Myeloid-derived growth factor regulates neutrophil motility in interstitial tissue damage.

Neutrophil recruitment to tissue damage is essential for host defense but can also impede tissue repair. The cues that differentially regulate neutrophil responses to tissue damage and infection remain unclear. Here, we report that the paracrine factor myeloid-derived growth factor (MYDGF) is induced by tissue damage and regulates neutrophil motility to damaged, but not infected, tissues in zebrafish larvae. Depletion of MYDGF impairs wound healing, and this phenotype is rescued by depleting neutrophils. Live imaging and photoconversion reveal impaired neutrophil reverse migration and inflammation resolution in mydgf mutants. We found that persistent neutrophil inflammation in tissues of mydgf mutants was dependent on the HIF-1α pathway. Taken together, our data suggest that MYDGF is a damage signal that regulates neutrophil interstitial motility and inflammation through a HIF-1α pathway in response to tissue damage.

Updating specific PCR primer for detection of Cryptocaryon irritans from reared Larimichthys polyactis.

Artificial breeding of small yellow croaker (Larimichthys polyactis) was recently achieved, providing a bright future for its commercial farming. In May 2019, a disease outbreak occurred among small yellow croakers in an aquaculture farm near Xiangshan Bay, charactering by white spots spotted on the surface of fish skin, gills and fins. The parasite was preliminarily identified as Cryptocaryon irritans based on morphological feature of the parasite and the symptoms on fish. However, the previously published specific primer pairs failed to confirm the existence of C. iriitans. Six nucleotides mismatches were discovered after mapping specific forward primer back to targeted gene. Therefore, an updated PCR specific primer was developed within the 9th highly variable region of 18S rRNA gene and conserved in all C. irritans sequences available in GenBank database. The specificity was verified in silico by Primer-BLAST against GenBank nucleotide. Laboratory cultured ciliates (Mesanophrys, Pseudokeronopsis and Uronema) as well as natural microbial community samples collected from sea water and river water was used as negative control to verify the specificity of the primer in situ. Besides, tank transfer method was used to evaluate the treatment of the parasite infection. By tank transfer method, 2.00 ± 0.61 out of 10 fish that already sever infected were successfully survived after 8 days treatment, meanwhile the control group died out at d 6. More loss to the treatment group during first five days was observed and may attribute to the combined effect from infection and stress the recent domesticated fish suffered during rotation. Therefore, tank transfer method was also effective to prevent small yellow croaker from further infection, however the loss of the small yellow croaker suffered from stress during rotation also needs to be carefully concerned. In conclusion, this study reported the first diagnose of C. irritans infection on small yellow croaker, provided updated specific primer to detect C. irritans infection on fish body and reported the effect of tank transfer on small yellow croaker treatment.

Live Imaging of Chemokine Receptors in Zebrafish Neutrophils During Wound Responses.

Leukocyte guidance by chemical gradients is essential for immune responses. Neutrophils are the first cells to be recruited to sites of tissue damage where they execute crucial antimicrobial functions. Their trafficking to these loci is orchestrated by several inflammatory chemoattractants, including chemokines. At the molecular level, chemoattractant signaling is regulated by the intracellular trafficking of the corresponding receptors. However, it remains unclear how subcellular changes in chemokine receptors affect leukocyte migration dynamics at the cell and tissue level. Here we describe a methodology for live imaging and quantitative analysis of chemokine receptor dynamics in neutrophils during inflammatory responses to tissue damage. These tools have revealed that differential chemokine receptor trafficking in zebrafish neutrophils coordinates neutrophil clustering and dispersal at sites of tissue damage. This has implications for our understanding of how inflammatory responses are self-resolved. The described tools could be used to understand neutrophil migration patterns in a variety of physiological and pathological settings and the methodology could be expanded to other signaling receptors.

Natural hybridization reveals incompatible alleles that cause melanoma in swordtail fish.

The establishment of reproductive barriers between populations can fuel the evolution of new species. A genetic framework for this process posits that "incompatible" interactions between genes can evolve that result in reduced survival or reproduction in hybrids. However, progress has been slow in identifying individual genes that underlie hybrid incompatibilities. We used a combination of approaches to map the genes that drive the development of an incompatibility that causes melanoma in swordtail fish hybrids. One of the genes involved in this incompatibility also causes melanoma in hybrids between distantly related species. Moreover, this melanoma reduces survival in the wild, likely because of progressive degradation of the fin. This work identifies genes underlying a vertebrate hybrid incompatibility and provides a glimpse into the action of these genes in natural hybrid populations.

