Successful Treatment of an Acinar Pancreatic Carcinoma in an Inland Bearded Dragon (Pogona vitticeps): A Case Report.

An adult, 362 g, male, intact inland bearded dragon (Pogona vitticeps) was admitted to a veterinary clinic due to a temporary cloacal prolapse and a two-week history of reduced overall condition and forage intake. Physical examination revealed an approximately 2 × 1 cm round-shaped, rigid intracoelomic tissue mass. Multiple sand deposits were present on the cloacal mucous membranes, though no signs of cloacal prolapse were present. The lizard was otherwise responsive but showed reduced body tension and movement behavior. Initial fecal examination revealed a high-grade oxyuriasis. A 2 × 1.5 cm sized intracoelomic, well-vascularized, round-shaped mass was subsequently visualized by ultrasonography. After a two-day stabilization therapy, the intracoelomic mass was removed by performing a standard ventral coeliotomy under general anesthesia. Histopathological examination of the excised mass revealed an acinar pancreatic adenocarcinoma with infiltration of the peritumorous connective soft tissue. The lizard remained at the clinic for a further seven days. Its postsurgical condition improved slowly. However, the lizard started regular forage intake 10 days after surgery, and general behavior enhanced constantly within the following three weeks. The animal was presented for a follow-up six weeks after surgery, showing bright and alert behavior with no signs of disease or illness. The lizard was re-examined 20 months after the initial presentation due to a reduced overall condition and reduced food intake. Blood chemistry evaluation revealed markedly decreased protein parameters, and moderate ascites was identified ultrasonographically. A distinct association with the preceding neoplastic disease could not be made, and the lizard returned to its regular condition under supportive therapy within three weeks. To the authors' knowledge, this is the first report of successful treatment of a pancreatic carcinoma in a bearded dragon.

Spontaneous Lethal Outbreak of Influenza A Virus Infection in Vaccinated Sows on Two Farms Suggesting the Occurrence of Vaccine-Associated Enhanced Respiratory Disease with Eosinophilic Lung Pathology.

Influenza A virus (IAV) infections in swine are usually subclinical, but they can reach high morbidity rates. The mortality rate is normally low. In this study, six vaccinated, spontaneously deceased sows revealed IAV infection and enhanced neutrophilic bronchopneumonia with unexpectedly large numbers of infiltrating eosinophils. The purpose of this study was to characterize these lung lesions with special emphasis on the phenotypes of inflammatory cells, the presence of eosinophilic peroxidase (EPO), and neutrophil extracellular traps (NETs). The number of Sirius red-stained eosinophils was significantly higher in the lungs of IAV-infected sows compared to healthy pigs, indicating a migration of eosinophils from blood vessels into the lung tissue stimulated by IAV infection. The detection of intra- and extracellular EPO in the lungs suggests its contribution to pulmonary damage. The presence of CD3+ T lymphocytes, CD20+ B lymphocytes, and Iba-1+ macrophages indicates the involvement of cell-mediated immune responses in disease progression. Furthermore, high numbers of myeloperoxidase-positive cells were detected. However, DNA-histone-1 complexes were reduced in IAV-infected sows, leading to the hypothesis that NETs are not formed in the IAV-infected sows. In conclusion, our findings in the lungs of IAV-infected vaccinated sows suggest the presence of so far unreported field cases of vaccine-associated enhanced respiratory disease.

Flowmetry and spectrophotometry for the assessment of intestinal viability in horses with naturally occurring strangulating small intestinal lesions.

