Preliminary study of proton magnetic resonance spectroscopy to assess bone marrow adiposity in the third metacarpus or metatarsus in Thoroughbred racehorses.

BACKGROUND: Magnetic resonance spectroscopy (MRS) has been used to investigate metabolic changes within human bone. It may be possible to use MRS to investigate bone metabolism and fracture risk in the distal third metacarpal/tarsal bone (MC/MTIII) in racehorses. OBJECTIVES: To determine the feasibility of using MRS as a quantitative imaging technique in equine bone by using the 1H spectra for the MC/MTIII to calculate fat content (FC). STUDY DESIGN: Observational cross-sectional study. METHODS: Limbs from Thoroughbred racehorses were collected from horses that died or were subjected to euthanasia on racecourses. Each limb underwent magnetic resonance imaging (MRI) at 3 T followed by single-voxel MRS at three regions of interest (ROI) within MC/MTIII (lateral condyle, medial condyle, proximal bone marrow [PBM]). Percentage FC was calculated at each ROI. Each limb underwent computed tomography (CT) and bone mineral density (BMD) was calculated for the same ROIs. All MR and CT images were graded for sclerosis. Histology slides were graded for sclerosis and proximal marrow space was calculated. Pearson or Spearman correlations were used to assess the relationship between BMD, FC and marrow space. Kruskal-Wallis tests were used to check for differences between sclerosis groups for BMD or FC. RESULTS: Eighteen limbs from 10 horses were included. A negative correlation was identified for mean BMD and FC for the lateral condyle (correlation coefficient = -0.60, p = 0.01) and PBM (correlation coefficient = -0.5, p = 0.04). There was a significant difference between median BMD for different sclerosis grades in the condyles on both MRI and CT. A significant difference in FC was identified between sclerosis groups in the lateral condyle on MRI and CT. MAIN LIMITATIONS: Small sample size. CONCLUSIONS: 1H Proton MRS is feasible in the equine MC/MTIII. Further work is required to evaluate the use of this technique to predict fracture risk in racehorses.

Cross-species oncogenomics offers insight into human muscle-invasive bladder cancer.

BACKGROUND: In humans, muscle-invasive bladder cancer (MIBC) is highly aggressive and associated with a poor prognosis. With a high mutation load and large number of altered genes, strategies to delineate key driver events are necessary. Dogs and cats develop urothelial carcinoma (UC) with histological and clinical similarities to human MIBC. Cattle that graze on bracken fern also develop UC, associated with exposure to the carcinogen ptaquiloside. These species may represent relevant animal models of spontaneous and carcinogen-induced UC that can provide insight into human MIBC. RESULTS: Whole-exome sequencing of domestic canine (n = 87) and feline (n = 23) UC, and comparative analysis with human MIBC reveals a lower mutation rate in animal cases and the absence of APOBEC mutational signatures. A convergence of driver genes (ARID1A, KDM6A, TP53, FAT1, and NRAS) is discovered, along with common focally amplified and deleted genes involved in regulation of the cell cycle and chromatin remodelling. We identify mismatch repair deficiency in a subset of canine and feline UCs with biallelic inactivation of MSH2. Bovine UC (n = 8) is distinctly different; we identify novel mutational signatures which are recapitulated in vitro in human urinary bladder UC cells treated with bracken fern extracts or purified ptaquiloside. CONCLUSION: Canine and feline urinary bladder UC represent relevant models of MIBC in humans, and cross-species analysis can identify evolutionarily conserved driver genes. We characterize mutational signatures in bovine UC associated with bracken fern and ptaquiloside exposure, a human-linked cancer exposure. Our work demonstrates the relevance of cross-species comparative analysis in understanding both human and animal UC.

Phaeochromocytoma associated with cardiomyopathy and leukocytoclastic vasculitis in a dog.

Phaeochromocytomas are rare tumours of the adrenal medulla that can be associated with various presentations. Many of the better characterized clinical signs, including weakness, tachycardia and tachypnoea, are attributable to excessive and unregulated catecholamine secretion from functional tumours. In addition to catecholamine-induced cardiomyopathy and vasospasm, the invasive nature of phaeochromocytomas can lead to occlusion of the caudal vena cava contributing to systemic cardiovascular compromise. In humans, leukocytoclastic vasculitis is a rarely reported manifestation of catecholamine excess associated with phaeochromocytomas. We now describe a dog that had an invasive unilateral phaeochromocytoma with histological evidence of myocardial damage, consistent with catecholamine-induced cardiomyopathy, and leukocytoclastic vasculitis of small vessels in a range of tissues. We conclude that catecholamine excess may have played a role in the pathogenesis of vasculitis in this case. To the best of our knowledge, this is the first documented association between phaeochromocytoma and leukocytoclastic vasculitis in a non-human species.

