Taxonomy on three novel species of Sphaeromyxa Thélohan 1892 (Myxozoa, Bivalvulida, Sphaeromyxidae) with insight into the evolution of the genus.

Three new myxosporeans of the genus Sphaeromyxa Thélohan 1892 were discovered from the coastal waters off Xiamen in the East China Sea and characterized based on morphological and SSU rDNA data. Sphaeromyxa photopectoralis sp. n. was described from Photopectoralis bindus, and Sphaeromyxa sebastisca sp. n. was described infecting both Sebastiscus marmoratus (type-host) and Scorpaenopsis cirrosa. These two species are morphologically consistent with the "balbianii" group, possessing straight myxospores and truncated ends, but are distinct from one another genetically and by myxospore dimensions. A third myxosporean infecting Siganus fuscescens was described as Sphaeromyxa xiamenensis sp. n., and this species is morphologically consistent with the "incurvata" group, bearing arcuate myxospores with rounded ends. The molecular phylogeny and estimated rRNA secondary structure suggest that marine sphaeromyxids are probably derived from freshwater myxidiids, and "incurvata" and "balbianii" groups might each represent independent evolutionary lineages. The present study also shows that S. limocapitis phylogenetically nested in "incurvata" group.

Identification of four new Kudoa spp. (Myxozoa: Myxosporea: Multivalvulida) in commercial fishes collected from South China Sea, Atlantic Ocean, and Bering Sea by integrated taxonomic approach.

Members of the myxosporean genus Kudoa are defined as having a myxospore with four or more shell valves (SVs) and a corresponding number of polar capsules (PCs). Here, we employed integrated taxonomic approaches to four kudoid isolates from Acentrogobius chlorostigmatoides and Konosirus punctatus from the South China Sea, off Guangdong, Pentanemus quinquarius from the Southeast Atlantic Ocean, off West African coast, and Atheresthes stomias from the Bering Sea, off Alaska, and concluded that all these kudoids were novel species, named Kudoa acentrogobia n. sp., Kudoa guangdongensis n. sp., Kudoa iidae n. sp., and Kudoa aburakarei n. sp., respectively. Kudoa guangdongensis n. sp., forming pseudocysts in the trunk muscle myofibers of K. punctatus, had large-sized tripod myxospores with three wing-like SV extensions and three PCs (one prominent PC and two rudimentary PCs). Phylogenetic analyses based on the 18S and 28S ribosomal RNA gene (rDNA) demonstrated its affinity to the genus Kudoa, not to Unicapsula characterized by a myxospore with one prominent PC and two rudimentary PCs, suggesting the atypical nature of this new species in the context of myxospore morphology. Three other kudoid isolates had four SVs and PCs in a semi-quadrated, ray-like, or cruciform myxospore, respectively, forming pseudocysts in the trunk muscle myofibers. Kudoa iidae n. sp. forming pseudocysts in the muscles of P. quinquarius from Southeast Atlantic Ocean had unique myxospores with ray-like form, showing close morphological resemblance to Kudoa rayformis, which were recorded from the muscle of Scomberomorus sierra from the Pacific Ocean off Panama. These two species had a phylogenetic relationship of morphological convergence, evolving separately in different sea areas. It is fairly difficult to differentiate K. acentrogobia n. sp. and K. aburakarei n. sp., prevalent in their host fishes, from kudoid species with similar myxospore morphology (e.g., Kudoa nova and Kudoa thyrsites, respectively), but distinct in phylogeny from known Kudoa spp. Combined taxonomic identification of multivalvulid myxosporeans based on both morphological criteria of isolated myxozoans and their molecular characterization could disclose their real biodiversity and phylogeny.

Characterization of Myxidium spinibarba sp. nov. (Cnidaria, Myxosporea, Myxidiidae) from Spinibarbus sinensis (Bleeker, 1871) in Chongqing China.

In the present study, we described a new species of Myxidium Bütschli, 1882, obtained from the gallbladder of Spinibarbus sinensis (Bleeker, 1871) from the Jialing River in Chongqing, China. Myxidium spinibarba sp. nov. was identified based on morphological and SSU rDNA sequence data. The mature myxospores were fusiform in valvular view and ovoid in sutural view, with somewhat protrusive poles and mean dimensions (all in µm) of 11.8 ± 0.5 (10.6-12.4) in length and 6.1 ± 0.5 (5.5-7.2) in width. The polar capsules were pyriform and equal in size with mean dimensions of 3.6 ± 0.4 (3.0-4.4) in length and 3.0 ± 0.2 (2.7-3.2) in width. The new species was distinct from related species of Myxidium in its morphology and molecular characteristics. Phylogenetic analysis indicated the clustering of species based on the presence or absence of valvular striations. Moreover, myxospore morphology, rather than the host environment, played an important role in the partial phylogenetic clustering.

