Publications by authors named "Jorge Del-Pozo"

37 Publications

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

BMC Genomics 2021 Mar 6;22(1):156. Epub 2021 Mar 6.

The Roslin Institute and Royal (Dick) School of Veterinary Sciences, The University of Edinburgh, Edinburgh, UK.

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 (h = 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.
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http://dx.doi.org/10.1186/s12864-021-07443-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7936450PMC
March 2021

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

Vaccines (Basel) 2021 Feb 17;9(2). Epub 2021 Feb 17.

VIM, UVSQ, INRAE, Université Paris-Saclay, Domaine de Vilvert, 78352 Jouy-En-Josas, France.

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.
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http://dx.doi.org/10.3390/vaccines9020163DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7922653PMC
February 2021

Xanthogranulomatous Pituitary Adenoma in a Dog with Typical Hyperadrenocorticism.

J Comp Pathol 2020 Oct 11;180:115-121. Epub 2020 Oct 11.

Royal (Dick) School of Veterinary Studies, University of Edinburgh, Easter Bush, Midlothian, UK.

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.
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http://dx.doi.org/10.1016/j.jcpa.2020.09.006DOI Listing
October 2020

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

Dis Aquat Organ 2020 Aug 6;140:119-128. Epub 2020 Aug 6.

Center for Agricultural Biotechnology, Kasetsart University, Kamphaeng Saen Campus, Nakhon Pathom 73140, Thailand.

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.
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http://dx.doi.org/10.3354/dao03500DOI Listing
August 2020

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

J Vet Diagn Invest 2020 Jan 18;32(1):142-146. Epub 2019 Nov 18.

Moredun Research Institute, Pentlands Science Park, Scotland, UK (Herrero, Dagleish, Thompson).

is a microsporidian associated with gill disease in farmed Atlantic salmon (). 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 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 ( < 0.05) and H&E ( < 0.05), and GT more than H&E ( < 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.
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http://dx.doi.org/10.1177/1040638719887707DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7003232PMC
January 2020

Changes in distribution, morphology and ultrastructure of chloride cell in Atlantic salmon during an AGD infection.

J Fish Dis 2019 Oct 20;42(10):1433-1446. Epub 2019 Aug 20.

Institute of Aquaculture, School of Natural Sciences, University of Stirling, Stirling, UK.

Amoebic gill disease (AGD) is emerging as one of the most significant health challenges affecting farmed Atlantic salmon in the marine environment. It is caused by the amphizoic amoeba Neoparamoeba perurans, with infestation of gills causing severe hyperplastic lesions, compromising overall gill integrity and function. This study used histology, transmission electron microscopy (TEM), immunohistochemistry and transcript expression to relate AGD-associated pathological changes to changes in the morphology and distribution of chloride cells (CCs) in the gills of Atlantic salmon (Salmo salar L.) showing the progression of an AGD infection. A marked reduction in numbers of immunolabelled CCs was detected, and a changing pattern in distribution and morphology was closely linked with the level of basal epithelial hyperplasia in the gill. In addition, acute degenerative ultrastructural changes to CCs at the lesion site were observed with TEM. These findings were supported by the early-onset downregulation of Na /K -ATPase transcript expression. This study provides supportive evidence that histological AGD lesion assessment was a good qualitative tool for AGD scoring and corresponded well with qPCR genomic Paramoeba perurans quantification. Ultrastructural changes induced in salmon CCs as a result of AGD are reported here for the first time.
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http://dx.doi.org/10.1111/jfd.13073DOI Listing
October 2019

Response to the letter to the editor "Pneumoperitoneum should be investigated".

Vet Radiol Ultrasound 2019 11 16;60(6):754. Epub 2019 Aug 16.

The Royal (Dick) School of Veterinary Studies and The Roslin Institute, The University of Edinburgh, Roslin, UK.

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http://dx.doi.org/10.1111/vru.12802DOI Listing
November 2019

A study of residual lesions in horses that recovered from clinical signs of chronic equine dysautonomia.

J Vet Intern Med 2019 Sep 22;33(5):2302-2311. Epub 2019 Jul 22.

Royal (Dick) School of Veterinary Studies and The Roslin Institute, The University of Edinburgh, Midlothian, United Kingdom.

Background: Equine dysautonomia (ED) causes degeneration and loss of autonomic neurons. Approximately 50% of chronic cases recover, but it is unclear how they survive neuronal loss.

Objectives: To assess lesions, autonomic neuron numbers, interstitial cells of Cajal (ICC), and neurodegeneration in recovered cases.

Animals: Thirteen cases (group ED), euthanized 10.3 ± 5.2 (1-16) years from diagnosis and 6 age-matched controls (group C).

