Journal of Veterinary Diagnostic Investigation

Avian influenza, caused by the avian influenza A virus (IAV), threatens poultry and public health. H6 subtype avian IAV is a low-pathogenic virus with hosts ranging from poultry and wild birds to mammals. H6 persists latently in poultry, which enables silent transmission and cross-species risk. The few fluorescence quantification assays that exist for H6 are mostly multiplexed. We developed a rapid, sensitive, efficient, monoplex fluorescence reverse-transcription qPCR (RT-qPCR) assay for H6 IAV. Specific primers and a TaqMan-MGB probe were designed based on the conserved hemagglutinin (HA) gene region of H6 IAVs from the GISAID database. The reaction components and conditions were optimized, and the assay was evaluated for specificity, sensitivity, and reproducibility. The optimized assay had excellent specificity, with no cross-reactivity with other avian viruses, including IAV subtypes H1–5, H7, H9, and H10, Newcastle disease virus, infectious bronchitis virus, fowl adenovirus, infectious laryngotracheitis virus, chicken anemia virus, Mycoplasmopsis (Mycoplasma) gallisepticum, and M. synoviae. Our method had a detection limit of 8.2 × 10⁰ copies/μL, which is 1,000 times more sensitive than conventional RT-PCR. The intra- and inter-assay CVs for all tested concentrations were both <1.5%, indicating good reproducibility. When applied to clinical swab samples, the sensitivity of our fluorescence RT-qPCR assay was 98.8% and specificity was 96.2% compared with traditional virus isolation. Our method could provide strong technical support for the early detection, monitoring, and prevention of H6 subtype IAV infection.

Between 2008 and 2024, fowl adenovirus (FAdV) genotypes were determined by hexon gene sequencing for 1,362 samples: 1,234 from 9 Canadian provinces and 128 samples from the United States. Most genotyped samples were from Ontario (681), followed by Alberta (243), Nova Scotia (116), British Columbia (77), Quebec (58), Saskatchewan (21), Manitoba (20), Newfoundland (16), and Prince Edward Island (2). Most samples (1,285) were related to inclusion body hepatitis (IBH); 77 samples were submitted for other reasons. Four FAdV genotypes (FAdV2, FAdV8a, FAdV8b, FAdV11) were associated with IBH-related submissions. Between 2008 and 2014, the most common strains associated with IBH outbreaks were FAdV11 and FAdV8a. However, since 2015, the identity of FAdVs involved in IBH outbreaks has shifted, with FAdV8b becoming the most frequent IBH-associated strain, largely displacing FAdV8a and FAdV11. In a much smaller group of 77 samples from non-IBH submissions, 10 FAdV genotypes were detected: FAdV1, FAdV2, FAdV3, FAdV4, FAdV6, FAdV7, FAdV8a, FAdV8b, FAdV9, and FAdV11. Although FAdV4 is a recognized causative agent of hepatitis–hydropericardium syndrome worldwide, no association with clinical disease was reported in the birds included in our study. Our comprehensive 17-y analysis of FAdV circulation patterns will support the development of control measures and vaccination programs to reduce the impact of FAdV-related outbreaks.

Chronic wasting disease (CWD) continues to be detected across the United States and globally; enhanced detection is critical for disease management. Silica nanoparticles (SiNPs) have shown promise in reducing time-to-detection for the real-time quaking-induced conversion (rtQuIC) assay in white-tailed deer (WTD) retropharyngeal lymph nodes (RPLNs). We aimed to document such decreased time-to-detection in 3 Wyoming, USA, cervid species. Additionally, we investigated maximum slope (max slope) as a metric of differentiating CWD status by rtQuIC testing, and how the receiver operating characteristic (ROC) may be useful for setting thresholds for QuIC outcomes. We performed rtQuIC testing, with and without SiNPs, on postmortem RPLNs from 39 WTD (Odocoileus virginianus), 40 mule deer (O. hemionus), and 40 Rocky Mountain elk (Cervus canadensis nelsoni). To measure effects of using SiNPs in the rtQuIC assay (nano-rtQuIC), median time-to-thresholds (tTh) for each sample replicate from QuIC and nano-rtQuIC was obtained using ROC thresholds. We found that nano-rtQuIC decreased the median tTh by 4.9, 5.3, and 3.6 h in WTD, mule deer, and elk, respectively. When using nano-rtQuIC, test sensitivity decreased by 5% in elk and by 4.8% in mule deer RPLN samples, whereas test sensitivity increased in WTD from 83.3% to 95.2%, indicating inhibition under the 50°C rtQuIC condition for WTD. Mechanisms of inhibition of rtQuIC by WTD RPLNs, comparatively by species, are unknown, but SiNPs and max slope analysis helped optimize rtQuIC test results. Interlaboratory validation and testing in a broader range of biological cervid samples would be useful for confirming these initial findings.