Fin damage and growth parameters relative to stocking density and feeding method in intensively cultured European perch (Perca fluviatilis L.).

We evaluated the relationship of stocking density to survival, growth performance and fin condition of European perch Perca fluviatilis with hand feeding and self-feeders. Hand-fed perch (body weight 19.1 ± 5.1 g and total length 107 ± 9 mm) were reared at 0.5, 1.0, 1.5 and 2.0 fish/L. Self-feeding perch (body weight 25.4 ± 3.9 g and total length 128 ± 7 mm) were reared at stocking densities of 0.6, 1.0 and 1.4 fish/L. Pond-reared perch served as a comparison group for fin damage assessment. We found no differences in survival rate among stocking densities with either feeding method. Hand-fed fish displayed the highest weight gain and SGR at stocking density of 0.5 fish/L. The self-feeding fish showed a non-linear association of weight gain with stocking density with the highest growth at 1.0 fish/L. Fin length was noticeably greater in pond-reared fish compared with RAS-reared fish regardless of feeding method. In both experiments, fin length relative to standard length showed a negative relationship with stocking density, with pectoral fins showing the greatest effect. Fin condition deteriorated with increasing stocking density, and growth was highest at 0.5 and 1.0 fish/L in hand-fed and self-feeding fish, respectively.

Inhibition of ErbB kinase signalling promotes resolution of neutrophilic inflammation.

Neutrophilic inflammation with prolonged neutrophil survival is common to many inflammatory conditions, including chronic obstructive pulmonary disease (COPD). There are few specific therapies that reverse neutrophilic inflammation, but uncovering mechanisms regulating neutrophil survival is likely to identify novel therapeutic targets. Screening of 367 kinase inhibitors in human neutrophils and a zebrafish tail fin injury model identified ErbBs as common targets of compounds that accelerated inflammation resolution. The ErbB inhibitors gefitinib, CP-724714, erbstatin and tyrphostin AG825 significantly accelerated apoptosis of human neutrophils, including neutrophils from people with COPD. Neutrophil apoptosis was also increased in Tyrphostin AG825 treated-zebrafish in vivo. Tyrphostin AG825 decreased peritoneal inflammation in zymosan-treated mice, and increased lung neutrophil apoptosis and macrophage efferocytosis in a murine acute lung injury model. Tyrphostin AG825 and knockdown of egfra and erbb2 by CRISPR/Cas9 reduced inflammation in zebrafish. Our work shows that inhibitors of ErbB kinases have therapeutic potential in neutrophilic inflammatory disease.

ECM alterations in Fndc3a (Fibronectin Domain Containing Protein 3A) deficient zebrafish cause temporal fin development and regeneration defects.

Fin development and regeneration are complex biological processes that are highly relevant in teleost fish. They share genetic factors, signaling pathways and cellular properties to coordinate formation of regularly shaped extremities. Especially correct tissue structure defined by extracellular matrix (ECM) formation is essential. Gene expression and protein localization studies demonstrated expression of fndc3a (fibronectin domain containing protein 3a) in both developing and regenerating caudal fins of zebrafish (Danio rerio). We established a hypomorphic fndc3a mutant line (fndc3awue1/wue1) via CRISPR/Cas9, exhibiting phenotypic malformations and changed gene expression patterns during early stages of median fin fold development. These developmental effects are mostly temporary, but result in a fraction of adults with permanent tail fin deformations. In addition, caudal fin regeneration in adult fndc3awue1/wue1 mutants is hampered by interference with actinotrichia formation and epidermal cell organization. Investigation of the ECM implies that loss of epidermal tissue structure is a common cause for both of the observed defects. Our results thereby provide a molecular link between these developmental processes and foreshadow Fndc3a as a novel temporal regulator of epidermal cell properties during extremity development and regeneration in zebrafish.

Clinical pathologies of bone fracture modelled in zebrafish.