Methods: Prospective, case control; routine post mortem examination, neuron counts in peripheral and enteric ganglia and immunohistochemical assessment of neural networks (Protein gene product [PGP] 9.5), ICC (c-kit), and neurodegeneration (beta-amyloid precursor protein and ubiquitin) in intestine.

Results: Postmortem findings in group ED were small intestinal dilation (4/12, 33%) and muscular hypertrophy (4/12, 33%), and gastric mucosal hypertrophy (3/11, 27%) and ulceration (4/11, 36%). Neuron density was lower in group ED (mean 39% lower for cranial cervical ganglion [P < .001], median 44% lower in celiacomesenteric ganglion [P = .01]). In intestine, neuronal depletion was worst in ileum (median 100% lower in submucosal plexus [P < .001], 91% lower in myenteric plexus [P = .004]). Group ED had less PGP 9.5 staining in ileal myenteric plexus (mean 66% lower [P = .04]) and circular muscle (median 75% lower [P = .006]). In ileum, there was less c-kit staining in myenteric plexus (median 57% lower [P = .02]) but not muscularis externa. Beta-amyloid precursor protein and ubiquitin results were not indicitive of neurodegeneration.

Conclusions And Clinical Importance: Intact ICC in muscularis externa might help maintain motility after neuronal loss. Treatment supporting ICC function warrants investigation.
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http://dx.doi.org/10.1111/jvim.15567DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6766533PMC
September 2019

Role of ectodysplasin signalling in middle ear and nasal pathology in rat and mouse models of hypohidrotic ectodermal dysplasia.

Dis Model Mech 2019 04 25;12(4). Epub 2019 Apr 25.

Developmental Biology Division, Roslin Institute and The Royal (Dick) School of Veterinary Studies, University of Edinburgh, Edinburgh EH25 9RG, UK

Patients with mutations in the ectodysplasin receptor signalling pathway genes - the X-linked ligand ectodysplasin-A (), the receptor or the receptor adapter - have hypohidrotic ectodermal dysplasia (HED). In addition to having impaired development of teeth, hair, eccrine sweat glands, and salivary and mammary glands, HED patients have ear, nose and throat disease. The mouse strains ( ) and ( ) have rhinitis and otitis media due to loss of submucosal glands in the upper airway. We report that prenatal correction of EDAR signalling in mice with the agonist anti-EDAR antibody rescues the auditory-tube submucosal glands and prevents otitis media, rhinitis and nasopharyngitis. The sparse- and wavy-haired () rat strain carries a mutation in the gene and has similar cutaneous HED phenotypes to mouse models. We report that auditory-tube submucosal glands are smaller in the homozygous mutant than those in unaffected heterozygous rats, and that this predisposes them to otitis media. Furthermore, the pathogenesis of otitis media in the rat HED model differs from that in mice, as otitis media is the primary pathology, and rhinitis is a later-onset phenotype. These findings in rodent HED models imply that hypomorphic as well as null mutations in EDAR signalling pathway genes may predispose to otitis media in humans. In addition, this work suggests that the recent successful prenatal treatment of X-linked HED (XLHED) in humans may also prevent ear, nose and throat disease, and provides diagnostic criteria that distinguish HED-associated otitis media from chronic otitis media with effusion, which is common in children.
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http://dx.doi.org/10.1242/dmm.037804DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6505480PMC
April 2019

Leprosy in red squirrels in the UK.

Vet Rec 2019 03;184(13):416

University of Melbourne, Parkville VIC 3010, Melbourne, Australia.

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http://dx.doi.org/10.1136/vr.l1385DOI Listing
March 2019

Chronic otitis media is initiated by a bulla cavitation defect in the FBXO11 mouse model.

Dis Model Mech 2019 03 21;12(3). Epub 2019 Mar 21.

Developmental Biology Division, Roslin Institute and The Royal (Dick) School of Veterinary Studies, University of Edinburgh, Edinburgh EH25 9RG, UK

Auditory bulla cavitation defects are a cause of otitis media, but the normal cellular pattern of bulla mesenchyme regression and its failure are not well understood. In mice, neural-crest-derived mesenchyme occupies the bulla from embryonic day 17.5 (E17.5) to postnatal day 11 (P11) and then regresses to form the adult air-filled bulla cavity. We report that bulla mesenchyme is bordered by a single layer of non-ciliated epithelium characterized by interdigitating cells with desmosome cell junctions and a basal lamina, and by gene expression and laminin staining of the basal lamina. At P11-P12, the mesenchyme shrinks: mesenchyme-associated epithelium shortens, and mesenchymal cells and extracellular matrix collagen fibrils condense, culminating in the formation of cochlea promontory mucosa bordered by compact non-ciliated epithelial cells. is a candidate disease gene in human chronic otitis media with effusion and we report that a bulla cavitation defect initiates the pathogenesis of otitis media in the established mouse model ( ). Persistent mesenchyme in bullae has limited mesenchymal cell condensation, fibrosis and hyperplasia of the mesenchyme-associated epithelium. Subsequent modification forms fibrous adhesions that link the mucosa and the tympanic membrane, and this is accompanied by dystrophic mineralization and accumulation of serous effusion in the bulla cavity. Mouse models of bulla cavitation defects are important because their study in humans is limited to post-mortem samples. This work indicates new diagnostic criteria for this otitis media aetiology in humans, and the prospects of studying the molecular mechanisms of murine bulla cavitation in organ culture.
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http://dx.doi.org/10.1242/dmm.038315DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6451434PMC
March 2019

British Red Squirrels Remain the Only Known Wild Rodent Host for Leprosy Bacilli.

