In 2023, high pathogenicity avian influenza (HPAI) H5N1 outbreaks caused mass mortality events affecting pinnipeds throughout South America. Here, we present the clinical, pathological, immunohistochemical, and molecular findings associated with HPAI H5N1 infection in 3 South American sea lions (Otaria flavescens) and 1 southern elephant seal (Mirounga leonina) stranded during the outbreak in Chubut, Argentina. Neurological signs were observed in 2 sea lions, with 1 also exhibiting respiratory distress. Necropsies were conducted on all 4 animals. Neuropathology revealed mild-to-severe lymphohistiocytic (2/4) to mixed neutrophilic (2/4) meningoencephalitis with neuronal necrosis, neuronophagia, glial cell proliferation, multifocal hemorrhage, and perivascular cuffing in all animals. Novel findings for HPAI H5N1-infected pinnipeds included choroid plexitis (2/4) and myelitis (2/4). Immunohistochemistry for viral nucleoprotein was positive in 3 of 4 animals and involved neurons (3/3), glial cells (3/3), and ependymal cells (2/3). Viral-related systemic findings included multifocal necrotizing myocarditis in the elephant seal and multifocal necrotizing placentitis in a sea lion, with HPAI virus detected within fetal tissues. Pulmonary lesions were minimal and were limited to multifocal necrosis of bronchial glands in 1 sea lion. HPAI H5 clade 2.3.4.4b virus was confirmed by polymerase chain reaction and sequencing in all animals. These findings indicate a predominance of central nervous system involvement in HPAI H5N1-infected pinnipeds and expand the recognized spectrum of virus-associated lesions, identifying novel tissue tropisms and transmission routes.

Cetacean morbillivirus (CeMV) drives recurrent unusual-mortality events, yet surveillance is uneven where laboratory capacity is limited. We developed a portable reverse transcription-insulated isothermal PCR (RT-iiPCR) assay targeting a conserved phosphoprotein (P)-gene segment and evaluated analytical performance, tissue-level clinical sensitivity, and concordance with reverse transcription-quantitative PCR (RT-qPCR) under low-copy conditions that resemble challenging strandings. Using synthetic RNA, RT-iiPCR achieved 100% detection from 62,560 to 513 copies µL-1, and probit analysis estimated a 95% limit of detection (LOD95) of 139 copies µL-1. Clinical sensitivity was assessed with two spiking regimes (RNA added after or before extraction) in cerebrum and lung. Singleplex RT-iiPCR maintained 100% positivity to approximately cycle threshold (Ct) 33 irrespective of spiking order, indicating that near-limit failures are governed by template scarcity rather than extraction loss. Duplex RT-iiPCR co-amplifying β2-microglobulin (B2M) provided process control with a slight sensitivity cost, sustaining perfect detection to ~Ct 30-31. Across low-copy panels, agreement with RT-qPCR was substantial (overall κ = 0.68-0.76) and very good in cerebrum (singleplex κ = 0.85; duplex κ = 0.87), while duplex lung showed lower concordance (κ = 0.55) driven solely by Ct >33 false-negative calls, with no false positives. The assay detected five lineages (dolphin morbillivirus [DMV], pilot whale morbillivirus [PWMV], beaked whale morbillivirus [BWMV], Guiana DMV [GDMV], and Fraser's DMV [FDMV]) in formalin-fixed, paraffin-embedded tissues archived up to 28 years, and sequence alignments indicate expected coverage of additional lineages. Lyophilized reagents, compact hardware, and a quick, simple workflow support deployment in hot, humid, resource-limited settings. A strain-agnostic, field-ready RT-iiPCR can underpin transboundary CeMV surveillance, enable rapid carcass triage and sequencing, and provide early warning where diagnostic gaps currently exist.

