Classical BSE dismissed as the cause of CWD in Norwegian red deer despite strain similarities between both prion agents.

The first case of CWD in a Norwegian red deer was detected by a routine ELISA test and confirmed by western blotting and immunohistochemistry in the brain stem of the animal. Two different western blotting tests were conducted independently in two different laboratories, showing that the red deer glycoprofile was different from the Norwegian CWD reindeer and CWD moose and from North American CWD. The isolate showed nevertheless features similar to the classical BSE (BSE-C) strain. Furthermore, BSE-C could not be excluded based on the PrPSc immunohistochemistry staining in the brainstem and the absence of detectable PrPSc in the lymphoid tissues. Because of the known ability of BSE-C to cross species barriers as well as its zoonotic potential, the CWD red deer isolate was submitted to the EURL Strain Typing Expert Group (STEG) as a BSE-C suspect for further investigation. In addition, different strain typing in vivo and in vitro strategies aiming at identifying the BSE-C strain in the red deer isolate were performed independently in three research groups and BSE-C was not found in it. These results suggest that the Norwegian CWD red deer case was infected with a previously unknown CWD type and further investigation is needed to determine the characteristics of this potential new CWD strain.

Effective detection of BRAFV595E mutation in canine urothelial and prostate carcinomas using immunohistochemistry.

Canine urothelial carcinoma (UC) and prostate carcinoma (PC) frequently exhibit the BRAFV595E mutation, akin to the BRAFV600E mutation common in various human cancers. Since the initial discovery of the BRAF mutation in canine cancers in 2015, PCR has been the standard method for its detection in both liquid and tissue biopsies. Considering the similarity between the canine BRAFV595E and human BRAFV600E mutations, we hypothesized that immunohistochemistry (IHC) using a BRAFV600E-specific antibody could effectively identify the canine mutant BRAFV595E protein. We tested 122 canine UC (bladder n = 108, urethra n = 14), 21 PC, and benign tissue using IHC and performed digital droplet PCR (ddPCR) on all 122 UC and on 14 IHC positive PC cases. The results from ddPCR and IHC were concordant in 99% (135/136) of the tumours. Using IHC, BRAFV595E was detected in 72/122 (59%) UC and 14/21 (65%) PC. Staining of all benign bladder and prostate tissues was negative. If present, mutant BRAF staining was homogenous, with rare intratumour heterogeneity in three (4%) cases of UC. Additionally, the BRAFV595E mutation was more prevalent in tumours with urothelial morphology, and less common in glandular PC or UC with divergent differentiation. This study establishes that BRAFV600-specific IHC is a reliable and accurate method for detecting the mutant BRAFV595E protein in canine UC and PC. Moreover, the use of IHC, especially with tissue microarrays, provides a cost-efficient test for large-scale screening of canine cancers for the presence of BRAF mutations. This advancement paves the way for further research to define the prognostic and predictive role of this tumour marker in dogs and use IHC to stratify dogs for the treatment with BRAF inhibitors.

Sodium iodide symporter immunolabelling as a predictor of clinical iodide uptake in canine thyroid carcinoma: A preliminary study.

Thyroid follicular tumours may take up iodide via the sodium-iodide symporter. Knowledge of iodide uptake could then allow treatment with I-131 in dogs with high-risk tumours. The objective of this study was to determine the relationship between clinically detectable iodide uptake (as determined by scintigraphy and/or thyroxine concentrations) and sodium iodide symporter immunohistochemical labelling on histologically fixed thyroid tumours. Nineteen dogs were identified who were diagnosed with thyroid carcinoma and underwent surgery from November 2017 to July 2021. All had recorded thyroid hormone concentrations and were hyperthyroid and/or underwent preoperative nuclear imaging using planar scintigraphy (technetium-99m or I-123), or I-124 PET-CT. All dogs subsequently underwent surgery to remove the thyroid mass. Twenty-two tumours were submitted for histopathologic analysis immediately following surgery, which confirmed a diagnosis of thyroid carcinoma for each tumour. Images and/or thyroid hormone concentrations were reviewed for the included cases, and tumours were sorted into an avid/functional group (group 1) and a non-avid/functional group (group 2). The tumour tissues were re-examined histologically using sodium iodide symporter (NIS) immunohistochemistry (IHC). Group 1 contained 15 avid/functional tumours. Twelve of these tumours had membranous NIS IHC labelling. Group 2 contained 7 non-avid tumours. One of these tumours had membranous NIS IHC labelling. This resulted in an overall sensitivity and specificity for identification of avid/functional tumours with membranous NIS of 80.0% and 85.7%, respectively. NIS IHC may predict ion trapping in canine follicular thyroid tumours. Further studies using iodide-based imaging are warranted to better determine the clinical utility of this diagnostic modality.

