Equine gamma herpesvirus presence and viral load are not associated with equine glandular gastric disease.

OBJECTIVE: To investigate the role of equine herpesvirus-2 (EHV-2) and equine herpesvirus-5 (EHV-5) in equine glandular gastric disease (EGGD) by visualizing and quantifying these gamma herpesviruses in EGGD-affected and normal glandular gastric mucosa of horses. A secondary objective was to describe the histopathological abnormalities in the equine gastric glandular mucosa in horses with EGGD. ANIMALS: 29 horses (n = 21 postmortem and 8 gastroscopy) categorized as normal (11), EGGD (12), or both EGGD and equine squamous gastric disease (6). METHODS: Glandular gastric mucosal samples were collected from horses by gastroscopy or postmortem. Histopathology and in situ hybridization targeting EHV-2 and EHV-5 were performed on grossly normal and abnormal glandular gastric mucosa. The number of in situ hybridization-positive cells per millimeter squared of tissue was calculated. Evaluators were blinded to groups. RESULTS: Glandular gastric tissues from horses without EGGD had higher viral loads in the mucosa than normal or abnormal tissues from EGGD horses. There was no difference in viral loads for EHV-2 or EHV-5 between grossly or endoscopically normal to abnormal gastric tissues within horses with EGGD. Lymphocytic plasmacytic gastritis was the most common histopathological abnormality, with only 3 horses having mucosal disruption (glandular ulcer or erosion). CLINICAL RELEVANCE: Equine gamma herpesviruses are unlikely to play a role in the pathophysiology of EGGD. EGGD is frequently inflammatory with occasional mucosal disruption (ulcer or erosion).

Proteomic profiling of membrane vesicles from Mycobacterium avium subsp. paratuberculosis: Navigating towards an insilico design of a multi-epitope vaccine targeting membrane vesicle proteins.

Bacteria typically produce membrane vesicles (MVs) at varying levels depending on the surrounding environments. Gram-negative bacterial outer membrane vesicles (OMVs) have been extensively studied for over 30 years, but MVs from Gram-positive bacteria only recently have been a focus of research. In the present study, we isolated MVs from Mycobacterium avium subsp. paratuberculosis (MAP) and analyzed their protein composition using LC-MS/MS. A total of 316 overlapping proteins from two independent preparations were identified in our study, and topology prediction showed these cargo proteins have different subcellular localization patterns. When MVs were administered to bovine-derived macrophages, significant up-regulation of pro-inflammatory cytokines was observed via qRT-PCR. Proteome functional annotation revealed that many of these proteins are involved in the cellular protein metabolic process, tRNA aminoacylation, and ATP synthesis. Secretory proteins with high antigenicity and adhesion capability were mapped for B-cell and T-cell epitopes. Antigenic, Immunogenic and IFN-γ inducing B-cell, MHC-I, and MHC-II epitopes were stitched together through linkers to form multi-epitope vaccine (MEV) construct against MAP. Strong binding energy was observed during the docking of the 3D structure of the MEV with the bovine TLR2, suggesting that the putative MEV may be a promising vaccine candidate against MAP. However, in vitro and in vivo analysis is required to prove the immunogenic concept of the MEV which we will follow in our future studies. SIGNIFICANCE: Johne's disease is a chronic infection caused by Mycobacterium avium subsp. paratuberculosis that has a potential link to Crohn's disease in humans. The disease is characterized by persistent diarrhea and enteritis, resulting in significant economic losses due to reduced milk yield and premature culling of infected animals. The dairy industry in the United States alone experiences losses of approximately USD 250 million due to Johne's disease. The current vaccine against Johne's disease is limited by several factors, including variable efficacy, limited duration of protection, interference with diagnostic tests, inability to prevent infection, and logistical and cost-related challenges. Nevertheless, a multiepitope vaccine design approach targeting M. avium subsp. paratuberculosis has the potential to overcome these challenges and offer improved protection against Johne's disease.

