Edson A. Adriano

Molecular phylogeny of the Myxobolus and Henneguya genera with several new South American species.

The present study consists of a detailed phylogenetic analysis of myxosporeans of the Myxobolus and Henneguya genera, including sequences from 12 Myxobolus/Henneguya species, parasites of South American pimelodids, bryconids and characids. Maximum likelihood and maximum parsimony analyses, based on 18 S rDNA gene sequences, showed that the strongest evolutionary signal is the phylogenetic affinity of the fish hosts, with clustering mainly occurring according to the order and/or family of the host. Of the 12 South American species studied here, six are newly described infecting fish from the Brazilian Pantanal wetland. Henneguya maculosus n. sp. and Myxobolus flavus n. sp. were found infecting both Pseudoplatystoma corruscans and Pseudoplatystoma reticulatum; Myxobolus aureus n. sp. and Myxobolus pantanalis n. sp. were observed parasitizing Salminus brasiliensis and Myxobolus umidus n. sp. and Myxobolus piraputangae n. sp. were detected infecting Brycon hilarii.

Henneguya pseudoplatystoma n. sp. causing reduction in epithelial area of gills in the farmed pintado, a South American catfish: Histopathology and ultrastructure

The present study is part of an ongoing investigation into the characteristics of Myxozoan parasites of Brazilian freshwater fish and was carried out using morphology, histopathology and electron microscopy analysis. A new Myxosporea species (Henneguya pseudoplatystoma) is described causing an important reduction in gill function in the farmed pintado (a hybrid fish from a cross between Pseudoplatystoma corruscans and Pseudoplatystoma fasciatum), which is a commercially important South American catfish. From a total of 98 pintado juveniles from fish farms in the states of São Paulo and Mato Grosso do Sul (Brazil), 36 samples (36.7%) exhibited infection of the gill filaments. Infection was intense, with several plasmodia occurring on a same gill filament. The plasmodia were white and measured up to 0.5mm in length; mature spores were ellipsoidal in the frontal view, measuring 33.2+/-1.9 microm in total length, 10.4+/-0.6 microm in body length, 3.4+/-0.4 microm in width and 22.7+/-1.7 microm in the caudal process. The polar capsules were elongated, measuring 3.3+/-0.4 microm in length and 1.0+/-0.1 microm in width and the polar filaments had six to seven turns. Histopathological analysis revealed the parasite in the connective tissue of the gill filaments and lamella. No inflammatory process was observed, but the development of the plasmodia reduced the area of functional epithelium. Ultrastructural analyses revealed a single plasmodial wall, which was in direct contact with the host cells and had numerous projections in direction of the host cells as well as extensive pinocytotic canals. A thick layer (2-6 microm) of fibrous material and numerous mitochondria were found in the ectoplasm. Generative cells and the earliest stage of sporogenesis were seen more internally. Advanced spore developmental stages and mature spores were found in the central portion of the plasmodia.

Myxobolus cordeiroi n. sp., a parasite of Zungaro jahu (Siluriformes: Pimelodiade) from Brazilian Pantanal: Morphology, phylogeny and histopathology☆

This work is part of an ongoing investigation into the characteristics of Myxozoan parasites of freshwater fish in Brazil and was carried out using morphology, histopathology and molecular analysis. A new Myxosporea species (Myxobolus cordeiroi) is described infecting the jaú catfish (Zungaro jahu). Fifty jaú specimens were examined and 78% exhibited plasmodia of the parasite. The plasmodia were white and round, measuring 0.3-2.0mm in diameter and the development occurred in the gill arch, skin, serosa of the body cavity, urinary bladder and eye. The spores had an oval body and the spore wall was smooth. Partial sequencing of the 18S rDNA gene resulted in a total of 505bp and the alignment of the sequences obtained from samples in different organs revealed 100% identity. In the phylogenetic analysis, the Myxobolus species clustered into two clades-one primarily parasites of freshwater fish and the other primarily parasites of marine fish. M. cordeiroi n. sp. was clustered in a basal position in the freshwater fish species clade. The histological analysis revealed the parasite in the connective tissue of the different infected sites, thereby exhibiting affinity to this tissue. The plasmodium was surrounded by an outer collagen capsule of fibers with distinct orientation from the adjacent connective tissue and an inner layer composed of delicate collagen fibrils-more precisely reticular fibers. The development of the parasite in the cornea and urinary bladder caused considerable stretching of the epithelium.

