Challenge for macrophages and mast cells of Chelon ramada to counter an intestinal microparasite, Myxobolus mugchelo (Myxozoa).

Thinlip mullet Chelon ramada is the most abundant mullet species found in the Comacchio lagoons (northern Adriatic Sea, Italy). Histological and ultrastructural sections of the intestine of C. ramada showed that over 83% of 48 mullets were infected with the intestinal parasite Myxobolus mugchelo (Myxozoa). In histological sections, plasmodia of M. mugchelo containing mature spores were situated closer to mucosal folds and were surrounded by numerous mast cells (MCs). Mature spores, generally oval in shape, were observed in the paracellular space among the enterocytes or within them. Near the infected epithelial cells, several MCs, rodlet cells and few neutrophils occurred. In intestinal epithelium, large cells resembling macrophages, some with spores of M. mugchelo inside, were observed. These macrophage-like cells were foamy and possessed elongate striated granules. The number of MCs and macrophages in the intestinal epithelium was significantly higher in parasitized fish. In some parasitized intestines, portions of epithelium were displaced by spores, or the spores were observed inside the damaged enterocytes. Immunohistochemical analysis of C. ramada infected or uninfected intestinal tissue revealed the presence of histamine, serotonin (5-HT), leu-enkephalin and inducible-nitric oxide synthase in epithelial macrophages. Several epithelial cells positive to proliferating cell-nuclear antigen were also observed in the proximity of the macrophages. The current study is the first to record the occurrence of intraepithelial macrophages which engulf myxozoan spores. A hypothesis on migration of spores from pancreas via intestinal wall to gut lumen is presented.

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.

Histopathological and ultrastructural studies of Myxobolus turpisrotundus from allogynogenetic gibel carp Carassius auratus gibelio in China.

During an ongoing systematic survey on species diversity of myxozoans parasitising allogynogenetic gibel carp Carassius auratus gibelio (Bloch) in China, plasmodia were detected in the fins, lip, jaw, gill chamber, gill arches, operculum and oral cavity of infected fish. Combining the morphological and molecular data, the present species was identified as Myxobolus turpisrotundus Zhang, Wang, Li et Gong, 2010. Histopathological examination revealed that despite infecting different organs, M. turpisrotundus always occurred in dermis, demonstrating its affinity to this tissue. Histopathological effect of M. turpisrotundus on the host is relatively mild except parasites in the gill arches producing compression of the adipose tissue and heavy adductor muscles deformation with lymphohistiocytic infiltrates. In addition, the plasmodia in different sites were with the same complex structure arrangement: cup-like cells with unknown derivation, a thin collagenous fibril layer, areolar connective tissue, basement membrane and host epithelial cell. Ultrastructural analysis showed that the parasite has monosporic pansporoblast and sporogenesis followed the usual pattern of most of the myxosporeans.

Myxobolus cerebralis (Hofer) infection risk in native cutthroat trout Oncorhynchus clarkii (Richardson) and its relationships to tributary environments in the Yellowstone Lake Basin.

Conservation of native species is challenged by the introduction of non-native pathogens and diseases into aquatic and terrestrial environments worldwide. In the Yellowstone Lake basin, Yellowstone National Park, the invasive parasite causing salmonid whirling disease Myxobolus cerebralis (Hofer) has been identified as one factor contributing to population declines of native Yellowstone cutthroat trout Oncorhynchus clarkii bouvieri (Jordan & Gilbert). In 2002 and 2003, we examined relationships between the stream environment and severity of M. cerebralis infection in native trout. Coefficients of variation of environmental features were calculated to examine variability. Ten years later, we reassessed infection levels at 22 tributaries broadly across the system. Results of principal component analysis (PCA) of physical features (2003) were negatively correlated with infection severity, mostly in lower jaw cartilage of cutthroat trout, and PCA of chemical features (and temperature) correlated with infection severity in cranial cartilage. Pelican Creek, where M. cerebralis prevalence and severity was high 2002-2003, remained high in 2012. We did not find evidence that the parasite had dispersed further within the system. Variable environmental features (physiological stress) across short spatiotemporal scales within a stream or season may possibly predispose salmonids to infection in the wild and facilitate parasite establishment.

