Evaluation of Ki-67, goblet cell and MUC2 mucin RNA expression in dogs with lymphoplasmacytic and granulomatous colitis.

Intestinal mucus barrier disruption may occur with chronic inflammatory enteropathies. The lack of studies evaluating mucus health in dogs with chronic colitis arises from inherent challenges with assessment of the intestinal mucus layer. It is therefore unknown if reduced goblet cell (GBC) numbers and/or mucin 2 (MUC2) expression, which are responsible for mucus production and secretion, correlate with inflammation severity in dogs with granulomatous colitis (GC) or lymphocytic-plasmacytic colitis (LPC). It is undetermined if Ki-67 immunoreactivity, which has been evaluated in dogs with small intestinal inflammation, similarly correlates to histologic severity in GC and LPC. Study objectives included comparing Ki-67 immunoreactivity, GBC population and MUC2 expression in dogs with GC, LPC and non-inflamed colon; and exploring the use of ribonucleic acid (RNAscope®) in-situ hybridization (ISH) to evaluate MUC2 expression in canine colon. Formalin-fixed endoscopic colonic biopsies were obtained from 48 dogs over an eight-year period. A blinded pathologist reviewed all biopsies. Dogs were classified into the GC (n=19), LPC (n=19) or no colitis (NC) (n=10) group based on final histopathological diagnosis. Ki-67 immunohistochemistry, Alcian-Blue/PAS staining to highlight GBCs, and RNAscope® ISH using customized canine MUC2-targeted probes were performed. At least five microscopic fields per dog were selected to measure Ki-67 labelling index (KI67%), GBC staining percentage (GBC%) and MUC2 expression (MUC2%) using image analysis software. Spearman's correlation coefficients were used to determine associations between World Small Animal Veterinary Association histologic score (WHS) and measured variables. Linear regression models were used to compare relationships between WHS with KI67%, GBC%, and MUC2%; and between GBC% and MUC2%. Median WHS was highest in dogs with GC. Median KI67% normalised to WHS was highest in the NC group (6.69%; range, 1.70-23.60%). Median GBC% did not correlate with colonic inflammation overall. Median MUC2% normalised to WHS in the NC group (10.02%; range, 3.05-39.09%) was two- and three-fold higher than in the GC and LPC groups respectively. With increased colonic inflammation, despite minimal changes in GBC% overall, MUC2 expression markedly declined in the LPC group (-27.4%; 95%-CI, -49.8, 5.9%) and mildly declined in the GC and NC groups. Granulomatous colitis and LPC likely involve different pathways regulating MUC2 expression. Decreased MUC2 gene expression is observed in dogs with chronic colitis compared to dogs without colonic signs. Changes in MUC2 expression appear influenced by GBC activity rather than quantity in GC and LPC.

Exploring the pathology of liver, kidney, muscle, and stomach of fledgling seabirds associated with plastic ingestion.

There remain significant gaps in knowledge about 'sub-lethal' impacts of plastic ingestion, particularly chronic impacts on cells, tissues, or organs. Few studies have applied traditional animal health tools, such as histopathology, to assess physiological damage to wildlife, with fewer still providing information on the dosage or exposure to plastics needed to elicit negative effects. Our study seeks to investigate a common hypothesis in plastic pollution research; that an increasing plastics burden will have an impact on an animal's health, examining two wild species with high levels of environmental exposure to plastic through their diet. Here we assess the histopathology of the muscle, upper digestive tract, liver and kidney of two seabird species that are known to be commonly exposed to plastic, comparing exposed and non-exposed individuals. Fledgling seabirds showed histopathological evidence of cumulative pressures such as starvation, disease, and endoparasite burden. However, we observed no evidence of chronic harm that could be explicitly linked to the plastics. We found one case of haemorrhage, reaffirming that large/sharp plastic foreign bodies may cause acute physical damage. Given the numerous interacting pressures on the health of fledging seabirds, including exposure to plastic, this study highlights the need to scrutinise plastic-animal interactions and research though a One Health lens.

Survey of the Clinical, Cytological, and Histopathological Features Associated with Neoplasms in Captive Avian Species in Melbourne, Australia.