BACKGROUND: Ancillary diagnostic methods to enhance the accuracy of viability assessment have not been established for use in clinical practice. OBJECTIVES: To assess intestinal microperfusion measured by Laser Doppler Flowmetry and Spectrophotometry (LDFS) in naturally occurring small intestinal strangulations of different origins and to compare this between viable and non-viable segments. STUDY DESIGN: Prospective clinical trial. METHODS: Forty horses undergoing colic surgery for naturally occurring small intestinal strangulations were included. Tissue oxygen saturation (tSO2), haemoglobin (tHB) and blood flow (tBF) were determined by LDFS before and after release of the strangulation. Intestinal biopsies were taken in cases that underwent intestinal resection or intraoperative euthanasia and assessed using a semi-quantitative mucosal injury score (MIS). The LDFS measurements were compared between the different categories of strangulation causes and histopathological injury using parametric and non-parametric tests (p < 0.05). RESULTS: Strangulations by pedunculated lipomas had lower tBF (13.9 ± 18 arbitrary units [AU]) than epiploic foramen entrapments (65.2 ± 61 AU; CI -1.697 to -0.2498; p = 0.005). Segments with MIS > 5 showed lower tBF during strangulation than segments with MIS < 4 (mean difference 61.1 AU; CI -1.119 to -0.07361; p = 0.03). This did not differ significantly following release of strangulation. Furthermore, there was a positive correlation between the inflammatory cell count and tBF during strangulation (r 0.34; CI 0.01 to 0.60; p = 0.04). The tSO2 and tHB did not differ between the different categories of lesions or injury. MAIN LIMITATIONS: No biopsies could be taken from the intestinal segments that did not undergo resection. The duration of strangulation could not reliably be ascertained. CONCLUSIONS: Blood flow measurements in naturally occurring strangulating lesions show a varying degree of ischaemia in different causes of strangulation. Intestinal blood flow measurements prior to release of the strangulation could potentially contribute to the identification of mucosal injury, yet a high individual variability and other contributing factors need to be considered.

Melan-A immunolabeling in canine extramedullary plasmacytomas.

Histologic diagnosis of less well-differentiated cases of canine extramedullary plasmacytomas (CEMPs) may require immunohistochemical confirmation to discriminate these tumors from other round cells tumors including lymphoma, cutaneous histiocytoma, and amelanotic melanomas. CEMPs are characterized by widespread immunoreactivity for multiple myeloma 1 (MUM1) antigen and λ light chains, while the melanocytic marker melan-A has been reported to yield negative results. Here, 33 randomly selected CEMPs, 20 melanocytomas, and 20 malignant melanomas were immunohistochemically tested for MUM1, melan-A, and PNL2. In addition, CEMPs were examined for PAX5, E-cadherin, CD3, CD18, CD20, S100, as well as λ and κ light chain immunoreactivity. All CEMPs were characterized by labeling for MUM1 and λ light chain, as well as variable immunopositivity for the remaining antibodies. Notably, 13 cases of CEMPs (39.4%) exhibited immunolabeling for melan-A. Melanocytic tumors immunolabeled for melan-A (40/40; 100%) and PNL2 (34/40; 85%). An unexpected cytoplasmic immunoreactivity for MUM1 was observed in 2 melanocytic tumors. Summarized, MUM1 or melan-A immunomarkers alone are not sufficient to differentiate between CEMPs and amelanotic melanomas and should be part of a larger immunopanel including λ light chain, CD20, and PNL2.

Measuring tissue oxygen saturation in the orad intestinal segment during equine colic surgery may aid in predicting the occurrence of postoperative ileus.

OBJECTIVE: To assess the histological injury and intestinal microperfusion measured by laser Doppler flowmetry and spectrophotometry (LDFS) of the small intestine orad to a strangulation during colic surgery. ANIMALS: Horses with naturally occurring small intestinal strangulations undergoing colic surgery were included. METHODS: In this prospective clinical trial, intestinal tissue oxygen saturation (tSO2) and tissue blood flow (tBF) were measured by LDFS orad to the strangulation following release of the strangulation (n = 18). The number of horses with postoperative reflux (POR) and the cases that survived until discharge were compared between groups using Fisher's exact test (P < .05). Intestinal biopsies were taken in cases that underwent intestinal resection or intraoperative euthanasia (n = 28). Measurements were compared between injured and noninjured segments with a Mann-Whitney U or t test. RESULTS: The tSO2 and tBF of the orad intestine were lower than previously reported in healthy horses. Horses with low tSO2 of < 35% were significantly more likely to suffer from POR (6/6 cases) compared to cases with tSO2 > 69% (1/6). The number of horses that survived were not statistically different between these groups (2/6 and 6/6). All horses with mucosal injury developed POR (6/6), which was significantly more likely compared to horses without mucosal injury (3/13). No significant difference in tSO2 or tBF could be found between the segments with and without histological injury. CLINICAL RELEVANCE: The results suggest that measuring tSO2 in the orad segment during colic surgery may aid in predicting postoperative issues.