Reduced bone quality or mineral density predict susceptibility to fracture and also attenuate subsequent repair. Bone regrowth is also compromised by bacterial infection, which exacerbates fracture site inflammation. Because of the cellular complexity of fracture repair, as well as genetic and environmental influences, there is a need for models that permit visualisation of the fracture repair process under clinically relevant conditions. To characterise the process of fracture repair in zebrafish, we employed a crush fracture of fin rays, coupled with histological and transgenic labelling of cellular responses; the results demonstrate a strong similarity to the phased response in humans. We applied our analysis to a zebrafish model of osteogenesis imperfecta (OI), which shows reduced bone quality, spontaneous fractures and propensity for non-unions. We found deficiencies in the formation of a bone callus during fracture repair in our OI model and showed that clinically employed antiresorptive bisphosphonates can reduce spontaneous fractures in OI fish and also measurably reduce fracture callus remodelling in wild-type fish. The csf1ra mutant, which has reduced osteoclast numbers, also showed reduced callus remodelling. Exposure to excessive bisphosphonate, however, disrupted callus repair. Intriguingly, neutrophils initially colonised the fracture site, but were later completely excluded. However, when fractures were infected with Staphylococcus aureus, neutrophils were retained and compromised repair. This work elevates the zebrafish bone fracture model and indicates its utility in assessing conditions of relevance to an orthopaedic setting with medium throughput.This article has an associated First Person interview with the first author of the paper.

Fin healing and regeneration in sturgeon.

Pectoral fin healing in fin spines and rays were examined in juvenile Atlantic sturgeon Acipenser oxyrinchus oxyrinchus following three different sampling techniques (n = 8-9 fish per treatment): entire leading fin spine removed, a 1-2 cm portion removed near the point of articulation, or a 1-2 cm portion removed from a secondary fin ray. Also, to determine whether antibiotic treatment influences healing, an additional group of fish (n = 8) was not given an injection of an oxytetracycline (OTC)-based antibiotic following removal of the entire leading fin spine. Following fin sampling, fish from different treatments were mixed equally between three large (4,000 I) recirculating systems and fin-ray healing and mortality were monitored over a 12 month period. To assess healing, blood samples were collected at 4 months to measure immune system responses, radiographs were taken at 4, 8 and 12 months to assess the degree of calcification in regions of damaged fins and fins were analyzed histologically at 12 months. Fish grew from a mean weight of 1.8 to 3.2 kg during the experiment and survival was near 100% in all treatments, with only one fish dying of unknown causes. Leukocyte counts, an indication of health status and survival were similar among treatments and in groups with or without antibiotic injection. Radiographs revealed mineralization took longer in fish with the entire leading fin spine removed and was the slowest near the point of articulation, presumably due to the greater structural support for the pectoral fin at this location. Histological sampling indicated spines and rays had similar healing patterns. Following injury, an orderly matrix of collagen bundles and many evenly spaced scleroblasts were present, transitioning to Sharpey fibres, with concentric layers forming lamellar bone. Healing and mineralization were characterized as periosteal osteogenesis and included embedded osteocytes surrounded by an osteoid seam. Chondroid formation was apparent in a few fractures not associated with treatments. The duration of time for external wound healing and internal mineralization of spines and rays depended on the fin treatment, with the slowest healing observed in fish with the most tissue removed, the entire leading fin spine.

Injury of Adult Zebrafish Expressing Acvr1lQ204D Does Not Result in Heterotopic Ossification.

Fibrodysplasia Ossificans Progressiva (FOP) is a rare, autosomal dominant genetic disorder in humans characterized by the gradual ossification of fibrous tissues, including skeletal muscle, tendons, and ligaments. In humans, mutations in the Type I BMP/TGFß family member receptor gene, ACVR1, are associated with FOP. Zebrafish acvr1l, previously known as alk8, is the functional ortholog of human ACVR1. We previously created and characterized the first adult zebrafish model for FOP by generating animals harboring heat shock-inducible mCherry-tagged constitutively active Acvr1l (Q204D). Since injury is a known trigger for heterotopic ossification (HO) development in human FOP patients, in this study, we investigated several injury models in Acvr1lQ204D-expressing zebrafish and the subsequent formation of HO. We performed studies of Activin A injection, cardiotoxin (CTX) injection, and caudal fin clip injury. We found that none of these methods resulted in HO formation at the site of injury. However, some of the cardiotoxin-injected and caudal fin-clipped animals did exhibit HO at distant sites, including the body cavity and along the spine. We describe these results in the context of new and exciting reports on FOP, and discuss future studies to better understand the etiology and progression of this disease.