The scientific understanding of cetacean pathology has advanced significantly in recent decades. However, data concerning the health status of members of the family Kogiidae remains scarce. This study presents a comprehensive pathological assessment and determination of causes of death in 45 stranded kogiids, comprising 35 Kogia breviceps and 10 K. sima, along the coasts of the Canary Islands between 1999 and 2018. Causes of death (CD) were classified as natural (30/45; 66.6%) or anthropogenic (11/45; 24.4%), while the cause remained undetermined in four cases (9%). Among natural causes, the most prevalent etiologies included trauma (13/30; 43.3%), infectious (7/30; 23.3%), parasitic (5/30; 16.6%), cardiomyopathy (3/30; 10%), malnutrition (1/30; 3.3%), and fetal distress (1/30; 3.3%). Anthropogenic causes were vessel collisions (7/11; 63.6%), interactions with fishing activities (2/11; 18.2%), and foreign body-associated pathology (2/11; 18.2%). Notably, intra- and interspecific traumatic interactions were frequently identified in this cohort, and variable degrees of gross and histologic cardiomyopathic changes were observed in 68.9% of individuals (31/45) without evident sex bias. Septicemia caused by Clostridium perfringens and C. tertium was confirmed in three cases. Additionally, cervical gill slit adenitis by Crassicauda sp. (15/45; 33%) and parasitic gastritis by Anisakis sp. (27/45; 60%) were frequent findings. These findings provide novel insights into the pathologic spectrum affecting kogiids and enhance the current understanding of their health status, with implications for diagnostic protocols, stranding response, and conservation strategies.

The adrenal glands are central regulators of endocrine function and stress physiology, yet detailed species-specific anatomical baselines remain limited in cetaceans. This study provides a comprehensive gross, histological, morphometric, and ultrastructural characterization of the adrenal glands in 55 short-beaked common dolphins (Delphinus delphis) examined postmortem in the Canary Islands. Adrenal glands were evaluated macroscopically and microscopically, and histological corticomedullary ratios were calculated from mid-transverse sections. Associations with body length, sexual maturity, and cause-of-death category were assessed statistically. Transmission electron microscopy was used to characterize cortical and medullary cellular ultrastructure. Adrenal weight showed a positive correlation with body length. The histological corticomedullary ratio showed no lateral asymmetry but differed significantly between sexually immature and mature individuals, indicating ontogenetic remodeling of adrenal architecture. In contrast, the corticomedullary ratio did not differ significantly between adult dolphins that died from acute events and those that died following more progressive pathological conditions. Ultrastructural analysis identified characteristic steroidogenic cortical cells and two distinct chromaffin cell populations in the medulla. These findings establish the first integrated anatomical baseline for the adrenal gland in Delphinus delphis, providing essential reference data for comparative anatomy, veterinary pathology, and the interpretation of endocrine-related findings in cetaceans.

Fisheries bycatch is an important source of direct human-induced mortality for marine mammals worldwide. Despite its significance, monitoring and quantifying bycatch mortality remains challenging. The examination of stranded animals can provide key information to assess the importance of bycatch mortality, contributing to understanding its impact on populations, and identifying the fisheries responsible. However, the utility of data from strandings remains underexplored. Based on an expert consultation using a questionnaire carried out under the auspices of the International Council for the Exploration of the Sea Working Group on Marine Mammal Ecology, this study assessed the potential of stranding networks in Western Europe to support the assessment of marine mammal population status and impacts of stressors. The present paper (the second of two) focuses on information on bycatch collected from strandings, and its utility for monitoring and assessing bycatch mortality. Responses from 45 organizations in 19 countries indicated that extensive information on marine mammal bycatch mortality is collected and can sometimes be used to identify the species or populations most affected by bycatch, reveal spatio-temporal trends in bycatch mortality, and determine the fisheries and/or fishing gear involved. However, the survey identified several limitations that hinder the contribution of stranding data to bycatch impact monitoring and assessment. There is a need to strengthen data collection and reporting, especially for certain taxa and regions, and for wider adoption and harmonization of best practise protocols for diagnosing causes of mortality and understanding its context, e.g. by determining health status, estimating demographic parameters, helping to identify the fishing gears responsible for bycatch mortality, and appropriately communicating the degree of uncertainty associated with interpretations. Carcass drift models could enable stranding data to be used to estimate total bycatch mortality and contribute to assessing the impact on populations. The need for adequate baseline funding for stranding networks to carry out their activities and for streamlined data reporting was widely recognized. Achieving coordinated, harmonized, large-scale and long-term data and samples collection following best practices will be essential to ensure that data from strandings on bycatch mortality can be integrated into bycatch impact assessments to inform management.