Feline coronavirus (FCoV) infects both domestic and wild felids and has the potential to cause feline infectious peritonitis (FIP), a progressive and often fatal systemic disease. Although rapid diagnosis and treatment are crucial in cases of FIP, conventional reverse-transcription quantitative real-time PCR (RT-qPCR) requires RNA extraction and specialized equipment, limiting its use for timely testing in general veterinary practice. We evaluated the performance of a direct RT-qPCR method using the PicoGene PCR1100 system (GoFoton, Ibaraki, Japan), which omits the RNA extraction step and delivers results within ~40 min. Compared with FCoV culture supernatants and extracted RNA, we estimated the limit of detection of this direct RT-qPCR method to be 150 copies/reaction—a detection sensitivity equivalent to that of conventional RT-qPCR targeting the FCoV 3′-UTR. We observed no cross-reactivity with other feline viruses or SARS-CoV-2. We subsequently analyzed 28 pleural and abdominal effusions collected from cats suspected of having FIP to compare the direct RT-qPCR method with the conventional approach. The sensitivity of the direct RT-qPCR method was 95.5% (95% CI: [78.2, 99.2]) and the specificity was 100% (95% CI: [61.0, 100.0]), which supports the use of the PCR1100 system as a rapid and user-friendly point-of-care tool for the detection of FCoV RNA in effusion samples.

White spot syndrome virus (WSSV; family Nimaviridae; taxon species White spot syndrome virus) is a major viral pathogen that poses a significant threat to the global shrimp industry, with early detection being the most effective strategy for disease control. We developed a CRISPR-Cas12a–based dual-target detection assay for WSSV, specifically targeting the VP28 gene (gene product is a major envelope protein) and WSSV366 (a latency-associated gene), optimized using Indian WSSV isolates. Our CRISPR RNAs for both targets had high efficiency, and we evaluated the assay using fluorescence-based and lateral flow strip (LFS) endpoint detection. In fluorescence assays, the Cr-WSSV assay (without recombinase polymerase amplification, RPA) detected WSSV at 3 × 10⁵ copies/μL; RPA integration significantly enhanced sensitivity, allowing detection at as low as 20 and 200 copies for VP28 and WSSV366, respectively, with 100% specificity. We developed a CRISPR-based LFS assay with optimized FAM-biotin reporter concentrations of 100 nM and 250 nM, yielding robust and reproducible results for improved field applicability. Performance evaluation confirmed lack of cross-reactivity to other WOAH-listed shrimp pathogens, while maintaining detection limits of 20 and 200 copies of VP28 and WSSV366. Clinical validation further demonstrated that the RPA-Cr-WSSV-LFS assay successfully detected WSSV366 even in VP28-negative samples, underscoring the importance of detecting WSSV366 in latent infections. Our rapid, cost-effective, and highly sensitive CRISPR-Cas–based assay enhances WSSV surveillance and biosecurity in shrimp aquaculture by incorporating structural and latency-associated gene markers, making it a promising alternative to conventional molecular testing.

The reliability of assumptions made about prevalence for pooling optimization varies greatly among target pathogens and surveillance strategies. When prevalence is unknown and difficult to anticipate, surveillance programs risk generating additional costs if pooling is suboptimal. Different methods of approximating optimal pool size (OPS) vary in precision of optimization, required sampling information, and the logistical demands placed on a laboratory workflow. Hence, it can be unclear how to assess compatibility between pooling optimization methods and the priorities of a surveillance program, sampling practices for the target population, and infection dynamics of the target pathogen. Our aim was to determine the relative performance in maximizing testing economy and cost reduction in different surveillance programs by simulating different pooling optimization methods on data from 280 submissions for bovine viral diarrhea virus (BVDV) surveillance (Nebraska Veterinary Diagnostic Center) and 111 submissions for Theileria orientalis surveillance (Virginia Tech Animal Laboratory Services). True prevalence, OPS, and historical prevalence were determined for each submission, and different optimization methods using fixed pool sizes, historical prevalence, and prevalence estimation testing were trialed on the data through Monte Carlo simulations. Contrasting results were observed between the 2 target pathogens, with historical prevalence being the most reliable optimization method for BVDV and the least reliable method for T. orientalis, which required significantly more tests than truly optimized pooling (p <0.05). Our results demonstrate the need to consider the interplay of infection dynamics, sampling practices, and surveillance priorities when selecting a pooling optimization approach.

