Histopathological, morphological, and molecular characterization of fish-borne zoonotic parasite Eustrongylides Excisus infecting Northern pike (Esox lucius) in Iran.

Eustrongylides excisus is a fish-borne zoonotic parasite known to infect various fish species, including Northern pike (Esox Lucius). This nematode, belonging to the family Dioctophymatidae, has a complex life cycle involving multiple hosts. This study aimed to investigate the occurrence of Eustrongylides nematodes in Northern pike (E. Lucius) collected from Mijran Dam (Ramsar, Iran). Between June and October 2023, an investigation was conducted on Northern pike from Mijran Dam in Ramsar, Iran, following reports of reddish parasites in their muscle tissues. Sixty fish were examined at the University of Tehran, revealing live parasites in the muscles, which were then analyzed microscopically and preserved for a multidisciplinary study. The skeletal muscle tissues of 85% (51/60) of fish specimens were infected by grossly visible larvae which were microscopically identified as Eustrongylides spp. In histopathological examination, the lesion was composed of encapsulated parasitic granulomatous myositis. Microscopically, the cystic parasitic granulomas compressed the adjacent muscle fibers, leading to their atrophy and Zenker's necrosis. Moreover, epithelioid macrophages, giant cells and mononuclear inflammatory cells were present around the larvae and between the muscle fibers. Finally, a molecular analysis by examining the ITS gene region, revealed that they belong to the species E. excisus. Eustrongylidiasis in northern Iran necessitates further research into the biology, epidemiology, and control of Eustrongylides nematodes, focusing on various hosts. This study is the first to comprehensively characterize E. excisus in Northern pike in Ramsar, Iran, raising concerns about possible zoonotic transmission.

Henneguya michiganensis n. sp. (Cnidaria: Myxosporea) from the gills of muskellunge Esox masquinongy Mitchill(Esociformes: Esocidae).

Henneguya Thélohan, 1892 is the second most species rich genus of myxozoans, with reports from freshwater and marine fish worldwide. In the Great Lakes region of North America, muskellunge Esox masquinongy is an important game fish species that serves as an apex predator in the ecosystems of many inland lakes. The myxozoan fauna of esocid fish, especially muskellunge, remains largely understudied. During fish health assessments, muskellunge were examined for parasitic infections and myxozoan pseudocysts were observed on gill clip wet mounts. When ruptured under pressure, the intralamellar pseudocysts released thousands of myxospores consistent with those of the genus Henneguya. The myxospores were 67.3-96.6 (79.1 ± 5.9) µm in total length. The spore body was 18.6-22.6 (20.9 ± 1.0) µm × 5.4-6.9 (6.3 ± 0.4) µm in valvular view and 3.5-4.0 (3.8 ± 0.3) µm wide in sutural view. The two pyriform polar capsules positioned at the anterior of the spore body were 6.4-7.7 (7.0 ± 0.4) µm × 1.8-2.1 (2.0 ± 0.1) µm and each contained a tightly coiled polar filament with 9-10 turns. Two tapering caudal processes extended from the posterior of the spore body and were 47.3-75.6 (58.3 ± 5.8) µm in length. Histologically, large intralamellar polysporic plasmodia were surrounded by plump pillar cells and a distinct layer of plasma. Mild inflammation was present peripherally, with small numbers of necrotic germinative cells and intraplasmodial phagocytes internally. Ribosomal 18S rRNA gene sequence data were obtained from three gill pseudocysts. The three ~2000-bp sequences were identical, but shared no significant similarity with any publicly available sequence data. Phylogenetic analyses demonstrated sequence data from this Henneguya fell within a well-supported clade of Henneguya spp. reported from northern pike Esox lucius in Europe. Based on the distinct morphological, histological and molecular data, this species is designated as Henneguya michiganensis n. sp. from muskellunge in Michigan, USA.

A new genus of Bucephalidae Poche, 1907 (Trematoda: Digenea) for three new species infecting the yellowtail pike, Sphyraena obtusata Cuvier (Sphyraenidae), from Moreton Bay, Queensland, Australia.

