Cutaneous angiomatosis-like presentation in koi carp (Cyprinus carpio koi): Clinical-pathological investigations.

The skin represents an indicator of an animal's health status. Causes of cutaneous diseases in fish most often trace back to biological agents. However, fish skin diseases can also arise from a complex interaction of infectious and non-infectious causes, making it more difficult to identify a specific aetiology. In the period between April and September of the years 2019-2022, four koi carp (Cyprinus carpio koi) from two European countries presented with multifocal, irregularly round, few mm to 1 cm, variably raised cutaneous reddened areas. The fish displayed good general condition. Cutaneous samples, investigated by microbiological and molecular methods and microscopy, did not indicate a primary pathogenic agent. Gross and histological findings of the cutaneous biopsies were consistent with a multifocal/reactive process centred on dermal vessels. The histological features were reminiscent of angiomatosis, a benign proliferative condition affecting the dermal vessels of mammals, including human patients. The clinical-pathological presentation and the dermatologic condition that affected the koi carp are discussed and compared with the veterinary and human literature.

A molecular insight into the lipid changes of pig Longissimus thoracis muscle following dietary supplementation with functional ingredients.

In this work, the Longissimus thoracis pig skeletal muscle was used as a model to investigate the impact of two different diets, supplemented with n-3 polyunsaturated fatty acids from extruded linseed (L) and polyphenols from grape skin and oregano extracts (L+P), on the lipidomic profile of meat. A standard diet for growing-finishing pigs (CTRL) was used as a control. Changes in lipids profile were investigated through an untargeted lipidomics and transcriptomics combined investigation. The lipidomics identified 1507 compounds, with 195 compounds fitting with the MS/MS spectra of LipidBlast database. When compared with the CTRL group, the L+P diet significantly increased 15 glycerophospholipids and 8 sphingolipids, while the L diet determined a marked up-accumulation of glycerolipids. According to the correlations outlined between discriminant lipids and genes, the L diet may act preventing adipogenesis and the related inflammation processes, while the L+P diet promoted the expression of genes involved in lipids' biosynthesis and adipogenic extracellular matrix formation and functioning.

Histological assessment of systemic granulomatosis progression in meagre Argyrosomus regius during cage ongrowing phase.

Meagre Argyrosomus regius is a potential candidate for Mediterranean aquaculture diversification, although several health issues still represent important bottlenecks for its sustainable production, including systemic granulomatosis (SG). To evaluate the SG progression in meagre during a 10 mo period of cage ongrowing, a histopathological investigation was carried out on 108 meagre fed 3 different diets (commercial pellets, hydrated commercial pellets and defrosted sardines). Histological sections of the gills and visceral organs were examined and lesions referable to SG scored from 1-3 according to the severity of the granulomatosis. The kidney and liver were the most affected organs, showing the highest percentage of positivity for granulomas and severity of lesions along the whole observation period. Using a statistical mixed model (GLMM) followed by odds ratio analysis, an effect of diet and temperature was found: the severity of liver and digestive tract SG scores decreased in the Cage 3 group (defrosted sardines) and with increasing temperature (p < 0.05, negative estimates, odds ratio <1). These observations, in accordance with the literature, suggest that SG in meagre could be related to nutritional-metabolic factors with the possible influence of environmental factors such as temperature.

Histological and immunohistochemical characterization of 17 gonadal tumours in koi carp (Cyprinus carpio koi).

Reports on abdominal tumours in koi carp are scarce and most are from the gonads. Their histological diagnosis is challenging due to the occurrence of mixed populations of neoplastic cells and the few availability of cross-reactive antibodies in fish tissues. The present study aims to provide a histopathological characterization of seventeen gonadal tumours, enriched by a wide antibody panel (vimentin, CD117, placental alkaline phosphatase-PLAP, AE1/AE3 cytokeratin, E-cadherin, proliferating cell nuclear antigen-PCNA, müllerian-inhibiting substance-MIS, GATA4 and Inhibin-α) applied on whole and tissue microarray (TMA) sections. Abdominal enlargement was associated with tumours filling 30%-80% of the abdominal cavity; frequently, the gonads had been completely replaced by neoplastic tissue. Twelve cases were characterized as sex cord-stromal tumours (SCSTs), three as germ cell tumours (GCTs), one as mixed germ cell sex cord-stromal tumour (MGCSCST) and one as carcinoma. By immunohistochemistry, PLAP enabled confirmation of GCTs, ovarian carcinoma and the objective identification of a further cell component in 8 out of the 12 SCSTs that were reclassified as mixed tumours. The use of an immunohistochemical panel can help in refining the histological diagnosis, but the morphological diagnosis still represents the main tool for the characterization of these tumours in koi carp.

