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The objective of the study was to describe the clinical, imaging, surgical and histological findings in a dog with Rathke's cleft cyst of the pituitary gland. A 6-year-old, female, neutered Staffordshire bull terrier was presented with an acute onset of abnormal behaviour. Magnetic resonance imaging of the skull showed a pituitary mass of 12.9 mm (height) × 8.8 mm (width) × 10.2 mm (length) with a pituitary height/brain area value of 0.73 (reference

Gross morphology of healthy and degenerated intervertebral discs (IVDs) is largely similar in horses as in dogs and humans. For further comparison, the biochemical composition and the histological and biochemical changes with age and degeneration were analyzed in 41 warmblood horses. From 33 horses, 139 discs and 2 fetal vertebral columns were evaluated and scored histologically. From 13 horses, 73 IVDs were assessed for hydration, DNA, glycosaminoglycans, total collagen, hydroxyl-lysyl-pyridinoline, hydroxylysine, and advanced glycation end-product (AGE) content. From 7 horses, 20 discs were assessed for aggrecan, fibronectin, and collagen type 1 and 2 content. Histologically, tearing of the nucleus pulposus (NP) and cervical annulus fibrosus (AF), and total histological score (tearing and vascular proliferation of the AF, and chondroid metaplasia, chondrocyte-like cell proliferation, presence of notochordal cells, matrix staining, and tearing of the NP) correlated with gross degeneration. Notochordal cells were not seen in IVDs of horses. Age and gross degeneration were positively correlated with AGEs and a fibrotic phenotype, explaining gross degenerative changes. In contrast to dogs and humans, there was no consistent difference in glycosaminoglycan content and hydration between AF and NP, nor decrease of these variables with age or degeneration. Hydroxylysine decrease and collagen 1 and AGEs increase were most prominent in the NP, suggesting degeneration started in the AP. In caudal cervical NPs, AGE deposition was significantly increased in grossly normal IVDs and total collagen significantly increased with age, suggesting increased biomechanical stress and likelihood for spinal disease in this part of the vertebral column.

An 11-year-old, English cocker spaniel was presented with subacute progressive signs of vestibular ataxia, tetraparesis, left-sided proprioceptive deficits, positional ventrolateral strabismus of the right eye and a right-sided menace deficit. Magnetic resonance imaging of the cranium and cranial cervical spinal cord revealed multifocal T2 hyper-/T1-hypointense intradural lesions with dural tail signs and intra-axial and intramedullary extension. Medical treatment resulted in initial improvement before deterioration was noticed. Cytological examination results of computed tomography-guided fine-needle aspiration biopsy of the C1–C2 lesion were consistent with mesenchymal neoplasia. Three days later, after progressive clinical deterioration, euthanasia was performed. Postmortem examination and subsequent histological examination of the brainstem and spinal cord revealed multifocal, strongly infiltrative growth of neoplastic cells with areas of pseudo-rosette formation by cylindrical neoplastic cells with moderately large, oval nuclei in addition to areas of spindle-shaped neoplastic cells with meningothelial whorls. The final diagnosis was a papillary (grade III) meningioma with multifocal leptomeningeal spread.

Rabies is caused by infection with a lyssavirus. Bat rabies is of concern for both public health and bat conservation. The current method for lyssavirus prevalence studies in bat populations is by oral swabbing, which is invasive for the bats, dangerous for handlers, time‐consuming and expensive. In many situations, such sampling is not feasible, and hence, our understanding of epidemiology of bat rabies is limited. Faeces are usually easy to collect from bat colonies without disturbing the bats and thus could be a practical and feasible material for lyssavirus prevalence studies. To further explore this idea, we performed virological analysis on faecal pellets and oral swabs of seven serotine bats (Eptesicus serotinus) that were positive for European bat 1 lyssavirus in the brain. We also performed immunohistochemical and virological analyses on digestive tract samples of these bats to determine potential sources of lyssavirus in the faeces. We found that lyssavirus detection by RT‐qPCR was nearly as sensitive in faecal pellets (6/7 bats positive, 86%) as in oral swabs (7/7 bats positive, 100%). The likely source of lyssavirus in the faeces was virus excreted into the oral cavity from the salivary glands (5/6 bats positive by immunohistochemistry and RT‐qPCR) or tongue (3/4 bats positive by immunohistochemistry) and swallowed with saliva. Virus could not be isolated from any of the seven faecal pellets, suggesting the lyssavirus detected in faeces is not infectious. Lyssavirus detection in the majority of faecal pellets of infected bats shows that this novel material should be further explored for lyssavirus prevalence studies in bats.

We tested samples collected from camels, camel workers, and other animals in Sudan and Qatar in 2015 and 2017 for evidence of Middle East respiratory syndrome coronavirus (MERS-CoV) infection. MERS-CoV antibodies were abundant in Sudan camels, but we found no evidence of MERS-CoV infection in camel workers, other livestock, or bats.