Three new species of the family Bucephalidae Poche, 1907 (Trematoda: Digenea) are described from the yellowtail pike, Sphyraena obtusata Cuvier (Sphyraenidae), from Moreton Bay, Queensland, Australia. The three species are morphologically consistent with the present broad concept of the genus Bucephalus Baer, 1827, but significant phylogenetic and ecological differences relative to the type-species of Bucephalus require the proposal of a new genus. Aenigmatrema n. g. is proposed for A. undecimtentaculatum n. sp. (type-species), A. inopinatum n. sp. and A. grandiovum n. sp. In addition, based on morphological, ecological and biogeographical similarities, we recombine two existing species of Bucephalus as Aenigmatrema kaku (Yamaguti, 1970) n. comb. and Aenigmatrema sphyraenae (Yamaguti, 1952) n. comb. Although the three species described in this study are extremely morphologically similar, they can be differentiated from each other, and from A. kaku and A. sphyraenae, morphometrically on the basis of egg size, tentacle number and a combination of the caecum and vitelline field lengths. Complete ITS2 rDNA, partial 28S rDNA and partial cox1 mtDNA sequence data were generated for the three new species, which formed a well-supported clade in all 28S phylogenetic analyses. An expanded phylogenetic tree for the subfamily Bucephalinae Poche, 1907 is presented, demonstrating unresolved issues with the morphology-based taxonomy of the subfamily. The three largest genera, Bucephalus, Rhipidocotyle Diesing, 1858 and Prosorhynchoides Dollfus, 1929 remain extensively polyphyletic, indicating the need for significant further systematic revision.

Parasites and Metals in Walleye (Sander vitreus) and Northern Pike (Esox lucius) from Boreal Montreal Lake (Saskatchewan, Canada): Assessment of Human Health Risks.

Industrial activities conducted in Northern Canada have raised concerns among Indigenous communities regarding wildlife contamination and potential consequences for human health. Therefore, an investigation on the chemical (metals/metalloids) and biological (parasite) burden of adult walleye (Sandervitreus) and northern pike (Esoxlucius) from Montreal Lake, Saskatchewan, was conducted to assess health risks related to fish consumption. Dissection revealed that both fishes displayed typical parasite communities, with Eubothrium sp. (Cestoda) and Raphidascarisacus (Nematoda) occurring the most frequently. None of the identified parasite species were infectious to humans. Concentrations of most inorganic contaminants in fish muscle were low and both walleye and pike can be considered healthy components of a balanced diet. However, due to slightly elevated mercury concentrations, excessive daily consumption of these fishes is not recommended, as mercury exposure over time may lead to adverse health effects.

Multiple isotope analyses of the pike tapeworm Triaenophorus nodulosus reveal peculiarities in consumer-diet discrimination patterns.

Previous studies of dietary isotope discrimination have led to the general expectation that a consumer will exhibit enriched stable isotope levels relative to its diet. Parasite-host systems are specific consumer-diet pairs in which the consumer (parasite) feeds exclusively on one dietary source: host tissue. However, the small numbers of studies previously carried out on isotopic discrimination in parasite-host (ΔXP-HT) systems have yielded controversial results, showing some parasites to be isotopically depleted relative to their food source, while others are enriched or in equilibrium with their hosts. Although the mechanism for these deviations from expectations remains to be understood, possible influences of specific feeding niche or selection for only a few nutritional components by the parasite are discussed. ΔXP-HT for multiple isotopes (δ13C, δ15N, δ34S) were measured in the pike tapeworm Triaenophorus nodulosus and two of its life-cycle fish hosts, perch Perca fluviatilis and pike Esox lucius, within which T. nodulosus occupies different feeding locations. Variability in the value of ΔXP-HT calculated for the parasite and its different hosts indicates an influence of feeding location on isotopic discrimination. In perch liver ΔXP-HT was relatively more negative for all three stable isotopes. In pike gut ΔXP-HT was more positive for δ13C, as expected in conventional consumer-diet systems. For parasites feeding on pike gut, however, the δ15N and δ34S isotope values were comparable with those of the host. We discuss potential causes of these deviations from expectations, including the effect of specific parasite feeding niches, and conclude that ΔXP-HT should be critically evaluated for trophic interactions between parasite and host before general patterns are assumed.