Experimental Induction of Acute Acanthamoeba castellanii Keratitis in Cats.

Purpose: To develop a feline model of acute Acanthamoeba keratitis using methods that replicate natural routes of infection transmission. Methods: Corneal Acanthamoeba castellanii inoculation was performed by three methods: topical inoculation with Acanthamoeba solution following corneal abrasion, placement of a contaminated contact lens for 7 days, and placement of a contaminated contact lens for 7 days following corneal abrasion. Sham inoculations with parasite-free medium and sterile contact lenses were also performed. Cats were monitored by ocular examination and in vivo corneal confocal microscopy for 21 days post-inoculation. Corneal samples were collected at intervals for microbiologic assessment, histopathology, and immunohistochemistry. Results: All cats in the corneal abrasion groups developed clinical keratitis. Clinical ocular disease was inconsistently detected in cats from the contaminated contact lens only group. Initial corneal lesions were characterized by multifocal epithelial leukocyte infiltrates. Ocular lesions progressed to corneal epithelial ulceration and diffuse stromal inflammation. After 14 days, corneal ulcerations resolved, and stromal inflammation consolidated into multifocal subepithelial and stromal infiltrates. Corneal amoebae were detected by culture, in vivo confocal microscopy, histopathology, and immunohistochemistry in cats with keratitis. Neutrophilic and lymphocytic keratoconjunctivitis with lymphoplasmacytic anterior uveitis were identified by histopathology. Coinfection with aerobic bacteria was detected in some, but not all, cats with keratitis. Ocular disease was not detected in the sham inoculation groups. Conclusions: Feline Acanthamoeba keratitis is experimentally transmissible by contaminated contact lenses and topical inoculation following corneal epithelial trauma. Translational Relevance: Experimentally induced acute Acanthamoeba keratitis in cats is clinically and histopathologically similar to its human counterpart.

Metacarpophalangeal Joint Pathology and Bone Mineral Density Increase with Exercise but Not with Incidence of Proximal Sesamoid Bone Fracture in Thoroughbred Racehorses.

Proximal sesamoid bone (PSB) fracture is the leading cause of fatal musculoskeletal injury in Thoroughbred racehorses in Hong Kong and the US. Efforts are underway to investigate diagnostic modalities that could help identify racehorses at increased risk of fracture; however, features associated with PSB fracture risk are still poorly understood. The objectives of this study were to (1) investigate third metacarpal (MC3) and PSB density and mineral content using dual-energy X-ray absorptiometry (DXA), computed tomography (CT), Raman spectroscopy, and ash fraction measurements, and (2) investigate PSB quality and metacarpophalangeal joint (MCPJ) pathology using Raman spectroscopy and CT. Forelimbs were collected from 29 Thoroughbred racehorse cadavers (n = 14 PSB fracture, n = 15 control) for DXA and CT imaging, and PSBs were sectioned for Raman spectroscopy and ash fraction measurements. Bone mineral density (BMD) was greater in MC3 condyles and PSBs of horses with more high-speed furlongs. MCPJ pathology, including palmar osteochondral disease (POD), MC3 condylar sclerosis, and MC3 subchondral lysis were greater in horses with more high-speed furlongs. There were no differences in BMD or Raman parameters between fracture and control groups; however, Raman spectroscopy and ash fraction measurements revealed regional differences in PSB BMD and tissue composition. Many parameters, including MC3 and PSB bone mineral density, were strongly correlated with total high-speed furlongs.

Radiomics Modeling of Catastrophic Proximal Sesamoid Bone Fractures in Thoroughbred Racehorses Using µCT.

