Two-step humane euthanasia in cave cockroaches

The need for evaluation of anxiety and pain

The study was conducted from August–September, 2024 at the Fish Seed Production Center and Fish Farm of the College of Fisheries Science, Nagpur, Maharashtra, India to systematically evaluate metabolic and stress-related responses of Labeo catla fingerlings under varying stocking densities to determine and standardize the optimal conditioning density that minimizes stress, reduces mortality, and enhances post-transport efficiency. In this study, Catla fingerlings (90.82±5.91 mm) were conditioned in a flow-through system with a water exchange rate of 6.5 l min-1 for the period of 24 h, during this, fingerlings were stocked at four densities: 1 fingerling l-1, 2 fingerlings l-1, 3 fingerlings l-1, and 4 fingerlings l-1. During conditioning, blood samples were collected for glucose, lactate, hemoglobin, and hematocrit estimation, while muscle samples were used for glycogen analysis following anesthesia with clove oil and humane euthanasia. Water quality parameters such as dissolved oxygen and total ammonia nitrogen were monitored to assess metabolic load under varying densities. The results showed density-dependent stress responses. 4 fingerlings l-1 exhibited significantly higher blood glucose and lactate, lower glycogen levels, and greater mortality (p<0.05), indicating elevated stress. While water quality remained stable overall, 4 fingerlings l-1 showed more DO depletion and total ammonia buildup. 3 fingerlings l-1 showed the best balance between stocking pressure and physiological stability, indicating it was the optimal conditioning density for Catla fingerlings in flow-through systems.

Many wild otariids (sea lions and fur seals) strand along the California coast annually with seizures following presumed exposure to harmful algal blooms (HABs). This is a long-term case study of a stranded subadult male Guadalupe fur seal (Arctocephalus townsendi) that stranded in 2001 and within days after admission began demonstrating seizure activity. The epilepsy was stabilized with antiepileptic drug (AED) therapy and advanced imaging was performed. Magnetic resonance (MR) scans showed signs of encephalopathy consistent with domoic acid (DA) toxicosis. This fur seal was deemed “non-releasable” and over the following 19 years, he received supportive veterinary care and AED treatment at a permitted facility. In the summer of 2020, the fur seal showed progressive behavioral and functional deterioration whereupon humane euthanasia was performed. Post-mortem MR and diffusion tensor images (DTI) were acquired. A volumetric comparison with historic scans showed evidence of severe progressive unilateral right hippocampal atrophy. Histopathology confirmed severe chronic right and mild left hippocampal atrophy. The progressive degenerative brain changes demonstrate how adaptive brain function can be with neuronal atrophy secondary to DA exposure. This case highlights some considerations and decision-making processes needed for the rehabilitation and placement of wild stranded otariids with acquired epilepsy secondary to DA toxicity.

Euthanasia of Food Animals

This case report describes the identification of multiple soft tissue sarcomas in the pharyngeal region of a 5-year-old Quarter Horse mare. Diagnostic work-up included physical examination, radiography, ultrasonography, endoscopic examination of upper airways and guttural pouch, and post mortem examination with histopathology. Humane euthanasia was indicated due to the chronicity of the condition, prognosis and financial constraints.

An orphaned 4-mo-old female mountain lion cub ( Puma concolor ) was captured along the coastline in Montaña de Oro State Park in Los Osos, California, USA. Following suspicion that the cub was visually impaired, ophthalmic examination revealed diffuse bilateral retinal atrophy. Due to a poor prognosis, humane euthanasia was elected. Necropsy and histopathological findings were consistent with photoreceptor degeneration. Based on the cub's signalment, history, and histopathology, a genetic or nutritional etiology was suspected, with the former etiology more strongly supported. To the authors' knowledge, this is the first report of photoreceptor degeneration in a wild felid and should be considered in cases of blindness.