Trans-cortical vessels in the mouse temporal bulla bone are a means to recruit myeloid cells in chronic otitis media and limit peripheral leukogram changes.

Chronic otitis media, inflammation of the middle ear, is a sequel to acute otitis media in ∼8% of children. Chronic otitis media with effusion is the most common cause of childhood deafness and is characterised by effusion of white blood cells into the auditory bulla cavity. Skull flat bones have trans-cortical vessels which are responsible for the majority of blood flow in and out of the bone. In experimental models of stroke and aseptic meningitis there is preferential recruitment of myeloid cells (neutrophils and monocytes) from the marrow in skull flat bones. We report trans-cortical vessels in the mouse temporal bone connect to the bulla mucosal vasculature and potentially represent a means to recruit myeloid cells directly into the inflamed bulla. The mutant mouse strains Junbo (Mecom Jbo/+ ) and Jeff (Fbxo11 Jf/+ ) develop chronic otitis spontaneously; Mecom Jbo/+ mice have highly cellular neutrophil (90%) rich bulla exudates whereas Fbxo11 Jf/+ mice have low cellularity serous effusions (5% neutrophils) indicating differing demand for neutrophil recruitment. However we found peripheral leukograms of Mecom Jbo/+ and Fbxo11 Jf/+ mice are similar to their respective wild-type littermate controls with healthy bullae and infer preferential mobilization of myeloid cells from temporal bulla bone marrow may mitigate the need for a systemic inflammatory reaction. The cytokines, chemokines and haematopoietic factors found in the inflamed bulla represent candidate signalling molecules for myeloid cell mobilization from temporal bone marrow. The density of white blood cells in the bulla cavity is positively correlated with extent of mucosal thickening in Mecom Jbo/+ , Fbxo11 Jf/+ , and Eda Ta mice and is accompanied by changes in epithelial populations and bone remodelling. In Mecom Jbo/+ mice there was a positive correlation between bulla cavity WBC numbers and total bacterial load. The degree of inflammation varies between contralateral bullae and between mutant mice of different ages suggesting inflammation may wax and wane and may be re-initiated by a new wave of bacterial infection. Clearance of white blood cells and inflammatory stimuli from the bulla cavity is impaired and this may create a pro-inflammatory feedback loop which further exacerbates otitis media and delays its resolution.

AMPK deficiency in smooth muscles causes persistent pulmonary hypertension of the new-born and premature death.

AMPK has been reported to facilitate hypoxic pulmonary vasoconstriction but, paradoxically, its deficiency precipitates pulmonary hypertension. Here we show that AMPK-α1/α2 deficiency in smooth muscles promotes persistent pulmonary hypertension of the new-born. Accordingly, dual AMPK-α1/α2 deletion in smooth muscles causes premature death of mice after birth, associated with increased muscularisation and remodeling throughout the pulmonary arterial tree, reduced alveolar numbers and alveolar membrane thickening, but with no oedema. Spectral Doppler ultrasound indicates pulmonary hypertension and attenuated hypoxic pulmonary vasoconstriction. Age-dependent right ventricular pressure elevation, dilation and reduced cardiac output was also evident. KV1.5 potassium currents of pulmonary arterial myocytes were markedly smaller under normoxia, which is known to facilitate pulmonary hypertension. Mitochondrial fragmentation and reactive oxygen species accumulation was also evident. Importantly, there was no evidence of systemic vasculopathy or hypertension in these mice. Moreover, hypoxic pulmonary vasoconstriction was attenuated by AMPK-α1 or AMPK-α2 deletion without triggering pulmonary hypertension.

Usefulness of the pancreas as a prime target for histopathological diagnosis of Tilapia parvovirus (TiPV) infection in Nile tilapia, Oreochromis niloticus.