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.

Quantification of CD3, FoxP3, and granzyme B immunostaining in canine renal cell carcinoma.

Tumor-infiltrating lymphocyte (TIL) density plays an important role in anti-tumor immunity and is associated with patient outcome in various human and canine malignancies. As a first assessment of the immune landscape of the tumor microenvironment in canine renal cell carcinoma (RCC), we retrospectively analyzed clinical data and quantified CD3, FoxP3, and granzyme B immunostaining in formalin-fixed paraffin-embedded tumor samples from 16 dogs diagnosed with renal cell carcinoma treated with ureteronephrectomy. Cell density was low for all markers evaluated. Increased numbers of intratumoral FoxP3 labelled (+) cells, as well as decreased granzyme B+: FoxP3+ TIL ratio, were associated with poor patient outcomes. Our initial study of canine RCC reveals that these tumors are immunologically cold and Tregs may play an important role in immune evasion.

Evaluation of species-specific polyclonal antibodies to detect and differentiate between Neospora caninum and Toxoplasma gondii.

Neosporosis and toxoplasmosis are major causes of abortion in livestock worldwide, leading to substantial economic losses. Detection tools are fundamental to the diagnosis and management of those diseases. Current immunohistochemistry (IHC) tests, using sera raised against whole parasite lysates, have not been able to distinguish between Toxoplasma gondii and Neospora caninum. We used T. gondii and N. caninum recombinant proteins, expressed in Escherichia coli and purified using insoluble conditions, to produce specific polyclonal rabbit antisera. We aimed to develop species-specific sera that could be used in IHC on formalin-fixed, paraffin-embedded (FFPE) tissue sections to improve the diagnosis of ruminant abortions caused by protozoa. Two polyclonal rabbit sera, raised against recombinant proteins, anti-Neospora-rNcSRS2 and anti-Toxoplasma-rTgSRS2, had specificity for the parasite they were raised against. We tested the specificity for each polyclonal serum using FFPE tissue sections known to be infected with T. gondii and N. caninum. The anti-Neospora-rNcSRS2 serum labeled specifically only N. caninum-infected tissue blocks, and the anti-Toxoplasma-rTgSRS2 serum was specific to only T. gondii-infected tissues. Moreover, tissues from 52 cattle and 19 sheep previously diagnosed by lesion profiles were tested using IHC with our polyclonal sera and PCR. The overall agreement between IHC and PCR was 90.1% for both polyclonal anti-rNcSRS2 and anti-rTgSRS2 sera. The polyclonal antisera were specific and allowed visual confirmation of protozoan parasites by IHC, but they were not as sensitive as PCR testing.

PD-L1 mRNA and protein expression in canine mammary carcinomas: Correlation with histopathological grade and molecular markers.

Programmed death ligand 1 (PD-L1) is an immune checkpoint molecule that plays a crucial role in regulating antitumor immune responses. Canine mammary carcinomas (CMCs) are common tumors of dogs. Despite extensive studies on the heterogeneity of CMCs, there is still a lack of effective precision therapies for the treatment of CMCs. In this study, we aimed to investigate the correlation between PD-L1 mRNA and protein expression in CMCs and explore its association with histopathological grade and molecular markers, including the estrogen receptor, epidermal growth factor receptor 2, and cytokeratin 5/6 (CK5/6). Formalin-fixed paraffin-embedded samples were evaluated for PD-L1 mRNA expression using RNA in situ hybridization and PD-L1 protein expression using immunohistochemistry. We observed no substantial correlation between PD-L1 mRNA and protein expression in CMCs; however, PD-L1 mRNA levels were significantly higher in grade 3 than in grade 1 tumors (P = .001). In addition, we observed a positive correlation between PD-L1 protein expression and CK5/6 expression in CMCs (P = .032). These findings suggest that PD-L1 expression in CMCs is heterogeneous and may be regulated post-transcriptionally. Further studies are needed to explore the prognostic and therapeutic implications of PD-L1 expression in different molecular subtypes of CMCs and their potential as predictive biomarkers for immunotherapy.