An evaluation of a double-tailed deformity in a coral-reef surgeonfish Acanthurus nigrofuscus (Acanthuridae) using micro-computed tomography.

X-ray micro-computed tomography scans were used to examine the caudal-fin structure of an unusual double-tailed deformity in an adult brown surgeonfish Acanthurus nigrofuscus from the Great Barrier Reef. In both this case and in a similar double-tailed deformity in a juvenile Tomini surgeonfish Ctenochaetus tominiensis from the Philippines, the caudal fin was duplicated along the dorsoventral axis. Detailed examination of the A. nigrofuscus specimen revealed that the deformity was associated with duplication and reflection of the hypural plates and the posterior vertebrae, yet the fish survived to adulthood, indicating that the effects of duplication on survival may be limited.

Abnormal nuclear morphology is independent of longevity in a zmpste24-deficient fish model of Hutchinson-Gilford progeria syndrome (HGPS).

Lamin is an intermediate protein underlying the nuclear envelope and it plays a key role in maintaining the integrity of the nucleus. A defect in the processing of its precursor by a metalloprotease, ZMPSTE24, results in the accumulation of farnesylated prelamin in the nucleus and causes various diseases, including Hutchinson-Gilford progeria syndrome (HGPS). However, the role of lamin processing is unclear in fish species. Here, we generated zmpste24-deficient medaka and evaluated their phenotype. Unlike humans and mice, homozygous mutants did not show growth defects or lifespan shortening, despite lamin precursor accumulation. Gonadosomatic indices, blood glucose levels, and regenerative capacity of fins were similar in 1-year-old mutants and their wild-type (WT) siblings. Histological examination showed that the muscles, subcutaneous fat tissues, and gonads were normal in the mutants at the age of 1 year. However, the mutants showed hypersensitivity to X-ray irradiation, although p53target genes, p21 and mdm2, were induced 6 h after irradiation. Immunostaining of primary cultured cells from caudal fins and visualization of nuclei using H2B-GFP fusion proteins revealed an abnormal nuclear shape in the mutants both in vitro and in vivo. The telomere lengths were significantly shorter in the mutants compared to WT. Taken together, these results suggest that zmpste24-deficient medaka phenocopied HGPS only partially and that abnormal nuclear morphology and lifespan shortening are two independent events in vertebrates.

Morphological and ssrDNA sequence based molecular characterization of a novel Thelohanellus species (Myxosporea: Myxobolidae) infecting the fins of Goldfish, Carassius auratus L. with special reference to its histopathological alteration.

A new species of myxozoan, Thelohanellus goldi n. sp. is described using morphological and molecular data, parasitizing the fin filaments from 18 of 25 host specimens (72.5%) of Carassius auratus collected from different ornamental fish farms of India. Mature spore of the new species were oval to spherical in frontal view having rounded posterior ends and tapering anterior end measures 8.7-10.26 (9.50) × 4.10-7.89 (5.84) µm. The single large polar capsule, round to oval in shape but slightly pointed at the anterior end measuring 4.91-7.63 (5.60) × 2.3-3.1 (2.96) µm and located just below the anterior end of the spore. Polar filament only at distal end with 5-6 loose coils. 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 18S rDNA gene resulted in a total of 2124 bp fragment of newly obtained small subunit ribosomal RNA gene sequence of the new species which Exhibit 93-95% homogeneity with other closely related species available in GenBank. The BLAST search and high genetic diversity of distance matrix of Myxobolus sp. did not properly match with any available sequences in GenBank and make sister clade with Thelohanellus caudatus and Thelohanellus habibpuri in the Thelohanellus clade including most of Thelohanellus spp. The study of evolutionary history enables us to understand the evolution of modern species and supports some uncertain topologies which are being presented regarding the morphometric analysis. The severity of myxozoan infection has been assessed in this article by observing the histopathological changes of fins of the C. auratus along with the diversity, distribution and taxonomic description of the new Thelohanellus species with their new host and locality records.

Zebrafish akt2 is essential for survival, growth, bone development, and glucose homeostasis.