Tilapia parvovirus (TiPV) is an emerging virus reportedly associated with disease and mortality in farmed tilapia. Although previous descriptions of histopathological changes are available, the lesions reported in these are not pathognomonic. Here, we report Cowdry type A inclusion bodies (CAIB) in the pancreas as a diagnostic histopathological feature found in adult Nile tilapia naturally infected with TiPV. This type of inclusion body has been well-known as a histopathological landmark for the diagnosis of other parvoviral infections in shrimp and terrestrial species. Interestingly, this lesion could be exclusively observed in pancreatic acinar cells, both in the hepatopancreas and pancreatic tissue along the intestine. In situ hybridization (ISH) using a TiPV-specific probe revealed the intranuclear presence of TiPV DNA in multiple tissues, including the liver, pancreas, kidney, spleen, gills and the membrane of oocytes in the ovary. These findings suggest that although TiPV can replicate in several tissue types, CAIB manifest exclusively in pancreatic tissues. In addition to TiPV, most diseased fish were co-infected with Streptococcus agalactiae, and presented with multifocal granulomas secondary to this bacterial infection. Partial genome amplification of TiPV was successful and revealed high nucleotide identity (>99%) to previously reported isolates. In summary, this study highlights the usefulness of pancreatic tissue as a prime target for histopathological diagnosis of TiPV in diseased Nile tilapia. This pattern may be critical when determining the presence of TiPV infection in new geographic areas, where ancillary testing may not be available. TiPV pathogenesis in this landmark organ warrants further investigation.

Immunization of Nile Tilapia (Oreochromis niloticus) Broodstock with Tilapia Lake Virus (TiLV) Inactivated Vaccines Elicits Protective Antibody and Passive Maternal Antibody Transfer.

Tilapia lake virus (TiLV), a major pathogen of farmed tilapia, is known to be vertically transmitted. Here, we hypothesize that Nile tilapia (Oreochromis niloticus) broodstock immunized with a TiLV inactivated vaccine can mount a protective antibody response and passively transfer maternal antibodies to their fertilized eggs and larvae. To test this hypothesis, three groups of tilapia broodstock, each containing four males and eight females, were immunized with either a heat-killed TiLV vaccine (HKV), a formalin-killed TiLV vaccine (FKV) (both administered at 3.6 × 106 TCID50 per fish), or with L15 medium. Booster vaccination with the same vaccines was given 3 weeks later, and mating took place 1 week thereafter. Broodstock blood sera, fertilized eggs and larvae were collected from 6-14 weeks post-primary vaccination for measurement of TiLV-specific antibody (anti-TiLV IgM) levels. In parallel, passive immunization using sera from the immunized female broodstock was administered to naïve tilapia juveniles to assess if antibodies induced in immunized broodstock were protective. The results showed that anti-TiLV IgM was produced in the majority of both male and female broodstock vaccinated with either the HKV or FKV and that these antibodies could be detected in the fertilized eggs and larvae from vaccinated broodstock. Higher levels of maternal antibody were observed in fertilized eggs from broodstock vaccinated with HKV than those vaccinated with FKV. Low levels of TiLV-IgM were detected in some of the 1-3 day old larvae but were undetectable in 7-14 day old larvae from the vaccinated broodstock, indicating a short persistence of TiLV-IgM in larvae. Moreover, passive immunization proved that antibodies elicited by TiLV vaccination were able to confer 85% to 90% protection against TiLV challenge in naïve juvenile tilapia. In conclusion, immunization of tilapia broodstock with TiLV vaccines could be a potential strategy for the prevention of TiLV in tilapia fertilized eggs and larvae, with HKV appearing to be more promising than FKV for maternal vaccination.

The EDA-deficient mouse has Zymbal's gland hypoplasia and acute otitis externa.

In mice, rats, dogs and humans, the growth and function of sebaceous glands and eyelid Meibomian glands depend on the ectodysplasin signalling pathway. Mutation of genes encoding the ligand EDA, its transmembrane receptor EDAR and the intracellular signal transducer EDARADD leads to hypohidrotic ectodermal dysplasia, characterised by impaired development of teeth and hair, as well as cutaneous glands. The rodent ear canal has a large auditory sebaceous gland, the Zymbal's gland, the function of which in the health of the ear canal has not been determined. We report that EDA-deficient mice, EDAR-deficient mice and EDARADD-deficient rats have Zymbal's gland hypoplasia. EdaTa mice have 25% prevalence of otitis externa at postnatal day 21 and treatment with agonist anti-EDAR antibodies rescues Zymbal's glands. The aetiopathogenesis of otitis externa involves infection with Gram-positive cocci, and dosing pregnant and lactating EdaTa females and pups with enrofloxacin reduces the prevalence of otitis externa. We infer that the deficit of sebum is the principal factor in predisposition to bacterial infection, and the EdaTa mouse is a potentially useful microbial challenge model for human acute otitis externa.