Despite the popularity of elasmobranchs in zoos and aquariums, data on hematological values for health monitoring remain limited, particularly for wild populations. In this study, reference intervals for key hematological parameters and blood pH were established from blood samples of 49 adult females of the Critically Endangered Spiny Butterfly Ray (Gymnura altavela) collected across three shallow beaches in the Canary Islands. Additional data were obtained from four adults maintained under human care. To our knowledge, this is the first report of hematological and blood pH reference intervals for free-ranging G. altavela. Lymphocytes (56

Canine leishmaniosis caused by Leishmania infantum commonly induces marked dysproteinemia, yet restricted or apparently monoclonal gammopathies may complicate interpretation of serum protein electrophoresis (SPE). This study characterized SPE and immunofixation findings in 38 clinically sick dogs with confirmed canine leishmaniosis and evaluated interobserver agreement in gammopathy classification. SPE and immunofixation using anti-human reagents was performed on serum from 38 sick dogs with confirmed clinical leishmaniosis and without evidence of plasma cell neoplasia. The SPE data alone and a combined SPE and immunofixation (IF) study were interpreted by 4 reviewers. Inter-observer agreement for presence of an acute phase response and gammopathy pattern by electrophoresis alone or with immunofixation was assessed using Gwet's AC1 and quadratic weighted AC2. Change in agreement with addition of immunofixation was assessed. All dogs demonstrated hypergammaglobulinemia and decreased albumin-to-globulin ratios. Hyperproteinemia and hypoalbuminemia were common, and hyperbetaglobulinemia was frequently observed. Immunofixation labeling was predominantly IgG with minimal IgA and IgM detection. Most cases had an IgG polyclonal or restricted polyclonal gammopathy. Interobserver agreement for gammopathy classification using SPE alone was high (AC2 = 0.902), but disagreements spanned the diagnostic spectrum in some cases. The addition of immunofixation significantly improved agreement (AC2 = 0.944; P = 0.034) and reduced extreme interpretive divergence. No case was unanimously classified as monoclonal, and incorporation of immunofixation reduced suspicion for monoclonality in cases initially considered equivocal. Dogs clinically affected for canine leishmaniosis exhibit predominantly IgG-driven polyclonal hypergammaglobulinemia. Together, SPE and IF provide a more complete characterization of gammopathy patterns in dogs with leishmaniosis, particularly when SPE reveals restricted or monoclonal-like abnormalities that could otherwise raise concern for plasma cell neoplasia.

Post-mortem computed tomography (PMCT) has become an increasingly important tool in cetacean stranding investigation, providing reusable, three-dimensional data of internal anatomical structures. Here, we present Cet.CT-Bank, a PMCT imaging dataset derived from stranded cetaceans recovered along the Canary Islands coast. The dataset comprises eight odontocete specimens representing six cetacean species, scanned using standardized CT protocols and stored as DICOM files with complete acquisition and reconstruction metadata. Each case is accompanied by automatically generated validation reports, ensuring traceability and reproducibility of the imaging data. Biological and pathological information is provided within this data descriptor and its supplementary materials to support integrative interpretation. Cet.CT-Bank is intended as an open, reusable imaging resource for morphological research, veterinary pathology, forensic investigation, and the development and validation of computational image analysis methods, including applications in computational image analysis, with potential relevance for future artificial intelligence-based approaches.

This study characterizes the pathological findings in Atlantic Cory's shearwater (Calonectris borealis) chicks from the Canary Islands associated with illegal hunting cases. A retrospective analysis of necropsy archives, including imaging techniques (X-rays and computed tomography) and histopathological examinations, was conducted on twenty juvenile shearwaters. The study revealed significant cranioencephalic trauma as the primary cause of death in both poaching events. Differences in the methods used by the perpetrators were noted, with variations in skull fractures and associated injuries. These findings highlight the importance of understanding local hunting techniques and sociocultural factors in forensic investigations. Diagnostic imaging, standardized necropsy, and histopathology are essential tools for the forensic investigation of illegal hunting in veterinary sciences.