Swim Bladder Disorders in Koi Carp (Cyprinus carpio).

Swim bladder disorders and consequent buoyancy problems are encountered in ornamental fish, including koi carp. Nevertheless, beyond clinical and pharmacological management, they are largely underdiagnosed. In this study, nine koi carp showing abdominal swelling and abnormal swimming behavior were investigated. Clinical approach, varying from case to case, included ultrasonographic and X-ray investigations, bacteriological analysis of the collected fluid, antimicrobial susceptibility pattern, and possibly histological analysis. Diagnostic imaging, corroborating gross examination, documented swim bladder deformation/dislocation and serous fluid within the swim bladder chambers of most animals. Bacteria belonging to the Aeromonas hydrophila/caviae group and Shewanella xiamenensis were identified. S. xiamenensis strains showed a sensibility to all tested molecules except for one strain, which was resistant to tetracycline and cyprofloxacin. Antibiotic treatment succeeded in the full recovery of three cases in which S. xiamemensis infection was detected. Chronic aerocystitis was histologically documented where tissue was available. The swim bladder histopathological findings highlighted a chronic process that had compromised the quality of life of the animals. A multidisciplinary clinical-pathological and microbiological approach is highly suggested to recognize swim bladder conditions as early as possible, aiming to drive medical intervention and raising the chances of fish survival.

Multifactorial Causes of Chronic Mortality in Juvenile Sturgeon (Huso huso).

This investigation focused on an episode of chronic mortality observed in juvenile Huso huso sturgeons. The examined subjects underwent pathological, microbiological, molecular, and chemical investigations. Grossly severe body shape deformities, epaxial muscle softening, and multifocal ulcerative dermatitis were the main observed findings. The more constant histopathologic findings were moderate to severe rarefaction and disorganization of the lymphohematopoietic lymphoid tissues, myofiber degeneration, atrophy and interstitial edema of skeletal epaxial muscles, and degeneration and atrophy of the gangliar neurons close to the myofibers. Chemical investigations showed a lower selenium concentration in affected animals, suggesting nutritional myopathy. Other manifestations were nephrocalcinosis and splenic vessel wall hyalinosis. Septicemia due to bacteria such as Aeromonas veronii, Shewanella putrefaciens, Citrobacter freundii, Chryseobacterium sp., and pigmented hyphae were found. No major sturgeon viral pathogens were detected by classical methods. Next-generation sequencing (NGS) analysis confirmed the absence of viral pathogens, with the exception of herpesvirus, at the order level; also, the presence of Aeromonas veronii and Shewanella putrefaciens was confirmed at the family level by the metagenomic classification of NGS data. In the absence of a primary yet undetected biological cause, it is supposed that environmental stressors, including nutritional imbalances, may have led to immune system impairment, facilitating the entry of opportunistic bacteria and mycotic hyphae.

Sequential Treatment of a Large Pituitary Corticotroph Neoplasm and Associated Neurological Signs in a Dog.

No standardized treatment guidelines are reported in veterinary medicine for dogs with large pituitary corticotroph neoplasms causing neurological signs, and such dogs usually have a short overall survival. When these dogs undergo pituitary surgery and the tumor regrows there are few reports of subsequent treatments. A 7 yr old male Maltese diagnosed with pituitary-dependent hypercortisolism developed seizures in conjunction with a large pituitary corticotroph adenoma and underwent transsphenoidal hypophysectomy. After 3 yr of clinical remission, hypercortisolism recurred, and trilostane therapy was initiated. One year later, the dog developed new neurological signs and computed tomography revealed regrowth of a large pituitary mass that was then treated with radiation therapy. The dog lived disease-free for 3 more yr. At postmortem examination, a more aggressive pituitary neoplasm than the one examined at the time of surgery was found, which is suggestive of malignant transformation into a carcinoma despite the absence of convincing metastasis.