BACKGROUND: Chronic hepatitis (CH) in dogs is common and has the tendency to progress to liver cirrhosis (LC). Circulating microRNAs might have the potential as markers for disease progression. OBJECTIVES: To investigate whether concentration of specific microRNAs in serum correlate with the stage and grade of CH in Labrador Retrievers. ANIMALS: Twenty-two Labrador Retrievers with histological CH (n = 8), LC (n = 7), and normal liver (NL, n = 7). METHODS: In this retrospective study, serum concentrations of miR-122, miR-29a, miR-133a, miR-181b, and miR-17-5p were measured by quantitative real-time PCR and evaluated using univariate linear regression in dogs. A multivariate model was fit including the grade of hepatitis and the stage of fibrosis. RESULTS: Of the 5 microRNAs, only circulating miR-122 and miR-29a were significantly associated with the grade of hepatitis and the stage of fibrosis. A positive correlation was identified between the grade of hepatitis with miR-122 (rs = 0.79, P

Histiocytic sarcoma is an aggressive hematopoietic malignancy of mature tissue histiocytes with a poorly understood etiology in humans. A histologically and clinically similar counterpart affects flat-coated retrievers (FCRs) at unusually high frequency, with 20% developing the lethal disease. The similar clinical presentation combined with the closed population structure of dogs, leading to high genetic homogeneity, makes dogs an excellent model for genetic studies of cancer susceptibility. To determine the genetic risk factors underlying histiocytic sarcoma in FCRs, we conducted multiple genome-wide association studies (GWASs), identifying two loci that confer significant risk on canine chromosomes (CFA) 5 (Pwald = 4.83x10-9) and 19 (Pwald = 2.25x10-7). We subsequently undertook a multi-omics approach that has been largely unexplored in the canine model to interrogate these regions, generating whole genome, transcriptome, and chromatin immunoprecipitation sequencing. These data highlight the PI3K pathway gene PIK3R6 on CFA5, and proximal candidate regulatory variants that are strongly associated with histiocytic sarcoma and predicted to impact transcription factor binding. The CFA5 association colocalizes with susceptibility loci for two hematopoietic malignancies, hemangiosarcoma and B-cell lymphoma, in the closely related golden retriever breed, revealing the risk contribution this single locus makes to multiple hematological cancers. By comparison, the CFA19 locus is unique to the FCR and harbors risk alleles associated with upregulation of TNFAIP6, which itself affects cell migration and metastasis. Together, these loci explain ~35% of disease risk, an exceptionally high value that demonstrates the advantages of domestic dogs for complex trait mapping and genetic studies of cancer susceptibility., Author summary We have identified two regions of the canine genome that explain a striking 35% of risk for developing histiocytic sarcoma in FCRs. The disease is uniformly lethal, affects 20% of FCRs, and parallels a cancer of the same name in humans. Both regions harbor genes involved in cell migration and cancer-related pathways. The first includes variants in regulatory regions at the tumor suppressor PIK3R6 locus that are strongly associated with histiocytic sarcoma and likely confer risk for other hematopoietic cancers. FCRs with risk alleles at the second locus demonstrate increased expression of TNFAIP6, which correlates with poor prognosis in multiple human cancers. In identifying genomic differences between affected and unaffected dogs, we advance our understanding of both canine and human health biology and set the stage for the development of diagnostic and therapeutic strategies.

Objectives To describe the signalment, ophthalmic examination findings, and follow-up of dogs affected with a previously unreported retrocorneal pigmentary lesion. Materials and methods Retrospective record evaluation spanning 2009-2019. Results Retrocorneal pigmentary lesions were described in 34 patients (46 eyes). German Shepherds (n = 7), Jack Russel terriers (n = 5), and terrier crosses (n = 4) made up 16/34 (47.1%) of the cases. The mean age was 13.5 years (range 1.4-14.2 years), and 16/30 (53.3%) dogs were female. Most dogs were affected unilaterally (22/34 (64.7%)), the others bilaterally, and 5/34 (14.7%) were referred for it while the others were incidentally diagnosed. The lesions affected the ventral, peripheral, inner cornea and had a round/undulated leading edge. The number of corneal clock hours affected was known for 41/46 (89.1%) eyes and involved 1-3 clock hours in 32/41 (78.1%) eyes, 4-6 in 6/41 (14.6%), 7-9 in 2/41 (4.9%), and 10 in 1/41 (2.4%). The central cornea was affected in 9/46 (19.6%) eyes, and in 5/9 (55.6%), the median corneal clarity score was G2 (scale: G0-G4). The commonest additional findings included free-floating uveal cysts (11/34 dogs, 32.4%), cataracts (6/34 dogs, 17.6%), and primary glaucoma (5/34 dogs, 14.7%). Gonioscopy was available in 16/34 (47.1%) dogs and was normal except in primary glaucoma cases. Follow-up was documented in 13/34 (38.2%) dogs with a mean follow-up of 17 months (range: 5-26 months). Lesion progression was documented in 6/13 (46.2%) dogs. Conclusions Retrocorneal pigmentation occurs as a slowly progressive lesion of older dogs that could impact vision. Histological studies of affected eyes are warranted.