As one of three akt isoforms, akt2 plays a key role in the regulation of widely divergent cellular processes in mammals. However, its role and underlying mechanisms in zebrafish remain largely unknown. To elucidate the function of akt2 in zebrafish, we generated zebrafish lacking akt2 gene via CRISPR/Cas9 technology. Akt2-null zebrafish exhibit partial lethality and severe growth deficiency, which is different from those observed in akt2-null mice. Furthermore, akt2-null zebrafish display deficiency in fin ray development, but their cartilage is not affected. Similar to observations in akt2-null mice, akt2-null zebrafish display impaired glucose homeostasis. However, in contrast to that in akt2-null mice, insulin level is lower in akt2-null zebrafish, implicating the symptoms of type I diabetes exhibited in akt2-null zebrafish. In addition, transcriptome analysis reveals that the genes involved in metabolism and osteogenesis are disturbed in akt2-null zebrafish. Taken together, these data not only support an important role of akt2 in zebrafish survival, growth, bone development and glucose homeostasis, but also suggest that akt2 has divergent functions between mice and zebrafish, even though they are evolutionarily conserved.

Quantitative assessment of the regenerative and mineralogenic performances of the zebrafish caudal fin.

The ability of zebrafish to fully regenerate its caudal fin has been explored to better understand the mechanisms underlying de novo bone formation and to develop screening methods towards the discovery of compounds with therapeutic potential. Quantifying caudal fin regeneration largely depends on successfully measuring new tissue formation through methods that require optimization and standardization. Here, we present an improved methodology to characterize and analyse overall caudal fin and bone regeneration in adult zebrafish. First, regenerated and mineralized areas are evaluated through broad, rapid and specific chronological and morphometric analysis in alizarin red stained fins. Then, following a more refined strategy, the intensity of the staining within a 2D longitudinal plane is determined through pixel intensity analysis, as an indicator of density or thickness/volume. The applicability of this methodology on live specimens, to reduce animal experimentation and provide a tool for in vivo tracking of the regenerative process, was successfully demonstrated. Finally, the methodology was validated on retinoic acid- and warfarin-treated specimens, and further confirmed by micro-computed tomography. Because it is easily implementable, accurate and does not require sophisticated equipment, the present methodology will certainly provide valuable technical standardization for research in tissue engineering, regenerative medicine and skeletal biology.

Acute and chronic toxicity of the benzoylurea pesticide, lufenuron, in the fish, Colossoma macropomum.

Lufenuron is a benzoylurea insecticide that interfere in chitin synthesis in insects. Although lufenuron is widely used in agriculture and aquaculture, rare are studies described that relates to possible toxic effects in fish. This work aimed to evaluate acute and chronic toxic effects of benzoylurea pesticide (lufenuron) on biological parameters of Colossoma macropomum (Tambaqui). In the acute test, juveniles of Tambaqui were divided into control group and five experimental groups with exposure from 0.1 to 0.9 mg/L of lufenuron for 96 h. Animals were also submitted to chronic toxicity test for four months in concentrations of 0.1 and 0.3 mg/L of lufenuron, the concentration used in the treatment of ectoparasites in fish and 50% of LC50 96 h, respectively. The presence of hemorrhages was observed in eyes, fins and operculum of fish exposed to 0.7 and 0.9 mg/L of lufenuron. Histological analysis showed changes in the morphology of fish gills submitted to acute toxicity test, as lamellar aneurysm and blood congestion inside lamellae. Lufenuron promoted damage in fish retina as in ability to respond to stimuli in photoreceptors and in ON-bipolar cells in acute test. In chronic test, blood glucose analysis and morphometric parameters showed no significant differences (p > 0.05). In general, Tambaqui exhibited behaviors associated with stress when exposed to lufenuron. Thus, lufenuron showed several toxic effects in relation to biological parameters in Tambaqui. This concerns about the use and discard of lufenuron, and indicates the requirement of environmental actions to prevent potential contamination of aquatic biota.

Spatial Distribution of Gyrodactylus salmonis (Monogenea) on the Body of Captive Fingerling Oncorhynchus mykiss, Including Attachment Within the Olfactory Chamber.

Gyrodactylus salmonis is a common ectoparasite on the fins and body of North American salmonids in fresh water. In this study, the spatial distribution of G. salmonis on 60 captive hatchery-reared rainbow trout, Oncorhynchus mykiss , is reported. The highest parasite densities occurred on 5 × 5-mm(2) sections of the dorsal fin followed by the trunk, other fins, and the olfactory chamber, with the lowest densities on the head. The finding of infections within the olfactory chamber of 93% of the fish was unexpected. One possibility is that such infections represented spillover from high-density infrapopulations that occur on the skin and fins. However, this possibility is unlikely, because worm densities at various sites along the body surface of infected fish did not correlate with densities within the olfactory chamber. The parasite conceivably enters the chamber either via water incurrent or by crawling in from the head and subsequently remaining at this site to feed and reproduce. Results from scanning electron microscopy are consistent with physical modification to the olfactory epithelium associated with the attachment/reattachment of the opisthaptor and epithelial grazing.