Front Vet Sci 2019 1;6. Epub 2019 Feb 1.

Global Health Institute, Federal Institute of Technology in Lausanne, Lausanne, Switzerland.

Eurasian red squirrels in the British Isles are the most recently discovered animal reservoir for the leprosy bacteria and . Initial data suggest that prevalence of leprosy infection is variable and often low in different squirrel populations. Nothing is known about the presence of leprosy bacilli in other wild squirrel species despite two others (Siberian chipmunk [], and Thirteen-lined ground squirrel []) having been reported to be susceptible to experimental infection with . Rats, a food-source in some countries where human leprosy occurs, have been suggested as potential reservoirs for leprosy bacilli, but no evidence supporting this hypothesis is currently available. We screened 301 squirrel samples covering four species [96 Eurasian red squirrels, 67 Eastern gray squirrels (), 35 Siberian chipmunks, and 103 Pallas's squirrels ()] from Europe and 72 Mexican white-throated woodrats () for the presence of and using validated PCR protocols. No DNA from leprosy bacilli was detected in any of the samples tested. Given our sample-size, the pathogen should have been detected if the prevalence and/or bacillary load in the populations investigated were similar to those found for British red squirrels.
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http://dx.doi.org/10.3389/fvets.2019.00008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6367869PMC
February 2019

Balancing selection at a premature stop mutation in the myostatin gene underlies a recessive leg weakness syndrome in pigs.

PLoS Genet 2019 01 30;15(1):e1007759. Epub 2019 Jan 30.

The Roslin Institute and Royal (Dick) School of Veterinary Studies, The University of Edinburgh, Midlothian, United Kingdom.

Balancing selection provides a plausible explanation for the maintenance of deleterious alleles at moderate frequency in livestock, including lethal recessives exhibiting heterozygous advantage in carriers. In the current study, a leg weakness syndrome causing mortality of piglets in a commercial line showed monogenic recessive inheritance, and a region on chromosome 15 associated with the syndrome was identified by homozygosity mapping. Whole genome resequencing of cases and controls identified a mutation causing a premature stop codon within exon 3 of the porcine Myostatin (MSTN) gene, similar to those causing a double-muscling phenotype observed in several mammalian species. The MSTN mutation was in Hardy-Weinberg equilibrium in the population at birth, but significantly distorted amongst animals still in the herd at 110 kg, due to an absence of homozygous mutant genotypes. In heterozygous form, the MSTN mutation was associated with a major increase in muscle depth and decrease in fat depth, suggesting that the deleterious allele was maintained at moderate frequency due to heterozygous advantage (allele frequency, q = 0.22). Knockout of the porcine MSTN by gene editing has previously been linked to problems of low piglet survival and lameness. This MSTN mutation is an example of putative balancing selection in livestock, providing a plausible explanation for the lack of disrupting MSTN mutations in pigs despite many generations of selection for lean growth.
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http://dx.doi.org/10.1371/journal.pgen.1007759DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6370237PMC
January 2019

The effect of micronutrient supplementation on growth and hepatic metabolism in diploid and triploid Atlantic salmon (Salmo salar) parr fed a low marine ingredient diet.

Comp Biochem Physiol B Biochem Mol Biol 2019 Jan 24;227:106-121. Epub 2018 Oct 24.

Institute of Aquaculture, University of Stirling, Stirling FK9 4LA, UK.