Coelozoic parasite of the family Ceratomyxidae (Myxozoa, Bivalvulida) described from motile vermiform plasmodia found in Hemiodus unimaculatus Bloch, 1794.

Myxozoans of the family Ceratomyxidae are common coelozoic parasites of marine, anadromous, and freshwater fish, and may also be found, less frequently, parasitizing the tissue of these hosts. The diversity and ecology of the freshwater species of the genus Ceratomyxa have been poorly investigated, leading to a knowledge gap that restricts the understanding of the distribution and prevalence of this group of parasites. In the present study, parasites were found inside vermiform plasmodia, characterised by oscillatory movements in the characiform species Hemiodus unimaculatus. The crescent-shaped and elongated spores, perpendicular to the suture line, have a mean length of 28.9 ± 2.7 µm and width of 2.6 ± 0.1 µm, with two symmetrical oval polar capsules, 1.9 ± 0.3 µm in length and 1.7 ± 0.2 µm in width, containing polar filaments with three or four coils, located near the central suture, with symmetrical lateral elongations 14.3 ± 1.1 µm in length and binucleate amoeboid sporoplasm. The integrated comparative analysis of the morphological characteristics and partial SSU rRNA sequences supported the identification of a new species of coelozoic Ceratomyxa, found in the gallbladder of H. unimaculatus, from the Tocantins basin, in the municipalities of Estreito and Imperatriz in eastern Brazilian Amazonia.The new species was denominated Ceratomyxa fonsecai n. sp.

Ceratomyxa mennani n. sp. (Myxosporea: Bivalvulida) parasitizing the gallbladder of the dusky grouper Epinephelus marginatus (Serranidae) from Tunisian waters.

Ceratomyxa mennani n. sp. is a new coelozoic Ceratomyxa species found in the gallbladder of Epinephelus marginatus from the Gulf of Tunis, Tunisia. Mature plasmodia were disporic, ovoid in shape measuring 9-12 µm in width and 11-14 µm in length. Mature myxospores were slightly crescent-shaped with almost straight posterior margin, measuring 5.8 ± 0.2 (5.4-6.1) µm in length and 12.7 ± 0.3 (11.9-13.0) µm in thickness. The two valves were unequal with rounded ends. Polar capsules were spherical, equal in size with 2.1 ± 0.2 (1.9-2.6) µm in diameter. The binucleated sporoplasm filled the entire cavity of the myxospore. Molecular analysis of SSU rDNA sequences indicated that C. mennani n. sp. was distinct from all other Ceratomyxa sequences in GenBank. Phylogenetic analysis revealed that C. mennani n. sp. clustered with Ceratomyxa species infecting Epinephelinae fishes. Seasonal prevalence of infection over one year was significantly higher in winter and the lowest in autumn. This is the third report of Ceratomyxa species infecting the gallbladder of Epinephelus marginatus from Tunisia and the first study to include molecular data.

Effect of Epigallocatechin Gallate on Viability of Kudoa septempunctata.

Kudoa septempunctata have been reported as a causative agent for acute transient gastrointestinal troubles after eating raw olive flounder (Paralichthys olivaceus). It raised public health concerns and quarantine control in several countries. Quantitative evaluation on viability of K. septempunctata is crucial to develop effective chemotherapeutics against it. A cytometry using fluorescent stains was employed to assess effect of three compounds on viability of K. septempunctata. Epigallocatechin gallate reduced markedly viability of K. septempunctata at 0.5 mM or more, and damaged K. septempunctata spores by producing cracks.

Skin nodules associated with parasitism with Henneguya sp. (Cnidaria: Myxosporea) in the neotropical fish Cyphocharax modestus.