Over a 3.5-year period, tissue samples from 141 companion and aviary birds with clinically suspected, naturally occurring solid neoplasms were collected via surgical biopsy (n = 53) or at necropsy examination (n = 88) from a population of birds presenting to an avian veterinary clinic in Melbourne, Australia. Neoplastic processes were identified in 73.7% (n = 104) of samples, with 83.7% (n = 87) being interpreted as malignant neoplasia and 16.3% (n = 17) being interpreted as benign neoplasia. The overall estimated prevalence of neoplasia in the study population (n = 5125) was 2.45% (95% confidence interval, 2-2.97%). The gastrointestinal and reproductive tracts were the most commonly affected systems. The most common presenting signs were nonspecific and included lethargy, coelomic distension, and inappetence. In 59 cases, fine-needle aspirates (FNAs) and impression smears were also obtained and evaluated cytologically. The accuracy of cytology for detecting neoplasia in birds and for determining whether a neoplastic process was benign or malignant was assessed by using histopathology as the "gold standard." There was complete agreement between the cytological and histopathological diagnoses in 72.8% (43/59) of cases. Cytology correctly identified 87.5% of these cases (35/40) as malignant neoplasms and 55.6% (5/9) as benign processes. There was no significant difference between the use of cytology and histopathology for the detection of malignant neoplasia in birds (P = 0.185). The accuracies of FNAs and impression smears for examining avian tumors were also compared. Overall, the best cell preservation was obtained by performing impression smears from tissues, with 62.2% (n = 28) returning high cellularity for cytological examination, compared to 53.8% (n = 14) when samples were obtained by FNA. This study provides an overview of the types and prevalence of neoplasms in a captive bird population from Australia, correlates physical examination findings with tumor types, and provides evidence that cytology is a reliable preliminary diagnostic tool for detecting neoplasia in birds.

A Multicentre Epidemiologic Study of Sudden and Unexpected Death in Adult Cats and Dogs in Australia.

Sudden and unexpected death (SUD) is a common reason for animals to undergo post-mortem examination. There is limited literature examining the causes of SUD in cats and dogs, and no research specific to Australia. The purpose of this study was to investigate the epidemiology and pathology of SUD in cats and dogs in a multicentric study across Australia. Retrospective post-mortem reports of SUD in cats and dogs were obtained from four veterinary schools in Australia distributed across four states. The frequency of SUD between institutes ranged from 2.1% to 6.5%. Dogs composed the majority of the study population (76%), and males outnumbered females, particularly in the feline subpopulation. After necropsy, 37% of SUD remained cause unknown, the largest category in both cats and dogs. When cause was identified, cardiovascular disease was most common in both species, followed by gastrointestinal disease in dogs, and trauma in cats. In dogs, multinomial logistic regression identified age as a risk factor significantly associated with the four largest categories of SUD. This study identified causes of SUD in Australian cats and dogs, including novel causes not previously reported. Further, this study revealed a higher rate of unsolved SUD in Australia than can be found in the literature from other countries.

Oil spill causes mass mortality of sea snakes in the Gulf of Oman.

Oil spills in the marine environment inflict significant impacts on a wide diversity of marine fauna. Despite the abundance of literature describing these impacts on numerous species, no studies describe the impacts on sea snakes. In this study we report, for the first time, details of an oil spill which caused mass mortality of sea snakes. In this study, 39 sea snake mortalities from the Gulf of Oman, in particular, the coast of Kalba, Sharjah, UAE, were examined. The investigated sea snakes belong to four different species (Hydrophis platurus, H. lapemoides, H. spiralis and H. ornatus). The majority (84.6%) of sea snakes were observed to have oil covering 75-100% of their bodies. The majority (91.4%) of sea snakes were also observed with oil covering their snouts and eyes. A large proportion (25.8, 41.4 and 34.5%) of sea snakes were observed with oil in their mouth, esophagus and stomach.

Genetic characterisation of Echinocephalus spp. (Nematoda: Gnathostomatidae) from marine hosts in Australia.

We genetically characterised larval and adult specimens of species of Echinocephalus Molin, 1858 (Gnathostomatidae) collected from various hosts found within Australian waters. Adult specimens of Echinocephalus were collected from a dasyatid stingray [Pastinachus ater (Macleay); n = 2] from Moreton Bay, Queensland and larvae from a hydrophiine sea snake [Hydrophis peronii (Duméril); n = 3] from Cape York Peninsula, Queensland, from an octopus (Octopus djinda Amor & Hart; n = 3) from Fremantle, Western Australia and from a lucinid bivalve [Codakia paytenorum (Iredale); n = 5] from Heron Island, Queensland Australia. All nematode samples were identified morphologically and genetically characterised using the small subunit nuclear ribosomal DNA (SSU). Some morphological differences were identified between previous studies of Echinocephalus spp. and those observed herein but the significance of these differences remains unresolved. Molecular phylogenetic analyses revealed that larval Echinocephalus sp. from H. peronii and C. paytenorum in Australia were very similar (with strong nodal support) to larval Echinocephalus sp. infecting two fish species from Egypt, Saurida undosquamis (Richardson) (Synodontidae) and Pagrus pagrus (Linnaeus) (Sparidae). The SSU sequences of larval Echinocephalus sp. from O. djinda and adults from P. ater formed a well-supported clade with that of adult E. overstreeti Deardorff and Ko, 1983 from the Port Jackson shark, Heterodontus portusjacksoni (Meyer), as well as that of the larval Echinocephalus sp., from the common carp (Cyprinus carpio Linnaeus) from Egypt. This study extends the intermediate host range of Echinocephalus larvae by including a sea snake for the first time. Findings of this study highlight the importance of genetic characterisation of larval and adult specimens of Echinocephalus spp. to resolve the current difficulties in the taxonomy of this genus.