Vitamin D deficiency, defined as low serum 25-hydroxyvitamin D (25D), is a highly prevalent finding in dogs with several disease states. Significant analytical variability between 25D methods of analysis in humans is well-documented; it is unknown if this variability exists in dogs. Our primary goal was to evaluate agreement of 2 chemiluminescent immunoassays (CLIA1, CLIA2); 2 liquid chromatography with tandem mass spectrometry (LC-MS/MS1, LC-MS/MS2) assays, from 3 laboratories; and a point-of-care lateral flow assay (LFA) across a wide range of expected 25D results in dogs. The null hypothesis was that all methods of analysis would have clinically acceptable agreement (<25 nmol/L difference). The tests were assessed for agreement via Bland–Altman analysis, Passing–Bablok regression, and Lin correlation coefficients. In Bland–Altman analysis, all inter-test comparisons exceeded the a priori clinically acceptable ≤25 nmol/L mean difference, except for LC-MS/MS1 vs. CLIA1 (mean difference −7.2 nmol/L). The range of mean bias across all test comparisons was −186% (LC-MS/MS2 vs. CLIA2) to 254% (CLIA2 vs. LFA). Tests of comparable methodology did not produce more similar results. The mean bias between the 2 CLIA assays was 54%, and the mean bias between the 2 LC-MS/MS tests was 125%. The significant analytical variability of canine 25D tests suggest that they cannot be used interchangeably, and caution should be used when comparing 25D results presented in canine studies from different laboratories and methods of analysis.

The intracellular bacterium, Coxiella burnetii, causes Q fever in humans and coxiellosis in animals. Cell-mediated immunity (CMI) has been shown to be important in Q fever; however, few studies have investigated the role of CMI in coxiellosis in cattle. Therefore, we aimed to optimize a whole-blood cytokine recall assay (CRA) and to evaluate C. burnetii–specific interferon-gamma (IFNγ) and interleukin 10 (IL10) responses in naturally infected cattle. Duplicate blood samples were collected from cows in an endemically infected dairy herd (as determined by serologic and molecular testing as well as through microbiologic culture) and subsequently transported at ambient temperature or on ice before stimulation with phase 1 and 2 C. burnetii antigen at various concentrations for 24 or 48 h; IFNγ and IL10 were measured in supernatant by ELISA. Using the optimized assay, the cytokine response was measured at calving and ~8 wk post-calving. The effect was assessed using generalized linear models and Spearman correlation coefficient (ρ). The greatest mean cytokine responses were elicited when blood samples were transported at ambient temperature and stimulated for 48 h with the highest C. burnetii antigen concentration. There was a significantly greater mean IFNγ (2.04 times) and IL10 (1.86 times) response at 8 wk post-calving compared to calving. At 8 wk post-calving, the correlation between cytokine response to phase 1 and 2 C. burnetii was ρ = 0.84 for IFNγ and ρ = 0.91 for IL10. Ultimately, our optimization study will guide informed use of the CRA in cattle research, allowing effective investigation of CMI in coxiellosis.

Nosemosis, caused by Vairimorpha (Nosema) ceranae or V. (Nosema) apis, is the main fungal disease affecting the Western honey bee (Apis mellifera). We evaluated the use of histology in the diagnosis of disease, identified the histologic patterns, and compared the efficacy of different staining techniques. We sampled 10 hives, collecting ~80 bees per hive. Spore counts were performed on 60 bees per sample using a hemocytometer in accordance with the standard procedure. Slides of whole bees were produced from the remaining bees, stained with 15 different techniques, and observed under a light microscope at 400×. Infection in the ventriculus was graded using hematoxylin–phloxine–saffron stain; prevalence and severity of the infection were determined; and an intra-class coefficient (ICC) was calculated to correlate the histologic results with the standard counting method. Based on contrast, specificity, and sensitivity, we found hot Gram chromotrope and Ziehl–Neelsen stains offered the best approach for highlighting Vairimorpha spores. These stains were optimized to find the ideal staining times for Vairimorpha by testing different immersion durations in key steps to enhance spore contrast. There was a notable association between histologic observations and spore count, with an ICC of 0.74 (95% CI [0.36, 0.91]) and 0.82 (95% CI [0.54, 0.93]) for the percentage of infected bees and histologic grade, respectively. Lesions included distension of ventricular epithelial cells, intracellular microsporidia, reduced ciliation, and disintegration of the peritrophic membrane. No spores were detected in extra-ventricular organs.