Two novel myxosporean species (Myxosporea: Bivalvulida), Myxobolus marumotoi n. sp. and Cardimyxobolus japonensis n. sp., from the dark sleeper, Odontobutis obscura, in Japan.

Two new myxosporean species, Myxobolus marumotoi n. sp. and Cardimyxobolus japonensis n. sp. (Myxozoa: Myxosporea: Bivalvulida), are described from the dark sleeper, Odontobutis obscura, in Japan, based on their morphological and molecular characterizations. Plasmodia of M. marumotoi n. sp. (Myxobolidae) grew in the myofiber of trunk muscles, forming pseudocysts. The rounded spore was relatively large, measuring 13.3-15.0 µm (average 13.8) in length, 14.2-15.0 µm (14.6) in width, and 10.0-11.7 µm (10.8) in thickness, with two subspherical polar capsules of 7.9-9.6 µm (8.4) in length by 5.4-6.3 µm (5.9) in width (n = 15). The polar capsules were directed toward the apex of the spore, packing five to six spirals of the polar filament. Plasmodia of C. japonensis n. sp. (Ortholineidae) were surrounded by thin fibrous tissue, forming cysts in the lamina propria of the alimentary tract. The spore was ovoid, wider than long, in valvular view and spindle-shaped in sutural view. It measured 8.8-10.4 µm (9.4) in length, 11.3-12.5 µm (11.9) in width, and 5.2-6.7 µm (5.8) in thickness, with two ovoid polar capsules of 4.2-5.0 µm (4.7) in length by 2.9-3.8 µm (3.3) in width (n = 15). The shell valves of spores often showed a flattened anterior border and semicircular posterior border, and the two polar capsules were directed toward opposite lateral sides. In addition, the sporoplasm contained an iodinophilous vacuole. Almost complete small-subunit (SSU) rDNA sequences, except for primer flanking regions, were obtained for both species; 1,996 bp long for the former and 1,588 bp long for the latter. On phylogenetic trees based on the SSU rDNA sequences of representative species of Bivalvulida, M. marumotoi n. sp. and C. japonensis n. sp. formed a distinct branch in the Henneguya/Myxobolus clade or near but outside this clade, respectively. This study is the first report of the genetic characterization for the genus Cardimyxobolus.

Myxobolus oralis sp. n. (Myxosporea: Bivalvulida) infecting the palate in the mouth of gibel carp Carassius auratus gibelio (Cypriniformes: Cyprinidae).

During a survey on the myxosporean fauna of gibel carp Carassius auratus gibelio (Bloch) in China, a species of Myxobolus Bütschli, 1882 that did not conform to any known species was found. The species is characterised by the presence of round to ellipsoidal plasmodia of 2.6-4.0 mm in diameter in the palate of host. Mature spores are obovate in frontal view and lemon-shaped in lateral view, with the following range, mean and standard deviation of dimensions: 10.8-12.8 µm (11.7 ± 0.4 µm) long, 8.2-9.9 µm (8.9 ± 0.4 µm) wide and 6.0-7.5 µm (6.8 ± 0.3 µm) thick. Two polar capsules are pyriform, 4.0-5.5 µm (4.8 ± 0.3 µm) long by 2.9-3.6 µm (3.0 ± 0.2 µm) wide. Polar filaments are coiled, with 5 to 6 turns. A small proportion of spores possesses a short caudal process. Scanning electron microscopy revealed discoid spores with a low sutural ridge and middle bulge. The small subunit ribosomal DNA sequence of this species did not match any available sequences in GenBank. Phylogenetically, this species is sister to M. nielii (Nie et Li, 1973) and M. hearti Chen, 1998 in a Henneguya-Myxobolus clade with robust support. Given the morphological and molecular differences between this species and other Myxobolus species, we propose the name Myxobolus oralis sp. n. for this parasite from gibel carp.