The effects of low marine ingredient diets supplemented with graded levels (L1, L2, L3) of a micronutrient package (NP) on growth and metabolic responses were studied in diploid and triploid salmon parr. Diploids fed L2 showed significantly improved growth and reduced liver, hepatic steatosis, and viscerosomatic indices, while fish fed L3 showed suppressed growth rate 14 weeks post feeding. In contrast, dietary NP level had no effect on triploid performance. Whole body mineral composition, with exception of copper, did not differ between diet or ploidy. Whole fish total AAs and N-metabolites showed no variation by diet or ploidy. Free circulating AAs and white muscle N-metabolites were higher in triploids than diploids, while branch-chained amino acids were higher in diploids than triploids. Diploids had higher whole body α-tocopherol and hepatic vitamins K and K than triploids. Increased tissue B-vitamins for niacin and whole-body folate with dietary NP supplementation were observed in diploids but not triploids, while whole body riboflavin was higher in diploids than triploids. Hepatic transcriptome profiles showed that diploids fed diet L2 was more similar to that observed in triploids fed diet L3. In particular, sterol biosynthesis pathways were down-regulated, whereas cytochrome P450 metabolism was up-regulated. One‑carbon metabolism was also affected by increasing levels of supplementation in both ploidies. Collectively, results suggested that, for optimised growth and liver function, micronutrient levels be supplemented above current National Research Council (2011) recommendations for Atlantic salmon when fed low marine ingredient diets. The study also suggested differences in nutritional requirements between ploidy.
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http://dx.doi.org/10.1016/j.cbpb.2018.10.004DOI Listing
January 2019

Canine and feline emphysematous gastritis may be differentiated from gastric emphysema based on clinical and imaging characteristics: Five cases.

Vet Radiol Ultrasound 2019 Mar 11;60(2):136-144. Epub 2018 Oct 11.

The Royal (Dick) School of Veterinary Studies and Roslin Institute, The University of Edinburgh, Roslin, EH25 9RG, UK.

Gastric pneumatosis is an imaging finding defined as the presence of gas foci in the gastric wall. In humans, this imaging feature can result from one of two separate clinical entities: life-threatening emphysematous gastritis or clinically benign gastric emphysema. This retrospective case series study describes the clinical and imaging features in five animals diagnosed with spontaneous gastric pneumatosis without gastric dilatation-volvulus. Three canine and two feline cases of spontaneous gastric pneumatosis were identified on radiographic and ultrasonographic examinations. In addition to gastric pneumatosis, one dog and two cats presented concomitant systemic signs such as lethargy, hematemesis, anemia, or leukocytosis. Two dogs remained asymptomatic or presented mild gastrointestinal signs. Portal gas was described in two dogs and one cat, and pneumoperitoneum in one dog. These features were not considered clinically significant. The dog and two cats with systemic signs were euthanized due to clinical deterioration and diagnosed with emphysematous gastritis. The gastric pneumatosis of both dogs without systemic signs resolved while on medical management without antibiotic therapy. These latter cases were interpreted as consistent with gastric emphysema. Findings from the current study indicated that gastric pneumatosis can occur without gastric dilatation-volvulus in cats and dogs and that a combination of clinical and imaging characteristics may help to differentiate between potentially life-threatening emphysematous gastritis and relatively benign gastric emphysema. More studies are needed to determine the etiology and risk factors associated with these conditions.
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http://dx.doi.org/10.1111/vru.12691DOI Listing
March 2019

Nebulisation of synthetic lamellar lipids mitigates radiation-induced lung injury in a large animal model.

Sci Rep 2018 09 6;8(1):13316. Epub 2018 Sep 6.

The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Edinburgh, United Kingdom.

Methods to protect against radiation-induced lung injury (RILI) will facilitate the development of more effective radio-therapeutic protocols for lung cancer and may provide the means to protect the wider population in the event of a deliberate or accidental nuclear or radiological event. We hypothesised that supplementing lipid membranes through nebulization of synthetic lamellar lipids would mitigate RILI. Following pre-treatment with either nebulised lamellar lipids or saline, anaesthetised sheep were prescribed fractionated radiotherapy (30 Gray (Gy) total dose in five 6 Gy fractions at 3-4 days intervals) to a defined unilateral lung volume. Gross pathology in radio-exposed lung 37 days after the first radiation treatment was consistent between treatment groups and consisted of deep red congestion evident on the pleural surface and firmness on palpation. Consistent histopathological features in radio-exposed lung were subpleural, periarteriolar and peribronchial intra-alveolar oedema, alveolar fibrosis, interstitial pneumonia and type II pneumocyte hyperplasia. The synthetic lamellar lipids abrogated radiation-induced alveolar fibrosis and reduced alpha-smooth muscle actin (ASMA) expression in radio-exposed lung compared to saline treated sheep. Administration of synthetic lamellar lipids was also associated with an increased number of cells expressing dendritic cell-lysosomal associated membrane protein throughout the lung.
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http://dx.doi.org/10.1038/s41598-018-31559-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6127301PMC
September 2018

Ultrasound and computed tomography of sacculitis and appendicitis in a rabbit.

Vet Radiol Ultrasound 2018 Sep 2;59(5):E56-E60. Epub 2018 Feb 2.

Royal (Dick) School of Veterinary Studies and Roslin Institute, The University of Edinburgh, Roslin, UK.