Myelolipomas are benign neoplastic lesions composed of adipose and hematopoietic tissue. Excluding dogs and cats, myelolipomas have been extensively described in the spleen of cheetahs (Acinonyx jubatus), the liver of Goeldi’s monkeys (Callimico goeldii), and birds in the order Psittaciformes. To better describe the species diversity and the clinical and pathologic relevance of myelolipomas, we conducted a detailed literature review and a retrospective multi-institutional study of myelolipomas, excluding cases diagnosed in dogs and cats. A total of 52 cases from 27 different species were diagnosed or reported (65% mammals and 35% birds) from 8 institutions. Notably, 18 of 27 (67%) of the animal species diagnosed with myelolipoma in our study lacked previous documentation in the literature. In mammals, myelolipomas were diagnosed in the spleen, liver, and adrenal gland, and less commonly in lymph nodes, mesentery, broad uterine ligament, and subcutis; most often, these proliferations were incidental findings at autopsy unrelated to animal death or euthanasia. In birds, the most frequently affected location was the liver, followed by the kidney and celom, occasionally resulting in antemortem clinical disease and adverse outcomes. Myelolipomas had clinical and pathologic relevance in 9 of 52 (17%) cases including birds and mammals. Osseous metaplasia was found within the myelolipomas in 2 of 52 (4%) cases. We conclude that myelolipomas are present in more species than previously documented and most cases are probably underreported. The clinical relevance of a myelolipoma can be determined by postmortem examination, and depends on the species affected, the anatomic location, and the size of the lesion.

Two Actinobacillus pleuropneumoniae (APP) isolates from clinical cases of porcine pleuropneumonia in Japan, positive for ApxIA, ApxIIA, and ApxIVA, were nontypeable using the agar gel diffusion (AGD) test but positive in the capsular serovar 1–specific PCR assay. Nucleotide sequence analysis revealed that gene clusters involved in the biosynthesis of the capsular polysaccharide (CPS) and lipopolysaccharide O-polysaccharide of the isolates were identical to those of serovar 1 reference strain 4047. The main difference found in the CPS loci was a loss of 7 nucleotides at the 3′-end of the cps1D gene in the atypical isolates, which is responsible for the defect in CPS production. Consistent with the serologic and molecular findings, transmission electron microscopic analysis confirmed the absence of detectable capsular material in the 2 atypical isolates. Collectively, our results suggest that this type of APP, defective in CPS production, may severely hamper serologic typing of the pathogen.

The genus Salmonella, particularly Salmonella enterica subsp. enterica serovars Choleraesuis and Typhimurium, poses significant challenges to swine production and leads to economic losses from conditions such as septicemia and enterocolitis. We evaluated the effects of experimental infection with Salmonella Typhimurium on clinical signs and anatomopathologic outcomes in pigs. Twenty 90-d-old pigs were divided into 2 groups: G1 received an oral inoculum of 10⁸ cfu of Salmonella Typhimurium; G2 served as a control. Pigs were monitored clinically for 30 d; postmortem examinations and microbiologic analyses were conducted. No significant differences were found in rectal temperature or weight between groups; however, diarrhea episodes were noted in the challenged group starting on day 5 post-inoculation. Isolates of Salmonella Typhimurium were detected intermittently in the challenged group; all positive samples came from pigs without diarrhea. Macroscopic lesions in G1 pigs included button-shaped ulcers in the ileocecal region, enlarged or hemorrhagic mesenteric lymph nodes, and hyperplasia of lymphoid tissue in the colon.

Streptococcus lutetiensis is a gram-positive organism in the non-enterococcal Lancefield group D Streptococcus bovis/S. equinus complex. S. lutetiensis has been reported as a cause of septicemia, endocarditis, and meningitis in humans, mastitis in dairy cows, and systemic streptococcosis in sea otters. A 3-mo-old reticulated giraffe (Giraffa reticulata) with a history of suspected failure of passive transfer (FPT) was submitted for postmortem examination, revealing bacterial septicemia, fibrinosuppurative polyarthritis, and embolic glomerulonephritis, with intravascular gram-positive cocci and microthrombosis in multiple organs. Bacterial culture and matrix-assisted laser desorption/ionization–time-of-flight mass spectrometry detected S. lutetiensis. S. lutetiensis septicemia was likely the result of FPT in this giraffe calf.

Canine extramedullary plasmacytomas are typically benign tumors of the skin, oral cavity, and alimentary tract that are cured by surgical excision. This tumor is rarely metastatic and aggressive. We report an unusual plasmacytoma in a dog that had been presented because of dyspnea. Aside from evidence of pleural effusion, no cutaneous lesions or other abnormalities were detected on physical examination. Nearly 2 L of pleural fluid were removed by thoracocentesis, and a sample was submitted for cytologic examination. The pleural fluid had increased protein and cell concentrations, with a predominance of individualized, large, round, atypical cells. Those cells frequently had Russell body–like intracytoplasmic structures, as seen on microscopic examination of modified Wright-stained concentrated slide preparations. Together, these findings were strongly supportive of a neoplastic plasma cell exudate. Immunohistochemical (CD3, CD20, MUM1, IgG, λ light chain) staining and B-cell PCR for antigen receptor rearrangement analysis performed on formalin-fixed, paraffin-embedded pleural cell-pellet sections confirmed a novel, clonal, IgG lambda extramedullary plasmacytoma with Mott cell differentiation that was most likely metastatic from a non-cutaneous primary site. Metastatic plasma cell neoplasia with voluminous serous cavity effusion carries a grave prognosis in humans, but has not been reported previously in dogs, to our knowledge.