A new myxozoan species, Henneguya sp., is described based on material from skin of Cyphocharax modestus. Mature myxospores are were elongate and ellipsoidal, measuring 21.4 ±â€¯1.2 (19.4-23.2) µm in total length, 5.1 ±â€¯0.3 (4.5-5.8) µm in width, 11.9 ±â€¯0.5 (10.9-12.7) µm in body length and 9.6 ±â€¯0.7 (8.4-10.5) µm in length of the caudal process. The polar capsules were elongated and had unequal sizes, with length of 5.1 ±â€¯0.4 (4.5-6.0) µm and 5.6 ±â€¯0.4 (4.9-6.3) µm for smaller and larger respectively and width of 1.8 ±â€¯0.2 (1.4-2.0) µm. The larger polar capsule had 8 turns in polar filament while the smaller polar capsule had 5 turns in polar filament. The macroscopic analysis revealed the presence of large nodules, which were located before and after the dorsal fin of the hosts. The histopathological analysis showed the development of nodules filled with plasmodia, surrounded by loose connective tissue, developed in the dermis of the skin. Many cysts containing countless spores, as well as free spores, were located in the dermis and hypodermis of the hosts, causing the disorganization of the connective tissue that is responsible for the support. This is the first record of a Henneguya species in C. modestus.

A new myxozoan species Henneguya unitaeniata sp. nov. (Cnidaria: Myxosporea) on gills of Hoplerythrinus unitaeniatus from Mato Grosso State, Brazil.

On the basis of morphological and molecular analyses, a new myxozoan parasite is described from the gills of the fish Hoplerythrinus unitaeniatus, collected in the municipality of Nova Xavantina, Mato Grosso State, Brazil. Plasmodia of Henneguya unitaeniata sp. nov. were oval and whitish and were found surrounded by collagen fibers forming plasmodia wall between gill filaments on the gill arch. The spores were ellipsoidal with two similar polar capsules. Morphometric analysis showed a total spore mean length of 23.8 ± 1.5 µm, spore body mean length of 14.5 ± 0.7 µm, caudal appendage mean length of 10.3 ± 1.4 µm, thickness mean length of 4.3 ± 0.3 µm, polar capsule mean length of 4.2 ± 0.5 µm, polar capsule mean width of 1.8 ± 0.3 µm, spore mean width of 4.8 ± 0.4 µm, and 4-5 polar filament coils. Phylogenetic analysis showed Henneguya unitaeniata sp. nov. as a basal species in a subclade formed by myxozoans that parasitize bryconid fishes.

Sinuolinea capsularis (Myxosporea: Sinuolineidae) Isolated from Urinary Bladder of Cultured Olive Flounder Paralichthys olivaceus.

Sinuolinea capsularis Davis, 1917 is myxosporean that infect the urinary system of the host fish. Insufficient morphological and molecular data of S. capsularis exits, and it is therefore difficult to make an accurate identification of the parasite. We tried a series of morphological and molecular analysis to identify an myxosporean isolated from urinary bladder of cultured olive flounder, Paralichthys olivaceus, from Jeju island in the Republic of Korea. Some of them were observed under a light microscope and SEM, and remain samples were used molecular and phylogenetic analysis. Mature spores were subspherical, measuring 13.9±0.6 µm in length and 13.8±0.8 µm in width. Two spherical polar capsules on opposite sides in the middle of the spore had a diameter range of 4.3±0.4 µm. Scanning electron microscopy revealed that spores a severely twisted the suture line. By the morphological comparison and analysis, it was identified as S. capsularis. In addition, we obtained the partial 18S rDNA of S. capsularis and first registered it in NCBI. Phylogenetic analysis showed that S. capsularis clustered with Zschokkella subclade infecting the urinary system of marine fish, and it supported the infection site tropism effect on phylogeny of marine myxosporeans as well as the origin of Sinuolinea is not monophyly.

Identification of a new myxosporean parasite Thelohanellus indiana n. sp. (Myxosporea: Myxobolidae) isolated from three major organs of goldfish, Carassius auratus L. highlighted with its morphological and SSU rDNA sequence based molecular description.