Ocular teratoid medulloepithelioma in a northern red-shouldered macaw: case report and literature review.

A 4-mo-old northern red-shouldered macaw (Diopsittaca nobilis) was admitted to the veterinary hospital of the Arruda Câmara Zoo, in the State of Paraiba, Brazil, for investigation of an orbital mass. Given rapid progression and lack of response to treatment, the bird was euthanized, and an autopsy was performed. Histologically, the mass consisted of a retrobulbar invasive tumor characterized by tubular and rosette-like structures, with interspersed heteroplastic tissues, such as aggregates of neuroglial cells and islands of hyaline cartilage. The tumor was immunopositive for pancytokeratin, GFAP, NSE, and S100. These findings were compatible with an ocular teratoid medulloepithelioma, a neoplasm best described in humans but also reported rarely in young cockatiels and African Grey parrots.

Shapes of pathology

ABSTRACT: The shape is one of the key features of a lesion and a pathologist must be able to identify and interpret these forms in the context of any gross and microscopic changes. One of the principles of adult learning is to engage the learner with previously understood information. If, when presenting material that is new, a connection with something the student already has familiarity with, the learning process is accelerated. As the learners are already familiar with shapes they have encountered throughout their pre-pathology learning, these can be used to hasten the incorporation and understanding of lesions. This paper describes various shapes that are used in describing lesions in veterinary pathology.

RESUMO: A forma é uma das principais características de uma lesão. Um patologista deve ser capaz de identificar e interpretar essas formas no contexto de quaisquer alterações macroscópicas ou microscópicas. Um dos princípios da aprendizagem de adultos é envolver o aluno com informações previamente compreendidas. Se, ao apresentar um material novo, se estabelece uma conexão com algo que o aluno já conhece, o processo de aprendizagem torna-se mais rápido e eficiente. Como os alunos já estão familiarizados com as formas que encontraram ao longo de sua aprendizagem pré-patologia, elas podem ser usadas para acelerar a incorporação e a compreensão das lesões. Este artigo descreve várias formas que são usadas na descrição de lesões em patologia veterinária.

Shapes of pathology / As formas da patologia

The shape is one of the key features of a lesion and a pathologist must be able to identify and interpret these forms in the context of any gross and microscopic changes. One of the principles of adult learning is to engage the learner with previously understood information. If, when presenting material that is new, a connection with something the student already has familiarity with, the learning process is accelerated. As the learners are already familiar with shapes they have encountered throughout their pre-pathology learning, these can be used to hasten the incorporation and understanding of lesions. This paper describes various shapes that are used in describing lesions in veterinary pathology.(AU)

A forma é uma das principais características de uma lesão. Um patologista deve ser capaz de identificar e interpretar essas formas no contexto de quaisquer alterações macroscópicas ou microscópicas. Um dos princípios da aprendizagem de adultos é envolver o aluno com informações previamente compreendidas. Se, ao apresentar um material novo, se estabelece uma conexão com algo que o aluno já conhece, o processo de aprendizagem torna-se mais rápido e eficiente. Como os alunos já estão familiarizados com as formas que encontraram ao longo de sua aprendizagem pré-patologia, elas podem ser usadas para acelerar a incorporação e a compreensão das lesões. Este artigo descreve várias formas que são usadas na descrição de lesões em patologia veterinária.(AU)

Histoplasmosis and multicentric lymphoma in a Nubian goat.