Respiratory disease and increased mortality occurred in minks on two farms in the Netherlands, with interstitial pneumonia and SARS-CoV-2 RNA in organ and swab samples. On both farms, at least one worker had coronavirus disease-associated symptoms before the outbreak. Variations in mink-derived viral genomes showed between-mink transmission and no infection link between the farms. Inhalable dust contained viral RNA, indicating possible exposure of workers. One worker is assumed to have attracted the virus from mink.

Highlights • IBV-QX vaccine and virulent progenitor have comparable RNA titers in the trachea. • IBV-QX vaccine shows delayed induction of lesions in the respiratory tract. • IBV-QX vaccine has reduced ability to disseminate from the site of inoculation to other target tissues. • IBV-QX vaccine displays no nephropathogenicity in SPF broilers. • Ascending virus transport seems to contribute to kidney infection., Infectious bronchitis (IB) is a highly contagious respiratory disease of poultry, caused by the avian coronavirus infectious bronchitis virus (IBV). Currently, one of the most relevant genotypes circulating worldwide is IBV-QX (GI-19), for which vaccines have been developed by passaging virulent QX strains in embryonated chicken eggs. Here we explored the attenuated phenotype of a commercially available QX live vaccine, IB Primo QX, in specific pathogens free broilers. At hatch, birds were inoculated with QX vaccine or its virulent progenitor IBV-D388, and postmortem swabs and tissues were collected each day up to eight days post infection to assess viral replication and morphological changes. In the trachea, viral RNA replication and protein expression were comparable in both groups. Both viruses induced morphologically comparable lesions in the trachea, albeit with a short delay in the vaccinated birds. In contrast, in the kidney, QX vaccine viral RNA was nearly absent, which coincided with the lack of any morphological changes in this organ. This was in contrast to high viral RNA titers and abundant lesions in the kidney after IBV D388 infection. Furthermore, QX vaccine showed reduced ability to reach and replicate in conjunctivae and intestines including cloaca, resulting in significantly lower titers and delayed protein expression, respectively. Nephropathogenic IBVs might reach the kidney also via an ascending route from the cloaca, based on our observation that viral RNA was detected in the cloaca one day before detection in the kidney. In the kidney distal tubular segments, collecting ducts and ureter were positive for viral antigen. Taken together, the attenuated phenotype of QX vaccine seems to rely on slower dissemination and lower replication in target tissues other than the site of inoculation.

Objective To determine whether surgical removal of urachal anomalies improves the outcomes of dogs with recurrent lower urinary tract disease (LUTD) and bacterial urinary tract infection (BUTI). Study design Retrospective study. Animals Thirty-three dogs with urachal anomalies and recurrent LUTD or BUTI. Methods Medical records of dogs with LUTD or BUTI and a diagnosis of urachal anomaly treated by partial cystectomy were reviewed. A minimum follow-up of 9 months was required for inclusion. Results Median age at onset of clinical signs was 12 months (range, 1 month to 10 years). Urachal anomalies were detected with histopathology in 20 of 28 (71%) dogs. At a median follow-up of 22 months (range, 9-114), 21 of 28 (64%) dogs were free of signs of LUTD. Nine (27%) dogs exhibited reduced signs of LUTD; in three (9%) dogs, no clinical improvement was observed. Among the 25 dogs with confirmed preoperative BUTI, 22 clinically improved with surgery. Conclusion Partial cystectomy reduced the long-term severity of clinical signs and risk of recurrence of LUTD or BUTI in dogs with confirmed or suspected urachal anomalies. Clinical significance Partial cystectomy should be considered as an adjunct to the treatment of LUTD and BUTI in dogs.

Leprosy is a human infectious disease caused by Mycobacterium leprae or Mycobacterium lepromatosis that can also occur in animals and even manifest as zoonosis. Recently, both mycobacteria were detected in red squirrels (Sciurus vulgaris) from the British Isles. To further explore the presence of leprosy-bacilli in North-West Europe, we screened Belgian and Dutch squirrels. Tissue samples from 115 animals tested by qPCR were negative for both pathogens. No molecular or pathological evidence was found of the presence of these zoonotic pathogens in North-West Europe.

Background Carriage of Mycoplasma pneumoniae (Mp) in the nasopharynx is considered a prerequisite for pulmonary infection. It is interesting to note that Mp carriage is also detected after infection. Although B cells are known to be involved in pulmonary Mp clearance, their role in Mp carriage is unknown. Methods In this study, we show in a mouse model that Mp persists in the nose after pulmonary infection, similar to humans. Results Infection of mice enhanced Mp-specific immunoglobulin (Ig) M and IgG levels in serum and bronchoalveolar lavage fluid. However, nasal washes only contained elevated Mp-specific IgA. These differences in Ig compartmentalization correlated with differences in Mp-specific B cell responses between nose- and lung-draining lymphoid tissues. Moreover, transferred Mp-specific serum Igs had no effect on nasal carriage in B cell-deficient μMT mice, whereas this enabled μMT mice to clear pulmonary Mp infection. Conclusions We report the first evidence that humoral immunity is limited in clearing Mp from the upper respiratory tract.