[Ecological and Biochemical Aspects of Parasite-Host Interactions in Transformed Aquatic Bodies: A Case Study of the Cestode Triaenophorus nodulosus and Its Host, the Northern Pike Esox lucius].

The lysosomal enzyme activities of the cestode Triaenophorus nodulosus and its host, the pike, in-aquatic bodies with different degrees of technogenic transformation (Northern Karelia, Russia) have been studied. As has been shown, iron-ore waste causes an increase in the acid phosphatase, nuclease, and beta-galactosidase activities of the host and a decrease in its beta-glucosidase and cathepsin D activities. As a rule, the changes in the same cestode enzyme activities are the opposite. With a decrease in the technogenic load, most of the studied characteristics display the trend of approaching the corresponding values observed in a clean lake. It is assumed that the host plays a leading role in the biochemical adaptation of the parasite and its host to mineral environmental pollution.

Rhipidocotyle fennica (Digenea: Bucephalidae) from Anodonta anatina and pike Esox lucius in Lithuania.

Ribosomal DNA sequences of Rhipidocotyle sp. adults from Esox lucius were shown to be identical to sequences of larval Rhipidocotyle fennica, occurring in Anodonta anatina in Lake Vilkoksnis, Lithuania. Morphological features and host specificity of this adult worm correspond with that, determinate in the first description of R. fennica in Finland. These data give the first evidence that a viable population of R. fennica exists in east central Europe. Bucephalus polymorphus which was reported in unionids in all previous publications is probably R. fennica.

[Population biology of cestode genus Triaenophorus in natural and man-made water bodies].

A comparative study of the frequencies of occurrence and distribution of the abundances of pike parasites, tapeworms of the genus Triaenophorus, was carried out in natural lakes and water bodies contaminated by the Kostomuksha mining and concentration mill (Northern Karelia). We demonstrate that the wide presence of T. crassus and T. nodulosus in natural northern lakes is due to the diversity of aquatic organisms and the structure of trophic relations. The abundance of both species is modeled by the negative binomial distribution. Relations in the host-parasite system are stable. Estimates of the parameter k of the negative binomial distribution reflect the changes in the interactions in the host-parasite system. T. nodulosus has survived in the man-made water body storing the highly mineralized recirculated water from the mill. Disturbance-related distinctions were detected in the parameters of the T. nodulosus abundance distribution in the host population. In the Okunevoye Lake, where the water discharged from the tailings dump is more diluted, the infection rates in pike did not differ from those in undisturbed water bodies.

[An example of anthropogeneous succession in the ecosystem of the Verkhnetulomsky Water Reservoir (the Tuloma River Basin, the Kola region)].

The destruction of initial biocenoses in the ecosystem of the Verkhnetulomsky Water Reservoir caused deep succession processes. As a result of the loss of the amphipod group in the benthos, replacement of the trophic links in fishes took place in the biocenosis of the reservoir. Benthos feeders changed over to the feeding on zooplankton and, as a result, became the constituent of the predators'diet. The structure of parasitic systems has changed. The parasites using amphipods as intermediate hosts have been and, on the contrary, the abundance of parasites infesting fishes through feeding of the latter on zooplankton has increased. Among the parasites, Triaenophorus crassus Forel became the cause of the appearance of stable triaenophorosis focus.

The neuroactive substances and associated muscle system in Rhipidocotyle campanula (Digenea, Bucephalidae) from the intestine of the pike Esox lucius.