Fish mortality and poor growth in surviving fish contribute substantial losses to the ornamental fish farms of India and revealed an infection of a new myxosporidian parasite Thelohanellus indiana n. sp. which has become one of the most important limiting factors for successful aquaculture management. The parasite infects Carassius auratus, an Indian goldfish, described on the basis of myxospores morphology and amplification of a part of 18 S rDNA gene. Three major attaching site of fish body have been explored for showing the location of attachment for the parasites. The whitish cysts of the parasites are about 2.5-3.5 mm contains large amount of lemon shaped mature myxospores measuring 12.1-15.2 (13.8) × 7.5-8.8 (8) µm. A single round or elliptical polar capsule located only at the anterior pole of the spore having 6.2-7.2 (6.8) × 3.3-4.7 (4.0) µm in diameter. The morphological characters have been assessed by both the light and scanning electron microscope. The most differentiating feature from closely related species was carried out by morpho-taxonomic affinities with previously described species which are tremendously supported by molecular taxonomy by partial sequencing of the 18 S rDNA gene resulted in a total of 2101 bp fragment of newly obtained SSU rRNA gene sequence of the new species which exhibit 79-91% homogeneity with other closely related species available in GenBank. The BLAST search of Thelohanellus sp. did not matches with any available sequences in GenBank and the phylogenetic analysis revealed that the novel species were sister to T. habibpuri and T. caudatus, in the Thelohanellus clade and form a closest neighboring branch as a subclade in phylogenetic tree from which the new Thelohanellus parasite is being placed. Both the branches are originating from monophyletic clade that are strongly supported by bootstrap values which indicate clearly about independent position of T. indiana n. sp.

Molecular phylogeny of the gill parasite Henneguya (Myxosporea: Myxobolidae) infecting Astyanax lacustris (Teleostei: Characidae) from fish farm in Brazil.

Molecular data of Henneguya chydadea Barassa, Cordeiro and Arana, 2003, found in the gill filaments of Astyanax lacustris bred in fish farm in the State of Mato Grosso do Sul, Brazil was obtained in order to estimate their phylogenetic position among other platysporines myxosporean. The prevalence of the parasite was 28.1% and the range intensity was 1-3 plasmodia per fish. The shape and measurements of mature myxospores were consistent with the characteristics previously defined to H. chydadea. The SSU rDNA sequence of the myxospores of H. chydadea resulted in a total of 1405 nucleotides, and this sequence did not match any of the myxozoan available in the GenBank. Phylogenetic analysis showed H. chydadea within the clade of histozoic myxosporeans and closed together with Henneguya rotunda and Myxobolus pantanalis reported in the gill arch and fins and gill filaments of Salminus brasiliensis respectively. Nonetheless, the SSU rDNA sequences of H. chydadea, H. rotunda and M. pantanalis have only 85.2% and 84.4% similarity, respectively. This is the first molecular study of a Henneguya species that parasitizes a fish belonging to the genus Astyanax in South America. The importance of myxosporeans introduction to new locations along with infected cultured host is emphasized.

18S and 28S rDNA identity and phylogeny of two novel myxosporeans infecting gills of cyprinid carps inhabiting a cold water wetland in northern India.

The sequences of 18S and 28S rDNA have been extensively used as molecular markers to resolve phylogenetic relationships among various myxosporeans infecting fishes. Two novel myxosporeans have been identified during the present study namely, Henneguya latiusii sp. nov. infecting gills of Crossocheilus latius and Myxobolus atkinsoni sp. nov. infecting gills of Labeo rohita inhabiting Ranjit Sagar Wetland, Punjab (India). These myxosporeans formed plasmodia in the gill arch and gill lamellae respectively and infection was moderate (gill plasmodium index, GPI: 2). Myxospores of both the species possessed distinct morphological and morphometric characteristics hence validated as new species. Phylogenetically, H. latiusii sp. nov. was closest to H. doneci infecting gill filaments of Carassius auratus gibelio, with 96% and 74% similarity in 18S and 28S rDNA respectively. Similarly, M. atkinsoni sp. nov. showed close homogeneity of 90% with M. turpisrotundus and 83% with M. nielii infecting gills of Carassius auratus. The phylogenetic analyses of species of Myxobolus and Henneguya showed a tendency to cluster according to the order or family of the host, tissue and geographical location. This study is the first report on LSU gene marker (28S rDNA) standardized on myxozoans infecting fresh water fishes from Indian subcontinent.

Morphology and molecular phylogeny of Ortholinea mullusi sp. nov. (Myxozoa) in Mullus barbatus from the Black Sea.