Summary The effect of exposure to different UVb compact lamps on the vitamin D status of growing bearded dragons (Pogona vitticeps) was studied. Forty-two newly hatched bearded dragons (

Summary Hepatic steatosis is a highly prevalent liver disease, yet research is hampered by the lack of tractable cellular and animal models. Steatosis also occurs in cats, where it can cause severe hepatic failure. Previous studies demonstrate the potential of liver organoids for modeling genetic diseases. To examine the possibility of using organoids to model steatosis, we established a long-term feline liver organoid culture with adult liver stem cell characteristics and differentiation potential toward hepatocyte-like cells. Next, organoids from mouse, human, dog, and cat liver were provided with fatty acids. Lipid accumulation was observed in all organoids and interestingly, feline liver organoids accumulated more lipid droplets than human organoids. Finally, we demonstrate effects of interference with β-oxidation on lipid accumulation in feline liver organoids. In conclusion, feline liver organoids can be successfully cultured and display a predisposition for lipid accumulation, making them an interesting model in hepatic steatosis research., Highlights • Feline organoids can be cultured from fresh and frozen liver and from needle biopsy • A feline-specific liver culture medium allows organoid expansion up to 32 passages • When fed fatty acids, feline liver organoids store more lipids than human organoids • Feline liver organoids can be used to model hepatic steatosis, In this study Kruitwagen and colleagues establish and characterize a feline liver organoid culture, which has adult stem cell properties and can be differentiated toward hepatocyte-like cells. They propose liver organoids as a tool to model hepatic steatosis and show that feline liver organoids accumulate more lipids than human organoids when provided with excess fatty acids.

During autumn–winter 2016–2017, highly pathogenic avian influenza A(H5N8) viruses caused mass die-offs among wild birds in the Netherlands. Among the ≈13,600 birds reported dead, most were tufted ducks (Aythya fuligula) and Eurasian wigeons (Anas penelope). Recurrence of avian influenza outbreaks might alter wild bird population dynamics.

International audience; Harbour porpoises (Phocoena phocoena) are the most prevalent cetaceans in the North Sea. The fecal viral flora of 21 harbour porpoises stranded along the Dutch coastline was analyzed by a metagenomics approach. Sequences of a novel cetacean mastadenovirus, designated harbour porpoise adenovirus 1 (HpAdV-1), were detected. The sequence of a 23-kbp genomic region, spanning the conserved late region, was determined using primer walking. Phylogenetic analysis indicated that HpAdV-1 is most closely related to bottlenose dolphin adenovirus and clusters with Cetartiodactyla adenoviruses. The prevalence of HpAdV-1 was low (2.6%) based on targeted PCR-screening of the intestinal contents of 151 harbour porpoises stranded between 2010 and 2013.

Finch trichomonosis in Europe is caused by a Trichomonas gallinae subtype A1 strain, considered to be clonal because lacking genetic heterogeneity in partial genotyping. The disease recently emerged and has been associated with a 66% reduction of the British breeding greenfinch (Chloris chloris) population. In contrast, in the Netherlands, where trichomonosis was detected in 2009, the breeding greenfinch population continued to grow in subsequent years. This study aimed to elucidate whether this discrepancy in population trends is because Trichomonas infection in Dutch greenfinches is associated with less severe disease, i.e., disease being less fatal. Therefore, it characterized and quantified trichomonosis in a convenience sample of greenfinches found dead and examined post-mortem between 2009 and 2017 and compared results to published data from Great Britain. Trichomonads were detected by cytology, histology, or culture in 95/101 greenfinches. The birds with trichomonads all had microscopic lesions in the upper digestive tract consistent with trichomonosis, indicating the trichomonads caused disease. The occurrence of significant lesions due to other causes was low. Some greenfinches with trichomonosis showed no macroscopic lesions. These birds showed significantly less ulceration of the mucosa and less extensive heterophil infiltration, but extent of macrophage infiltration and presence of bacteria was similar to that of birds with macroscopic lesions, and significant lesions due to other causes were equally rare. Therefore, trichomonosis was considered similarly fatal in both groups. The frequency of fatal trichomonosis in the Dutch greenfinches did not differ significantly from that reported from Great Britain. Partial genotyping of the ITS1-5,8S-ITS2 and Fe-hydrogenase regions of T. gallinae was performed to detect genetic heterogeneity, that could indicate the presence of other, possibly less virulent, strains. In 60/63 samples there was full alignment of sequences with the clonal strain of T. gallinae subtype A1. The remaining three samples had the same single synonymous nucleotide difference in the Fe-hydrogenase region; however, pathology is these three was identical to the others. Collectively, the results provide no clear evidence for less severe disease as explanation for the discrepancy in census data trends. We conclude that trichomonosis is a threat concealed in Dutch breeding greenfinch census data.