To describe the clinical findings and case progression in a dog presenting with severe systemic inflammatory response, refractory shock, progressive metabolic acidosis, and respiratory failure that was ultimately diagnosed with hypertrophic osteodystrophy (HOD). A 4-month-old male intact Mastiff presented with a 24-hour history of lethargy and generalized ostealgia. On examination, the dog was recumbent, febrile, and tachycardic with pain on palpation of the abdomen, right femur, and mandible. Appendicular joint radiographs showed changes consistent with osteochondrosis and ulnar-retained cartilaginous cores, with no overt evidence of HOD. Initial treatment included IV fluid therapy, multimodal analgesia, and broad-spectrum antimicrobials. Vasopressor therapy was initiated following hemodynamic decompensation. Synovial fluid cytological analysis and culture revealed nonseptic suppurative inflammation and no bacterial growth, respectively. Blood and urine cultures also yielded no growth. Viscoelastic testing was consistent with hypercoagulability. The dog initially had a metabolic acidosis with appropriate respiratory compensation that progressed to a mixed metabolic and respiratory acidosis despite aggressive therapies that included antimicrobials, vasopressors, positive inotropes, and corticosteroids. Humane euthanasia was elected approximately 32hours after admission. Necropsy yielded a diagnosis of HOD. This is the first report detailing the occurrence of refractory shock and hypercoagulability associated with HOD in a dog without evidence of another identified comorbidity. HOD should be considered in any young, large-breed dog with generalized ostealgia and signs of systemic illness, even in the absence of classic radiographic abnormalities. Further investigation of coagulation status in dogs with HOD and a secondary systemic inflammatory response is warranted.

A 6-year-old male neutered boxer mix canine was presented for a one-month history of dyschezia, hematochezia, and constipation. Colonoscopy and endoscopic biopsies revealed non-specific lymphoplasmacytic, eosinophilic colitis. Despite pursuing various therapies over a 3.5-month clinical course (including hypoallergenic diet, antibiotics, prokinetics, laxatives, and anti-inflammatory glucocorticoids), the patient's condition did not improve. Two and a half months after initial presentation, the patient developed circumferential proctitis with multiple draining tracts and obstipation. Humane euthanasia and postmortem examination were elected. Gross and histological findings revealed locally extensive pyogranulomatous coloproctitis with many intralesional PAS-positive, GMS-negative 30-40 μm in diameter, hyaline, pauciseptate, irregularly branching fungal hyphae, hyphal bodies or chlamydospores and 25-45 μm in diameter thick-walled zygospores. Fungal culture of fluid from the draining tracts was performed, and DNA sequence analysis of the ITS and partial LSU of the nuclear ribosomal RNA genes were used to identify and confirm a novel species, Basidiobolus arizonensis. Basidiobolus spp. are saprobes in the order Basidiobolales and most commonly cause granulomatous infections of the skin, respiratory tract, and gastrointestinal tract in veterinary species and humans. To the authors' knowledge, this is the first report of novel Basidiobolus arizonensis causing localized coloproctitis in a dog.

An 8-month-old male Yorkshire terrier was presented for ingestion of 800 mg of an over-the-counter caffeine supplement. Clinical signs included extreme tachycardia, facial fasciculation, coma/stupor and flailing. Due to the lack of response to medical therapies, humane euthanasia was elected. Microscopically, necrotic neurons were scattered throughout the hippocampus, olfactory cortex, pyriform lobe, amygdala, and basal nuclei, with relative sparing of the caudate nuclei. In addition, mild skeletal myocyte necrosis and mural necrosis of cardiac arterioles in the left and right ventricles were noted. This is the first report of the microscopic lesions associated with caffeine intoxication in a dog.

A 7-year-old, 153.0-kg American Miniature mare presented for evaluation of keratoconjunctivitis of the right eye (OD). A superior palpebral conjunctival mass and stromal keratitis were diagnosed. The incisional biopsy diagnosis was a presumptive corneal hemangiosarcoma. Transpalpebral enucleation was performed, and histopathologic evaluation confirmed angiosarcoma of the conjunctiva, cornea, and extraocular muscles. The horse developed progressive epistaxis and orbital swelling following surgery. A systemic workup was performed 3 months after enucleation, revealing regrowth within the orbit and marked cranial cervical lymphomegaly, suggestive of metastasis. Humane euthanasia was performed, and necropsy confirmed a locally invasive periorbital tumor with metastasis to the submandibular tissue, submandibular lymph node, and thoracic inlet. Histopathologic evaluation of necropsy specimens revealed polygonal to spindle neoplastic cells lining neoplastic vascular channels lacking erythrocytes. Immunohistochemically, the neoplastic cells labeled strongly positive for PROX-1, vimentin, CD-31, VEGF, weakly positive for factor VIII-related antigen, and negative for collagen IV. Based on the clinical, histological, and immunohistochemical features of this tumor, a primary ocular lymphangiosarcoma with metastasis was diagnosed.