Proximal sesamoid bone (PSB) fractures are the most common musculoskeletal injury in race-horses. X-ray CT imaging can detect expressed radiological features in horses that experienced catastrophic fractures. Our objective was to assess whether expressed radiomic features in the PSBs of 50 horses can be used to develop machine learning models for predicting PSB fractures. The µCTs of intact contralateral PSBs from 50 horses, 30 of which suffered catastrophic fractures, and 20 controls were studied. From the 129 intact µCT images of PSBs, 102 radiomic features were computed using a variety of voxel resampling dimensions. Decision Trees and Wrapper methods were used to identify the 20 top expressed features, and six machine learning algorithms were developed to model the risk of fracture. The accuracy of all machine learning models ranged from 0.643 to 0.903 with an average of 0.754. On average, Support Vector Machine, Random Forest (RUS Boost), and Log-regression models had higher performance than K-means Nearest Neighbor, Neural Network, and Random Forest (Bagged Trees) models. Model accuracy peaked at 0.5 mm and decreased substantially when the resampling resolution was greater than or equal to 1 mm. We find that, for this in vitro dataset, it is possible to differentiate between unfractured PSBs from case and control horses using µCT images. It may be possible to extend these findings to the assessment of fracture risk in standing horses.

Plakoglobin and High-Mobility Group Box 1 Mediate Intestinal Epithelial Cell Apoptosis Induced by Clostridioides difficile TcdB.

Clostridioides difficile infection (CDI) is the leading cause of antibiotic-associated intestinal disease, resulting in severe diarrhea and fatal pseudomembranous colitis. TcdB, one of the essential virulence factors secreted by this bacterium, induces host cell apoptosis through a poorly understood mechanism. Here, we performed an RNA interference (RNAi) screen customized to Caco-2 cells, a cell line model of the intestinal epithelium, to discover host factors involved in TcdB-induced apoptosis. We identified plakoglobin, also known as junction plakoglobin (JUP) or γ-catenin, a member of the catenin family, as a novel host factor and a previously known cell death-related chromatin factor, high-mobility group box 1 (HMGB1). Disruption of those host factors by RNAi and CRISPR resulted in resistance of cells to TcdB-mediated and mitochondrion-dependent apoptosis. JUP was redistributed from adherens junctions to the mitochondria and colocalized with the antiapoptotic factor Bcl-XL. JUP proteins could permeabilize the mitochondrial membrane, resulting in the release of cytochrome c. Our results reveal a novel role of JUP in targeting the mitochondria to promote the mitochondrial apoptotic pathway. Treatment with glycyrrhizin, an HMGB1 inhibitor, resulted in significantly increased resistance to TcdB-induced epithelial damage in cultured cells and a mouse ligated colon loop model. These findings demonstrate the critical roles of JUP and HMGB1 in TcdB-induced epithelial cell apoptosis. IMPORTANCE Clostridioides difficile infection (CDI) is the leading cause of hospital-acquired diarrhea. Toxins, especially TcdB, cause epithelial cell apoptosis, but the underlying cell death mechanism is less clear. Through an apoptosis-focused RNAi screen using a bacterium-made small interfering (siRNA) library customized to a human colonic epithelial cell model, we found a novel host factor, plakoglobin (γ-catenin), as a key factor required for cell apoptosis induced by TcdB. Plakoglobin targets and permeabilizes mitochondria after stimulation by TcdB, demonstrating a hitherto underappreciated role of this catenin family member in the apoptosis of intestinal epithelial cells. We also found a previously known cell death-related chromatin factor, HMGB1, and explored the inhibition of HMGB1 for CDI therapy in vivo.

Mediastinal Serous Cavity Mesothelioma in Two Dogs.