Middle East respiratory syndrome coronavirus (MERS-CoV) uses the S1(B) domain of its spike protein to bind to dipeptidyl peptidase 4 (DPP4), its functional receptor, and its S1(A) domain to bind to sialic acids. The tissue localization of DPP4 in humans, bats, camelids, pigs, and rabbits generally correlates with MERS-CoV tropism, highlighting the role of DPP4 in virus pathogenesis and transmission. However, MERS-CoV S1(A) does not indiscriminately bind to all α2,3-sialic acids, and the species-specific binding and tissue distribution of these sialic acids in different MERS-CoV-susceptible species have not been investigated. We established a novel method to detect these sialic acids on tissue sections of various organs of different susceptible species by using nanoparticles displaying multivalent MERS-CoV S1(A). We found that the nanoparticles specifically bound to the nasal epithelial cells of dromedary camels, type II pneumocytes in human lungs, and the intestinal epithelial cells of common pipistrelle bats. Desialylation by neuraminidase abolished nanoparticle binding and significantly reduced MERS-CoV infection in primary susceptible cells. In contrast, S1(A) nanoparticles did not bind to the intestinal epithelium of serotine bats and frugivorous bat species, nor did they bind to the nasal epithelium of pigs and rabbits. Both pigs and rabbits have been shown to shed less infectious virus than dromedary camels and do not transmit the virus via either contact or airborne routes. Our results depict species-specific colocalization of MERS-CoV entry and attachment receptors, which may be relevant in the transmission and pathogenesis of MERS-CoV. IMPORTANCE MERS-CoV uses the S1(B) domain of its spike protein to attach to its host receptor, dipeptidyl peptidase 4 (DPP4). The tissue localization of DPP4 has been mapped in different susceptible species. On the other hand, the S1(A) domain, the N-terminal domain of this spike protein, preferentially binds to several glycotopes of α2,3-sialic acids, the attachment factor of MERS-CoV. Here we show, using a novel method, that the S1(A) domain specifically binds to the nasal epithelium of dromedary camels, alveolar epithelium of humans, and intestinal epithelium of common pipistrelle bats. In contrast, it does not bind to the nasal epithelium of pigs or rabbits, nor does it bind to the intestinal epithelium of serotine bats and frugivorous bat species. This finding supports the importance of the S1(A) domain in MERS-CoV infection and tropism, suggests its role in transmission, and highlights its potential use as a component of novel vaccine candidates.

Morphology of the equine cervical intervertebral disc is different from that in humans and small companion animals and published imaging data are scarcely available. The objectives of this exploratory, methods comparison study were (a) to describe MRI features of macroscopically nondegenerated and degenerated intervertebral discs (b) to test associations between spinal location and macroscopic degeneration or MRI‐detected annular protrusion and between MRI‐detected annular protrusion and macroscopic degeneration, and (c) to define MRI sequences for characterizing equine cervical intervertebral disc degeneration. Ex vivo MRI of intervertebral discs was performed in 11 horses with clinical signs related to the cervical region prior to macroscopic assessment. Mixed‐effect logistic regression modeling included spinal location, MRI‐detected annular protrusion, and presence of macroscopic degeneration with “horse” as random effect. Odds ratio and 95% confidence interval were determined. Reduced signal intensity in proton density turbo SE represented intervertebral disc degeneration. Signal voids due to presence of gas and/or hemorrhage were seen in gradient echo sequences. Presence of macroscopic intervertebral disc degeneration was significantly associated with spinal location with odds being higher in the caudal (C5 to T1) versus cranial (C2 to C5) part of the cervical vertebral column. Intervertebral discs with MRI‐detected annular protrusion grades 2‐4 did have higher odds than with grade 1 to have macroscopic degeneration. It was concluded that MRI findings corresponded well with gross macroscopic data. Magnetic resonance imaging of the equine cervical intervertebral disc seems to be a promising technique, but its potential clinical value for live horses needs to be explored further in a larger and more diverse population of horses.

Since Galileo's days the effect of size on the anatomical characteristics of the structural elements of the body has been a subject of interest. However, the effects of scaling at tissue level have received little interest and virtually no data exist on the subject with respect to the osteochondral unit in the joint, despite this being one of the most lesion-prone and clinically relevant parts of the musculoskeletal system. Imaging techniques, including Fourier transform infrared imaging, polarized light microscopy and micro computed tomography, were combined to study the response to increasing body mass of the osteochondral unit. We analyzed the effect of scaling on structural characteristics of articular cartilage, subchondral plate and the supporting trabecular bone, across a wide range of mammals at microscopic level. We demonstrated that, while total cartilage thickness scales to body mass in a negative allometric fashion, thickness of different cartilage layers did not. Cartilage tissue layers were found to adapt to increasing loads principally in the deep zone with the superficial layers becoming relatively thinner. Subchondral plate thickness was found to have no correlation to body mass, nor did bone volume fraction. The underlying trabecular bone was found to have thicker trabeculae (r=0.75, p