A 9-month-old neutered male rabbit was referred for lethargy, anorexia, and gastrointestinal stasis. Routine hematology, serum biochemistry, and diagnostic imaging were performed. Computed tomography revealed a wall thickening of the sacculus rotundus and appendix, which was further confirmed on abdominal ultrasound. Full thickness biopsies were collected with histopathology diagnosing a chronic multifocal heterophilic granulomatous sacculitis and appendicitis. The patient was treated medically and at 6 weeks follow-up, clinical signs and intestinal changes had completely regressed. Inflammation of the sacculus rotundus and appendix should be considered as a cause of gastrointestinal stasis in rabbits.
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http://dx.doi.org/10.1111/vru.12602DOI Listing
September 2018

Use of Salmon Cardiac Primary Cultures (SCPCs) of different genotypes for comparative kinetics of mx expression, viral load and ultrastructure pathology, after infection with Salmon Pancreas Disease Virus (SPDV).

Fish Shellfish Immunol 2018 Jan 2;72:181-186. Epub 2017 Nov 2.

Royal Dick School of Veterinary Sciences, University of Edinburgh, UK.

In vitro fish based models have been extensively applied in human biomedical research but, paradoxically, less frequently in the research of fish health issues. Farmed Atlantic salmon can suffer from several viral conditions affecting the heart. Therefore, species-specific, cardiac in vitro models may represent a useful tool to help further understanding and management of these diseases. The mechanisms underlying genotype based resistance are complex and usually rely on a combined effect of elements from both the innate and adaptive immune response, which are further complicated by external environmental factors. Here we propose that Salmon Cardiac Primary Cultures (SCPCs) are a useful tool to investigate these mechanisms as the basis for genotypic differences between Atlantic salmon families in susceptibility to cardiotropic viral disease. Using SCPCs produced from two different commercially available Atlantic salmon embryonated ova (Atlantic Ova IPN sensitive" (S) and "Atlantic QTL-innOva IPN/PD" (R)), the influence of host genotype on the viral load and mx expression following Salmon Pancreas Disease Virus infection was assessed over a 15 day period. Both R and S SCPCs groups were successfully infected. A measurable difference between groups of viral nsP1 and host antiviral mx gene expression was observed (i.e. a later, but larger onset of mx expression in the R group). Mx expression peaks were followed by a decrease in viral nsP1 in both groups. Additionally, ultrastructural examination of infected SCPCs allowed the description of degenerative changes at the individual cell level. The SCPC model presents some advantages, over current fish cell culture monolayers and in vivo material, such as the presence of different cell components normally present in the target organ, as well as the removal of a layer of functional complexity (acquired immunity), making it possible to focus on tissue specific, early innate immune mechanisms. These preliminary results highlight the importance of considering genetic origin when selecting the fish source for the production of SCPCs, as well as their usefulness as screening tools for assessment of genotypic differences in disease resistance.
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http://dx.doi.org/10.1016/j.fsi.2017.10.059DOI Listing
January 2018

Characterization of Triptolide-Induced Hepatotoxicity by Imaging and Transcriptomics in a Novel Zebrafish Model.

Toxicol Sci 2017 10;159(2):380-391

Edinburgh University/BHF Centre for Cardiovascular Science, The Queen's Medical Research Institute, Edinburgh EH16?4TJ, UK.

Triptolide is a vine extract used in traditional Chinese medicines and associated with hepatotoxicity. In vitro data suggest that inhibition of RNA synthesis may be the mechanism of toxicity. For studying drug-induced liver injury the zebrafish has experimental, practical and financial advantages compared with rodents. The aim of this study was to explore the mechanism of triptolide toxicity using zebrafish as the model system. The effect of triptolide exposure on zebrafish larvae was determined with regard to mortality, histology, expression of liver specific microRNA-122 and liver volume. Fluorescent microscopy was used to track toxicity in the Tg(-2.8lfabp:GFP)as3 zebrafish line. Informed by microscopy, RNA-sequencing was used to explore the mechanism of toxicity. Triptolide exposure resulted in dose-dependent mortality, a reduction in the number of copies of microRNA-122 per larva, hepatocyte vacuolation, disarray and oncotic necrosis, and a reduction in liver volume. These findings were consistent across replicate experiments. Time-lapse imaging indicated the onset of injury was 6 h after the start of exposure, at which point, RNA-sequencing revealed that 88% of genes were down-regulated. Immune response associated genes were up-regulated in the triptolide-treated larvae including nitric oxide synthase. Inhibition of nitric oxide synthase increased mortality. Triptolide induces hepatotoxicity in zebrafish larvae. This represents a new model of drug-induced liver injury that complements rodents. RNA sequencing, guided by time-lapse microscopy, revealed early down-regulation of genes consistent with previous invitro studies, and facilitated the discovery of mechanistic inflammatory pathways.
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http://dx.doi.org/10.1093/toxsci/kfx144DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5837554PMC
October 2017

Atlantic salmon cardiac primary cultures: An in vitro model to study viral host pathogen interactions and pathogenesis.