Following treatment for pneumonia, a 1-y-old female Nubian goat was presented because of a persistent fever for 3 mo and peripheral lymphadenopathy for 1 mo. Cytology and histology of the superficial cervical and prefemoral lymph nodes demonstrated a moderate-to-marked "left-shifted" lymphoid population, suggestive of lymphoma, and extremely rare extracellular, 2-4 µm, oval, basophilic yeast, consistent with Histoplasma capsulatum. On immunohistochemistry, >95% of the lymphocytes demonstrated positive cytoplasmic and membranous immunoreactivity for CD3. Histoplasma spp. urine antigen and serum antibody testing were positive and negative, respectively. Panfungal PCR and sequencing of DNA extracted from scrolls of formalin-fixed, paraffin-embedded tissue yielded matches to H. capsulatum with 99-100% identity. Given the poor prognosis and persistent pyrexia, the animal was euthanized. Postmortem examination confirmed concurrent multicentric, intermediate-size, T-cell, lymphoblastic lymphoma and histoplasmosis; lesions consistent with intestinal coccidiosis and suspected pulmonary Rhodococcus equi were also noted. Although dimorphic fungi have been described previously in goats, lesions of Histoplasma spp. had not been documented in this species, to our knowledge. Given the low disease burden, it is suspected that the lymphoma was primary, leading to an immunocompromised state and development of secondary, opportunistic infections.

Chronic excess fluoride uptake contributes to degenerative joint disease (DJD): Evidence from six marsupial species.

One of the manifestations of chronic fluoride toxicosis in mammals is skeletal fluorosis, which can include lesions of degenerative joint disease (DJD). Although DJD lesions have been less commonly studied than bone or dental lesions in relation to the pathology and epidemiology of fluoride toxicosis, there have been multiple independent studies in various species that have concluded that there appears to be an effect. The mechanisms by which fluoride affects the joints are not clear, but our data provide evidence that chronic excess dietary fluoride intake contributes to DJD. Our study is the first to specifically address the association between fluoride exposure and DJD in multiple species of free-ranging mammals. We describe levels of DJD in six marsupial species (Macropus giganteus, Notamacropus rufogriseus, Wallabia bicolor, Phascolarctos cinereus, Trichosurus vulpecula and Pseudocheirus peregrinus) inhabiting high and low fluoride environments. Lesions occurred to varying extents in all species, and lesion distribution varied with biomechanical differences in gait. In addition, we show an association (independent of age) between increasing bone fluoride concentration (as a measure of fluoride exposure) and increasing prevalence of moderate and severe DJD in five species of marsupial, which we propose does not persist at the highest levels of fluoride exposure due to selective survival bias.

Postmortem examination of Australian sea snakes ( Hydrophiinae): Anatomy and common pathologic conditions.

There is limited published information about disease in wild sea snakes and no standardized guideline for postmortem examination of sea snakes. Identifying causes of morbidity and mortality of marine vertebrate species has been pivotal to understanding disease factors implicated in stranding events and assisting with the formulation of conservation plans. Additionally, postmortem findings can provide valuable information on life history traits and the ecology of these reclusive species. Sick, moribund, or dead sea snakes are intermittently washed ashore along Australian and international beaches and provide an opportunity to examine a subset of the population and identify causes of population decline. We present an illustrated description of sea snake anatomy and describe a systematic approach to postmortem examination of sea snakes. We describe common pathologic conditions identified from clinical and postmortem examinations of stranded Australian sea snakes from southeast Queensland. Notable pathologic conditions include traumatic injury, inflammatory conditions, parasitic infections, and neoplasia.

Skeletal fluorosis in marsupials: A comparison of bone lesions in six species from an Australian industrial site.

In this study we explored the prevalence, type, location and severity of skeletal lesions in six species of Australian marsupial (Macropus giganteus, Notamacropus rufogriseus, Wallabia bicolor, Phascolarctos cinereus, Trichosurus vulpecula and Pseudocheirus peregrinus) from high and low-fluoride environments. Lesions occurred to varying extents in all species, and lesion distribution varied with biomechanical differences in gait and mastication. Bone fluoride levels increased with severity of periosteal hyperostosis. The mean bone fluoride concentration of individuals lacking hyperostosis (across all species, from both high and low-fluoride environments) was 1100±260µgF-/g dry bone, compared to 4300±1200µgF-/g and 6300±1200µgF-/g in those with mild and severe grade hyperostosis, respectively. Multivariable modelling showed that the probability of observing a lesion varied across species, anatomical location, age and bone fluoride concentration (in a non-linear manner). The pathological changes reported in the marsupials are consistent with the range of fluoride-related lesions described in other mammals, and biomechanical differences among the studied marsupial species offer some explanation for the degree of interspecific variability in prevalence, type, anatomical location, and severity of the lesions.

Ultrastructural and Molecular Characterisation of an Heterosporis-Like Microsporidian in Australian Sea Snakes (Hydrophiinae).