The mediastinal serous cavity (MSC)-well documented but seldom recognized in the caudal mediastinum-is embryologically derived from the omental bursa. Mesothelioma arising from the MSC in two dogs is described. Both dogs presented with acute life-threatening hemorrhagic pleural effusion. Contrast computed tomography revealed a large solitary spherical-to-cylindrical tumor in the caudal mediastinum with variably thick, contrast-enhancing walls with lobular to frond-like proliferations that arose circumferentially and projected internally into a single, variably sized, fluid-attenuating lumen. The wall and lumen corresponded to the serous membrane and serous cavity of the MSC. Surgical exploration confirmed that both tumors arose from the mediastinum. Both had similar histologic findings, and special stains were necessary for definitive diagnosis. The tumor was nonresectable in one dog, and it was euthanized intraoperatively. The other survived 7 mo. An MSC mesothelioma should be considered a possible cause of hemothorax in dogs that may be detected on thoracic radiography and computed tomography. Differential diagnoses include esophageal foreign body or neoplasm, paraesophageal diaphragmatic hernia, MCS empyema, and pulmonary adenocarcinoma, with thoracic computed tomography helping to rule out foreign body and diaphragmatic hernia. For confirmed neoplasms, histochemistry and immunohistochemistry should be performed to differentiate between mesothelioma and pulmonary adenocarcinoma.

Proximal sesamoid bone microdamage is localized to articular subchondral regions in Thoroughbred racehorses, with similar fracture toughness between fracture and controls.

OBJECTIVE: To determine whether proximal sesamoid bone (PSB) microdamage and fracture toughness differ between Thoroughbred racehorses sustaining PSB fracture and controls. STUDY DESIGN: Cadaveric case-control. ANIMALS: Twenty-four Thoroughbred racehorses (n = 12 PSB fracture, n = 12 control). METHODS: Proximal sesamoid bones were dissected, and gross pathological changes and morphological measurements were documented. High-speed exercise history data were evaluated. Microdamage was assessed in fracture, fracture-contralateral limb (FXCL) and control PSBs using whole bone lead uranyl acetate (LUA) staining with micro-CT imaging or basic fuchsin histological analysis. Fracture toughness mechanical testing was carried out in 3-point-bending of microbeams created from PSB flexor cortices. Data were analyzed using ordinal logistic and linear regression models. RESULTS: Microdamage was detected most commonly in the articular subchondral region of PSBs via LUA micro-CT and basic fuchsin histology. There were no differences in microdamage between FXCL and control PSBs. Fracture toughness values were similar for FXCL (1.31 MPa√m) and control (1.35 MPa√m) PSBs. Exercise histories were similar except that horses sustaining fracture spent a greater percentage of their careers in rest weeks. CONCLUSION: Microdamage was detected in the articular region of PSBs but was not greater in horses sustaining catastrophic PSB fracture. Fracture toughness of PSB flexor cortices did not differ between FXCL and control PSBs. CLINICAL SIGNIFICANCE: Although uncommon, microdamage is localized to the articular region of Thoroughbred racehorse PSBs. Catastrophic PSB failure is not associated with lower PSB flexor cortex fracture toughness.

Bacterial culture and immunohistochemical detection of bacteria and endotoxin in cats with suppurative cholangitis-cholangiohepatitis syndrome.

OBJECTIVE: To characterize the frequency and type of bacterial infection by culture- and immunohistochemical (IHC)-based methods and determine the impact of infection on clinical features and survival time in cats with suppurative cholangitis-cholangiohepatitis syndrome (S-CCHS). ANIMALS: 168 client-owned cats with S-CCHS (cases). PROCEDURES: Clinical features, bacterial culture results, culture-inoculate sources, and survival details were recorded. Cases were subcategorized by comorbidity (extrahepatic bile duct obstruction, cholelithiasis, cholecystitis, ductal plate malformation, biopsy-confirmed inflammatory bowel disease, and biopsy-confirmed pancreatitis) or treatment by cholecystectomy or cholecystoenterostomy. Culture results, bacterial isolates, Gram-stain characteristics, and IHC staining were compared among comorbidities. Lipoteichoic acid IHC staining detected gram-positive bacterial cell wall components, and toll-like receptor expression IHC reflected pathologic endotoxin (gram-negative bacteria) exposure. RESULTS: Clinical features were similar among cases except for more frequent abdominal pain and lethargy in cats with positive culture results and pyrexia, abdominal pain, and hepatomegaly for cats with polymicrobial infections. Bacteria were cultured in 93 of 135 (69%) cats, with common isolates including Enterococcus spp and Escherichia coli. IHC staining was positive in 142 of 151 (94%) cats (lipoteichoic acid, 107/142 [75%]; toll-like receptor 4, 99/142 [70%]). With in-parallel interpretation of culture and IHC-based bacterial detection, 154 of 166 (93%) cats had bacterial infections (gram-positive, 118/154 [77%]; gram-negative, 111/154 [72%]; polymicrobial, 79/154 [51%]). Greater frequency of bacterial isolation occurred with combined tissue, bile, and crushed cholelith inoculates. Infection and gram-positive bacterial isolates were associated with significantly shorter long-term survival times. CLINICAL RELEVANCE: S-CCHS was associated with bacterial infection, pathologic endotoxin exposure, and frequent polymicrobial infection in cats. Combined tissue inoculates improved culture detection of associated bacteria.