A rare case of ocular and testicular T-cell lymphoma in a hermaphrodite koi carp (Cyprinus carpio Linnaeus 1758): clinical, histopathological, and immunohistochemical study.

BACKGROUND: Lymphatic neoplasia may occur in various types, such as lymphoma, lymphosarcoma, lympholeukemia, and plasmacytoid leukemia. Lymphoma, defined as a malignant tumour of lymphoid tissue, has been found in a number of fish families including Esocidae and Salmonidae. However, the occurrence of lymphoma is rare in those belonging to the Cyprinidae. A final diagnosis of ocular and testicular T-cell lymphoma in the present study was based on the clinical signs, morphology, and texture of the tumour masses in the macroscopic and microscopic examinations. In addition, histopathological and immunohistochemical findings corresponded to T-cell lymphoma characteristics. CASE PRESENTATION: A 2-year-old hermaphrodite koi carp (Cyprinus carpio Linnaeus 1758) with a large ocular mass and severe exophthalmia in the right eye was referred to the Ornamental Fish Clinic in October 2020. Under anesthesia, enucleation was performed. 57 days after enucleation of the right eye, exophthalmia in the left eye was discovered. 221 days after surgery, the fish was discovered to be dead. At necropsy, a large soft tissue mass attached to the left testis was discovered. There were also small whitish nodules on the surface of the liver. Histopathology revealed a hypercellular ocular mass with scant connective tissue. The sections also revealed multifocal hemorrhages, round to ovoid neoplastic cells, mild-to-moderate anisokaryosis and anisocytosis, and mitotic figures. Basophilic neoplastic cells were found in blood vessels within the testicular mass, raising the possibility of systemic spread. The liver showed microscopic metastasis with morphologic similarities to the ocular and testicular tumors. The neoplastic cells infiltrating the left and right eyes as well as the testicular mass were immunohistochemically positive for CD3 but negative for CD20. The masses were diagnosed as T-cell lymphoma based on histopathological and immunohistochemical findings. CONCLUSIONS: This case report provides the first evidence of clinical, histopathological, morphological, and immunohistochemical findings of an ocular and testicular T-cell lymphoma in a hermaphrodite koi carp (Cyprinus carpio) in Iran.

Gastroesophageal tube of the Iguana iguana (Iguanidae): histological description, histochemical and immunohistochemical analysis of 5-HT and SS cells / Tubo gastroesofágico da Iguana iguana (Iguanidae): descrição histológica, histoquímica e análise imunohistoquímica de células 5-HT e SS

The work aims were to describe the histological and histochemical structure of the gastroesophageal tube of Iguana iguana and verify the occurrence and distribution of immunoreactive serotonin (5-HT) and somatostatin (SS) cells. Fragments of the gastrointestinal tract (GIT) of five iguanas were which underwent standard histological and immunohistochemistry technique. Immunoreactive cells for 5-HT and SS were quantified using the STEPanizer. The oesophagus has ciliated columnar pseudostratified epithelium with staining Alcian blue (AB) + and goblet cells highly reactive to periodic acid Schiff (PAS). In the cervical oesophagus, the numerical density of 5-HT cells per unit area (QA [5-HT cells]/µm2) was 4.6x10-2 ± 2.0 and celomatic oesophagus presented QA = 4.0x10-2 ± 1.0. The epithelium of the stomach is simple columnar, PAS and AB +. The cranial and middle regions of the stomach presented (QA [5-HT cells]/µm2) = 6.18x10-2 ± 3.2 and the caudal region, QA = 0.6x10-2 ± 0.2. The SS cells were only observed in the caudal stomach, with numerical density (QA [SS cells]/µm2) = 1.4x10-2 ± 0.9 In I. iguana, variation was observed in terms of the distribution of mucus secretions and the pattern of occurrence of serotonin and somatostatin-secreting enteroendocrine cells in the TGI, which possibly will result in an interspecific adaptive response.