Functional analysis finds differences on the muscle transcriptome of pigs fed an n-3 PUFA-enriched diet with or without antioxidant supplementations.

Supplementing pig diets with n-3 polyunsaturated fatty acids (n-3 PUFA) may produce meat products with an increased n-3 fatty acid content, and the combined antioxidants addition could prevent lipid oxidation in the feed. However, to date, the effects of these bioactive compounds at the molecular level in porcine skeletal muscle are mostly unknown. This study aimed to analyse changes in the Longissimus thoracis transcriptome of 35 pigs fed three diets supplemented with: linseed (L); linseed, vitamin E and Selenium (LES) or linseed and plant-derived polyphenols (LPE). Pigs were reared from 80.8 ± 5.6 kg to 151.8 ± 9.9 kg. After slaughter, RNA-Seq was performed and 1182 differentially expressed genes (DEGs) were submitted to functional analysis. The L vs LES comparison did not show differences, while L vs LPE showed 1102 DEGs and LES vs LPE 80 DEGs. LPE compared to the other groups showed the highest number of up-regulated genes involved in preserving muscle metabolism and structure. Results enlighten that the combined supplementation of bioactive lipids (n-3 PUFA from linseed) with plant extracts as a source of polyphenols increases, compared to the only addition of linseed, the expression of genes involved in mRNA metabolic processes and transcriptional regulation, glucose uptake and, finally, in supporting muscle development and physiology. These results improve the knowledge of the biological effect of bioactive compounds in Longissimus thoracis muscle, and sustain the growing interest over their use in pig production.

Effect of diets supplemented with linseed alone or combined with vitamin E and selenium or with plant extracts, on Longissimus thoracis transcriptome in growing-finishing Italian Large White pigs.

BACKGROUND: Supplementing farm animals diet with functional ingredients may improve the nutritional quality of meat products. Diet composition has been also demonstrated to influence the gene expression with effect on biological processes and pathways. However, the knowledge on the effect of nutrients at the molecular level is scant. In particular, studies on the effects of antioxidants and polyphenols dietary supplementation have been investigated mainly in rodents, and only scarcely in farm animals so far. RNA-Seq with next-generation sequencing is increasingly the method of choice for studying changes in the transcriptome and it has been recently employed also in pig nutrigenomics studies to identify diet-induced changes in gene expression. The present study aimed to investigate the effect of diets enriched with functional ingredients (linseed, vitamin E and plant extracts) on the transcriptome of pig Longissimus thoracis to elucidate the role of these compounds in influencing genes involved in muscle physiology and metabolism compared to a standard diet. RESULTS: Eight hundred ninety-three significant differentially expressed genes (DEGs) (FDR adjusted P-value ≤ 0.05) were detected by RNA-Seq analysis in the three diet comparisons (D2-D1, D3-D1, D4-D1). The functional analysis of DEGs showed that the diet enriched with n-3 PUFA from linseed (D2) mostly downregulated genes in pathways and biological processes (BPs) related to muscle development, contraction, and glycogen metabolism compared to the standard diet. The diet supplemented with linseed and vitamin E/Selenium (D3) showed to mostly downregulate genes linked to oxidative phosphorylation. Only few genes involved in extracellular matrix (ECM) organization were upregulated by the D3. Finally, the comparison D4-D1 showed that the diet supplemented with linseed and plant extracts (D4) upregulated the majority of genes compared to D1 that were involved in a complex network of pathways and BPs all connected by hub genes. In particular, IGF2 was a hub gene connecting protein metabolism, ECM organization, immune system and lipid biosynthesis pathways. CONCLUSION: The supplementation of pig diet with n-3 PUFA from linseed, antioxidants and plant-derived polyphenols can influence the expression of a relevant number of genes in Longissimus thoracis muscle that are involved in a variety of biochemical pathways linked to muscle function and metabolism.