Leptospira borgpetersenii Leucine-Rich Repeat Proteins Provide Strong Protective Efficacy as Novel Leptospiral Vaccine Candidates.

Leucine-rich repeat (LRR) proteins are advocated for being assessed in vaccine development. Leptospiral LRR proteins were identified recently in silico from the genome of Leptospira borgpetersenii serogroup Sejroe, the seroprevalence of leptospiral infections of cattle in Thailand. Two LRR recombinant proteins, rKU_Sej_LRR_2012M (2012) and rhKU_Sej_LRR_2271 (2271), containing predicted immunogenic epitopes, were investigated for their cross-protective efficacies in an acute leptospirosis model with heterologous Leptospira serovar Pomona, though, strains from serogroup Sejroe are host-adapted to bovine, leading to chronic disease. Since serovar Pomona is frequently reported as seropositive in cattle, buffaloes, pigs, and dogs in Thailand and causes acute and severe leptospirosis in cattle by incidental infection, the serogroup Sejroe LRR proteins were evaluated for their cross-protective immunity. The protective efficacies were 37.5%, 50.0%, and 75.0% based on the survival rate for the control, 2012, and 2271 groups, respectively. Sera from 2012-immunized hamsters showed weak bactericidal action compared to sera from 2271-immunized hamsters (p < 0.05). Therefore, bacterial tissue clearances, inflammatory responses, and humoral and cell-mediated immune (HMI and CMI) responses were evaluated only in 2271-immunized hamsters challenged with virulent L. interrogans serovar Pomona. The 2271 protein induced prompt humoral immune responses (p < 0.05) and leptospiral tissue clearance, reducing tissue inflammation in immunized hamsters. In addition, protein 2271 and its immunogenic peptides stimulated splenocyte lymphoproliferation and stimulated both HMI and CMI responses by activating Th1 and Th2 cytokine gene expression in vaccinated hamsters. Our data suggest that the immunogenic potential renders rhKU_Sej_LRR_2271 protein a promising candidate for the development of a novel cross-protective vaccine against animal leptospirosis.

Determination of correlation of proximal sesamoid bone osteoarthritis with high-speed furlong exercise and catastrophic sesamoid bone fracture in Thoroughbred racehorses.