Concurrent Detection of a Papillomatous Lesion and Sequence Reads Corresponding to a Member of the Family Adintoviridae in a Bell's Hinge-Back Tortoise (Kinixys belliana).

An adult male Bell's hinge-back tortoise (Kinixys belliana) was admitted to a veterinary clinic due to a swelling in the oral cavity. Physical examination revealed an approximately 2.5 × 1.5 cm sized, irregularly shaped tissue mass with villiform projections extending from its surface located in the oropharyngeal cavity. An initial biopsy was performed, and the lesion was diagnosed as squamous papilloma. Swabs taken for virological examination tested negative with specific PCRs for papillomavirus and herpesvirus. Further analysis of the oropharyngeal mass via metagenomic sequencing revealed sequence reads corresponding to a member of the family Adintoviridae. The tissue mass was removed one week after the initial examination. The oral cavity remained unsuspicious in follow-up examinations performed after one, five and twenty weeks. However, a regrowth of the tissue was determined 23 months after the initial presentation. The resampled biopsy tested negative for sequence reads of Adintoviridae. Conclusively, this report presents the diagnostic testing and therapy of an oral cavity lesion of unknown origin. The significance of concurrent metagenomic determination of adintovirus sequence reads within the tissue lesion is discussed.

Nanobodies to multiple spike variants and inhalation of nanobody-containing aerosols neutralize SARS-CoV-2 in cell culture and hamsters.

The ongoing threat of COVID-19 has highlighted the need for effective prophylaxis and convenient therapies, especially for outpatient settings. We have previously developed highly potent single-domain (VHH) antibodies, also known as nanobodies, that target the Receptor Binding Domain (RBD) of the SARS-CoV-2 Spike protein and neutralize the Wuhan strain of the virus. In this study, we present a new generation of anti-RBD nanobodies with superior properties. The primary representative of this group, Re32D03, neutralizes Alpha to Delta as well as Omicron BA.2.75; other members neutralize, in addition, Omicron BA.1, BA.2, BA.4/5, and XBB.1. Crystal structures of RBD-nanobody complexes reveal how ACE2-binding is blocked and also explain the nanobodies' tolerance to immune escape mutations. Through the cryo-EM structure of the Ma16B06-BA.1 Spike complex, we demonstrated how a single nanobody molecule can neutralize a trimeric spike. We also describe a method for large-scale production of these nanobodies in Pichia pastoris, and for formulating them into aerosols. Exposing hamsters to these aerosols, before or even 24 h after infection with SARS-CoV-2, significantly reduced virus load, weight loss and pathogenicity. These results show the potential of aerosolized nanobodies for prophylaxis and therapy of coronavirus infections.

Histological and immunohistochemical characterization of mucosa-associated lymphoid tissue and antigen-presenting cells in trachea and lung of cattle.