Intracranial sarcomas are rarely reported in dogs. A 9-y-old, castrated male mixed-breed dog was evaluated and euthanized because of a 1-wk history of lethargy, dullness, disorientation, right-sided head tilt, and circling to the right. Grossly, a pale-tan, firm, 1.5-cm mass effaced ~60% of the right-ventral aspect of the pons. Histologically, the mass was composed of elongate neoplastic cells with abundant cytoplasm and round-to-oval nuclei arranged in bundles and supported by collagenous stroma. Anisocytosis and anisokaryosis were moderate, with 11 mitoses in 2.37 mm² (10 FN22/40× fields). The neoplasm was well-demarcated with occasional areas of infiltration in the surrounding neuroparenchyma. Neoplastic cells had widespread cytoplasmic immunolabeling for vimentin, patchy cytoplasmic immunolabeling for myoglobin, weak cytoplasmic immunolabeling for desmin, and rare cytoplasmic immunolabeling for myogenin. Transmission electron microscopy (TEM) revealed bundles of oval-to-elongate neoplastic cells with small-to-medium cytoplasmic aggregates of rough endoplasmic reticulum cisterns. The centrally-to-peripherally located nuclei were round and had one or more compact-to-reticulate nucleoli. Collagen bundles were in intimate contact with the plasma membrane of neoplastic cells and formed the abundant extracellular matrix. Histologic, immunohistochemical, and ultrastructural findings were consistent with an encephalic fibrosarcoma. The myogenic differentiation suggested by immunohistochemistry could not be confirmed by TEM.

A 3-mo-old domestic shorthair cat was presented with multiple fractures. Bone morphology was normal radiographically, with no long bone deformity or increased bone translucency. A bone biopsy from the ilium was examined histologically, revealing that bone matrix in the trabeculae extended from the growth plate, but cartilage remained in the distal trabeculae. Osteoblasts were observed at the bone surface via immunohistochemical detection with an anti-RUNX2 antibody. Whole-genome sequencing identified a homozygous missense mutation (valine to methionine) in the zinc-dependent metalloprotease domain of BMP1, a gene associated with human osteogenesis imperfecta type 13. In silico analysis predicted that this mutation would disrupt BMP1 protein function, which could affect type I collagen processing. Our findings suggest that a missense mutation in BMP1 may cause feline osteogenesis imperfecta.

A 9-mo-old, castrated male Saint Bernard dog was presented for evaluation of periorbital swelling, severe uveitis, and secondary glaucoma. Concurrently, chest radiographs had evidence of pneumonia. Enucleation was performed after failure of aggressive medical management. Histopathology of the globe confirmed severe necrosuppurative panophthalmitis and periocular cellulitis with myriad intra- and extracellular bacteria forming long filamentous chains. The bacteria were gram-positive and GMS-positive but acid-fast–negative. Next-generation sequencing (NGS) was performed on formalin-fixed, paraffin-embedded (FFPE) tissue from the eye. We identified a bacterium in the Actinomycetaceae family with a 100% BLAST match, suggestive of the previously described Actinomyces catuli strain (CCUG 41709). Clinical improvement followed enucleation and continued medical management, leading to reduction of the periocular swelling and resolution of the lung disease. Uveitis is common in dogs and is the most common cause of glaucoma. In many cases of bacterial uveitis, the exact bacterial organisms remain unknown if culture is not performed before fixation. Actinomyces sp. should be considered in patients with severe endophthalmitis or panophthalmitis, especially with evidence of systemic disease. NGS on FFPE samples may be a useful tool for identifying infectious organisms, especially in cases in which culture is not an option.

The number of pet minipigs, also called micro- or mini-pigs, has increased. Although cardiovascular diseases, particularly congenital malformations, are well documented in industrial and laboratory minipigs, they have not been reported in pet minipigs, to our knowledge. Detailed cardiac examination in awake pigs is challenging and typically requires sedation or anesthesia, making awareness of potential cardiac conditions essential. We present clinical and pathology findings from 3 pet minipig autopsies: aortic dissection, myxomatous valve degeneration, and ventricular septal defect. These cases highlight the need for increased awareness and investigation of cardiovascular diseases in pet minipigs.