OBJECTIVE: To examine whether proximal sesamoid bone (PSB) articular cartilage and bone osteoarthritic changes or palmar osteochondral disease (POD) scores were associated with exercise history and catastrophic PSB fracture in Thoroughbred racehorses. SAMPLES: PSBs from 16 Thoroughbred racehorses (8 with and 8 without PSB fracture). PROCEDURES: Exercise history was collected, and total career high-speed furlongs was used as the measure of total exercise per horse. At necropsy, medial and lateral condyles of the third metacarpus from each forelimb were assigned a POD score, followed by imaging with micro-CT for evaluation of osteophyte size. Three investigators that were blinded to the type of PSB (fracture or no fracture) used the Osteoarthritis Research Society International (OARSI) scoring system to evaluate acellularity, chondrocyte necrosis, cartilage fibrillation, chondrone formation, safranin O stain uptake, and tidemark advancement of 1 central sagittal tissue section/PSB (4 PSBs/horse). Cartilage thickness and bone necrosis were scored on the basis of histologic examination. RESULTS: POD score, osteophyte size score, percentage of bone necrosis, tidemark advancement, chondrone formation, and total OARSI score were greater in horses with more accrued total career high-speed furlongs. Scores for POD, osteophyte size, fibrillation, acellularity, chondrone formation, and total OARSI were greater for horses with PSB fracture. CONCLUSIONS AND CLINICAL REVELANCE: OARSI scoring revealed that more advanced osteoarthritic changes strongly correlated with total career high-speed furlongs and PSB fracture. However, the effect of exercise was dominant, suggesting that exercise history will be important to include in future models that aim to assess risk factors for catastrophic PSB fracture.

Pathogenesis, MicroRNA-122 Gene-Regulation, and Protective Immune Responses After Acute Equine Hepacivirus Infection.

BACKGROUND AND AIMS: Equine hepacivirus (EqHV) is phylogenetically the closest relative of HCV and shares genome organization, hepatotropism, transient or persistent infection outcome, and the ability to cause hepatitis. Thus, EqHV studies are important to understand equine liver disease and further as an outbred surrogate animal model for HCV pathogenesis and protective immune responses. Here, we aimed to characterize the course of EqHV infection and associated protective immune responses. APPROACH AND RESULTS: Seven horses were experimentally inoculated with EqHV, monitored for 6 months, and rechallenged with the same and, subsequently, a heterologous EqHV. Clearance was the primary outcome (6 of 7) and was associated with subclinical hepatitis characterized by lymphocytic infiltrate and individual hepatocyte necrosis. Seroconversion was delayed and antibody titers waned slowly. Clearance of primary infection conferred nonsterilizing immunity, resulting in shortened duration of viremia after rechallenge. Peripheral blood mononuclear cell responses in horses were minimal, although EqHV-specific T cells were identified. Additionally, an interferon-stimulated gene signature was detected in the liver during EqHV infection, similar to acute HCV in humans. EqHV, as HCV, is stimulated by direct binding of the liver-specific microRNA (miR), miR-122. Interestingly, we found that EqHV infection sequesters enough miR-122 to functionally affect gene regulation in the liver. This RNA-based mechanism thus could have consequences for pathology. CONCLUSIONS: EqHV infection in horses typically has an acute resolving course, and the protective immune response lasts for at least a year and broadly attenuates subsequent infections. This could have important implications to achieve the primary goal of an HCV vaccine; to prevent chronicity while accepting acute resolving infection after virus exposure.

Equine leptospirosis: Experimental challenge of Leptospira interrogans serovar Bratislava fails to establish infection in naïve horses.

BACKGROUND: Little information is available about experimental inoculation of leptospirosis in horses and the pathogenicity of Leptospira interrogans serovar Bratislava in this host. OBJECTIVES: To determine the serological, clinical, pathological and haematological responses of horses to L. interrogans serovar Bratislava strain PigK151. STUDY DESIGN: Randomised controlled in vivo experiment. METHODS: Ten seronegative female foals were divided into 2 groups, control (n = 4) and challenged (n = 6). The challenged group received 1 × 109 leptospires divided equally between topical ocular and intraperitoneal injections. Blood and urine samples were analysed. The temperature was recorded daily for the first 9 days, then weekly. Sera were tested by microscopic agglutination test (MAT). Automated complete blood count, differential and chemistry panel were performed. Histopathological analysis was performed on sections of liver, kidney, urinary bladder, uterine body and pineal gland. Sample culturing was performed from blood, urine, liver, kidney, reproductive tract and vitreous humour. RESULTS: No pyrexia was noted. PCR and culture were negative from all samples. Differences between groups were found in CBC, differential counts and serum biochemistry panel (or profile), suggesting that leptospiral challenge triggered an inflammatory response. No evidence of leptospirosis was found from histopathological analysis. All challenged foals developed a humoral response. The MAT allowed the confirmation of the infecting serovar at a later stage, but it also revealed cross-reactive results that were further explained by genomic analysis. MAIN LIMITATIONS: This experimental challenge had two main limitations: (a) the results might have varied if another strain from the same serovar had been used and (b) the use of another route of infection and a higher bacterial dose might have achieved colonisation. CONCLUSIONS: Based on these findings, it may suggest that L. interrogans serovar Bratislava is neither pathogenic nor host-adapted serovar for horses, although these results might have varied if another strain from the same serovar had been used instead.