Os objetivos do trabalho foram descrever a estrutura histológica e histoquímica do tubo gastroesofágico da Iguana iguana e verificar a ocorrência e distribuição de células serotonina (5-HT) e somatostatina (SS) imunorreativas. Fragmentos do trato gastrointestinal (TGI) de cinco iguanas foram submetidos à técnica histológica e imunohistoquímica padrão. As células imunorreativas para 5-HT e SS foram quantificadas usando o STEPanizer. O esôfago apresenta epitélio pseudoestratificado colunar ciliado Alcian blue (AB) positivo, com células caliciformes altamente reativas ao ácido periódico de Schiff (PAS). No esôfago cervical, a densidade numérica de células 5-HT por unidade de área (QA [células 5-HT] / µm2) foi de 4.6x10-2 ± 2.0 e o esôfago celomático apresentou QA = 4.0x10-2 ± 1.0. O epitélio do estômago é colunar simples, PAS e AB positivo. As regiões cranial e média do estômago apresentaram (QA [células 5-HT] / µm2) = 6.18x10-2 ± 3.2 e a região caudal, QA = 0.6x10-2 ± 0.2. As células SS foram observadas apenas no estômago caudal, com densidade numérica (QA [células SS] / µm2) = 1.4x10-2 ± 0.9. Em I. iguana, foi observada variações em termos da distribuição das secreções de muco e padrão de ocorrência das células enteroendócrinas secretoras de serotonina e somatostatina no TGI, o que possivelmente reflete uma resposta adaptativa interespecifica.

Linfoma extranodal de células B em vesícula urinária de um cão / Extranodal B-cell lymphoma in the urinary bladder in a dog

O linfoma é uma neoplasia de alta recorrência na rotina oncológica de medicina veterinária. Pode ser classificado em linfoma Hodgking-liked, com raros casos descritos somente em felinos,e não Hodgking, sendo este segundo o mais comum, subdividido em linfomas B ou T. O objetivo deste trabalho foi relatar a conduta clínica, diagnóstica e terapêutica do caso de uma cadela, de 12 anos, sem raça definida, que manifestava disúria, prostração, dor abdominal e ao exame físico a presença de uma massa na região hipogástrica. Esta foi diagnosticada com linfoma de grandes células por meio de exames de citologia e biópsia, com solicitação do exame de imunoistoquímica que confirmou linfoma difuso de grandes células de imunofenótipo B. Sem o envolvimento de nenhum outro sistema, classificou-se como linfoma primário de bexiga extranodal. O animal passou pelo tratamento quimioterápico realizando nove sessões de quimioterapia pelo protocolo de CHOP, contudo devido ao agravamento do caso a paciente veio a óbito cerca de sete meses após o diagnóstico da doença. O caso estudado foi de extrema importância para a compreensão de linfomas primários de bexiga em razão da escassez de informações relacionadas na literatura. Ainda, o cão é um excelente modelo experimental de linfomas não Hodgking em humanos, consequentemente compreender essa doença em cães promove a evolução conjunta da medicina humana.