We report about the muscular system and the serotonergic and FMRFamidergic components of the nervous system of the Bucephalidae trematode, Rhipidocotyle campanula, an intestinal parasite of the pike. We use immunocytochemical methods and confocal scanning laser microscopy (CLSM). The musculature is identified by histochemical staining with fluorescently labeled phalloidin. The body wall musculature of R. campanula contains three layers of muscle fibres - the outer thin circular, intermediate longitudinal and inner diagonal muscle fibres running in two opposite directions. The digestive system of R. campanula possess of a well-developed musculature: radial, longitudinal and circular muscle elements are detected in the pharynx, circular and longitudinal muscle filaments seen in the oesophagus, and longitudinal and the circular muscle fibres were found in the intestinal wall. Specific staining indicating the presence of actin muscle filaments occurs in the cirrus sac localized in the posterior body region. The frontal region of anterior attachment organ, the rhynchus, in R. campanula is represented by radial muscle fibres. The posterior part of the rhynchus comprise of radial muscles forming the organ's wall, and several strong longitudinal muscle bundles. Serotonergic and FMRFamidergic structures are detected in the central and peripheral compartments of the nervous system of R. campanula, that is, in the paired brain ganglia, the brain commissure, the longitudinal nerve cords, and connective nerve commissures. The innervations of the rhynchus, pharynx, oesophagus and distal regions of the reproductive system by the serotonergic and FMRFamidergic nervous elements are revealed. We compare our findings obtained on R. campanula with related data for other trematodes.

The state of knowledge of the parasitic fauna of the pike Esox lucius Linnaeus, 1758 in Poland in the 50th anniversary of the publication of the Catalogue of Parasitic Fauna of Poland. Part II. Parasites of Cyclostomes and Fish by Jadwiga Grabda

Parasites of pike Esox lucius from freshwater and brackish water in Poland are listed. This checklist summarized data published in Catalogue of Parasitic Fauna of Poland. Part II. Parasites of Cyclostomes and Fish by Jadwiga Grabda and later data from original studies. Parasites are listed alphabetically in higher taxa with their location in host and distribution in Polish waters with references. Parasites belonging to the higher taxa as Protista ­ 11 records, Myxosporea ­ 6, Digenea ­ 17, Monogenea ­ 2, Cestoda ­ 9, Nematoda ­ 7, Acanthocephala ­ 7, Hirudinida ­ 5, Mollusca ­ unidentified glochidia, Crustacea ­ 6, were found. The most frequently noted parasites have been Azygia lucii, Diplostomum spp., Tylodelphys clavata, Tetraonchus monenteron, Triaenophorus nodulosus, Raphidascaris acus and Ergasilus sieboldi.

Isolation and partial structural characterization of new Kunitz-type trypsin inhibitors from the pike cestode Triaenophorus nodulosus.

The inhibitors produced by the parasitic worms successfully protect them from the host's proteases and are supposed to underlie the host-parasite specificity. Our previous study has shown that the extracts from the pike tapeworm Triaenophorus nodulosus inhibit host proteinases and commercial trypsin. We aimed to isolate and identify the components responsible for trypsin inactivation. After a two-step separation the molecular masses were measured by SE-HPLC. The sample proved to contain four fractions represented by polypeptides (1-45 kDa) and low-molecular hydrophobic compounds. According to SDS-PAGE analysis, the major polypeptides in the fractions displaying the highest inhibition had masses of 14.4 kDa. The study culminated in partial N-terminal amino acid sequence analysis with a further search for homology. The research revealed two novel Kunitz-type proteins potentially responsible for the inhibitory capacity of the tapeworms against trypsin. Our findings extend the list of cestodes relying on Kunitz-type proteins in the host-parasite molecular cross-talk.

Altered energetics and parasitism in juvenile northern pike (Esox lucius) inhabiting metal-mining contaminated lakes.

The objective of this study was to evaluate possible factors that could be contributing to altered bioenergetics of juvenile northern pike (Esox lucius) living in lakes receiving effluent from the Key Lake uranium mill in northern Saskatchewan, Canada. Although glycogen and triglycerides stores in liver and muscle were significantly greater in pike from exposure lakes compared to the reference, triglycerides stores of aquatic insects and spottail shiners that are prey items of juvenile pike showed no overall differences among lakes. Measures of parasitism, on the other hand, were negatively correlated with pike bioenergetics thereby reflecting a possible energetic cost of parasitism on reference lake fishes. The degree of infection, as measured by the abundance and biomass of intestinal parasites and the abundance of monogeneans on pike gills, was greatest in reference fishes and intermediate in low-exposure pike, whereas high-exposure fishes harbored no parasites.

[The study of non-organic substances content in Henneguya oviperda (Cohn, 1895) and Myxobolus pseudodispar (Gorbunova, 1936) myxospore valves].

