized by confocal microscopy. The expression of TLR2 and MUC2 in HT-29 cells was up-regulated in stimulation with the exosomes. We suggest that G. seoi-secreted exosomes offer a new point of view in the mechanism of worm expulsion from the host through enhancement of TLR2 and MUC2 expression.

Positive density-dependent growth supports costs sharing hypothesis and population density sensing in a manipulative parasite.

Parasites manipulate their hosts' phenotype to increase their own fitness. Like any evolutionary adaptation, parasitic manipulations should be costly. Though it is difficult to measure costs of the manipulation directly, they can be evaluated using an indirect approach. For instance, theory suggests that as the parasite infrapopulation grows, the investment of individual parasites in host manipulation decreases, because of cost sharing. Another assumption is that in environments where manipulation does not pay off for the parasite, it can decrease its investment in the manipulation to save resources. We experimentally infected rainbow trout Oncorhynchus mykiss with the immature larvae of the trematode Diplostomum pseudospathaceum, to test these assumptions. Immature D. pseudospathaceum metacercariae are known for their ability to manipulate the behaviour of their host enhancing its anti-predator defenses to avoid concomitant predation. We found that the growth rate of individual parasites in rainbow trout increased with the infrapopulation size (positive density-dependence) suggesting cost sharing. Moreover, parasites adjusted their growth to the intensity of infection within the eye lens where they were localized suggesting population density sensing. Results of this study support the hypothesis that macroparasites can adjust their growth rate and manipulation investment according to cost sharing level and infrapopulation size.

Malformations of the gill filaments of the ruffe Gymnocephalus cernuus (L.) (Pisces) caused by echinostomatid metacercariae.

In parasite surveys of fishes from Lake Balaton and its tributaries in Hungary, infections with metacercariae of a species of the digenean genus Echinochasmus (Trematoda: Echinostomatidae) were found in seven species of fish. In ruffe, Gymnocephalus cernuus, malformations of the gill filaments apparently caused by these infections were observed. These malformations were in the form of bifurcations of the filaments at about their mid-length. At the point where the filaments bifurcate, an Echinochasmus metacercaria was always embedded in the cartilaginous ray of the gill filament. All specimens of the ruffe were found to be infected by these metacercariae, and each ruffe specimen was infected by 30-300 metacercariae. Such a bifurcation was found in all of the ruffe specimens, but, apart from these gill malformations, the metacercariae produced only local changes in the cartilage. In the other six infected fish species, only local signs were observed in the cartilage. Experimental infections of chicks with metacercariae resulted in the finding of the sexual adult (marita) of an unidentified species of Echinochasmus. ITS sequences of the adult and metacercaria corresponded with each other, and also with a cercaria isolated from a gravel snail (Lithoglyphus naticoides), with a 99.5-100% similarity.

The life cycle of Neocladocystis intestinalis (Vaz, 1932) (Digenea: Cryptogonimidae), in Aylacostoma chloroticum (Prosobranchia: Thiaridae), and Salminus brasiliensis (Characiformes: Characidae), in Argentina.

The life cycle of Neocladocystis intestinalis (Vaz, 1932) was resolved experimentally. The prosobranchiate snail Aylacostoma chloroticum Hylton Scott (Thiaridae) collected in the Yacyretá Dam, Province of Misiones, Argentina, was found naturally infected with cercariae that possessed pigmented eye spots, 7 pairs of penetration glands, 12 pairs of flame cells, and a V-shaped, or Y-shaped excretory vesicle with very short stem. The cercariae developed in oval cysts, which were found on fin rays, and under scales of naturally and experimentally exposed tetragonopterid fish species and of experimentally exposed poecilid and prochilodont fish species. Adults were obtained experimentally from juvenile Salminus brasiliensis (Characidae), bred in captivity, and infected with metacercariae from albino Gymnocorymbus ternetzi (Tetragonopteridae), which had been exposed to emerging cercariae.