Evaluation of new leptospiral antigens for the diagnosis of equine leptospirosis: An approach using pan-genomic analysis, reverse vaccinology and antigenic selection.

BACKGROUND: The current gold standard diagnostic test for leptospirosis is the microscopic agglutination test (MAT), which has many drawbacks; therefore, the development of a better and easier serological test for leptospirosis is needed. OBJECTIVES: To apply reverse vaccinology (RV) and antigenic selection on the assortment of leptospiral targets and evaluate their potential for use as reagents for the diagnosis of equine leptospirosis. STUDY DESIGN: Cross-sectional study. METHODS: The antigenic selection parameters were: proteins with antigenicity score ≥0.5 (VaxiJen), at least one B cell epitope and size between 10 and 275 KDa. New leptospiral proteins were cloned, expressed and serologically screened against equine sera (n = 128) on a single analysis and comparative combinations. Sensitivity (Se) and specificity (Sp), accuracy, positive predictive value (PPV) and negative predictive value (NPV) were calculated. A BLAST with nucleotide and protein sequences was used to identify the serovar or species specificity. MAIN LIMITATIONS: This cross-sectional analysis had three main limitations: (a) The equine sera used in these tests were limited to sera submitted to the Animal Health Diagnosis Center and were only tested against seven serovars; (b) MAT results were considered being 'perfect', and the highest titre presented was considered being the infecting serovar, which may not hold true; (c) The strains used to represent the serovars and the limited number of different serovars and species included in the genetic analysis, which leads to the possibility that these proteins might be present in different species or serovars that perhaps would be seroprevalent in another geographic region. CONCLUSIONS: The new leptospiral antigens described in this research could increase the sensitivity and specificity of ELISA for detection of Leptospira exposure and the detection of leptospirosis in horses along with support from other clinical signs. Some of these new antigens might be used to improve the detection of infecting serovar.

Acanthamoeba sclerokeratitis in a cat.

CASE DESCRIPTION: A 12-year-old neutered male domestic shorthair cat with chronic anterior uveitis and secondary glaucoma of the right eye was examined for persistent blepharospasm 2 weeks after corneal debridement and grid keratotomy for nonhealing superficial ulcerative keratitis. CLINICAL FINDINGS: Examination of the right eye revealed a central superficial corneal ulcer associated with corneal epithelial and subepithelial infiltrates and mild aqueous flare. Structures consistent with amoeboid cysts and trophozoites were detected in the cornea by in vivo confocal microscopy. Suppurative keratitis was identified cytologically. An Acanthamoeba spp was isolated through culture and identified by a PCR assay of corneal specimens. TREATMENT AND OUTCOME: Symptomatic and antiamoebic (polyhexamethylene biguanide 0.02% ophthalmic solution) treatments were instituted. Over the following 6 weeks, the cat lost vision in the affected eye and lesions progressed to nonulcerative stromal keratitis associated with a dense paracentral corneal stroma ring infiltrate and anterior lens luxation. The globe was enucleated, and lymphoplasmacytic sclerokeratitis, anterior uveitis, and retinal detachment were noted. Acanthamoeba organisms were detected within the corneal stroma and anterior sclera with histologic and immunohistochemical stains. The amoebae were classified to the Acanthamoeba T4 genotype by DNA sequencing. The cat had no medical problems attributed to Acanthamoeba infection over 36 months after enucleation, until the cat was lost to follow-up. CLINICAL RELEVANCE: Naturally acquired Acanthamoeba sclerokeratitis is described in a cat for the first time. Acanthamoeba infection should be considered for cats with superficial corneal disease refractory to appropriate treatments and especially occurring after ocular trauma, including keratotomy.