Four sea snakes (two Hydrophis major, one Hydrophis platurus, one Hydrophis elegans) were found washed ashore on different beaches in the Sunshine Coast region and Fraser Island in Queensland, Australia between 2007-2013. Each snake had multiple granulomas and locally extensive regions of pallor evident in the hypaxial and intercostal musculature along the body. Lesions in two individuals were also associated with vertebral and rib fractures. Histological examination revealed granulomas scattered throughout skeletal muscle, subcutaneous adipose tissue and fractured bone. These were composed of dense aggregates of microsporidian spores surrounded by a mantle of macrophages. Sequences (ssrRNA) were obtained from lesions in three sea snakes and all revealed 99% similarity with Heterosporis anguillarum from the Japanese eel (Anguillarum japonica). However, ultrastructural characteristics of the organism were not consistent with those of previous descriptions. Electron microscopic examination of skeletal muscle revealed large cysts (not xenomas) bound by walls of fibrillar material (Heterosporis-like sporophorocyst walls were not detected). The cysts contained numerous mature microsporidian spores arranged in small clusters, sometimes apparently within sporophorous vesicles. The microspores were monomorphic, oval and measured 2.5-3.0 µm by 1.6-1.8 µm. They contained isofilar polar filaments with 11 (infrequently 9-12) coils arranged in two ranks. This is the first published report of a microsporidian infection in hydrophiid sea snakes. This discovery shows microsporidia with molecular affinities to Heterosporis anguillarum but ultrastructural characters most consistent with the genus Pleistophora (but no hitherto described species). Further studies are required to determine whether the microsporidian presented here belongs to the genus Heterosporis, or to a polymorphic species group as suggested by the recognition of a robust Pleistophora/Heterosporis clade by molecular studies. The gross and histological pathology associated with these infections are described.

Experimental manipulation reveals few subclinical impacts of a parasite community in juvenile kangaroos.

Large mammalian herbivores are commonly infected with gastrointestinal helminths. In many host species, these helminths cause clinical disease and may trigger conspicuous mortality events. However, they may also have subclinical impacts, reducing fitness as well as causing complex changes to host growth patterns and body condition. Theoretically, juveniles should experience significantly greater costs from parasites, being immunologically naive and undergoing a significant growth phase. The aims of our study were to quantify the subclinical effects of helminths in juvenile eastern grey kangaroos (Macropus giganteus), which commonly harbour large burdens of gastrointestinal nematodes and are susceptible to associated mass mortality during cold, wet conditions. We conducted a field experiment on a population of free-ranging kangaroos, removing nematodes from one group of juveniles using an anthelmintic treatment. We then compared growth parameters (body condition and growth rates) and haematological parameters of this group with an age-matched, parasitised (untreated) control group. Treated juvenile kangaroos had significantly higher levels of plasma protein (albumin) but, contrary to our predictions, showed negligible changes in all the other parameters measured. Our results suggest that juvenile kangaroos are largely unaffected by their gastrointestinal helminth burdens, and may be able to compensate for the costs of parasites.

A new concept to stimulate mucosal as well as systemic immunity by parenteral vaccination as applied to the development of a live attenuated Salmonella enterica serovar Dublin vaccine.

A new concept of slow "drip feeding" that enables activation of mucosal as well as systemic immunity following parenteral vaccination was demonstrated using Salmonella Dublin in a mouse model. The live vaccine candidate, N-RM25, generated from a wild S. Dublin strain utilising metabolic-drift (spontaneous chromosomal) mutations had a unique sensitivity to bile and restricted growth in the presence of a very low concentration of bile salts No. 3 (0.075% (w/v)) but also had the ability to survive in a high concentration (19.2%) of the substance. Following intraperitoneal administration with 10(7) cfu, N-RM25 colonised and survived (10(1)-10(3) cfu/g) in the liver and spleen of mice for over 24 days without causing disease. A small number of the mutant organisms also penetrated the gall bladder and gut, most likely via the enterohepatic circulation. N-RM25 induced significant levels of serum IgG, IgA and intestinal secretory IgA. A second metabolic-drift mutant (R-NM29) that was rapidly eliminated from the liver and spleen and highly unlikely to penetrate the gall bladder and gut, stimulated some systemic immunity, but induced no mucosal immunity because it did not reach the immune stimulation sites within the gut. In vaccine trials, N-RM25 was significantly more effective in eliminating the homologous challenge bacteria (S. Dublin wild strain FD436) from the internal organs and intestinal lumen when compared to R-NM29 and the negative control. N-RM25 prevented the development of systemic infection and produced 100% protection.