Rabbit haemorrhagic disease virus (RHDV) type 2 (GI.2/RHDV2/b) is an emerging pathogen in wild rabbits and in domestic rabbits vaccinated against RHDV (GI.1). Here we report the genome sequence of a contemporary RHDV2 isolate from the Netherlands and investigate the immunogenicity of virus-like particles (VLPs) produced in insect cells. RHDV2 RNA was isolated from the liver of a naturally infected wild rabbit and the complete viral genome sequence was assembled from sequenced RT-PCR products. Phylogenetic analysis based on the VP60 capsid gene demonstrated that the RHDV2 NL2016 isolate clustered with other contemporary RHDV2 strains. The VP60 gene was cloned in a baculovirus expression vector to produce VLPs in Sf9 insect cells. Density-gradient purified RHDV2 VLPs were visualized by transmission electron microscopy as spherical particles of around 30 nm in diameter with a morphology resembling authentic RHDV. Immunization of rabbits with RHDV2 VLPs resulted in high production of serum antibodies against VP60, and the production of cytokines (IFN-&gamma, and IL-4) was significantly elevated in the immunized rabbits compared to the control group. The results demonstrate that the recombinant RHDV2 VLPs are highly immunogenic and may find applications in serological detection assays and might be further developed as a vaccine candidate to protect domestic rabbits against RHDV2 infection.

Avian coronaviruses cause major global problems in the poultry industry. As causative agents of huge economic losses, the detection and understanding of the molecular determinants of viral tropism are of ultimate importance. Here, we set out to study those parameters and obtained in-depth insight into the virus-host interactions of guinea fowl coronavirus (GfCoV). Our data indicate that diversity in GfCoV viral attachment proteins results in differences in degrees of affinity for glycan receptors, as well as altered avidity for intestinal tract tissues, which might have consequences for GfCoV tissue tropism and pathogenesis in guinea fowls., Guinea fowl coronavirus (GfCoV) causes fulminating enteritis that can result in a daily death rate of 20% in guinea fowl flocks. Here, we studied GfCoV diversity and evaluated its phenotypic consequences. Over the period of 2014 to 2016, affected guinea fowl flocks were sampled in France, and avian coronavirus presence was confirmed by PCR on intestinal content and immunohistochemistry of intestinal tissue. Sequencing revealed 89% amino acid identity between the viral attachment protein S1 of GfCoV/2014 and that of the previously identified GfCoV/2011. To study the receptor interactions as a determinant for tropism and pathogenicity, recombinant S1 proteins were produced and analyzed by glycan and tissue arrays. Glycan array analysis revealed that, in addition to the previously elucidated biantennary di-N-acetyllactosamine (diLacNAc) receptor, viral attachment S1 proteins from GfCoV/2014 and GfCoV/2011 can bind to glycans capped with alpha-2,6-linked sialic acids. Interestingly, recombinant GfCoV/2014 S1 has an increased affinity for these glycans compared to that of GfCoV/2011 S1, which was in agreement with the increased avidity of GfCoV/2014 S1 for gastrointestinal tract tissues. Enzymatic removal of receptors from tissues before application of spike proteins confirmed the specificity of S1 tissue binding. Overall, we demonstrate that diversity in GfCoV S1 proteins results in differences in glycan and tissue binding properties. IMPORTANCE Avian coronaviruses cause major global problems in the poultry industry. As causative agents of huge economic losses, the detection and understanding of the molecular determinants of viral tropism are of ultimate importance. Here, we set out to study those parameters and obtained in-depth insight into the virus-host interactions of guinea fowl coronavirus (GfCoV). Our data indicate that diversity in GfCoV viral attachment proteins results in differences in degrees of affinity for glycan receptors, as well as altered avidity for intestinal tract tissues, which might have consequences for GfCoV tissue tropism and pathogenesis in guinea fowls.

8658860258318000Recently, genetic alterations in the genes encoding succinate dehydrogenase subunit B and D (SDHB and SDHD) were identified in pet dogs that presented with spontaneously arising pheochromocytomas (PCC) and paragangliomas (PGL, together PPGL), suggesting dogs might be an interesting comparative model for the study of human PPGL. To study whether canine PPGL resembled human PPGL, we investigated a series of 50 canine PPGLs by immunohistochemistry to determine the expression of synaptophysin (SYP), tyrosine hydroxylase (TH) and succinate dehydrogenase subunit A (SDHA) and B (SDHB). In parallel, 25 canine PPGLs were screened for mutations in SDHB and SDHD by Sanger sequencing. To detect large chromosomal alterations, single nucleotide polymorphism (SNP) arrays were performed for 11 PPGLs, including cases for which fresh frozen tissue was available. The immunohistochemical markers stained positive in the majority of canine PPGLs. Genetic screening of the canine tumors revealed the previously described variants in four cases, SDHB p.Arg38Gln (n = 1) and SDHD p.Lys122Arg (n = 3). Furthermore, the SNP arrays revealed large chromosomal alterations of which the loss of chromosome 5, partly homologous to human chromosome 1p and chromosome 11, was the most frequent finding (100% of the six cases with chromosomal alterations). In conclusion, canine and human PPGLs show similar genomic alterations, suggestive of common interspecies PPGL-related pathways.