Comparative Studies of Renin-Null Zebrafish and Mice Provide New Functional Insights.

BACKGROUND: The renin-angiotensin system is highly conserved across vertebrates, including zebrafish, which possess orthologous genes coding for renin-angiotensin system proteins, and specialized mural cells of the kidney arterioles, capable of synthesising and secreting renin. METHODS: We generated zebrafish with CRISPR-Cas9-targeted knockout of renin (ren-/-) to investigate renin function in a low blood pressure environment. We used single-cell (10×) RNA sequencing analysis to compare the transcriptome profiles of renin lineage cells from mesonephric kidneys of ren-/- with ren+/+ zebrafish and with the metanephric kidneys of Ren1c-/- and Ren1c+/+ mice. RESULTS: The ren-/- larvae exhibited delays in larval growth, glomerular fusion and appearance of a swim bladder, but were viable and withstood low salinity during early larval stages. Optogenetic ablation of renin-expressing cells, located at the anterior mesenteric artery of 3-day-old larvae, caused a loss of tone, due to diminished contractility. The ren-/- mesonephric kidney exhibited vacuolated cells in the proximal tubule, which were also observed in Ren1c-/- mouse kidney. Fluorescent reporters for renin and smooth muscle actin (Tg(ren:LifeAct-RFP; acta2:EGFP)), revealed a dramatic recruitment of renin lineage cells along the renal vasculature of adult ren-/- fish, suggesting a continued requirement for renin, in the absence of detectable angiotensin metabolites, as seen in the Ren1YFP Ren1c-/- mouse. Both phenotypes were rescued by alleles lacking the potential for glycosylation at exon 2, suggesting that glycosylation is not essential for normal physiological function. CONCLUSIONS: Phenotypic similarities and transcriptional variations between mouse and zebrafish renin knockouts suggests evolution of renin cell function with terrestrial survival.

Evaluation of a Harmonized Undergraduate Catalog for Veterinary Public Health and Food Hygiene Pedagogy in Europe.

Current and emerging veterinary public health (VPH) challenges raised by globalization, climate change, and industrialization of food production require the veterinarian's role to evolve in parallel and veterinary education to adapt to reflect these changes. The European Food Hygiene catalog was developed to provide a list of topics relevant to Day One Competencies in VPH. A study was undertaken to ensure that the catalog and teaching practices were pertinent to the work of public health veterinarians. Relevant stakeholders were consulted using questionnaires and semi-structured interviews. A long questionnaire was distributed to 49 academics teaching VPH in European veterinary schools to review topics listed in the catalog. Eighteen responses were received (36.7%), representing 12 European countries. There was general agreement that most topics were appropriate for the undergraduate VPH curriculum. A short questionnaire was distributed to 348 European veterinarians working in the industry. Twenty-four questionnaires (6.7%) were received, representing eight European countries. Despite the low participation rate, topics needing greater emphasis in the undergraduate curriculum included Hazard Analysis Critical Control Points (HACCP), food microbiology, and audits. Seven semi-structured interviews with public health veterinarians working in the UK identified the need for curricular changes including greater practical experience and a shift from a focus on meat inspection to risk management. This may be partly achieved by replacing traditional lectures with authentic case-based scenarios. The study findings can be used to inform the future direction to VPH education for veterinary students across Europe.

Efficacy of heat-killed and formalin-killed vaccines against Tilapia tilapinevirus in juvenile Nile tilapia (Oreochromis niloticus).