Lymphoma is a highly recurrent rate neoplasm in the oncology routine of veterinary medicine. It can be classified into Hodgking-like, rarely described just in felines, and non-Hodgking lymphoma, the latter being the most commun, subdivided into B-cell lymphoma and T-cell lymphoma. The objective of this study was to report the clinical and therapeutic conduct within the diagnosis procedures of a 12-years-old female dog, mixed breed, who manifested dysuria, prostation, abdominal pain and on the physical examination a mass in the hypogastric region was noticed. This was diagnosed as a large cell lymphoma by means cytology and biopsy, also immunohistochemistry was required which confirmed the diffuse large cell lymphoma of immunophenotyping B. Without any other sistem envolved, the neoplasm was classified as primary urinary bladder lymphoma extranodal. The animal underwent chemotherapy, performing nine sessions according to the Madison protocol, however, due to the worsening of the case, the patient died about seven months after the diagnosis of the disease. This case was extremely importante for the understanding of primary urinary bladder lymphomas due to the scarcity of informations in the literature. Also, dog is an excellent experi,emtal model of non Hodgking lymphomas in humans, thus understandig this disease in dogs promotes the joint evolution of human medicine.

Metastatic endometrioid carcinoma in a hybrid Leontopithecus sp.

Primary female reproductive neoplasms in Platyrrhines species are few reported. We present the gross, histological, and immunohistochemical findings of metastatic endometrioid carcinoma in the uterus, urinary bladder, jejunum, and rectum of a Leontopithecus sp. The neoplastic endometrial cells expressed strong cytoplasmic immunolabeling of cytokeratin 7.

Dynamics of epigenetic modifiers and environmentally sensitive proteins in a reptile with temperature induced sex reversal†.

The mechanisms by which sex is determined, and how a sexual phenotype is stably maintained during adulthood, have been the focus of vigorous scientific inquiry. Resources common to the biomedical field (automated staining and imaging platforms) were leveraged to provide the first immunofluorescent data for a reptile species with temperature induced sex reversal. Two four-plex immunofluorescent panels were explored across three sex classes (sex reversed ZZf females, normal ZWf females, and normal ZZm males). One panel was stained for chromatin remodeling genes JARID2 and KDM6B, and methylation marks H3K27me3, and H3K4me3 (Jumonji Panel). The other CaRe panel stained for environmental response genes CIRBP and RelA, and H3K27me3 and H3K4me3. Our study characterized tissue specific expression and cellular localization patterns of these proteins and histone marks, providing new insights to the molecular characteristics of adult gonads in a dragon lizard Pogona vitticeps. The confirmation that mammalian antibodies cross react in P. vitticeps paves the way for experiments that can take advantage of this new immunohistochemical resource to gain a new understanding of the role of these proteins during embryonic development, and most importantly for P. vitticeps, the molecular underpinnings of sex reversal.

Histopathological and immunohistochemical characteristics of malignant adenomyoepithelioma in a cat: case report / [Características histopatológicas e imuno-histoquímicas do adenomioepitelioma maligno em gata: relato de caso]

The malignant adenomyoepithelioma is a rare mammary tumor in women and uncommon in cats with only one report in this species. In this case report, the histopathological and immunohistochemical characteristics of six cases of malignant adenomyopithelioma in the feline mammary gland are described. Microscopic evaluation of tumors showed dense cellular neoplastic proliferation, composed of malignant myoepithelial and epithelial cells, formed by varied arrangements and presenting papillary, tubular and solid nest proliferation. Immunohistochemistry was performed for markers Ki67, Cox-2, RE, RP, p63 and HER-2. All cases were positive for p63, confirming the myoepithelial nature of neoplastic cells. The diagnosis of malignant adenomyopithelioma was made possible through the association between histopathological characteristics and immunohistochemical results.(AU)

O adenomioepitelioma maligno é uma neoplasia mamária rara em mulheres e incomum em gatas, possuindo apenas uma descrição nessa espécie. Neste relato de caso, são descritas as características histopatológicas e imuno-histoquímicas de seis casos de adenomioepitelioma maligno na glândula mamária felina. A avaliação microscópica dos tumores demonstrou proliferação neoplásica densamente celular, composta por células mioepiteliais e epiteliais malignas dispostas em padrão papilar, tubular e ninhos sólidos. Foi realizada técnica de imuno-histoquímica para os marcadores Ki67, Cox-2, RE, RP, p63 e HER-2. Todos os casos foram positivos para p63, confirmando a natureza mioepitelial das células neoplásicas. O diagnóstico de adenomioepitelioma maligno foi possível por meio da associação entre as características histopatológicas e os resultados de imuno-histoquímica.(AU)