Effect of dietary polyunsaturated fatty acid and antioxidant supplementation on the transcriptional level of genes involved in lipid and energy metabolism in swine.

Porcine fat traits depend mostly on the interaction between nutritional and genetic factors. However, the pathways and biological processes influenced by this interaction are still poorly known in pigs, although they can have a huge impact on meat quality traits. The present research provides new knowledge insight into the effect of four diets (D1 = standard diet; D2 = linseed supplementation; D3 = linseed, vitamin E and selenium supplementation; D4 = linseed and plant-derived polyphenols supplementation) on the expression of 24 candidate genes selected for their role in lipid and energy metabolism. The data indicated that 10 out of 24 genes were differentially expressed among diets, namely ACACA, ADIPOQ, ADIPOR1, CHREBP (MLXPL), ELOVL6, FASN, G6PD, PLIN2, RXRA and SCD. Results from the univariate analysis displayed an increased expression of ACACA, ADIPOQ, ADIPOR1, CHREBP, ELOVL6, FASN, PLIN2, RXRA and SCD in D4 compared to D2. Similarly, ACACA, ADIPOQ, ADIPOR1, ELOVL6 and SCD were highly expressed in D4 compared to D3, while no differences were observed in D2-D3 comparison. Moreover, an increased expression of G6PD and ELOVL6 genes in D4 compared to D1 was observed. Results from the multivariate analysis confirmed that D2 was not different from D3 and that ACACA, SCD and FASN expression made D4 different from D2 and D3. Comparing D4 and D1, the expression levels of ELOVL6 and ACACA were the most influenced. This research provides evidence that the addition of both n-3 PUFA and polyphenols, derived from linseed, grape-skin and oregano supplementation in the diets, stimulates the expression of genes involved in lipogenesis and in oxidative processes. Results evidenced a greater effect on gene expression of the diet added with both plant extracts and n-3 PUFA, resulting in an increased expression of genes coding for fatty acid synthesis, desaturation and elongation in pig Longissimus thoracis muscle.

Detection of Cyprinid Herpesvirus 1 DNA in cutaneous squamous cell carcinoma of koi carp (Cyprinus carpio).

BACKGROUND: Cutaneous squamous cell carcinoma (cSCC) is an uncommon disease affecting koi carp (Cyprinus carpio). Cutaneous papilloma (carp pox) is a benign epidermal proliferation reported in koi and has been shown to be caused by Cyprinid Herpesvirus 1 (CyHV1). HYPOTHESIS/OBJECTIVES: Histological, ultrastructural and molecular investigations were carried out aiming to investigate the aetiology of cSCC within archived tissue samples. ANIMALS: Surgical samples of masses located on the integument, fins and lips of 13 koi carp belonging to different private owners were included in this retrospective study. METHODS: CyHV1 DNA and RNA presence were investigated in five cSCC formalin-fixed paraffin-embedded tissue samples to recognize CyHV1 presence and its replication activity. RESULTS: All cases were histologically diagnosed as cSCC. The ultrastructural observations confirmed the squamous differentiation of neoplastic epithelial cells, which showed abundant tonofilament bundles and desmosomes. Although no virus particles were revealed ultrastructurally, the molecular investigation detected viral DNA in five epidemiologically unrelated cSCC. Viral transcript analysis revealed no evidence for viral replication in the tested cSCC, which could be consistent with latent infection. CONCLUSIONS AND CLINICAL IMPORTANCE: These findings illustrate the frequent association of carp cSCC with CyHV1, although a direct cause-effect relationship cannot be established at this time. Therefore, surveillance programmes should take into account the suspected viral origin of cSCC to better inform prevention and control of CyHV1 in the future.

Short communication: Alteration in blood parameters by enrofloxacin in juvenile lesser spotted dogfish (Scyliorhinus canicula, Linnaeus, 1758) after intramuscular injection.