Traditional treatments for heart failure (HF) with reduced ejection fraction (HFrEF) are ineffective at treating HF with preserved ejection fraction (HFpEF), establishing a need for more targeted therapeutic approaches. To date, the comparative vascular adaptions induced by these two distinct forms of HF have been largely unexplored. Therefore, the objective of this study was to compare HFrEF and HFpEF pathophysiological adaptions in both coronary and cerebral microvasculature in two clinically relevant swine models of HF. We hypothesized that HFrEF and HFpEF will induce differing pathophysiological adaptations within the microvasculature of the heart as well as peripheral tissues, such as the brain. Intact female Ossabaw swine (2 months old) were fed a Western Diet for 4 months to develop metabolic syndrome (HOMA-IR: HFrEF 2.94 ± 0.34; HFpEF 2.95 ± 0.59). At 6 months of age, animals were either subjected to 90 minutes ischemia followed by reperfusion (I/R) to induce a myocardial infarction (MI) recapitulating cardiometabolic HFrEF (38.0 ± 3.9% EF) or were aortic banded to induce pressure-overload with relevance to cardiometabolic HFpEF (61.5 ± 1.7% EF). Following humane euthanasia, coronary arterioles (139.5 ± 4.9 μm inner diameter) and middle cerebral artery second order pial arterioles (281.1 ± 16.7 μm) from each model were dissected and either cannulated and pressurized for myography experiments or flash frozen for RNA isolation, sequencing, and predictive pathway analysis. Smooth muscle dependent U46619 (thromboxane A2 agonist)-induced constriction of cerebral arterioles was not different between groups (HFrEF 46.9 ± 10.3%; HFpEF 50.7 ± 4.8%), however, smooth muscle dependent dilation to sodium nitroprusside (SNP, nitric oxide mimetic) was significantly reduced in HFrEF (37.1 ± 14.0%) compared to HFpEF (83.6 ± 4.6%), suggesting the dilatory capacity of peripheral vessels is impaired when EF% is reduced. Furthermore, RNA-sequencing comparison between HFrEF and HFpEF uncovered 35 differentially expressed genes in the coronary (n=3) and 23 differentially expressed genes in cerebral (n=3) arterioles (q ≤ 0.1). Ingenuity pathway analysis predicted upstream regulators that code for angiotensinogen, STAT3, and interferon-Ƴ were inhibited (z-score > -2.0) in the HFrEF model, which was associated with decreased proliferation of smooth muscle cells and vasculogenesis with increased organismal death and inflammation in the coronary arterioles. Furthermore, analysis of the cerebral arterioles predicted inhibition of angiotensinogen (z-score > -2.0) in the HFrEF model which was associated with increased fibroblast cell death, dedifferentiation of smooth muscle cells, and recruitment of macrophages. Together, these data reveal distinct pathophysiological functional and transcriptomic adaptions between HFrEF and HFpEF in the coronary and cerebral arterioles. Additionally, these data provide three candidate genes that regulate these pathophysiological processes and could be targeted for potential therapeutic intervention. Funding: REGENCOR and Department of Defense (DOD) W81XWH-18-1-0179. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

A 2.5‐year‐old male African hedgehog (Atelerix albiventris) presented with an intraoral, soft tissue mass overlying the left mandible. A computed tomography scan and subsequent fine needle aspirate suggested a malignant spindle cell tumour. The tumour was excised with a partial hemimandibulectomy. Histopathology demonstrated an incompletely excised spindle cell sarcoma. The hedgehog underwent adjuvant therapy consisting of intravenous carboplatin and oral lomustine, followed by palliative radiation therapy once tumour recurrence was noted on follow‐up surveillance. Radiation therapy was initially successful in decreasing tumour size, but the hedgehog re‐presented a month later acutely non‐ambulatory paraparetic with a distal right antebrachial mass. Diagnostics including radiographs and fine needle aspirate were consistent with metastatic neoplasia and humane euthanasia was elected.