Ultrastructure, surface topography, morphology and histological observations of a new parasitic cnidarian of the marbled swamp eel from the world's largest tropical wetland area, Pantanal, Brazil.

Myxosporeans are a diverse group of microscopic cnidarians of wide distribution that evolved into a parasitic lifestyle. A new myxosporean species, Myxobolus sp., is herein described infecting the mandible of wild specimens of Synbranchus marmoratus, caught in the world's largest tropical wetland area, Pantanal, Brazil. Light, scanning, transmission electron microscopy and histological observations unveiled detailed taxonomic information of the new myxosporean cnidarian. Ultrastructural analysis revealed a detailed description of plasmodia structures which can be used for comparison with plasmodia from other species of myxobolids. Both histological and ultrastructural observations evidenced a connective tissue capsule surrounding the plasmodia of Myxobolus sp. as a histopathological host reaction to the infection of this parasitic cnidarian. Histology showed that tissue tropism of the new myxosporean occurs in a well-defined part of the mandible, with development of plasmodia occurring in the epidermis layer. Mature myxospores from the valvular view featured an ovoid shape and had a short prolongation of the spore valves in the posterior end. Myxospores measured 22.7 ± 1.2 µm (21.5-23.9 µm) in length, 12.5 ± 0.4 µm (12.1-12.9 µm) in width and 11.3 ± 0.5 (10.8-11.8 µm) in thickness. Polar capsules were pyriform equally-sized and measuring 4.6 ± 0.3 µm (3.9-4.3 µm) in length and 2.9 ± 0.1 µm in width (2.8-3.0 µm). Finally, this study substantiates the still hidden myxosporean diversity from South America.

Influence of the seasonal and host related factors on the metazoan parasites of Chelon saliens (Mugilidae) in the Turkish coast of the Black Sea.

The leaping mullet Chelon saliens is one of the economically significant fish species and the revealing its parasite fauna in relation with some ecological and host related factors will provide new data for our current knowledge. A total of 165 leaping mullet were collected from Sinop coasts of the Black Sea in the period from September 2015 to August 2016 and investigated for parasites. Eleven parasite species including Myxobolus parvus, Myxobolus sp., Sphaerospora mugilis (Myxozoa), Ligophorus szidati, Solostamenides mugilis (Monogenea), Schikhobalotrema sparisomae, Saccocoelium tensum, Saccocoelium obesum (Digenea), Hysterothylacium aduncum (Nematoda), Neoechinorhynchus sp. (Acanthocephala) and Ergasilus lizae (Copepoda) have been identified. The overall infection prevalence, mean intensity and mean abundance values were 65.5%, 26.2 and 17.2, respectively. The overall infection prevalence was dominated by L. szidati, followed by M. parvus and Digenea-group. On the other hand, the overall mean intensity values were dominated by Digenea-group, followed by L. szidati and E. lizae, respectively, while the mean abundance values were dominated by L. szidati, followed by Digenea-group and E. lizae. The infection indices of all identified parasites were also calculated in relation with length classes and sex of fish as well as season and the differences were evaluated statistically. Seasonally significant differences in the infection prevalence and mean abundance were found for Digenea-group, Ligophorus szidati and Neoechinorhynchus sp. These differences were also significant in the length classes of Digenea-group and Ligophorus szidati. This study is the first investigation on seasonal and host related dynamics of parasites of C. saliens in the southern coasts of the Black Sea and all investigated factors were found to influence the infection indices of dominating parasite species.

Myxozoan infection in thinlip mullet Chelon ramada (Mugiliformes: Mugilidae) in the Sea of Galilee.