Here we describe 2 outbreaks of intoxication by Ricinus communis in cattle in Argentina. In outbreak 1, in 2010, 180 heifers were introduced to a paddock heavily invaded by R. communis. Thirty-two animals developed watery diarrhea, and 6 of them were drooling, and had constant chewing motions, blindness, incoordination, depression, and prostration. Four affected animals died 12–14 h after the onset of clinical signs; another died 4 d later. The surviving 27 animals were removed from the paddock and recovered. At autopsy, several organs were congested and hemorrhagic, and abundant pericarps, leaves, and seeds of R. communis were found in the rumen content. The main microscopic lesion was acute, diffuse, superficial necrotizing gastroenteritis, and intestinal congestion and hemorrhage. In outbreak 2, in 2013, severe neurologic signs were observed in 12 of 300 cows after being introduced into a corn paddock without grain production that had been severely invaded by R. communis. Affected animals were excited and had tremors, drooling, incoordination, and prostration. The herd was immediately transferred to another paddock, and all affected cows recovered without treatment. In outbreak 1, the clinical signs and lesions were characteristic of simultaneous poisoning by R. communis fruits, which contain ricin and cause mainly digestive signs and lesions, and by leaves and pericarps, which contain ricinine and cause nervous signs. In outbreak 2, clinical signs and the recovery of the animals suggest that the intoxication was caused by ricinine, which is present in the leaves of R. communis.

An outbreak of botulism occurred in March 2024 among horses at a Quarter Horse stud farm in Central-West Brazil. After ingesting baleage, 22 of 26 (85%) horses housed in stables and fed baleage became ill. The affected horses had dysphagia, muscular weakness, fasciculations, and progressive recumbency; 13 of 22 (59%) died within a few days. The diagnosis of type C botulism was established based on clinical and epidemiologic findings and confirmed by mouse bioassays, which indicated botulinum toxin type C in liver samples and intestinal contents. Furthermore, PCR testing identified toxigenic Clostridium botulinum in the baleage consumed by the horses.

A 4-y-old male Labrador Retriever was submitted for autopsy following radiographic examination by the owner, a veterinarian, which had revealed 3 embedded projectiles. Autopsy revealed a single entrance wound on the left flank. Using flap-by-flap dissection, the trajectory of the projectile was traced through the skin and lumbar musculature to its location where it had perforated the abdominal aorta, causing fatal acute hemoperitoneum. Remarkably, the copper-coated pellet (Diabolo) was found in the lumen of the right femoral artery, consistent with projectile embolism. No external trauma was present at that site. Embolization can occur when a projectile enters the vasculature and travels to a distal location, typically requiring low residual kinetic energy and a vessel of sufficient caliber. Although well documented in human forensic medicine, projectile embolism is exceedingly rare in veterinary cases. To our knowledge, embolization of a projectile in the femoral artery in a dog has not been reported previously. Our case highlights the importance of comprehensive radiographic imaging before autopsy and illustrates the diagnostic value of correlating radiologic findings with meticulous gross examination in veterinary forensic pathology cases.

An 11-y-old spayed female American rabbit (Oryctolagus cuniculus) was presented for autopsy with a left maxillary mass. Grossly, the mass was firm, pale-tan, and firmly adhered to the left maxillary bone. Similar masses were observed in the lungs, liver, and kidneys. Histologically, the mass was poorly demarcated, unencapsulated, highly infiltrative, and densely cellular, consisting of neoplastic cuboidal-to-polygonal cells arranged in acini and occasional tubules anchored in a delicate fibrous stroma. Neoplastic cells were positive for anti-keratin/cytokeratin AE1/AE3 antibody. Similar neoplastic cells were present in the lungs, liver, and kidneys. Histopathologic and immunohistochemical findings were consistent with an adenocarcinoma. Based on the anatomy and histology of the rabbit lacrimal gland and adjacent lacrimal gland structures, we concluded that this was an adenocarcinoma arising from the accessory lacrimal gland. To our knowledge, adenocarcinoma originating from the accessory lacrimal gland has not been documented previously in a rabbit.

Conidiobolus sp. causes conidiobolomycosis, an emerging invasive fungal disease that affects humans and animals, mainly in tropical regions. In sheep, the disease has a major economic impact. We report an outbreak of conidiobolomycosis on a sheep farm in the Brazilian Amazon, as well as give a brief review of the subject. Veterinary care was requested on a rural property on which 3 of 35 sheep had developed prostration, facial edema, and exophthalmos. Two of the sick animals died, and a third was referred to the Veterinary Hospital of the Federal University of Acre (Rio Branco, Acre, Brazil). After imaging tests, the animal was euthanized given the advanced clinical stage of the disease. The main autopsy findings were rhinosinusitis with caseous necrosis, destruction of the nasal turbinates, and pulmonary granulomas. Based on our histologic, immunologic, microbiologic, and molecular tests, the outbreak was confirmed to be caused by Conidiobolus lamprauges, a saprozoonotic agent that has not been reported previously in northern Brazil, to our knowledge.