The presence of bronchus-associated lymphoid tissue (BALT) and its structural components has been described in different healthy animal species and in animals with diseases of the respiratory tract. In contrast to normal mammals, BALT is absent in healthy human adult lungs, but has been found in the lungs of children. The histological characteristics of organized mucosa-associated lymphoid tissue (MALT), its subsets of immune cells and their in situ distribution in the lung of healthy subadult and adult cattle shows close similarities with BALT in humans and other animal species such as sheep, horses and pigs. This study clearly demonstrates that organized MALT also occurs in the tracheal mucosa of cattle. The absence of tracheal MALT and BALT in calves suggest that these structures are not constitutive. In the mucosa of bovine trachea, bronchi and bronchioli, MHC II+ and CD11c+ dendritic cells (DCs) are located in the epithelium and in the lamina propria mucosae. These DCs are already present in calves soon after birth. Examination of tangential epithelial sheets shows that in the bovine tracheal epithelium, like in man and rat, a dense network of MHC II+ and CD11c+ DCs exists and that their number is considerably higher than in conventional transverse sections. In the bovine tracheal and bronchial epithelium, MHC II+ DCs are extending their dendrites towards the lumen indicating that these DCs possibly are involved in sampling of luminal antigens. The presence of significantly higher numbers of MHC II+ DCs in the tracheal and bronchial/bronchiolar mucosa of older cattle in than in calves possibly results from local stimulation with exogenous antigens during postnatal life. Detection of DCs expressing the costimulatory molecules CD80 and CD86 in calves and cattle suggests maturation of DCs, which is most likely induced by stimulation with exogenous antigens.

T cell immunity ameliorates COVID-19 disease severity and provides post-exposure prophylaxis after peptide-vaccination, in Syrian hamsters.

Background: The emergence of novel SARS-CoV-2 variants that resist neutralizing antibodies drew the attention to cellular immunity and calls for the development of alternative vaccination strategies to combat the pandemic. Here, we have assessed the kinetics of T cell responses and protective efficacy against severe COVID-19 in pre- and post-exposure settings, elicited by PolyPEPI-SCoV-2, a peptide based T cell vaccine. Methods: 75 Syrian hamsters were immunized subcutaneously with PolyPEPI-SCoV-2 on D0 and D14. On D42, hamsters were intranasally challenged with 102 TCID50 of the virus. To analyze immunogenicity by IFN-γ ELISPOT and antibody secretion, lymphoid tissues were collected both before (D0, D14, D28, D42) and after challenge (D44, D46, D49). To measure vaccine efficacy, lung tissue, throat swabs and nasal turbinate samples were assessed for viral load and histopathological changes. Further, body weight was monitored on D0, D28, D42 and every day after challenge. Results: The vaccine induced robust activation of T cells against all SARS-CoV-2 structural proteins that were rapidly boosted after virus challenge compared to control animals (~4-fold, p<0.05). A single dose of PolyPEPI-SCoV-2 administered one day after challenge also resulted in elevated T cell response (p<0.01). The vaccination did not induce virus-specific antibodies and viral load reduction. Still, peptide vaccination significantly reduced body weight loss (p<0.001), relative lung weight (p<0.05) and lung lesions (p<0.05), in both settings. Conclusion: Our study provides first proof of concept data on the contribution of T cell immunity on disease course and provide rationale for the use of T cell-based peptide vaccines against both novel SARS-CoV-2 variants and supports post-exposure prophylaxis as alternative vaccination strategy against COVID-19.

Sox9, Hopx, and survivin and tuft cell marker DCLK1 expression in normal canine intestine and in intestinal adenoma and adenocarcinoma.