PLoS One 2017 20;12(7):e0181058. Epub 2017 Jul 20.

Royal Dick School of Veterinary Sciences - University of Edinburgh, Edinburgh, United Kingdom.

Development of Salmon Cardiac Primary Cultures (SCPCs) from Atlantic salmon pre-hatch embryos and their application as in vitro model for cardiotropic viral infection research are described. Producing SCPCs requires plating of trypsin dissociated embryos with subsequent targeted harvest from 24h up to 3 weeks, of relevant tissues after visual identification. SCPCs are then transferred individually to chambered wells for culture in isolation, with incubation at 15-22°. SCPCs production efficiency was not influenced by embryo's origin (0.75/ farmed or wild embryo), but mildly influenced by embryonic developmental stage (0.3 decline between 380 and 445 accumulated thermal units), and strongly influenced by time of harvest post-plating (0.6 decline if harvested after 72 hours). Beating rate was not significantly influenced by temperature (15-22°) or age (2-4 weeks), but was significantly lower on SCPCs originated from farmed embryos with a disease resistant genotype (F = 5.3, p<0.05). Two distinct morphologies suggestive of an ex vivo embryonic heart and a de novo formation were observed sub-grossly, histologically, ultra-structurally and with confocal microscopy. Both types contained cells consistent with cardiomyocytes, endothelium, and fibroblasts. Ageing of SCPCs in culture was observed with increased auto fluorescence in live imaging, and as myelin figures and cellular degeneration ultra-structurally. The SCPCs model was challenged with cardiotropic viruses and both the viral load and the mx gene expression were measurable along time by qPCR. In summary, SCPCs represent a step forward in salmon cardiac disease research as an in vitro model that partially incorporates the functional complexity of the fish heart.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0181058PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5519056PMC
September 2017

WT1 expression in breast cancer disrupts the epithelial/mesenchymal balance of tumour cells and correlates with the metabolic response to docetaxel.

Sci Rep 2017 03 27;7:45255. Epub 2017 Mar 27.

Institute of Genetics and Molecular Medicine, University of Edinburgh, Crewe Road South, Edinburgh, EH4 2XU, UK.

WT1 is a transcription factor which regulates the epithelial-mesenchymal balance during embryonic development and, if mutated, can lead to the formation of Wilms' tumour, the most common paediatric kidney cancer. Its expression has also been reported in several adult tumour types, including breast cancer, and usually correlates with poor outcome. However, published data is inconsistent and the role of WT1 in this malignancy remains unclear. Here we provide a complete study of WT1 expression across different breast cancer subtypes as well as isoform specific expression analysis. Using in vitro cell lines, clinical samples and publicly available gene expression datasets, we demonstrate that WT1 plays a role in regulating the epithelial-mesenchymal balance of breast cancer cells and that WT1-expressing tumours are mainly associated with a mesenchymal phenotype. WT1 gene expression also correlates with CYP3A4 levels and is associated with poorer response to taxane treatment. Our work is the first to demonstrate that the known association between WT1 expression in breast cancer and poor prognosis is potentially due to cancer-related epithelial-to-mesenchymal transition (EMT) and poor chemotherapy response.
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http://dx.doi.org/10.1038/srep45255DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5366898PMC
March 2017

Low-dose acetaminophen induces early disruption of cell-cell tight junctions in human hepatic cells and mouse liver.

Sci Rep 2017 01 30;7:37541. Epub 2017 Jan 30.

Hepatology Laboratory, University of Edinburgh, Royal Infirmary of Edinburgh, 49 Little France Crescent EH16 4SB, UK.

Dysfunction of cell-cell tight junction (TJ) adhesions is a major feature in the pathogenesis of various diseases. Liver TJs preserve cellular polarity by delimiting functional bile-canalicular structures, forming the blood-biliary barrier. In acetaminophen-hepatotoxicity, the mechanism by which tissue cohesion and polarity are affected remains unclear. Here, we demonstrate that acetaminophen, even at low-dose, disrupts the integrity of TJ and cell-matrix adhesions, with indicators of cellular stress with liver injury in the human hepatic HepaRG cell line, and primary hepatocytes. In mouse liver, at human-equivalence (therapeutic) doses, dose-dependent loss of intercellular hepatic TJ-associated ZO-1 protein expression was evident with progressive clinical signs of liver injury. Temporal, dose-dependent and specific disruption of the TJ-associated ZO-1 and cytoskeletal-F-actin proteins, correlated with modulation of hepatic ultrastructure. Real-time impedance biosensing verified in vitro early, dose-dependent quantitative decreases in TJ and cell-substrate adhesions. Whereas treatment with NAPQI, the reactive metabolite of acetaminophen, or the PKCα-activator and TJ-disruptor phorbol-12-myristate-13-acetate, similarly reduced TJ integrity, which may implicate oxidative stress and the PKC pathway in TJ destabilization. These findings are relevant to the clinical presentation of acetaminophen-hepatotoxicity and may inform future mechanistic studies to identify specific molecular targets and pathways that may be altered in acetaminophen-induced hepatic depolarization.
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http://dx.doi.org/10.1038/srep37541DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5278402PMC
January 2017

Detection of Tilapia Lake Virus in Clinical Samples by Culturing and Nested Reverse Transcription-PCR.