BACKGROUND: Focal seizures with fear as a primary ictal manifestation, their diagnostic challenges, and impact on quality of life are well described in human medicine. Reports focusing on ictal fear-like behavior in animals are scarce. OBJECTIVE: To describe the clinical and histopathological characteristics of a novel focal epilepsy in Boerboel dogs. ANIMALS: Five client-owned Boerboel littermates presented for evaluation of sudden episodes of severe fear-related behavior. METHODS: Clinical examination, complete blood cell count, routine blood biochemistry, and urinalysis were performed in all dogs. Magnetic resonance imaging (MRI) scans of the brain were performed in 3 affected Boerboels. In addition, in 2 affected Boerboels, metabolic screening, cerebrospinal fluid (CSF) analysis, and necropsy were performed. RESULTS: Onset of signs was 3 months of age in all affected Boerboels. All Boerboels howled loudly, had an extremely fearful facial expression and trembled during seizures. All affected Boerboels also had autonomic or motor signs. Results of laboratory investigations, diagnostic imaging, and metabolic screening were generally unremarkable. Histopathology showed moderate numbers of single large vacuoles in the perikaryon of neurons throughout the brain, specifically in the deeper cerebral cortical regions. Family history, pedigree analysis, and the homogenous phenotype were suggestive of autosomal recessive inheritance. CONCLUSIONS AND CLINICAL IMPORTANCE: The observed paroxysmal fear-related behavior represents a newly recognized hereditary focal epilepsy in dogs with distinctive clinical and histopathologic features. Veterinarians should be aware that sudden episodes of unusual behavior can represent focal epilepsy.

Objective To report the long‐term outcome of nine dogs treated for caudal cervical spondylomyelopathy (CCSM) with surgical spinal fusion. Study design Short case series. Animals Nine large‐breed dogs. Methods Medical records of dogs treated for disc‐associated CCSM (2013‐2016) were reviewed. The surgery objective was spinal distraction by implantation of a SynCage and fixation with two Unilock plates. Follow‐up included the Helsinki pain score questionnaire, neurological grading, radiography, computed tomography (CT), and micro‐CT (μCT) with subsequent histopathology (two dogs). Results Clinical follow‐up was obtained between 9 and 51 months (27.4 ± 13.4 months). The Helsinki pain score and neurological Griffith score improved (P

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.

Episodes of inflammation and pain are predominant features of arthritic joint diseases. Drug delivery systems (DDS) could reduce inflammation and pain long-term without chances of infection upon multiple injections. To allow for long-term evaluation of DDS, we modified a previously published acute arthritis model by extending follow-up periods between flare-ups. Unilateral synovial inflammation of the knee was induced by intra-articular injection of streptococcal cell wall peptidoglycan polysaccharide (PGPS), and flare-ups were induced by intravenous PGPS injections every 4 weeks for a total duration of 84 days. In PGPS-reactivated animals, joint swelling, pain behavior, post mortem synovitis, and osteophyte formation were notable features. Hepatitis, splenitis and inflammation of non-primed joints were observed as systemic side effects. To test the applicability of the modified arthritis model for long-term testing of DDS, the duration of anti-inflammatory and analgesic effects of a corticosteroid released from two different polymer-based platforms was evaluated. The current modified arthritis model has good applicability for testing of DDS for a prolonged period of time. Furthermore, the novel autoregulatory polyesteramide (PEA) microsphere platform releasing triamcinolone acetonide (TAA) was benchmarked against poly lactic-co-glycolic acid (PLGA) and reduced joint swelling and pain behavior more potently compared to TAA-loaded PLGA microspheres.

Objectives The aim of this study was to describe the cellular and stromal components of the hepatic progenitor cell niche in feline hepatic lipidosis (FHL). Methods Immunohistochemical staining for the progenitor/bile duct marker (K19), activated Kupffer cells (MAC387), myofibroblasts (alpha-smooth muscle actin [α-SMA]) and the extracellular matrix component laminin were used on seven liver biopsies of cats with FHL and three healthy cats. Double immunofluorescence stainings were performed to investigate co-localisation of different cell types in the hepatic progenitor cell (HPC) niche. Results HPCs, Kupffer cells, myofibroblasts and laminin deposition were observed in the liver samples of FHL, although with variability in the expression and positivity of the different immunostainings between different samples. When compared with the unaffected cats where K19 positivity and minimal α-SMA and laminin positivity were seen mainly in the portal area, in the majority of FHL samples K19 and α-SMA-positive cells and laminin positivity were seen also in the periportal and parenchymatous area. MAC387-positive cells were present throughout the parenchyma. Conclusions and relevance This is a preliminary morphological study to describe the activation and co-localisation of components of the HPC niche in FHL. Although the HPC niche in FHL resembles that described in hepatopathies in dogs and in feline lymphocytic cholangitis, the expression of K19, α-SMA, MAC387 and lamin is more variable in FHL, and a common pattern of activation could not be established. Nevertheless, when HPCs were activated, a spatial association between HPCs and their niche could be demonstrated.