Self-renewal of the intestinal epithelium originates from stem cells located at the crypt base. Upregulation of various stem cell markers in intestinal epithelial neoplasms indicates a potential role of stem cells in tumorigenesis. In this study, the immunoreactivity of potential intestinal stem cell markers (Sry box transcription factor 9 [Sox9], homeodomain-only protein [Hopx], survivin) and tuft cell marker doublecortin-like kinase 1 (DCLK1) in normal canine intestine and intestinal epithelial neoplasms was investigated. Formalin-fixed paraffin-embedded (FFPE) small and large intestine as well as intestinal neoplasms (55 colorectal adenomas [CRAs], 17 small intestinal adenocarcinomas [SICs], and 12 colorectal adenocarcinomas [CRCs]) were analyzed immunohistologically. Potential stem cell markers Sox9, Hopx, and survivin were detected in the crypts of normal canine small and large intestine. DCLK1+ tuft cells were present in decreasing numbers along the crypt-villus axis of the jejunum and rarely detectable in large intestine. In canine intestinal epithelial tumors, nuclear Sox9 immunoreactivity was detectable in 84.9% (CRA), 80% (CRC), and 77% of epithelial neoplastic cells (SIC). Hopx and survivin were expressed within cytoplasm and nuclei of neoplastic cells in benign and malignant tumors. DCLK1 showed a cytoplasmic reaction within neoplastic cells. The combined score of Hopx, DCLK1, and survivin varied among the examined cases. Overall, malignant tumors showed lower DCLK1 scores but higher Hopx scores in comparison with benign tumors. For survivin, no differences were detectable. In conclusion, stem cell markers Sox9, Hopx, and survivin were detectable at the crypt base and the immunoreactivity of Sox9, DCLK1, survivin, and Hopx was increased in canine intestinal adenomas and adenocarcinomas compared with normal mucosa.

Infections with highly pathogenic avian influenza A virus (HPAIV) H5N8 in harbor seals at the German North Sea coast, 2021.

In brain tissue of three harbor seals of the German North Sea coast, high virus loads of highly pathogenic avian influenza virus (HPAIV) H5N8 were detected. Identification of different virus variants indicates high exposure to HPAIV circulating in wild birds, but there is no evidence for H5 specific antibodies in healthy seals. Replication of avian viruses in seals may allow HPAIV to acquire mutations needed to adapt to mammalian hosts as shown by PB2 627K variants detected in these cases.

Hanoverian F/W-line contributes to segregation of Warmblood fragile foal syndrome type 1 variant PLOD1:c.2032G>A in Warmblood horses.

BACKGROUND: Warmblood fragile foal syndrome (WFFS) is a lethal condition detected in Warmblood horses. Its origin and association with performance traits and fertility among horse populations is unknown. OBJECTIVES: To validate the previously identified WFFS type 1 (WFFST1)-associated missense variant PLOD1:c.2032G>A and to investigate its distribution among various horses with particular focus on Hanoverian breed, as well as its pathomorphological picture. The study aimed at identifying the origin of the mutant allele and its correlation with performance and fertility traits in Warmblood horses. STUDY DESIGN: Retrospective case-control and association study. METHODS: WFFST1 variant was validated using whole genome sequencing (WGS) in 78 equids. In an affected foal with a homozygous mutant genotype, necropsy was performed. Skin samples were examined using histology and transmission electron microscopy. Pathway analysis was performed to trace back 81 genetic carriers to the most common recent ancestor. Furthermore, generalised linear model analysis was employed to test estimated breeding values (EBVs) for differences in performance and fertility traits among different genotypes in Hanoverian horses. RESULTS: WFFST1 variant had the lowest minor allele frequency among all variants detected in WGS data in the region of PLOD1. Further genotyping of this variant revealed allele frequencies of 0.14 in Hanoverian horses. Histological investigations of the WFFST1-affected foal showed loosely arranged collagen fibres in the dermis. Ultrastructurally, multifocal areas with degraded collagen fibrils and fibrillar plaques were detected. Further pathway analysis revealed a stallion from the Hanoverian sire F/W line as the most common recent ancestor of all tested genetic carriers. Furthermore, WFFST1 variant was found to be correlated with EBVs for gait-related traits as well as conformation and dressage. MAIN LIMITATIONS: Study evaluated carriers and cases only from Europe. CONCLUSIONS: This study provides a comprehensive evaluation of WFFST1 variant and traces it back to its potential origin.

Laryngeal chondritis as a differential for upper airway diseases in German sheep.