Tilapia tilapinevirus (also known as tilapia lake virus, TiLV) is considered to be a new threat to the global tilapia industry. The objective of this study was to develop simple cell culture-based heat-killed (HKV) and formalin-killed (FKV) vaccines for the prevention of disease caused by TiLV. The fish were immunized with 100 µl of either HKV or FKV by intraperitoneal injection with each vaccine containing 1.8 × 106 TCID50- inactivated virus. A booster vaccination was carried out at 21-day post-vaccination (dpv) using the same protocol. The fish were then challenged with a lethal dose of TiLV at 28 dpv. The expression of five immune genes (IgM, IgD, IgT, CD4 and CD8) in the head kidney and spleen of experimental fish was assessed at 14 and 21 dpv and again after the booster vaccination at 28 dpv. TiLV-specific IgM responses were measured by ELISA at the same time points. The results showed that both vaccines conferred significant protection, with relative percentage survival of 71.3% and 79.6% for HKV and FKV, respectively. Significant up-regulation of IgM and IgT was observed in the head kidney of fish vaccinated with HKV at 21 dpv, while IgM, IgD and CD4 expression increased in the head kidney of fish receiving FKV at the same time point. After booster vaccination, IgT and CD8 transcripts were significantly increased in the spleen of fish vaccinated with the HKV, but not with FKV. Both vaccines induced a specific IgM response in both serum and mucus. In summary, this study showed that both HKV and FKV are promising injectable vaccines for the prevention of disease caused by TiLV in Nile tilapia.

Ultrastructural insights into the replication cycle of salmon pancreas disease virus (SPDV) using salmon cardiac primary cultures (SCPCs).

Salmon pancreas disease virus (SPDV) has been affecting the salmon farming industry for over 30 years, but despite the substantial amount of studies, there are still a number of recognized knowledge gaps, for example in the transmission of the virus. In this work, an ultrastructural morphological approach was used to describe observations after infection by SPDV of an ex vivo cardiac model generated from Atlantic salmon embryos. The observations in this study and those available on previous ultrastructural work on SPDV are compared and contrasted with the current knowledge on terrestrial mammalian and insect alphaviral replication cycles, which is deeper than that of SPDV both morphologically and mechanistically. Despite their limitations, morphological descriptions remain an excellent way to generate novel hypotheses, and this has been the aim of this work. This study has used a target host, ex vivo model and resulted in some previously undescribed features, including filopodial membrane projections, cytoplasmic stress granules or putative intracytoplasmic budding. The latter suggests a new hypothesis that warrants further mechanistic research: SPDV in salmon may have retained the capacity for non-cytolytic (persistent) infections by intracellular budding, similar to that noted in arthropod vectors of other alphaviruses. In the notable absence of a known intermediate host for SPDV, the presence of this pattern suggests that both cytopathic and persistent infections may coexist in the same host. It is our hope that the ultrastructural comparison presented here stimulates new research that brings the knowledge on SPDV replication cycle up to a similar level to that of terrestrial alphaviruses.

Proteomic characterization of serum proteins from Atlantic salmon (Salmo salar L.) from an outbreak with cardiomyopathy syndrome.

Cardiomyopathy syndrome (CMS), caused by piscine myocarditis virus (PMCV), is a serious challenge to Atlantic salmon (Salmo salar L.) aquaculture. Regrettably, husbandry techniques are the only tool to manage CMS outbreaks, and no prophylactic measures are available at present. Early diagnosis of CMS is therefore desirable, preferably with non-lethal diagnostic methods, such as serum biomarkers. To identify candidate biomarkers for CMS, the protein content of pools of sera (4 fish/pool) from salmon with a CMS outbreak (3 pools) and from clinically healthy salmon (3 pools) was compared using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). Overall, seven proteins were uniquely identified in the sera of clinically healthy fish, while 27 proteins were unique to the sera of CMS fish. Of the latter, 24 have been associated with cardiac disease in humans. These were grouped as leakage enzymes (creatine kinase, lactate dehydrogenase, glycogen phosphorylase and carbonic anhydrase); host reaction proteins (acute-phase response proteins-haptoglobin, fibrinogen, α2-macroglobulin and ceruloplasmin; and complement-related proteins); and regeneration/remodelling proteins (fibronectin, lumican and retinol). Clinical evaluation of the suitability of these proteins as biomarkers of CMS, either individually or as part of a panel, is a logical next step for the development of early diagnostic tools for CMS.

Regional and organ-level responses to local lung irradiation in sheep.