Phylogenetic and host-parasite relationship analysis of Henneguya multiplasmodialis n. sp infecting Pseudoplatystoma spp. in Brazilian Pantanal wetland

A new species of the genus Henneguya (Henneguya multiplasmodialis n. sp.) was found infecting the gills of three of 89 specimens (3.3%) of Pseudoplatystoma corruscans and two of 79 specimens (2.6%) of Pseudoplatystoma reticulatum from rivers in the Pantanal wetland, Brazil. Partial sequencing of the 18S rDNA gene of the spores obtained from one plasmodium from the gills of P. corruscans and other one from the gills of P. reticulatum, respectively, resulted in a total of 1560 and 1147 base pairs. As the spores of H. multiplasmodialis n. sp. resemble those of Henneguya corruscans, which is also a parasite of P. corruscans, sequencing of the 18S rDNA gene of the spores of H. corruscans found on P. corruscans caught in the Brazilian Pantanal wetland was also provided to avoid any taxonomic pendency between these two species, resulting in 1913 base pairs. The sequences of H. multiplasmodialis n. sp. parasite of P. corruscans and P. reticulatum and H. corruscans did not match any of the Myxozoa available in the GenBank. The similarity of H. multiplasmodialis n. sp. obtained from P. corruscans to that from P. reticulatum was of 99.7%. Phylogeny revealed a strong tendency among Henneguya species to form clades based on the order and/or family of the host fish. H. multiplasmodialis n. sp. clustered in a clade with Henneguya eirasi and H. corruscans, which are also parasites of siluriforms of the family Pimelodidae and, together with the clade composed of Henneguya spp. parasites of siluriforms of the family Ictaluridae, formed a monophyletic clade of parasites of siluriform hosts. The histological study revealed that the wall of the plasmodia of H. multiplasmodialis n. sp. were covered with a stratified epithelium rich in club cells and supported by a layer of connective tissue. The interior of the plasmodia had a network of septa that divided the plasmodia into numerous compartments. The septa were composed of connective tissue also covered on both sides with a stratified epithelium rich in club cells. Inflammatory infiltrate was found in the tissue surrounding the plasmodia as well as in the septa.

Host–parasite–environment relationship, morphology and molecular analyses of Henneguya eirasi n. sp. parasite of two wild Pseudoplatystoma spp. in Pantanal Wetland, Brazil☆

A new myxosporean species, Henneguya eirasi n. sp., is described parasitizing the gill filaments of Pseudoplatystoma corruscans and Pseudoplatystoma fasciatum (Siluriformes: Pimelodidae) caught in the Patanal Wetland of the state of Mato Grosso, Brazil. The parasite formed white, elongated plasmodia measuring up to 3mm. Mature spores were ellipsoidal in the frontal view, measuring 37.1 ± 1.8 μm in total length, 12.9 ± 0.8 μm in body length, 3.4 ± 0.3 μm in width, 3.1 ± 0.1 μm in thickness and 24.6 ± 2.2 μm in the caudal process. Polar capsules were elongated and equal in size, measuring 5.4 ± 0.5 μm in length and 0.7 ± 0.1 μm in width. Polar filaments had 12-13 coils. Histopathological analysis revealed that the parasite developed in the sub-epithelial connective tissue of the gill filaments and the plasmodia were surrounded by a capsule of host connective tissue. The plasmodia caused slight compression of the adjacent tissues, but no inflammatory response was observed in the infection site. Ultrastructure analysis revealed a single plasmodial wall connected to the ectoplasmic zone through numerous pinocytotic canals. The plasmodial wall exhibited numerous projections and slightly electron-dense material was found in the ectoplasm next to the plasmodial wall, forming a line just below the wall. Partial sequencing of the 18S rDNA gene of H. eirasi n. sp. obtained from P. fasciatum resulted in a total of 1066 bp and this sequence did not match any of the Myxozoa available in the GenBank. Phylogenetic analysis revealed the Henneguya species clustering into clades following the order and family of the host fishes. H. eirasi n. sp. clustered alone in one clade, which was the basal unit for the clade composed of Henneguya species parasites of siluriform ictalurids. The prevalence of the parasite was 17.1% in both fish species examined. Parasite prevalence was not influenced by season, host sex or host size.

Phylogeny, ultrastructure, histopathology and prevalence of Myxobolus oliveirai sp. nov., a parasite of Brycon hilarii (Characidae) in the Pantanal wetland, Brazil

This paper presents the morphological, histological and ultrastructural characteristics of Myxobolus oliveirai sp. nov., a parasite of the gill filaments in Brycon hilarii from the Brazilian Pantanal. Out of 216 B. hilariispecimens examined (126 wild and 90 cultivated), 38.1% of wild specimens (n = 48) were infected. The parasites form elongated plasmodia primarily in the tip of gill filaments, reaching about 3 mm in length. A thorough comparison with all the Myxobolus species described from South American hosts, as well as nearly all the Myxobolus species described so far is provided. Partial sequencing of the 18S rDNA gene revealed a total of 1,527 bp. The Myxobolus species parasite of B. hilarii did not match any of the Myxozoa available in GenBank. In the phylogenetic analysis, M. oliveirai sp. nov. composed a monophyletic group with eight other species: five species of Myxobolus parasites of mugilid fishes, two parasites of pangasiid and one of centrarchid. Infection prevalence values of the parasite revealed no significant differences between wet and dry seasons or between males and females. The importance of the infection to the farming of the host species is emphasized.

Light, electron microscopy and histopathology of Myxobolus salminus n. sp., a parasite of Salminus brasiliensis from the Brazilian Pantanal.