Ultra- and micro-structural changes of respiratory tracts in SARS-CoV-2 infected Syrian hamsters.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic is causing a global crisis. It is still unresolved. Although many therapies and vaccines are being studied, they are still in their infancy. As this pandemic continues, rapid and accurate research for the development of therapies and vaccines is needed. Therefore, it is necessary to understand characteristics of diseases caused by SARS-CoV-2 through animal models. Syrian hamsters are known to be susceptible to SARS-CoV-2. They were intranasally inoculated with SARS-CoV-2. At 2, 4, 8, 12, and 16 days post-infection (dpi), these hamsters were euthanized, and tissues were collected for ultrastructural and microstructural examinations. Microscopic lesions were prominent in the upper and lower respiratory tracts from 2 and 4 dpi groups, respectively. The respiratory epithelium in the trachea, bronchiole, and alveolar showed pathological changes. Inflammatory cells including neutrophils, lymphocytes, macrophages, and eosinophils were infiltrated in/around tracheal lamina propria, pulmonary vessels, alveoli, and bronchiole. In pulmonary lesions, alveolar wall was thickened with infiltrated inflammatory cells, mainly neutrophils and macrophages. In the trachea, epithelial damages started from 2 dpi and recovered from 8 dpi, consistent with microscopic results, High levels of SARS-CoV-2 nucleoprotein were detected at 2 dpi and 4 dpi. In the lung, lesions were most severe at 8 dpi. Meanwhile, high levels of SARS-CoV-2 were detected at 4 dpi. Electron microscopic examinations revealed cellular changes in the trachea epithelium and alveolar epithelium such as vacuolation, sparse micro-organelle, and poor cellular margin. In the trachea epithelium, the number of cytoplasmic organelles was diminished, and small vesicles were prominent from 2 dpi. Some of these electron-lucent vesicles were filled with virion particles. From 8 dpi, the trachea epithelium started to recover. Because of shrunken nucleus and swollen cytoplasm, the N/C ratio of type 2 pneumocyte decreased at 8 and 12 dpi. From 8 dpi, lamellar bodies on type 2 pneumocyte cytoplasm were increasingly observed. Their number then decreased from 16 dpi. However, there was no significant change in type 1 pneumocyte. Viral vesicles were only observed in the cytoplasm of type 2 pneumocyte. In conclusion, ultra- and micro-structural changes presented in this study may provide useful information for SARS-CoV-2 studies in various fields.

Pathology, immunohistochemistry and molecular study of cattle screened by comparative tuberculin test for diagnosis latent form of Johne's disease.

Mycobacterium avium subsp. paratuberculosis causes Johne's disease in cattle and other ruminants. It is one of the most widespread in terms of prevalence and one of the most important diseases in the livestock industry in terms of economic importance. In the latent form of the disease, animals can act as carriers. The main purpose of this study was to evaluate avian tuberculin in a single intradermal comparative tuberculin test (SICTT), histopathology, immunohistochemistry, and molecular methods for diagnosis of infected cows with the latent form of the disease. Twenty-three asymptomatic cows and without post-mortem signs of the disease were selected using the single intradermal comparative tuberculin test and were tested. In this study, avian tuberculin in SICTT was able to in diagnosing the latent form of the disease. The results confirm immunohistochemistry and real-time PCR as a sensitive and reliable diagnostic complementary approach for detecting latent forms of Paratuberculosis.

Primary cell culture of canine corneal endothelial cells.