Enrofloxacin (EF) is a veterinary drug for respiratory, gastrointestinal and urinary tract infections. Parenteral administration at doses higher than the manufacturer recommended dosage has induced cartilage lesions in juvenile mammals and poultry. In elasmobranchs EF is commonly used for the treatment of infectious diseases, however only one study has been performed previously to evaluate pharmacokinetics and the potential chondrotoxicity of a fluoroquinolone in adult skates. In present study juvenile lesser spotted dogfish (Scyliorhinus canicula, Linnaeus, 1758) were treated with low (LD, 5mg/kg) and high dose (HD, 10mg/kg) of EF daily via intramuscular injection for 15 consecutive days, as indicated in a recognized treatment. Hematological parameters, oxidative stress and histology of vertebral cartilage were evaluated. No cartilage damage was observed. Hematological parameters evaluation underlined a significant (p=0.035), dose-dependent reduction in red blood cell count and in hematocrit (40% and 6%, respectively). Anyway, the biological significance of this reduction is doubtful, due to limited decrease observed. The evaluation of oxidative stress parameters underlined that glutathione levels, as well as glutathione S-transferase and catalase activities, were significantly increased in HD group animals, with respect to untreated and solvent control groups. Obtained data do not support a toxic effect of EF on dogfish cartilage, while concern arises from hematological and oxidative stress data.

Hernia of the swim bladder (aerocystocele) with concurrent mycotic granulomatous inflammation and swim bladder carcinoma in a wild mullet (Mugil cephalus).

We describe a hernia of the swim bladder, with a concurrent mycotic granulomatous inflammation, and carcinoma of the swim bladder in a wild mullet (Mugil cephalus) referred for an exophytic dorsal mass. Grossly, the mass was white, soft, and composed of multiple cystic gas-containing chambers connected by a funnel-shaped tissue segment to the coelomic swim bladder. Histologically, the mass was characterized by cysts of variable size, multifocally contiguous with the subepithelial rete mirabile, supported by abundant fibrous tissue. The skin covering the herniated swim bladder was focally ulcerated and replaced by abundant granulation tissue in which multiple scattered granulomas centered on pigmented fungal hyphae were observed. These granulomas were also seen in the remaining coelomic portion of the swim bladder as well as in the spleen, perivisceral pancreas, and peritoneal adipose tissue; the fungus was molecularly identified as Cladosporium spp. Focally, arising from the herniated swim bladder epithelium, an unencapsulated poorly demarcated, moderately cellular neoplasm, composed of islands, lobules, and acini of neoplastic epithelium, was found.

Hikui disease in nine koi carp (Cyprinus carpio): first description of a cutaneous perivascular wall tumour.

BACKGROUND: Hikui disease is a well known disfiguring disease of koi carp (Cyprinus carpio) primarily affecting fish with red pigmentation. It causes light orange to golden yellow, multifocal to coalescing raised patches, starting from the red cutaneous areas. Some cases respond to surgery or topical treatment, but recurrence is common. OBJECTIVES: To describe the clinical and pathological presentation of Hikui disease and its cause. ANIMALS: Nine affected koi carp belonging to private hobbyists. METHODS: Eight fish underwent surgery or biopsy; one was euthanized. Tissues were submitted for histology, immunohistochemistry and transmission electron microscopy. RESULTS: Five fish showed typical lesions of Hikui disease, whereas four fish showed an atypical presentation characterized by focal or multifocal, oedematous, dark red cutaneous plaques or nodules. Histology showed unencapsulated, infiltrating and densely cellular neoplasms composed of spindle cells arranged in bundles, rows and whorls frequently centred on capillaries. Immunohistochemistry for smooth muscle actin labelled neoplastic cells in all cases. Ultrastructure showed neoplastic cells with slender cytoplasmic processes encircling the capillaries, a thin basal membrane and occasional plasmalemmal vesicles. CONCLUSIONS AND CLINICAL IMPORTANCE: All of the data supported a neoplastic process producing perivascular wall tumours. Immunoreactivity to smooth muscle actin and the ultrastructural features were indicative of a pericyte origin (haemangiopericytoma). This is the first report dealing with Hikui disease that has achieved a conclusive diagnosis. The neoplastic nature of this condition suggests the potential usefulness of a surgical approach in the clinical management of less severe cases.