The horse hoof has evolved from the same appendage that now makes up the human middle finger, to bear up to 100% of a 500-kg animal, while taking up an area of less than only .05 m2. To meet these exceptionally high mechanical demands, the most distal bone (P3) is suspended in the hoof capsule through a remarkable adaptation of lamina tissue (in humans, this is the tissue underneath a fingernail). The P3 bone is encapsulated by the keratinous hoof (fingernail) in direct contact with the ground and the external environment. The lamina tissue is built to hold the bone so that the P3 bone "floats" within the hoof, providing cushion and impact dampening. When the lamina tissue begins to fail due to sickness, obesity, or age, the developed condition is referred to as laminitis, which causes the P3 bone to rotate downwards. The clinical impact of laminitis is pain, damage to the laminar tissue, and possible humane euthanasia. To diagnose laminitis, veterinarians currently take three radiographs of the horse hoof: from the lateral, ventral, and caudal views. The angle of the bone from the hoof wall in the lateral view is commonly used to diagnosis laminitis. In a healthy animal, this angle would be close to 0 and, in a horse with laminitis, this angle would be much larger. Typically, the other two views are mainly used to determine the overall health of the P3 bone, rather than to help diagnose laminitis. This limits the diagnostic ability of the veterinarian in the case of incomplete, missing, or inconclusive lateral radiographs. The goal of this study therefore was to develop an alternative method of diagnosing laminitis utilizing the ventral radiograph. It was hypothesized that the downward rotation of the P3 bone (as seen in the lateral view) would affect the shape of the P3 bone in the ventral radiograph. More specifically, if the change in the shape of the P3 bone in the ventral radiograph could be measured using the eccentricity of the bone, the ratio of the areas of the P3 bone to hoof, or the difference between the hoof and bone. These measurements were examined to determine if there was statistically significant difference between the healthy and pathological radiographs. The overall goal was divided into two aims: 1) determine if analysis of ventral view images could detect differences in known laminitis and healthy images, and 2) validate the new procedure. In the first aim, 15 sets of radiographs were studied from known cases, of which 7 subjects were diagnosed with laminitis and 8 were confirmed healthy. The angle of the bone from the hoof wall, the areas of the bone and hoof, and eccentricity of the bone were calculated for each case using custom written Matlab programs. It was determined, through statistical testing such as F-tests and 2-way ANOVAs, that eccentricity was the most viable diagnosing method among the three new methods being tested. To validate this diagnostic technique and satisfy the second aim, 35 blinded sets of radiographs were measured and diagnosed with the commonly used method and this new eccentricity-based method. These diagnoses were then compared to a veterinarian's official diagnoses. It was shown, through statistical testing, that eccentricity is an accurate diagnosing technique that exceeds the conventional practice in sensitivity, specificity, false positive rates, and false negative rates.

Background Novel cardiac valve prostheses require long-term studies assessing safety in animal models prior to clinical use. Although sheep are well-established preclinical models, the anatomy and morphology of the sheep heart impart unique challenges in the positioning and securement of transcatheter mitral valve replacement (TMVR) devices, resulting in early postoperative mortality. We developed a refined hybrid surgical approach to study the chronic in vivo performance of a novel TMVR device intended for regulatory approval. Methods Eleven adult sheep were enrolled in this study. Eight Abbott Next-Generation Cephea (Cephea) TMVR devices were implanted using a hybrid surgical approach, with the valve deployed and secured under direct visualization on cardiopulmonary bypass. For a control, three Medtronic Mosaic bioprostheses (Mosaic) were implanted with standard techniques. Sheep were survived for 20 weeks per regulatory guidance, followed by humane euthanasia and comprehensive necropsy. Results All sheep were successfully implanted and recovered from surgery, with 7/8 Cephea and 3/3 Mosaic animals surviving to the 20-week study endpoint. A surgical complication unrelated to TMVR performance resulted in one early death at 58 days post-operatively. At term, all Cephea valves performed comparably to controls, appearing well-healed within the annulus with competent, unobstructed leaflets. Conclusions Our novel hybrid surgical approach allowed for a comprehensive evaluation of the long-term performance of the Cephea, with 100% survival at 30 days and 88% at the 20-week endpoint. These results suggest that this approach may improve chronic survival in animal studies evaluating the safety of novel TMVR devices prior to clinical investigation.

Chondrosarcoma in a captive giant anteater (Myrmecophaga tridactyla).