J Clin Microbiol 2017 03 14;55(3):759-767. Epub 2016 Dec 14.

Department of Cell Research and Immunology, The George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel

Tilapia are an important group of farmed fish that serve as a significant protein source worldwide. In recent years, substantial mortality of wild tilapia has been observed in the Sea of Galilee and in commercial ponds in Israel and Ecuador. We have identified the etiological agent of these mass die-offs as a novel orthomyxo-like virus and named it tilapia lake virus (TiLV). Here, we provide the conditions for efficient isolation, culturing, and quantification of the virus, including the use of susceptible fish cell lines. Moreover, we describe a sensitive nested reverse transcription-PCR (RT-PCR) assay allowing the rapid detection of TiLV in fish organs. This assay revealed, for the first time to our knowledge, the presence of TiLV in diseased Colombian tilapia, indicating a wider distribution of this emerging pathogen and stressing the risk that TiLV poses for the global tilapia industry. Overall, the described procedures should provide the tilapia aquaculture industry with important tools for the detection and containment of this pathogen.
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http://dx.doi.org/10.1128/JCM.01808-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5328443PMC
March 2017

Red squirrels in the British Isles are infected with leprosy bacilli.

Science 2016 11;354(6313):744-747

Royal (Dick) School of Veterinary Studies and Roslin Institute, University of Edinburgh, Easter Bush Campus, Roslin, Scotland, UK.

Leprosy, caused by infection with Mycobacterium leprae or the recently discovered Mycobacterium lepromatosis, was once endemic in humans in the British Isles. Red squirrels in Great Britain (Sciurus vulgaris) have increasingly been observed with leprosy-like lesions on the head and limbs. Using genomics, histopathology, and serology, we found M. lepromatosis in squirrels from England, Ireland, and Scotland, and M. leprae in squirrels from Brownsea Island, England. Infection was detected in overtly diseased and seemingly healthy animals. Phylogenetic comparisons of British and Irish M. lepromatosis with two Mexican strains from humans show that they diverged from a common ancestor around 27,000 years ago, whereas the M. leprae strain is closest to one that circulated in Medieval England. Red squirrels are thus a reservoir for leprosy in the British Isles.
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http://dx.doi.org/10.1126/science.aah3783DOI Listing
November 2016

Unusual presentations of feline cowpox.

Vet Rec 2016 Oct;179(17):442-443

Batchelor, Davidson and Watson, Edinburgh EH4 3QP; e-mail:

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http://dx.doi.org/10.1136/vr.i5767DOI Listing
October 2016

Hypersensitivity to Culicoides midges causing seasonal dermatitis in sheep.

Vet Parasitol Reg Stud Reports 2016 Jun 8;3-4:53-56. Epub 2016 Jun 8.

Farm Animal Practice, Royal (Dick) School of Veterinary Studies, University of Edinburgh, Easter Bush, Roslin, Midlothian EH25 9RG, UK.

Culicoides midges are important as vectors of disease, as an irritant with severe effects on human outdoor activities in certain areas and as the cause of insect bite hypersensitivity in domestic animals (most notably horses). Here we report, for the first time, the confirmation of ovine hypersensitivity to Culicoides midges in Hebridean sheep suffering from seasonal allergic skin disease using intradermal allergen testing. The affected sheep formed 40% of the adults in the small flock, and this indication of a potentially high prevalence of a condition with welfare implications is of concern.
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http://dx.doi.org/10.1016/j.vprsr.2016.06.001DOI Listing
June 2016

Characterization of a Novel Orthomyxo-like Virus Causing Mass Die-Offs of Tilapia.

mBio 2016 Apr 5;7(2):e00431-16. Epub 2016 Apr 5.

Center for Infection and Immunity, Mailman School of Public Health, Columbia University, New York, New York, USA

Unlabelled: Tilapia are an important global food source due to their omnivorous diet, tolerance for high-density aquaculture, and relative disease resistance. Since 2009, tilapia aquaculture has been threatened by mass die-offs in farmed fish in Israel and Ecuador. Here we report evidence implicating a novel orthomyxo-like virus in these outbreaks. The tilapia lake virus (TiLV) has a 10-segment, negative-sense RNA genome. The largest segment, segment 1, contains an open reading frame with weak sequence homology to the influenza C virus PB1 subunit. The other nine segments showed no homology to other viruses but have conserved, complementary sequences at their 5' and 3' termini, consistent with the genome organization found in other orthomyxoviruses. In situ hybridization indicates TiLV replication and transcription at sites of pathology in the liver and central nervous system of tilapia with disease.