OBJECTIVE: To establish a primary cell culture and clarify the characteristics of canine corneal endothelial cells in vitro. PROCEDURES: The eyes were enucleated from dogs that were euthanized for reasons unrelated to this study. Enucleated canine eyes were dissected, and the intact corneas were isolated from the globes. Using enzymes, the corneal endothelial cells were dispersed from the cornea. The obtained canine corneal endothelial cells were cultured in a cell culture dish. Cultured corneal endothelial cells were morphologically evaluated using phase-contrast microscopy. Immunohistochemical analysis of the cultured cells, particularly of the corneal endothelial cell marker, zonula occludens-1 (ZO-1), Na+ /K+ -ATPase, and vimentin, was performed to clarify whether the cultured cells were actually corneal endothelial cells. Furthermore, the post-passage morphology of cultured cells was evaluated. RESULTS: Canine primary cultured corneal endothelial cells showed morphologically small, cobblestone-like structures. The isolated cells had proliferative ability in vitro and demonstrated positive expression of the corneal endothelial cell markers, ZO-1, Na+ /K+ -ATPase, and vimentin. However, repeated passages resulted in larger cell sizes as assessed by phase-contrast microscopy. Repeated passages also resulted in lower cell density. CONCLUSIONS: This study demonstrated the successful culture of canine corneal endothelial cells. This might enhance the understanding of corneal endothelial cell characteristics in dogs.

Enteric neuronal development in canine small intestine - an immunohistochemical study.

The development of the enteric nervous system (ENS) is still a valid and intensely studied issue. However, literature in the field has no data on this topic in the dog. The present investigations were performed in three groups of fetuses from mongrel dogs - from the third, sixth- -seventh, and ninth week of pregnancy - and in 3-5-day-old puppies (3 specimens for each age group). The tissues (the medial parts of the duodenum, jejunum, and ileum with the cecum and a small portion of the adjacent ascending colon) were cut using a cryostat and the sections were processed for single- and double-labeling immunohistochemistry using antisera against acetylated tubulin (AcTub), vesicular acetylcholine transporter (VAChT), nitric oxide synthase (NOS), vasoactive intestinal polypeptide (VIP), galanin (GAL), neuropeptide Y (NPY), substance P (SP), and calcitonin gene-related peptide (CGRP). In the 3-week-old fetuses, some oval cells invading the gut wall were found. From the seventh week of pregnancy onwards, two different enteric ganglia were present: submucosal and myenteric. The estimated number of nerve elements in the 9-week-old fetuses was much higher than that observed in the 6-7-week-old individuals. There was no significant difference in the estimated number of nerve structures between the 9-week-old fetuses and the 3-5-day-old puppies. The colonization pattern and the development of the ENS in the canine small intestine are very similar to those observed in other mam- mals. However, a few exceptions have been confirmed, regarding the time of appearance of the VIP-, GAL-, and CGRP-immunoreactive neurons, and their distribution in different portions of the canine bowel during development.

Light and electron-microscopic localization of CD9 and surfactant protein A and D in normal lungs of the horse.

The lung is a complex organ, and its physiology and immunology are regulated by various immune molecules and cells. Lung surfactant, a mixture of phospholipids and proteins produced by the bronchiolar and type II alveolar epithelial cells, is one such important player in lung physiology. Compared to knowledge about the biology of the surfactant in rodents and humans, only limited data are available on the surfactant in the horse. Although there are data linking levels of surfactant proteins with respiratory disease in the horse, there are no data on the cellular localization of surfactant protein A (SP-A) and surfactant protein D (SP-D). A member of the tetraspanin family of proteins, CD9 is a cell-signaling and adhesion protein and its expression has been detected in both normal and cancer cells, including those in the lung. Because there are no immunolocalization data on SP-A, SP-D, and CD9 in the normal lungs of the horse, our objective was to conduct a light and electron microscopic immunocytochemical study on normal lungs of the horse. The data showed SP-A and SP-D in bronchiolar epithelial and type II alveolar epithelial cells. These proteins were also localized in type I alveolar epithelial cells, pulmonary intravascular macrophages, and neutrophils, which is likely an outcome of endocytosis of the proteins by these cells. CD9 was present in the airway and vascular smooth muscle cells, endothelium, and blood cells, but not in the airway epithelium. These new data provide a baseline to further examine the expression and functions of SP-A, SP-D, and CD9 proteins in inflammation associated with respiratory diseases in the horse.