The analysis of organic and non-organic substances in the valves of long stored in water Henneguya oviperda (Gohn, 1895) and Myxobolus pseudodispar (Gorbunova, 1936) myxospores has been made using the cytochemical methods and atomic-absorptional spectroscopy. The high content of Si (up to 94 %) and the presence of Ca, Mg, Fe, Mn cations (Cu, Al, Ni, Cr in fewer amounts) have been detected.

Invalidity of three Palaearctic species of Triaenophorus tapeworms (Cestoda: Pseudophyllidea): evidence from morphometric analysis of scolex hooks.

A comparative study of the scolex hook morphology of five species of tapeworms of the genus Triaenophorus Rudolphi, 1793 (Cestoda: Pseudophyllidea), parasites of pikes (Esox lucius L. and E. reichertii Dybowski) in the Palaearctic Region, was carried out. Measurements of scolex hooks of 81 plerocercoids and 492 adults from different hosts and regions were compared using basic statistics and forward stepwise linear discriminant analysis. The shape of the scolex and that of tridental hooks were found to be suitable only for differentiation of the taxa with a similar shape of hooks, i.e. Triaenophorus nodulosus (Pallas, 1781) from T. amurensis Kuperman, 1968, and T. crassus Forel, 1868 from T. meridionalis Kuperman, 1968 and T. orientalis Kuperman, 1968, respectively. In contrast, discriminant analysis did not enable reliable separation of specimens of individual taxa of these two morphological groups due to high intraspecific variability and overlaps between species. This was reflected in low classification efficiencies (average 83%) of all species of the T. crassus group, whereas all T. amurensis specimens were misidentified as T. nodulosus. The new data also considerably enlarge (up to twofold) the size range of the species described by Kuperman in 1968, which invalidates suitability of the most important discriminant characteristic, the width of the basal plate, for delimitation of Triaenophorus species. Based on the present data, all Kuperman's taxa are considered to represent only distinct geographical populations of T. nodulosus and T. crassus. As a result, T. anmurensis is synonymized with T. nodulosus, whereas T. orientalis is considered to be a synonym of T. crassus. Previous synonymisation of T. ineridionalis with T. crassus, first proposed by Dubinina (1987), is also accepted.

[Indigenous microflora associated with the tegument of Triaenophorus nodulosus (Cestoda) and the intestine of its pike host].

Investigations of the indigenous microflora associated with the mucous intestines of fish and its cestode parasites have been for the first time carried out using the methods of transmission and scanning electron microscopy. New data on the bacterial biodiversity in the cestode and its fish host are obtained. Nanobacteria and spirochaetes are for the first time revealed in a fish host together with the previously known bacteria forming the intestinal microflora of fish. Spirochaetes were shown to be associated with the intestines of a pike host only, while nanobacteria cover abundantly the surface of the apical parts of the intestinal microvilli and the apical parts of the microtriches in the cestode tegument. The similarity of the bacterial floras associated with the apical surface of the parasite tegument and the intestine of the host should be noted. At the same time, deeper bacterial communities represented by obligate symbionts are specific. Thus, there is a normal indigenous microflora in cestodes, associated with the tegumental surface. This symbiotic microflora has specific morphological features and provides the balance of relations in the parasite-host system.

Prevalence of Diphyllobothrium latum (Cestoda: Diphyllobothriidae) plerocercoids in fish species from four Italian lakes and risk for the consumers.