Myxosporeans of the genus Ortholinea have a worldwide distribution and infect organs and tissues of exclusively marine fishes. Here we describe the morphological and molecular characteristics of Ortholinea mullusi sp. nov. parasitizing the urinary bladder and kidney tubules of red mullet Mullus barbatus collected from the coastal zone of Sinop in the Black Sea, Turkey. Polysporic plasmodia with immature spores were either elongate, 37.0 ± 4.5 SD (30-50) µm long and 45.0 ± 3.8 (40-55) µm wide, or were round, up to 100.0 µm in diameter. Mature, free spores were spherical in the frontal view and measured 9.3 ± 0.2 (9.0-9.7) µm in length, 8.7 ± 0.3 (8.2-9.3) µm in width and 7.7 ± 0.1 (7.5-7.9) µm in thickness. We observed 2 polar capsules of equal size, which measured 3.1 ± 0.1 (3.0-3.2) µm long by 2.5 ± 0.1 (2.4-2.6) µm wide, and the tips of the polar capsules were open towards the sutural line. The prevalence of infection by O. mullusi sp. nov. was 24.5%. Phylogenetic analysis based on nuclear small subunit ribosomal DNA (SSU rDNA) clearly suggested O. mullusi to be a new species, clustered within a lineage comprising O. labracis and O auratae. Pairwise nucleotide similarities and DNA distance values between O. mullusi sp. nov. and sister Ortholinea species also supported this suggestion.

Synopsis of the species of Ceratomyxa Thélohan, 1892 (Cnidaria, Myxosporea, Ceratomyxidae) described between 2007 and 2017.

A synopsis of the species of Ceratomyxa Thélohan, 1892 (Cnidaria, Myxosporea, Ceratomyxidae) described between 2007 and 2017 is presented, including 83 nominal species. For each species, the most important morphological and morphometric features are indicated in tabulated format. Included are also the site of infection within the host, and geographical location, plus a full bibliography of the original records of species. Molecular data (GenBank accession numbers) are also provided whenever possible.

Henneguya laseeae n. sp. from flathead catfish (Pylodictis olivaris) in the upper Mississippi River.

A novel species of Henneguya was isolated from flathead catfish (Pylodictis olivaris) captured in the upper Mississippi River near Lansing (Allamakee County), IA, and La Crosse (La Crosse County), WI. Designated Henneguya laseeae n. sp., this novel species is described using critical morphological features, histology, and 18S ribosomal RNA gene sequence. Ovoid cysts, ranging from 1200 to 1800 µm in width, tended to be at filament tips or in the distal third, often directly on the filament midline, but occasionally paramedian. Lanceolate-shaped myxospores were consistent with those of the genus Henneguya. The spore body was 16.2 ± 0.5 µm (mean ± standard deviation; range = 15.1-17.0 µm) in length, 6.0 ± 0.4 µm (5.1-6.6 µm) in width, and 4.7 ± 0.2 µm (4.4-4.9 µm) thick. The two polar capsules at the anterior of the spore body were 5.9 ± 0.3 µm (5.3-6.3 µm) in length and 1.8 ± 0.1 µm (1.6-2.1 µm) in width and contained six to seven turns in the polar filament. The caudal processes tapered to fine points and were 54.3 ± 2.9 µm (49.1-61.7 µm) in length. Total spore length was 70.4 ± 3.3 µm (64.5-79.4 µm). The spores and plasmodium of this species are of similar size and morphology to other species of Henneguya from ictalurid fishes. Additionally, the 18S rRNA gene sequences placed this isolate within a clade populated by Henneguya spp. from North American ictalurids. This is the first reported species of Henneguya from flathead catfish.

Observations on non-random distribution of spores of Henneguya spp. (Cnidaria: Myxosporea: Myxobolidae) within plasmodia.

Species of the cnidarian genus Henneguya Thélohan, 1892 (Myxosporea: Myxobolidae) are histozoic parasites commonly found in freshwater and, more rarely, in marine fish. The development of these parasites in fish tissues includes the formation of plasmodia within which occurs the sporogony originating spores with two caudal processes, which are usually randomly distributed within the plasmodia. In this report the authors present some cases of non-random distribution of the spores of six species of Henneguya within their plasmodia. Two different patterns of non-random distribution were found based on a literature survey. These patterns and their origin are discussed. Apparently this non-random distribution of the spores is due to both internal and external factors.

Description of a new Neoactinomyxum type actinosporean from the oligochaete Branchiura sowerbyi Beddard.

Actinosporean infection of oligochaetes living in the mud of a commercial gibel carp pond with myxosporean disease was studied. Six actinospore types were detected exclusively from the oligochaete Branchiura sowerbyi Beddard with very high prevalence (18%). Five out of the six types were identified as the same actinosporeans described in previous reports, the sixth actinosporean was identified as a new Neoactinomyxum type and described here based on morphological and molecular characterisation. Spore body of the actinospore was globular, much smaller than caudal processes. Three caudal processes were disc-like in apical view, hemispherical in side view, closer together and encircling the spore body. The number of sporoplasm cells was detected as eight in one specimen. The new actinosporean markedly differed from other Neoactinomyxum types in literature having much bigger caudal processes. DNA sequence analyses further confirmed the morphological identification, and revealed the actinosporean described here (KU641392) possessed less than 94% sequence similarity with myxozoans available in the GenBank database.