Mycobacterium celatum is a slow-growing, non-tuberculous mycobacterium with occasional branching morphology that has been reported mainly in immunocompromised human patients and rarely in other animal species. A 5-y-old female ferret was presented to a veterinary clinic with a history of anorexia and lethargy; peripheral lymphadenopathy, splenomegaly, and pneumonia were detected. Despite treatment, progressive deterioration of the ferret’s health led to euthanasia of the animal. Autopsy revealed numerous small 1–4-mm white nodules in the lung, spleen, and kidneys. Histologically, the kidney and lung nodules were pyogranulomas containing slender, elongate acid-fast bacilli with occasional branching. Bacterial culture was negative after 5 d of aerobic incubation. A PCR assay of kidney tissue was positive for Mycobacterium spp., with 100% DNA sequence similarity to M. celatum. M. celatum can cause systemic infection in humans and animals resembling tuberculoid mycobacterial infection. The diagnosis can be challenging due to cross-reactivity with tuberculosis-specific molecular assays and slow growth on bacterial culture. Although M. celatum has been reported elsewhere in ferrets more commonly than in other animals, M. celatum has not been reported previously in any animal species in North America, to our knowledge. M. celatum should be included as a potential pathogen in systemic mycobacterial infections, especially in ferrets.

Mucormycosis is an angioinvasive fungal infection caused by ubiquitous saprophytic fungi of the order Mucorales. In humans, the infection is generally caused by inhalation of spores or traumatic inoculation in cutaneous wounds, typically affecting immunocompromised patients. In animals, Mucorales infection is common in cattle, but is rare in the other species, including felids. No cases of mucormycosis have been reported to date in non-domestic felids, to our knowledge. We describe a case of mucormycosis in a 5-y-old female captive tiger (Panthera tigris). The affected tiger was part of a group of 9 subjects, 7 of which had serohemorrhagic nasal discharge, anorexia, and depression, followed by hemorrhagic diarrhea. Five of the tigers died and organs from one animal were sent for histologic examination, which revealed severe necrohemorrhagic enteritis, with periodic acid-Schiff– and Grocott methenamine silver–positive fungal hyphae. Mucor colonies were cultured from sawdust, which served as litter for the animals, indicating a possible source of infection through inhalation of spores and/or ingestion of contaminated sawdust. Mucormycosis is a possible differential for necrohemorrhagic enteritis in captive felids; careful storage and periodic assessment of the materials used as litter for these animals is recommended.

Feline herpesviral ulcerative dermatitis is associated with ulceration of the face and oral mucosa. Histologically, lesions consist of eosinophilic inflammation in the superficial dermis, with occasional eosinophilic intranuclear viral inclusion bodies in the epidermis, follicular epithelium, and associated glands. We describe a case of atypical felid alphaherpesvirus 1 (FeAHV1; family Orthoherpesviridae, taxon species Varicellovirus felidalpha1) dermatitis in a 6-y-old, castrated male, domestic shorthair cat in which lesions were located on the limbs and shoulder without facial or oral involvement. Histologic findings consisted of deep eosinophilic and plasmacytic dermatitis with vasculitis, thrombosis, and intranuclear viral inclusion bodies within macrophages. Immunohistochemistry (IHC) for FeAHV1 revealed viral antigen within macrophages in the deep dermis but not in epithelial cells. FeAHV1 was identified in the affected tissue via a consensus PCR assay targeting the DNA polymerase and the DNA packaging terminase subunit 1 for herpesviruses, followed by sequencing of the PCR amplicons. Similar lesions appeared on the right shoulder and right tarsal area ~7 mo after the first biopsy. Histologically, lesions were typical of feline ulcerative dermatitis and consisted of extensive areas of epidermal and follicular necrosis with ulceration and occasional intranuclear eosinophilic viral inclusion bodies in the epidermis and sebocytes. IHC in the second biopsy sample revealed FeAHV1 antigen within epithelial cells and macrophages near the ulcerated areas. Clinicians and pathologists should be aware of unusual manifestations of FeAHV1-associated ulcerative dermatitis in cats.

Nutritional myopathy (NM) is a sporadic disease of growing cattle, mostly associated with selenium (Se) deficiency, and characterized by necrosis of striated muscle. There is scant literature about NM in adult cattle. We describe 2 clusters of gastrocnemius and serratus ventralis NM associated with Se and copper (Cu) deficiency in adult beef cattle in Argentina. Cluster 1 included bilateral dropped hocks in 5 cows. Grossly, the gastrocnemius muscles of both hindlimbs had symmetrical hemorrhagic foci surrounded by pale-yellow areas. Cluster 2 included bilateral dorsal scapular displacement in a cow and a heifer. The animals of cluster 2 had diffusely elongated and pale-white serratus ventralis muscles. Microscopically, the affected muscles of all of the animals in the 2 clusters had polyphasic degeneration and necrosis. We observed below-normal blood glutathione peroxidase activity and plasma Cu concentrations in affected and unaffected animals of both clusters. Liver and kidney Se concentrations were normal, likely because the animals had been supplemented with this mineral soon before death. Liver Cu was below the normal range in all autopsied animals. Cu deficiency could favor oxidative damage produced by Se deficiency and interfere with collagen synthesis in the epimysium, predisposing the animal to muscle injuries. Although NM is more frequent in growing cattle, it should also be considered among the differential diagnoses for lameness in adult cattle.