Ultrasonographic and pathologic study of schwannoma in a Goldfish (Carassius auratus).

An adult Goldfish (Carassius auratus) developed an exophitic, multinodular, mass in the dorsal region involving the fin. The clinical and pathologic approach included ultrasonography, cytology, histopathology, immunohistochemistry, and ultrastructural evaluation. B-mode and color Doppler ultrasonographic findings showed an oval, well-defined mass with slightly inhomogeneous parenchymatous echotexture associated with an intense intralesional vascularization. Cytology and histology revealed neoplastic pleomorphic spindle cells arranged in a storiform pattern or in palisades typical of Antoni A pattern schwannoma. Moderate anisocytosis and anisokaryosis and occasional binucleation were also present. The neoplastic tissue deeply infiltrated the skeletal muscle of the dorsal region. Immunohistochemistry showed a diffuse cytoplasmic immunoreactivity of neoplastic cells to S100 protein and proliferating cell nuclear antigen (PCNA); glial fibrillary acidic protein was negative. The PCNA proliferation rate was 23.5% (calculated as the mean of 10 fields). Ultrastructurally, neoplastic cells were juxtaposed with parallel nuclei forming the typical palisade pattern and sharing cytoplasmic and nuclear features with human schwannoma. This is the first cytologic description supplied with echographic investigation of a malignant schwannoma in a fish species.

Proliferation, apoptosis, and fractal dimension analysis for the quantification of intestinal trophism in sole (Solea solea) fed mussel meal diets.

BACKGROUND: The evaluation of intestinal trophism, mainly the mucosal layer, is an important issue in various conditions associated with injury, atrophy, recovery, and healing of the gut. The aim of the present study was to evaluate the kinetics of the proliferation and apoptosis of enterocytes by immunohistochemistry and to assess the complexity of intestinal mucosa by fractal dimension (FD) analysis in Solea solea fed different experimental diets. RESULTS: Histomorphological evaluation of all intestinal segments did not show signs of degeneration or inflammation. Cell proliferation index and FD were significantly reduced with a diet high in mussel meal (MM; p = 0.0034 and p = 0.01063, respectively), while apoptotic index did not show any significant difference for the same comparison (p = 0.3859). Linear regression analysis between apoptotic index (independent variable) and FD (dependent variable) showed a statistically significant inverse relationship (p = 0.002528). Linear regression analysis between cell proliferation index (independent variable) and FD (dependent variable) did not show any significant correlation (p = 0.131582). CONCLUSIONS: The results demonstrated that diets containing increasing levels of mussel meal in substitution of fishmeal did not incite a hyperplastic response of the intestinal mucosa. The mussel meal, which is derived from molluscs, could mimic the characteristics of the sole's natural prey, being readily digestible, even without increasing the absorptive surface of intestinal mucosa. Interestingly, from this study emerged that FD could be used as a numeric indicator complementary to in situ quantification methods to measure intestinal trophism, in conjunction with functional parameters.

Ocular glioneuroma with medulloepitheliomatous differentiation in a goldfish (Carassius auratus).

An intraocular mass in the left eye causing chronic severe exophthalmia in an adult female goldfish (Carassius auratus) is described. The fish shared an aquarium with another goldfish found dead with gross and microscopic lesions consistent with mycobacteriosis. Histological examination of the left eye, histochemical (periodic acid-Schiff [PAS], Alcian blue, Ziehl-Neelsen) and immunohistochemical tests (glial fibrillary acidic protein, human neuronal protein, vimentin, and cytokeratin AE1/AE3) were carried out on the intraocular mass. Neoplastic cells forming an unencapsulated highly cellular proliferation partially covered by an intact corneal epithelium were stained with Alcian blue, which demonstrated an abundant hyaluronic acid-rich extracellular matrix. Multifocally, there were cyst-like dilatations bordered by neuroepithelial cells, which were PAS-positive. The complex neoplastic proliferation was composed of glial-like cells, neuronal-like cells (immunoreactive to glial fibrillary acidic protein and human neuronal protein, respectively) and neuroepithelium, which suggested a retinal origin.