BACKGROUND: Ovine laryngeal chondritis is a rare entity of sheep in the USA, Great Britain, New Zealand and Iceland, but has not been reported in Germany so far. Here, two German cases are reported. CASE PRESENTATION: Two rams showed severe and progressive signs of dyspnea. Endoscopically, a severe bilateral swelling of the larynx was identified in both rams. Due to poor prognosis and progression of clinical signs one ram was euthanized, while the other ram died overnight. In both cases, a necrosuppurative laryngitis and chondritis of arytenoid cartilages was found at necropsy. Fusobacterium necrophorum and Streptococcus ovis were isolated from the laryngeal lesion in one animal. CONCLUSIONS: This is the first report of ovine laryngeal chondritis in continental Europe. This entity should be considered a differential diagnosis for upper airway disease in sheep.

Characterization of abortion, stillbirth and non-viable foals homozygous for the Warmblood Fragile Foal Syndrome.

Warmblood fragile foal syndrome (WFFS) is a monogenetic defect with autosomal recessive inheritance. The WFFS homozygosity is non-compatible with extra-uterine life. Although as many as 15% of Warmblood horses are WFFS carriers, there has been little veterinary focus on this condition. The aim of this study was to determine outcomes and symptoms of clinical signs and pathological abnormalities during pregnancies when there were WFFS homozygous foetuses. Diagnostic material of 15 abortion or stillbirth cases with suspected diagnosis of WFFS was available for this study. Additionally, there were examinations in 37 cases where there were no indications of WFFS when submitted for routine diagnostic procedures. Foals in all cases were genotyped and external morphological defects were recorded. Amongst the 15 cases in which WFSS was suspected, there were 14 homozygous foetuses with the WFFS allele (WFFS/WFFS). Three heterozygous WFFS foetuses (N/WFFS) were detected in the cases submitted for routine diagnostic procedures. Of the 14 WFFS homozygous foetuses, 11 of mares had a gestation length of at least 320 days. Nine foals were born alive but died within a short time. Skin defects were obvious in 12 WFFS homozygous foals, and there was abnormal flexibility in the digital joints, flexed forelegs and incomplete closure of the abdominal wall in five, four, and one of the foals, respectively. In conclusion, the predominant manifestation of WFFS are death during the latter stages of gestation or live births with foals being non-viable. Losses in Warmblood horse breeding caused by WFFS are greater than previously assumed.

Canine primary jejunal and colonic epithelial cells predominantly express TLR5 and TLR9 but do not change TLR expression pattern after stimulation with certain Toll-like receptor ligands.

The intestinal mucosa is in contact with abundant luminal antigens and coordinates immune responses to differentiate commensals from pathogens. Intestinal epithelial cells (IECs) not only represent a physical barrier but also an immunologically important cell type that recognizes microbe-associated molecular patterns via Toll-like receptors (TLR). The importance of TLR expression has been elucidated for intestinal disorders in humans, mice and dogs. However, as knowledge about canine intestinal TLRs is mainly limited to the transcriptional level, the present study analyzed the protein expression of TLR2, TLR3, TLR4, TLR5 and TLR9 by primary canine IECs in the steady state and after stimulation with TLR ligands. This exhibited TLR5 and TLR9 to be predominantly expressed in canine IECs. TLR stimulation did not result in changes of the TLR expression pattern. Further studies are needed to elucidate whether this implicates hyporesponsiveness of canine IECs towards TLR stimulation under steady state conditions.

[Ocular dermoids and microphthalmia in a juvenile sheep]. / Okuläre Dermoide und Mikrophthalmie bei einem Jungschaf.

A 6-month-old Leine sheep was presented because of dermal tissue located on the left eye. During the first examination, the animal was clinically silent, apart from the deformed eye. A corneal and conjunctival dermoid and blindness of the left eye were diagnosed. Over a period of a year, the animal displayed conjunctivitis and inflammation of the affected eye. Furthermore, the sheep did not develop according to its age. During histopathological examination of the euthanized animal, microphthalmia and aphakia of the left eye were found in addition to the dermoids. Dermoids are described in humans and in different domestic animals. They can be combined with other congenital malformations. In sheep, dermoids are rarely diagnosed or reported in the literature.