In this report, we describe the morphology and histopathology of Myxobolus salminus n. sp., a parasite of the gill filaments of wild Salminus brasiliensis (dourado) from the Brazilian Pantanal. The small polysporic plasmodia were approximately 100 microm in diameter and the development was asynchronous. The mature spores were oval to pear shaped and had a smooth wall. The spore measurements were (mean+/-S.D., with range in parentheses): length 10.1+/-0.4 microm (9.6-10.5), width 6.1+/-0.4 microm (5.8-6.6) and thickness 5.0+/-0.6 microm (4.7-5.3). The polar capsules were elongated and of equal size: length 4.6+/-0.2 microm (4.3-4.8) and width 1.7+/-0.1 microm (1.5-1.9). The histological analysis revealed numerous plasmodia in the blood vessels of the gill filaments. The site of parasite development was the wall of the large-caliber blood vessel of the gill filament, with progressive growth towards the lumen, resulting in the obstruction of blood flow, congestion and perivascular edema. The ultrastructural study revealed that the plasmodial wall was composed of two membranes, had numerous pinocytic canals and was in direct contact with the basement membrane of the vessel. The development of the parasite was asynchronous, with mature spores, immature spores and young developmental stages randomly distributed throughout the plasmodium. The prevalence of the parasite was 4.4%, with male and female fish being infected.

Brevimulticaecum sp. (Nematoda: Heterocheilidae) larvae parasitic in freshwater fish in the Pantanal wetland, Brazil

During the study of metazoan parasites of freshwater fish in the Pantanal wetland (state of Mato Grosso do Sul, Brazil), third-stage larvae of Brevimulticaecum sp. were collected from the abdominal cavity, liver parenchyma, mesentery, skeletal muscles and stomach wall of the following species: Gymnotus inaequilabiatus, Hemisorubim platyrhynchos, Hoplias aff. malabaricus, Myleus levis, Pseudoplatystoma corruscans, Pygocentrus nattereri, Serrasalmus marginatus and Sorubim lima. The larvae exhibited the main characteristics of the genus Brevimulticaecum, such as short ventricular appendices and the position of excretory pore and excretory nucleus. This is the first report and description of larval specimens of Brevimulticaecum parasitic in these hosts, which are fish species distributed throughout South America.

Morphology, ultrastructure and phylogeny of Myxobolus curimatae n. sp. (Myxozoa: Myxosporea) a parasite of Prochilodus costatus (Teleostei: Prochilodontidae) from the São Francisco River, Brazil

Myxobolus curimatae n. sp. has been found infecting the gill filaments of Prochilodus costatus (Prochilodontidae) from the São Francisco River in the state of Minas Gerais, Brazil. The prevalence of the species was 18.7%. Mature spores were rounded from a frontal view, with elongated polar capsules of equal size, and had polar filaments with 9-10 turns. Ultrastructural analysis revealed that sporogenesis patterns followed those of other Myxobolus species. The plasmodium walls had numerous invaginations and protrusions, and few pinocytic channels. Numerous mitochondria, generative cells and young pansporoblasts were observed in the peripherical areas of the plasmodia, and mature spores were found in deeper layers. A layer of collagenic fibrils surrounded the plasmodia. The morphological data and molecular analysis of the 18S rDNA identified this parasite as a new species. The maximum likelihood phylogenetic tree showed M. curimatae n. sp., as a sister species of Thelohanellus marginatus, in a basal branch of the subclade composed by parasites with tropism to different organs and host families.

Morphological, ultrastructural and phylogenetic analyses of Myxobolus hilarii n. sp (Myxozoa, Myxosporea), a renal parasite of farmed Brycon hilarii in Brazil

Myxobolus hilarii n. sp. was described, based on morphology, histology, ultrastructure and 18S rDNA sequencing, infecting the kidney of Brycon hilarii (Valenciennes 1850) (Characiformes: Bryconidae) taken from fish farms in the state of São Paulo, Brazil. Thirteen specimens of B. hilarii were examined and 100% had round, white plasmodia in the kidney. The mature myxospores were rounded, measuring 11.5 ± 0.8 (9.8-13.4) μm in length, 11.0 ± 0.7 (9.7-12.4) μm in width and 7.6 ± 1.0 (6.7-9.0) μm in thickness. Polar capsules were elongated and of equal size, with 6.5 ± 0.4 (6.0-7.2) μm in length and 4.0 ± 0.2 (3.6-5.3) μm in width and their polar filaments had 5 to 7 coils. Histological analysis revealed plasmodial development in the renal tubules, causing compression and deformation of adjacent tissues and destruction of renal tubule cells. Ultrastructural analysis showed direct contact between the plasmodial wall and the host tissue and asynchronous plasmodial development. The phylogenetic analysis of South American myxobolids, based on 18S rDNA sequencing, showed the myxosporeans grouping into two main clades. M. hilarii n. sp. appears as sister species of Myxobolus piraputangae.