In recent years there has been a re-emergence of diphyllobothriasis by Diphyllobothrium latum (Cestoda: Diphyllobothriidae) in Italy, France and Switzerland, where in the past this fish-borne zoonosis was widespread and then virtually disappeared. A change in eating habits such as the consumption of raw/undercooked freshwater fish, has led to an increased risk for consumers of ingesting infective larvae of D. latum. A survey on the factors responsible for the re-emergence of human diphyllobothriasis in Italy was carried out from March 2013 to December 2014. The aim of this study was to assess the diffusion of D. latum plerocercoids in the fish populations of the sub-alpine lakes of Maggiore, Como, Iseo and Garda, updating the scarce historical data and assessing a preliminary "risk level" of the lacustrine environments and fish species under investigation. A total of 2228 fish belonging to 5 species, 690 from Lake Maggiore, 500 from Lake Como, 655 from Lake Iseo and 383 from Lake Garda were submitted to parasitological examination. The presence of D. latum plerocercoid larvae was detected in 6.6%, 25.4% and 7.6% of perch (Perca fluviatilis) from Lakes Maggiore, Como and Iseo respectively. The parasite was also present in pike (Esox lucius) with prevalence values ranging from 71.4 to 84.2% and in 3.6-3.8% of burbot (Lota lota) from Lakes Iseo and Como. Fish from Lake Garda were negative as well as sampled whitefish (Coregonus lavaretus) and shad (Alosa fallax lacustris). The results of this survey showed a widespread presence of D. latum plerocercoid larvae in Maggiore, Como and Iseo fish populations. Urban fecal contamination of water is still a key issue to be resolved, together with the improvement of communication with consumers regarding the best dietary habits and the most effective processes of parasite inactivation, required for the consumption of raw/undercooked fish caught in high-risk areas.

[The role of alpha-amylase of symbiotic microflora in digestion by lower cestodes and their fish hosts].

The symbiotic microflora associated with the digestive-transport surfaces of the pike intestine and the parasitic cestodes Triaenophorus nodulosus proved capable of the initial stages of carbohydrate hydrolysis mediated by alpha-amylase. The products of hydrolysis by alpha-amylase can be used by both the host and the parasite, which decreases energy expenditures of the macroorganisms. The levels of the bacterial alpha-amylase activity are comparable to those of the analogous enzyme absorbed on the mucosa of the intestine and on the cestode tegument, which indicates a considerable contribution of enzymes of the symbiotic microflora to digestion by the host and the parasite. Apparently, this contribution depends on the fish diet.

Pathogenicity of the gill artery worm Philometra obturans (Nematoda) in northern pike (Esox lucius) in southwest Finland.

The pathogenicity of female Philometra obturans (Prenant, 1886) was investigated. Fish samples were taken in May and June 2001 from Lake Littoistenjärvi, in southwest Finland. The intensity of infection varied between 1 and 4 worms, these being found in the afferent artery of the 2 outermost gill arches. The prevalence was 33.9% (n = 59). Host condition and tissue reactions were investigated through dissection and analysis of histopathology. The parasites obstructed the gill arteries, and the arterial wall was stretched around intact worms. Hypertrophy of the arterial wall was observed, especially when only disintegrating remains of the parasites were present. Infected pike were in a poorer condition after catch, transportation, and holding in aquaria and thus less tolerant to stress than uninfected pike. This was correlated with the number of infected gill arteries (P < 0.05). The results show that infection with this nematode has a detrimental effect on the host when combined with environmental or physiological stress and may result in host mortality.

Taxonomic problems, seasonality and histopathology of Henneguya creplini (Myxosporea) infection of the pikeperch Stizostedion lucioperca in Lake Balaton.

Plasmodia of a Henneguya species measuring 70-900 microns and exhibiting season-dependent stages of development were detected throughout a three-year study on gill myxosporosis of Lake Balaton pikeperch (Stizostedion lucioperca (L.)). Sixty-five out of 160 fish (41%) examined in the period of study were infected by the parasite. Infection was the most prevalent (48%) among pikeperch specimens exceeding 40 cm in length. The highest prevalence of infection (58%) was recorded in 1995-1996 while the lowest (30%) in 1996-1997. The youngest plasmodia appeared in April, and started to develop within the capillaries of the secondary lamellae of the gill filaments. The round or ellipsoidal plasmodia which continued their gradual growth in the subsequent months of the year achieved a size of 800-900 microns by the late autumn months, but remained in intralamellar location throughout the developmental cycle. Mature spores developed in the plasmodia by the end of winter. On the basis of their shape and size, the spores were identified as Henneguya creplini (Gurley, 1894). However, because of the uncertain taxonomy of species assigned to the genus Henneguya the taxonomic position of the parasite requires further study. The host reaction consisting of epithelial proliferation and granulation tissue formation starts around the infected secondary lamella only after the maturation of spores and the disruption of plasmodia.