Neisseria animaloris is considered to be a commensal of the canine and feline oral cavities. It is able to cause systemic infections in animals as well as humans, usually after a biting trauma has occurred. We recovered N. animaloris from chronically inflamed bite wounds on pectoral fins and tailstocks, from lungs and other internal organs of eight harbour porpoises. Gross and histopathological evidence suggest that fatal disseminated N. animaloris infections had occurred due to traumatic injury from grey seals. We therefore conclude that these porpoises survived a grey seal predatory attack, with the bite lesions representing the subsequent portal of entry for bacteria to infect the animals causing abscesses in multiple tissues, and eventually death. We demonstrate that forensic microbiology provides a useful tool for linking a perpetrator to its victim. Moreover, N. animaloris should be added to the list of potential zoonotic bacteria following interactions with seals, as the finding of systemic transfer to the lungs and other tissues of the harbour porpoises may suggest a potential to do likewise in humans.

Middle East respiratory syndrome coronavirus (MERS-CoV) transmission from dromedaries to humans has resulted in major outbreaks in the Middle East. Although some other livestock animal species have been shown to be susceptible to MERS-CoV, it is not fully understood why the spread of the virus in these animal species has not been observed in the field. In this study, we used rabbits to further characterize the transmission potential of MERS-CoV. In line with the presence of MERS-CoV receptor in the rabbit nasal epithelium, high levels of viral RNA were shed from the nose following virus inoculation. However, unlike MERS-CoV-infected dromedaries, these rabbits did not develop clinical manifestations including nasal discharge and did shed only limited amounts of infectious virus from the nose. Consistently, no transmission by contact or airborne routes was observed in rabbits. Our data indicate that despite relatively high viral RNA levels produced, low levels of infectious virus are excreted in the upper respiratory tract of rabbits as compared to dromedary camels, thus resulting in a lack of viral transmission.

Recently, protocols for amplicon based whole genome sequencing using Nanopore technology have been described for Ebola virus, Zika virus, yellow fever virus and West Nile virus. However, there is some debate regarding reliability of sequencing using this technology, which is important for applications beyond diagnosis such as linking lineages to outbreaks, tracking transmission pathways and pockets of circulation, or mapping specific markers. To our knowledge, no in depth analyses of the required read coverage to compensate for the error profile in Nanopore sequencing have been described. Here, we describe the validation of a protocol for whole genome sequencing of USUV using Nanopore sequencing by direct comparison to Illumina sequencing. To that point we selected brain tissue samples with high viral loads, typical for birds which died from USUV infection. We conclude that the low-cost MinION Nanopore sequencing platform can be used for characterization and tracking of Usutu virus outbreaks.

Four outbreaks of leukoencephalomyelopathy in colonies of SPF cats on a long‐term diet of irradiated dry cat food were observed in the Netherlands between 1989 and 2001. As a primary defect in myelin formation was suspected to be the cause of the disease and myelin consists mainly of lipids and their fatty acids, we investigated the fatty acid composition of the white matter of the spinal cord of affected and control cats and of irradiated and non‐irradiated food. The irradiated food had low levels of alpha‐linolenic acid compared to linoleic acid as well as a high total omega‐6:omega‐3 ratio of 7:1 in the irradiated and of 2:1 in the non‐irradiated food. The white matter of the spinal cord showed low levels of linoleic acid and absence of alpha‐linolenic acid in affected cats as well as absence of lignoceric and nervonic acid in both affected and control cats. These abnormalities in fatty acid composition of the white matter of the spinal cord may reflect an increased need for alpha‐linolenic acid as a substrate for longer chain omega‐3 fatty acids to compose myelin and thus indicate a particular species sensitivity to dietary deficiency in omega‐3 polyunsaturated fatty acids, particularly alpha‐linolenic acid in cats. Our findings indicate that abnormalities in fatty acid metabolism in myelin play an essential role in the pathogenesis of this acquired form of leukoencephalomyelopathy in cats.

Sequence-based typing of Francisella tularensis has led to insights in the evolutionary developments of tularemia. In Europe, two major basal clades of F. tularensis subsp. holarctica exist, with a distinct geographical distribution. Basal clade B.6 is primarily found in Western Europe, while basal clade B.12 occurs predominantly in the central and eastern parts of Europe. There are indications that tularemia is geographically expanding and that strains from the two clades might differ in pathogenicity, with basal clade B.6 strains being potentially more virulent than basal clade B.12. This study provides information on genotypes detected in the Netherlands during 2011–2017. Data are presented for seven autochthonous human cases and for 29 European brown hares (Lepus europaeus) with laboratory confirmed tularemia. Associated disease patterns are described for 25 European brown hares which underwent post-mortem examination. The basal clades B.6 and B.12 are present both in humans and in European brown hares in the Netherlands, with a patchy geographical distribution. For both genotypes the main pathological findings in hares associated with tularemia were severe (sub)acute necrotizing hepatitis and splenitis as well as necrotizing lesions and hemorrhages in several other organs. Pneumonia was significantly more common in the B.6 than in the B.12 cases. In conclusion, the two major basal clades present in different parts in Europe are both present in the Netherlands. In hares found dead, both genotypes were associated with severe acute disease affecting multiple organs. Hepatitis and splenitis were common pathological findings in hares infected with either genotype, but pneumonia occurred significantly more frequently in hares infected with the B.6 genotype compared to hares infected with the B.12 genotype.