Importance: The economic impact of worldwide trade in tilapia is estimated at $7.5 billion U.S. dollars (USD) annually. The infectious agent implicated in mass tilapia die-offs in two continents poses a threat to the global tilapia industry, which not only provides inexpensive dietary protein but also is a major employer in the developing world. Here we report characterization of the causative agent as a novel orthomyxo-like virus, tilapia lake virus (TiLV). We also describe complete genomic and protein sequences that will facilitate TiLV detection and containment and enable vaccine development.
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http://dx.doi.org/10.1128/mBio.00431-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4959514PMC
April 2016

Quantitative imaging of tissue sections using infrared scanning technology.

J Anat 2016 Jan 29;228(1):203-13. Epub 2015 Oct 29.

Roslin Institute, University of Edinburgh, Edinburgh, UK.

Quantification of immunohistochemically (IHC) labelled tissue sections typically yields semi-quantitative results. Visualising infrared (IR) 'tags', with an appropriate scanner, provides an alternative system where the linear nature of the IR fluorophore emittance enables realistic quantitative fluorescence IHC (QFIHC). Importantly, this new technology enables entire tissue sections to be scanned, allowing accurate area and protein abundance measurements to be calculated from rapidly acquired images. Here, some of the potential benefits of using IR-based tissue imaging are examined, and the following are demonstrated. Firstly, image capture and analysis using IR-based scanning technology yields comparable area-based quantification to those obtained from a modern high-resolution digital slide scanner. Secondly, IR-based dual target visualisation and expression-based quantification is rapid and simple. Thirdly, IR-based relative protein abundance QIHC measurements are an accurate reflection of tissue sample protein abundance, as demonstrated by comparison with quantitative fluorescent Western blotting data. In summary, it is proposed that IR-based QFIHC provides an alternative method of rapid whole-tissue section low-resolution imaging for the production of reliable and accurate quantitative data.
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http://dx.doi.org/10.1111/joa.12398DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4694169PMC
January 2016

Echinococcus multilocularis Detection in Live Eurasian Beavers (Castor fiber) Using a Combination of Laparoscopy and Abdominal Ultrasound under Field Conditions.

PLoS One 2015 13;10(7):e0130842. Epub 2015 Jul 13.

Veterinary Department and Conservation Programmes, Royal Zoological Society of Scotland, Edinburgh, United Kingdom; Scottish Society for the Prevention of Cruelty to Animals, National Wildlife Rescue Centre, Fishcross, United Kingdom.

Echinococcus multilocularis is an important pathogenic zoonotic parasite of health concern, though absent in the United Kingdom. Eurasian beavers (Castor fiber) may act as a rare intermediate host, and so unscreened wild caught individuals may pose a potential risk of introducing this parasite to disease-free countries through translocation programs. There is currently no single definitive ante-mortem diagnostic test in intermediate hosts. An effective non-lethal diagnostic, feasible under field condition would be helpful to minimise parasite establishment risk, where indiscriminate culling is to be avoided. This study screened live beavers (captive, n = 18 or wild-trapped in Scotland, n = 12) and beaver cadavers (wild Scotland, n = 4 or Bavaria, n = 11), for the presence of E. multilocularis. Ultrasonography in combination with minimally invasive surgical examination of the abdomen by laparoscopy was viable under field conditions for real-time evaluation in beavers. Laparoscopy alone does not allow the operator to visualize the parenchyma of organs such as the liver, or inside the lumen of the gastrointestinal tract, hence the advantage of its combination with abdominal ultrasonography. All live beavers and Scottish cadavers were largely unremarkable in their haematology and serum biochemistry with no values suspicious for liver pathology or potentially indicative of E. multilocularis infection. This correlated well with ultrasound, laparoscopy, and immunoblotting, which were unremarkable in these individuals. Two wild Bavarian individuals were suspected E. multilocularis positive at post-mortem, through the presence of hepatic cysts. Sensitivity and specificity of a combination of laparoscopy and abdominal ultrasonography in the detection of parasitic liver cyst lesions was 100% in the subset of cadavers (95%Confidence Intervals 34.24-100%, and 86.7-100% respectively). For abdominal ultrasonography alone sensitivity was only 50% (95%CI 9.5-90.6%), with specificity being 100% (95%CI 79.2-100%). For laparoscopy alone sensitivity was 100% (95% CI 34.2-100%), with specificity also being 100% (95% CI 77.2-100%). Further immunoblotting, PCR and histopathological examination revealed one individual positive for E. multilocularis, whilst the other individual was positive for Taenia martis.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0130842PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4500463PMC
April 2016