Malakoplakia is an uncommon granulomatous disease reported to affect mainly the urinary bladder and, less frequently, the urogenital tract, gastrointestinal tract, and lymph nodes. We describe cases of malakoplakia within the renal medulla of 2 dogs. Both dogs had a history of recurrent urinary tract infections with urine culture growth of Escherichia coli. Autopsy findings in dog 1 included ~60 tan-to-yellow, irregular nodules within the renal medulla, and tan-to-red, semi-firm nodules within the urinary bladder mucosa. Dog 2 had no gross changes within the kidneys but rather had ~20 red, semi-firm nodules along the surface of the apex of the urinary bladder. Histologically, the renal medulla of both dogs had sheets of foamy IBA1-positive macrophages with cytoplasmic granules and inclusions of intensely periodic acid–Schiff (PAS)-positive and occasional intracytoplasmic von Kossa– and Prussian blue–positive Michaelis–Gutmann bodies, consistent with malakoplakia. Fluorescence in situ hybridization and/or immunohistochemistry confirmed E. coli within macrophages. An underlying endocrinopathy could be a predisposing factor in the development of malakoplakia in older dogs.

Pneumothorax is reported infrequently in marine mammals, most often secondary to physical or barometric trauma. We describe tension pneumothorax associated with verminous bronchopneumonia in a juvenile white-beaked dolphin stranded in Québec, Canada. Autopsy findings included a violent escape of air upon opening the pleural cavity, and white nematode-filled cystic nodules extending into the subpleural space and the lung parenchyma. One of these cysts had ruptured on the pleural surface. Numerous nematodes were observed in the lumen of main and secondary bronchi. Mediastinal lymph nodes were also enlarged. Helminths were collected and processed for molecular species identification. Histologically, verminous bronchopneumonia was marked by massive infiltration of the airways by polymorphonuclear and histiocytic cells surrounding adult and larval nematodes. Regional lymph nodes also contained focal infiltrates of polymorphonuclear cells associated with larvae. The nematodes were identified as Halocercus lagenorhynchi according to their morphology and molecular characterization. We attributed the death of this dolphin calf to tension pneumothorax secondary to a massive infection by the lungworm H. lagenorhynchi.

We report here a case of lipid-rich abdominal fluid in a 31-y-old Arabian gelding with acute colic. Abdominocentesis yielded opaque, yellow-to-orange peritoneal fluid with an elevated triglyceride concentration (42 g/L, RI: <0.10 g/L; fluid:serum triglyceride ratio 11.4, RI: <1.0). A second abdominocentesis performed 3 d later revealed clear fluid, suggesting transient chyloabdomen. Despite medical management, the horse developed respiratory distress secondary to pharyngeal edema and was euthanized. Autopsy identified a large omental lipoma and severe ulcerative gastritis, but no effusion. We speculate that necrosis of the lipoma might have contributed to the lipid-rich fluid, although definitive evidence was lacking.

A 2-y-old spayed female mixed-breed dog was presented with a 3-mo history of seizures. Coalescing intra-axial complex cystic lesions in the right cerebral hemisphere identified on MRI were suggestive of hydatid cysts; however, PCR testing of a fecal sample for Echinococcus spp. was negative. The dog was euthanized after 3 mo of treatment due to worsening signs and was submitted for postmortem examination. Coalescing 0.5–3-cm cavitations effaced ~20% of the left and 40% of the right cerebral hemispheres, and contained numerous 3–5-mm long ovoid-to-elongate, soft, white-to-clear metacestodes. Similar structures extended into the subarachnoid space. Histology revealed multiple larval cestodes consistent with invaginated cysticerci present in bladder compartments. Cysticerci each had a scolex, convoluted invaginated spiral canal, and spinous tegument with numerous calcareous corpuscles. Within many of the cysticerci, visible armed rostella had refractile hooklets and muscular suckers. Light microscopic evaluation of whole cysticerci preserved in ethanol revealed rostellar hooks with blade-to-guard length and handle-to-guard length that were within RIs for Taenia crassiceps. Sequencing of DNA amplicons obtained via PCR confirmed 100% sequence identity to T. crassiceps. To our knowledge, canine neural cysticercosis attributed to T. crassiceps has not been reported previously. Our case highlights the successful integration of multiple diagnostic modalities in a case of canine neural cysticercosis.