MicroRNA 19a replacement partially rescues fin and cardiac defects in zebrafish model of Holt Oram syndrome.

Holt-Oram Syndrome (HOS) is an autosomal dominant heart-hand syndrome caused by mutations in the TBX5 gene, a transcription factor capable of regulating hundreds of cardiac-specific genes through complex transcriptional networks. Here we show that, in zebrafish, modulation of a single miRNA is sufficient to rescue the morphogenetic defects generated by HOS. The analysis of miRNA-seq profiling revealed a decreased expression of miR-19a in Tbx5-depleted zebrafish embryos compared to the wild type. We revealed that the transcription of the miR-17-92 cluster, which harbors miR-19a, is induced by Tbx5 and that a defined dosage of miR-19a is essential for the correct development of the heart. Importantly, we highlighted that miR-19a replacement is able to rescue cardiac and pectoral fin defects and to increase the viability of HOS zebrafish embryos. We further observed that miR-19a replacement shifts the global gene expression profile of HOS-like zebrafish embryos towards the wild type condition, confirming the ability of miR-19a to rescue the Tbx5 phenotype. In conclusion our data demonstrate the importance of Tbx5/miR-19a regulatory circuit in heart development and provide a proof of principle that morphogenetic defects associated with HOS can be rescued by transient miRNA modulation.

Long-lasting effects of dexamethasone on immune cells and wound healing in the zebrafish.

This study assessed the lasting impact of dexamethasone (DEX) exposure during early development on tissue repair capacity at later life stages (5, 14, and 24 days post fertilization [dpf]) in zebrafish larvae. Using the caudal fin amputation model, we show that prior exposure to DEX significantly delays but does not prevent wound healing at all life stages studied. DEX-induced impairments on wound healing were fully restored to normal levels with longer post amputation recovery time. Further analyses revealed that DEX mainly exerted its detrimental effects in the early phase (0-5 hours) of wound-healing process. Specifically, we observed the following events: (1) massive amount of cell death both by necrosis and apoptosis; (2) significant reduction in the number as well as misplacement of macrophages at the wound site; (3) aberrant migration and misplacement of neutrophils and macrophages at the wound site. These events were accompanied by significant (likely compensatory) changes in the expression of genes involved in tissue patterning, including up-regulation of FKBP5 6 hours post DEX exposure and that of Wnt3a and RARγ at 24 hours post amputation. Taken together, this study provides evidence that DEX exposure during early sensitive periods of development appears to cause permanent alterations in the cellular/molecular immune processes that are involved in the early phase of wound healing in zebrafish. These findings are consistent with previous studies showing that antenatal course of DEX is associated with immediate and lasting alterations of the immune system in rodent models and humans. Therefore, the current findings support the use of the larval zebrafish model to study the impact of stress and stress hormone exposure in immature organisms on health risks in later life.

Morphological and molecular characterization of Thelohanellus hoffmanni sp. nov. (Myxozoa) infecting goldfish Carassius auratus auratus.

A new species of the genus Thelohanellus Kudo, 1933 (Myxosporea, Bivalvulida) was isolated from the fins of goldfish Carassius auratus auratus (Linnaeus 1758). The fish had been imported from China by an Austrian retailer. Nodules from the margins of the fins contained pyriform myxospores with a singular polar capsule. In valvular view, the spores measured 12.2 µm in length and 6.4 µm in width. In sutural view, the thickness was 2.9 µm. The polar capsule measured 4.2 × 3.1 µm and contained a polar filament with 8 to 9 coils. Histological sections showed plasmodia of 0.2 to 4.0 mm diameter with the earlier developmental stages of the parasite in the periphery and the mature spores closer to the center. In the transmission electron microscope examination, the different developmental stages could be observed. Morphological data, host specificity, tissue tropism, and molecular analysis of the small subunit rDNA identify this parasite as a new species of Thelohanellus, which we have named Thelohanellus hoffmanni sp. nov.