Lung is a dose-limiting organ in radiotherapy. This may limit tumour control when effort is made in planning to limit the likelihood of radiation-induced lung injury (RILI). Understanding the factors that dictate susceptibility to radiation-induced pulmonary fibrosis will aid in the prevention and management of RILI, and may lead to more effective personalized radiotherapy treatment. As the interaction of regional and organ-level responses may shape the chronic consequences of RILI, we sought to characterise both aspects of the response in an ovine model. A defined volume of left pulmonary parenchyma was prescribed 5 fractions of 6 Gy within 14 days while the contralateral lung dose was constrained. Radiographic changes via computed tomography (CT) were documented to define differences in radio-exposed lung relative to non-exposed lung at d21, d63 and d171 (n = 2), and at d21, d147 and d227 (n = 2). Gross and histologic lung changes were evaluated in samples derived at necropsy examination to define the chronic pulmonary response to radiation. Irradiated lung demonstrated reduced radio-density and increased homogeneity as evidenced from texture based radiomic feature analysis, relative to the control lung. At necropsy, the radiation field was readily defined by pallor on the pleural surface, which was also evident on the cut surface of fixed lung specimens. The degree and homogeneity of pallor reflected the sparse presence of erythrocytes in alveolar septal capillaries of radiation-exposed lung. These changes contrasted with dilated and congested microvasculature in the contralateral control lung. Referencing data to measurements made in control lung volumes of sheep experiencing acute RILI indicated that interstitial collagen continues to deposit in the radio-exposed lung field. Overall lung vascularity increased during the chronic response, as evidenced by increased expression of endothelial cell marker (CD31); however, vascularity was consistently decreased in irradiated lung and was negatively correlated with lung collagen. Other organ-level responses included increased expression of alpha smooth muscle actin (ASMA), increased numbers of proliferating cells (Ki67 positive), and cells expressing the dendritic cell-lysosomal associated membrane protein (DC-LAMP) antigen. The chronic response to RILI in this model is effected at both the whole organ and local lung level. Whilst the long-term consequences of exposure to radiation involved the continued deposition of collagen in the radiation field, organ-level responses also included increased vascularization and increased expression of ASMA, Ki67 and DC-LAMP. Interrupting the interplay between these aspects may influence susceptibility to pulmonary fibrosis after radiotherapy. We advocate for the importance of large animal model systems in pursuing these opportunities to target local, organ-level and systemic mechanisms in parallel within the same subject over time.

Non-Lethal Sequential Individual Monitoring of Viremia in Relation to DNA Vaccination in Fish-Example Using a Salmon Alphavirus DNA Vaccine in Atlantic Salmon Salmo salar.

Traditionally, commercial testing for vaccine efficacy has relied on the mass infection of vaccinated and unvaccinated animals and the comparison of mortality prevalence and incidence. For some infection models where disease does not cause mortality this approach to testing vaccine efficacy is not useful. Additionally, in fish experimental studies on vaccine efficacy and immune response the norm is that several individuals are lethally sampled at sequential timepoints, and results are extrapolated to represent the kinetics of immune and disease parameters of an individual fish over the entire experimental infection period. In the present study we developed a new approach to vaccine testing for viremic viruses in fish by following the same individuals over the course of a DNA vaccination and experimental infection through repeated blood collection and analyses. Injectable DNA vaccines are particularly efficient against viral disease in fish. To date, two DNA vaccines have been authorised for use in fish farming, one in Canada against Infectious Haemorrhagic Necrotic virus and more recently one in Europe against Salmon Pancreatic Disease virus (SPDv) subtype 3. In the current study we engineered and used an experimental DNA vaccine against SPDv subtype 1. We measured viremia using a reporter cell line system and demonstrated that the viremia phase was completely extinguished following DNA vaccination. Differences in viremia infection kinetics between fish in the placebo group could be related to subsequent antibody levels in the individual fish, with higher antibody levels at terminal sampling in fish showing earlier viremia peaks. The results indicate that sequential non-lethal sampling can highlight associations between infection traits and immune responses measured at asynchronous timepoints and, can provide biological explanations for variation in data. Similar to results observed for the SPDv subtype 3 DNA vaccine, the SPDv subtype 1 DNA vaccine also induced an interferon type 1 response after vaccination and provided high protection against SPDv under laboratory conditions when fish were challenged at 7 weeks post-vaccination.

Investigating mechanisms underlying genetic resistance to Salmon Rickettsial Syndrome in Atlantic salmon using RNA sequencing.