Le poumon est un organe complexe, et sa physiologie et son immunologie sont régulées par diverses molécules et cellules immunitaires. Le surfactant pulmonaire, un mélange de phospholipides et de protéines produits par les cellules épithéliales bronchiolaires et alvéolaires de type II, est un acteur important de la physiologie pulmonaire. Par rapport aux connaissances sur la biologie du surfactant chez les rongeurs et les humains, seules des données limitées sont disponibles sur le surfactant chez le cheval. Bien qu'il existe des données reliant les niveaux de protéines du surfactant à une maladie respiratoire chez le cheval, il n'y a pas de données sur la localisation cellulaire de la protéine de surfactant A (SP-A) et de la protéine de surfactant D (SP-D). Membre de la famille des protéines tétraspanines, CD9 est une protéine de signalisation et d'adhésion cellulaire et son expression a été détectée dans les cellules normales et cancéreuses, y compris celles du poumon. Comme il n'y a pas de données d'immunolocalisation pour SP-A, SP-D et CD9 dans les poumons normaux du cheval, notre objectif était de mener une étude immunocytochimique au microscope optique et électronique sur les poumons normaux du cheval. Les données ont montré la présence de SP-A et SP-D dans les cellules épithéliales bronchiolaires et alvéolaires de type II. Ces protéines étaient également localisées dans les cellules épithéliales alvéolaires de type I, les macrophages intravasculaires pulmonaires et les neutrophiles, ce qui est probablement le résultat de l'endocytose des protéines par ces cellules. Le CD9 était présent dans les cellules des voies respiratoires et des muscles lisses vasculaires, l'endothélium et les cellules sanguines, mais pas dans l'épithélium des voies respiratoires. Ces nouvelles données fournissent une base de référence pour examiner plus à fond l'expression et les fonctions des protéines SP-A, SP-D et CD9 dans l'inflammation associée aux maladies respiratoires chez le cheval.(Traduit par Docteur Serge Messier).

Pathological and Immunohistochemical Microscopy of Natural Cases of Canine and Feline Neoplastic Mammary Lesions.

Mammary cancer is the second most common tumor worldwide. Small animal mammary neoplasms provide an outstanding model to study cancer in humans, as tumors in both share a similar environment, histopathologic features, and biological behavior. This study aims to investigate the percentage and microscopy of breast tumors in affected dogs and cats; its relationship to breed, age, and sex; and the immunohistochemical expression of estrogen receptor (ER), progesterone receptor (PR), Ki-67, and cytokeratin 8. Twenty-four females (12 dogs and 12 cats) and one male were examined from February 2018 to February 2020. The highest percentage of mammary neoplasia from the highest to the lowest manifested as tubular carcinoma, leiomyosarcoma, fibroadenoma, and cystic papillary carcinoma. The current study reported the second micropapillary invasive carcinoma in a male cat and the third lipid-rich carcinoma in a female cat. Although tubular carcinoma was the most common mammary neoplasm in cats, leiomyosarcoma was the most common in dogs. The immunohistochemical staining revealed diffuse and intense cytoplasmic immunoreactivity for cytokeratin 8 in lipid-rich carcinomas. However, moderate expression of ER in benign tumors and slight to moderate ER expression in malignant mammary lesions were reported. On the contrary, there was a negative PR expression in benign lesion. It could be concluded that a close relationship between ER expression and nuclear antigen Ki-67 was found.

Expression of DEP Domain-Containing 1B in Canine Lymphoma and Other Types of Canine Tumours.

Canine lymphoma is the most common haematological malignancy in dogs and is typically treated with multidrug chemotherapy. Most cases are at risk of relapse after several courses of chemotherapy and the oncogenic mechanism remains unknown. This study was aimed at identifying genes expressed in canine lymphoma by cDNA microarray. We found elevated expression of Dishevelled, EGL-10 and pleckstrin (DEP) domain-containing 1B (DEPDC1B) in canine lymphoma cells compared with cells and tissues from healthy dogs. Canine DEPDC1B protein was detected in 13 of 41 lymphoma specimens by immunohistochemistry, but was not detected in lymph nodes from normal dogs. Immunoreactive DEPDC1B protein was also detected in several other types of canine tumour. This is the first report documenting the association of DEPDC1B with canine cancer and the results suggest that DEPDC1B might serve as a potential marker or therapeutic target for canine malignancies.