Mullets (Mugilidae) are economically important fish in Israel. Two species of mugilids (i.e., the thinlip mullet Chelon ramada and the flathead grey mullet Mugil cephalus) have been stocked in the Sea of Galilee (Lake Kinneret) in order to increase fishermen's income and lake water quality. These catadromous species do not reproduce in the lake, consequently, fingerlings have been introduced every year since 1958. Following a survey of myxozoan infections in the Sea of Galilee, we described Myxobolus pupkoi n. sp. infecting the gill arches, and reported Myxobolus exiguus from visceral peritoneum and gall bladder of C. ramada. The prevalence of infection of both Myxobolus pupkoi n. sp. and M. exiguus were 11.5% (2/23). Our study indicates that the parasites infecting C. ramada belong to a lineage of myxozoans infecting mugilids. This result suggests that the infection took place in the Mediterranean Sea, where the fingerlings were caught, before their introduction into the Sea of Galilee. Since 2018 only farm-raised fingerlings have been introduced. We thus recommend to closely monitor the presence of these parasites in the future to determine if the presence of parasites disappear with the introduction of farm-raised fingerlings.

Correlation of environmental attributes with histopathology of native Yellowstone cutthroat trout naturally infected with Myxobolus cerebralis.

Infection by the invasive parasite Myxobolus cerebralis (causing whirling disease in salmonids) is strongly influenced by a stream's physico-chemical characteristics, which might affect host pathology. We examined whether environmental variables of a M. cerebralis-positive tributary to Yellowstone Lake, Yellowstone National Park, U.S.A., correlated with the histopathology of naturally infected native cutthroat trout Oncorhynchus clarkii bouvieri. Host inflammatory response and cranial cartilage lesions were the main correlates with whirling behavior. Canonical correlation analyses showed that the prevalence of trout with severe lesions in the cranial and jaw cartilages was highest in stream sites with a combination of high temperature and low specific conductivity. Our results reveal that environmental components can affect when and where a pathogen resides within the host, and manifestation of disease. Recognition of the synergism among environmental and histopathology factors most conducive to whirling disease will increase our prediction and detection abilities for M. cerebralis in salmonid hosts.

Morphological and molecular characterization of myxobolids (Cnidaria, Myxozoa) infecting cypriniforms (Actinopterygii, Teleostei) endemic to the Iberian Peninsula.

The Iberian Peninsula provides a unique freshwater ecosystem for native and endemic cypriniforms to thrive. Despite cypriniforms being hosts to multiple myxobolids worldwide, little research has been performed in this geographic location. In this study, the examination of three Iberian endemic cypriniforms showed that myxosporean richness in the Iberian Peninsula is underestimated, with three new and one known myxobolid species being reported based on morphological and molecular data (SSU). Myxobolus arcasii n. sp. is described from the kidney and gonads of the "bermejuela" Achondrostoma arcasii, M. duriensis n. sp. from the gills of the Northern straight-mouth nase Pseudochondrostoma duriense, and Thelohanellus paludicus n. sp. from the intestine of the Southern Iberian spined-loach Cobitis paludica. Myxobolus pseudodispar Gorbunova, 1936 is further reported from several organs of P. duriense, and from the spleen of A. arcasii. The occurrence of M. pseudodispar in endemic Iberian species reveals that host-shift followed its co-introduction with central European leuciscids into this geographic location. Several other myxobolids originally described from barbels in central Europe have also been reported from the Iberian endemic cypriniform Luciobarbus bocagei. Nonetheless, except for M. musculi, the identification of these myxobolids in L. bocagei is here shown to be dubious and require molecular confirmation. Phylogenetic analyses reveal M. arcasii n. sp. and M. duriensis n. sp. clustering within different lineages of leuciscid-infecting species, showing that myxobolids entered Leuciscidae as hosts multiple times during their evolution. Constituting the first myxobolid reported from the subfamily Cobitinae, Thelohanellus paludicus n. sp. stands alone in the tree topology.