BACKGROUND: Salmon Rickettsial Syndrome (SRS), caused by Piscirickettsia salmonis, is one of the primary causes of morbidity and mortality in Atlantic salmon aquaculture, particularly in Chile. Host resistance is a heritable trait, and functional genomic studies have highlighted genes and pathways important in the response of salmon to the bacteria. However, the functional mechanisms underpinning genetic resistance are not yet well understood. In the current study, a large population of salmon pre-smolts were challenged with P. salmonis, with mortality levels recorded and samples taken for genotyping. In parallel, head kidney and liver samples were taken from animals of the same population with high and low genomic breeding values for resistance, and used for RNA-Sequencing to compare their transcriptome profile both pre and post infection. RESULTS: A significant and moderate heritability (h2 = 0.43) was shown for the trait of binary survival. Genome-wide association analyses using 38 K imputed SNP genotypes across 2265 animals highlighted that resistance is a polygenic trait. Several thousand genes were identified as differentially expressed between controls and infected samples, and enriched pathways related to the host immune response were highlighted. In addition, several networks with significant correlation with SRS resistance breeding values were identified, suggesting their involvement in mediating genetic resistance. These included apoptosis, cytoskeletal organisation, and the inflammasome. CONCLUSIONS: While resistance to SRS is a polygenic trait, this study has highlighted several relevant networks and genes that are likely to play a role in mediating genetic resistance. These genes may be future targets for functional studies, including genome editing, to further elucidate their role underpinning genetic variation in host resistance.

Xanthogranulomatous Pituitary Adenoma in a Dog with Typical Hyperadrenocorticism.

Xanthogranuloma of the sellar region has been reported in both humans and animals. The lesion is rare, and its aetiology and pathogenesis are not fully understood. The association of sellar xanthogranuloma with an adenoma, known as xanthogranulomatous pituitary adenoma (XPA), is an extremely rare condition in humans and is usually associated with anterior pituitary insufficiencies, headache, vomiting and visual deficits. We present the first report of XPA in an animal. A 7-year-old male neutered Labrador Retriever was presented for investigation of progressive lethargy, vomiting and hyporexia, having been previously diagnosed with chronic kidney disease, pituitary-dependent hyperadrenocorticism and hypoparathyroidism. The dog was euthanized due to lack of response to medical treatment and post-mortem examination revealed XPA. Although rare, xanthogranulomatous lesions should be considered in patients with pituitary disease.

Simultaneous detection of scale drop disease virus and Flavobacterium columnare from diseased freshwater-reared barramundi Lates calcarifer.

Freshwater farming of barramundi Lates calcarifer in Thailand is a growing sector in aquaculture, but mortalities due to infectious diseases are still a major threat to this industry. In 2018, an episode of severe mortality in juvenile barramundi was noted in a freshwater earth pond site. Fish presented with severe gill necrosis, as well as severe cutaneous hemorrhages, scale loss, and discoloration at the base of dorsal fin (saddleback lesions). Histopathology revealed extensive necrosis of skeletal muscle and gill filaments, as well as basophilic inclusion bodies and megalocytosis in muscle, gill, liver, and kidney. Scale drop disease virus (SDDV) infection was subsequently confirmed by virus-specific semi-nested PCR. Further, DNA sequences of the viral major capsid protein (MCP) and ATPase genes had a respective homology of 99.85 and 99.92% with sequences of SDDV infecting barramundi in saltwater culture. Gill necrosis and saddleback lesions are not typical lesions associated with scale drop syndrome. Their presence was explained by Flavobacterium columnare isolation from the gill, followed by positive F. columnare-specific PCR. To our knowledge, this is the first report of SDDV-associated mortality in freshwater-farmed barramundi. Furthermore, this mortality presented as a concurrent infection with SDDV and F. columnare, with typical lesions of both infections.

Comparison of histologic methods for the detection of Desmozoon lepeophtherii spores in the gills of Atlantic salmon.

Desmozoon lepeophtherii is a microsporidian associated with gill disease in farmed Atlantic salmon (Salmo salar). Detection of the parasite in histologic tissue sections is challenging using common histochemical stains given that the small, widely distributed parasite spores typically occur individually or in small clusters. We compared the ability of 4 histologic methods to detect D. lepeophtherii spores in serial sections of Atlantic salmon gill tissue: hematoxylin and eosin (H&E), Gram-Twort (GT), calcofluor white (CW), and immunohistochemistry (IHC). Using CW as a benchmark to calculate a relative ratio, IHC consistently detected more spores than CW (median: 1.3), followed by GT (median: 0.2) and H&E (median: 0.1). IHC detected significantly more spores than GT (p < 0.05) and H&E (p < 0.05), and GT more than H&E (p < 0.05). We found significant underestimation of numbers of microsporidia spores in gill disease in Atlantic salmon using conventional histochemical stains and recommend the use of CW or IHC to detect the parasite in tissue sections.