Tetractinomyxon stages genetically consistent with Sphaerospora dicentrarchi (Myxozoa: Sphaerosporidae) found in Capitella sp. (Polychaeta: Capitellidae) suggest potential role of marine polychaetes in parasite's life cycle.

Known life cycles of myxosporean parasites have two hosts, but very few life cycles have been disclosed, especially in the marine environment. Sphaerospora dicentrarchi Sitjà-Bobadilla and Álvarez-Pellitero, 1992 is a systemic parasite from the European seabass, Dicentrarchus labrax (Linnaeus, 1758), a highly valuable commercial fish. It affects its health, leading to aquaculture production losses. During 2013 and 2014, an actinospore survey was conducted in a total of 5942 annelids collected from a fish farm in Algarve and from the Aveiro Estuary, in Portugal. A new tetractinomyxon actinospore was found in a capitellid polychaete, belonging to the genera Capitella collected at the fish farm. The tetractinomyxons were pyriform measuring 11·1 ± 0·7 µm in length and 7·2 ± 0·4 µm in width, and presented three rounded polar capsules measuring 2·4 ± 0·3 µm in diameter. The molecular analysis of the 18S rRNA gene sequences from the tetractinomyxons revealed a similarity of 100% with the DNA sequences deposited in the GenBank from S. dicentrarchi myxospores collected from the European seabass and the spotted seabass in the same fish farm and 99·9% similarity with the DNA sequence obtained from the myxospores found infecting the European seabass in the Aveiro Estuary. Therefore, the new tetractinomyxons are inferred to represent the actinospore phase of the S. dicentrarchi life cycle.

Description of new types of sphaeractinomyxon actinospores (Myxozoa: Myxosporea) from marine tubificid oligochaetes, with a discussion on the validity of the tetraspora and the endocapsa as actinospore collective group names.

Ten new types of sphaeractinomyxon actinospores are morphologically and molecularly described from the coelomic cavity of two marine oligochaete hosts, Limnodriloides agnes (Hrabe, 1967) and Tubificoides pseudogaster (Dahl, 1960), from Aveiro estuary, Portugal. The smallest sphaeractinomyxon type measured 17 µm (length) × 19 µm (width) × 19 µm (apical diameter), whereas the largest type measured 61 µm × 76 µm × 80 µm. While considering the 10 types of sphaeractinomyxon, it was found that the number of spores developing inside pansporocysts varied between 1, 2, 4, and 8. The total prevalence of infection was of 19 % for the two host species, with a maximum recorded for spring and summer (25-26 %). While considering each type of sphaeractinomyxon individually, it was found that the prevalence values ranged between 0.3 and 1.7 %. All described sphaeractinomyxons were most similar to Myxobolus species. The validity of the tetraspora and endocapsa collective group names is discussed.

Morphology, seasonality and phylogeny of Zschokkella trachini n. sp. (Myxozoa, Myxosporea) infecting the gallbladder of greater weever Trachinus draco (L.) from Tunisian waters.

We describe a new myxosporean species, Zschokkella trachini n. sp., infecting the gallbladder of greater weever Trachinus draco Linnaeus 1758 from Tunisia. This is the first record of Zschokkella species in T. draco. Plasmodia were polysporic producing six to eight mature spores; they were attached to the gallbladder epithelium or free floating in the bile. Mature spores were sub-ovoid in the frontal view, measured 15.2 ± 0.6 (14.4-16.0) µm in length and 9.8 ± 0.7 (9.0-10.8) µm in width. Two equal spherical polar capsules 4.0 ± 0.4 (3.6-4.5) µm in diameter, were located separately at the spore's extremities. The prevalence of infection ranged from 23.5 to 87.7 %. Morphological data and molecular analysis of the small subunit rDNA gene identified this parasite as a new species of Zschokkella. Neighbour joining clustered the parasite in a sub-clade containing other Zschokkella species parasiting the gallbladder of marine fish hosts, located within the coelozoic clade of the major freshwater clade. This is the second Zschokkella species reported from Tunisia.