Clinical, clinicopathologic, and hepatic histopathologic features associated with probable ketoconazole-induced liver injury in dogs: 15 cases (2015-2018).

OBJECTIVE: To characterize clinical, clinicopathologic, and hepatic histopathologic features and outcome for dogs with probable ketoconazole-induced liver injury. ANIMALS: 15 dogs with suspected ketoconazole-induced liver injury that underwent liver biopsy. PROCEDURES: Medical record data were summarized regarding signalment, clinical signs, clinicopathologic and hepatic histopathologic findings, concurrent medications, ketoconazole dose, treatment duration, and outcome. RESULTS: Median age and body weight were 8.2 years (range, 5 to 15 years) and 13.0 kg (28.6 lb; range, 8.2 to 38.0 kg [18.0 to 83.6 lb]), respectively. The most common breed was Cocker Spaniel (n = 5). All dogs received ketoconazole to treat cutaneous Malassezia infections. Median daily ketoconazole dose was 7.8 mg/kg (3.5 mg/lb; range, 4.4 to 26.0 mg/kg [2.0 to 11.8 mg/lb]), PO. Treatment duration ranged from 0.3 to 100 cumulative weeks (intermittent cyclic administration in some dogs); 6 dogs were treated for ≤ 10 days. Common clinical signs included lethargy, anorexia, and vomiting. All dogs developed high serum liver enzyme activities. Hepatic histopathologic findings included variable lobular injury, mixed inflammatory infiltrates, and conspicuous aggregates of ceroid-lipofuscin-engorged macrophages that marked regions of parenchymal damage. Five dogs developed chronic hepatitis, including 3 with pyogranulomatous inflammation. Of the 10 dogs reported to have died at last follow-up, survival time after illness onset ranged from 0.5 to 165 weeks, with 7 dogs dying of liver-related causes. CONCLUSIONS AND CLINICAL RELEVANCE: Findings for dogs with hepatotoxicosis circumstantially associated with ketoconazole treatment suggested proactive monitoring of serum liver enzyme activities is advisable before and sequentially after initiation of such treatment.

Developing mechanically robust, triazole-zwitterionic hydrogels to mitigate foreign body response (FBR) for islet encapsulation.

Zwitterionic hydrogels such as those based on polycarboxybetaine (PCB) or polysulfobetaine (PSB) have potential for various biomedical applications, due to their biocompatibility and low biofouling properties. However, the poor mechanical properties of zwitterionic hydrogels developed to date remain a challenge, severely limiting their practical uses. To improve the mechanical properties without compromising their zwitterionic feature or biocompatibility, we designed a new class of zwitterionic hydrogels by introducing triazole moieties into the hydrogel monomers that could form energy-dissipating π-π stacking. Compared to conventional zwitterionic hydrogels, the triazole-zwitterionic (TR-ZW) ones exhibited similarly excellent antifouling properties, but were much more mechanically robust with higher stretchability (250% tensile strain), better compression-resistance (89% compressive strain and 65% compression for at least 10 cycles without any crack) and better folding-resistance. In addition, upon subcutaneous implantation in mice, the TR-ZW hydrogels induced significantly lower foreign body responses (FBR) (i.e. less fibrosis and more blood vessel formation relative to a poly(2-hydroxyethyl methacrylate) hydrogel control). As an example of their potential applications, we showed the use of the TR-ZW hydrogels for islet encapsulation and transplantation and demonstrated diabetes correction up to ~1 month in mice in the convenient subcutaneous site.