Leishmania infantum is a vector-borne protozoan causing zoonotic visceral leishmaniasis in the Mediterranean basin, where dogs are the primary reservoir. However, infection in various domestic and wild mammals raises questions about their epidemiological roles. Reliable diagnosis in non-canine species remains challenging, as most serological assays are developed for dogs and cats. This study evaluated a commercially available multispecies rapid immunochromatographic test (Uranotest® Leishmania feline) for detecting anti-Leishmania antibodies in a diverse panel of domestic and non-domestic animals. A total of 186 serum samples from different species were analyzed, previously classified as seropositive (n = 65) or seronegative (n = 121) using an in-house ELISA as reference. Immunochromatographic rapid test performance was compared to ELISA results using Cohen’s kappa to assess agreement. Overall agreement reached 82.3

Cardiac arrhythmias are frequently detected during routine auscultation in athletic horses. Assessment of their clinical significance requires electrocardiography (ECG) to analyze waveforms and intervals. In recent years, portable ECG devices have been introduced to simplify electrocardiographic examination, offering advantages such as ease of use and the ability to review results via a smartphone. This study evaluated the utility and accuracy of a portable ECG device compared with a standard electrocardiograph. Simultaneous ECG recordings were obtained from 28 healthy adult horses at rest using a portable device (eKuore) and a standard electrocardiograph (Cardioline Vet600). Agreement between the two methods was assessed using Passing–Bablok regression and Bland–Altman analysis. All recordings were independently interpreted by two observers. A significant bias was observed between methods for heart rate, PR interval, and QRS duration, with higher values obtained using the portable device. Two arrhythmias were identified: second-degree atrioventricular block and atrial fibrillation. Agreement for arrhythmia detection was perfect (κ = 1.00), while agreement for heart rhythm classification (bradycardia, normal rhythm, or tachycardia) was good (κ = 0.73). In conclusion, the portable device demonstrated good diagnostic performance for arrhythmia detection during routine clinical examinations. However, further studies are required to establish reference intervals specific to this device.

Knowledge of the reference intervals for serum proteins of the European bison (Bison bonasus) can be of great help in the diagnosis of infectious and inflammatory diseases, which are very common in these animals; however, reference intervals for this species are unknown. A total of 131 serum samples were obtained from the European bison from different locations in Poland. Agarose gel electrophoresis (AGE) and capillary zone electrophoresis (CZE) were used to separate and identify protein fractions. In all sera, six fractions were identified: albumin, α1, α2, β1-, β2-, and γ-globulins by both analytical methods. Reference intervals (90% CI) were determined. Comparisons were made to find significant differences based on the sex, age or lifestyle of the animals. Concordance between the methods was assessed using Passing–Bablok regression, Bland–Altman analysis, and Lin’s concordance correlation. A higher percentage of α1-globulins was found in the females than in the males. Captive bison had significantly lower γ-globulin levels compared to free-living bison. The γ-globulin levels increased with the age of the animals. The Bland–Altman analysis demonstrated a systematic bias indicating no good agreement between methods, and only the values for albumin, β1-, β2-, γ-globulins, and total globulins were considered acceptable according to Lin’s concordance correlation. Both methods are adequate for identifying serum proteins, but they are not interchangeable, necessitating the use of separate reference intervals for each method.

Exploring the presence of endocrine disrupting compounds in marine organisms, particularly cetaceans, is crucial to evaluate contamination within the ocean's food web and the detection of synthetic hormones in cetaceans is a strong indication of anthropogenic pollution in marine ecosystems. Due to their characteristics, blubber samples are a key component in assessing the analysis of cetaceans, but this type of sample require precise and sensitive analytical methods. Despite some methodologies have been developed for the analysis of natural steroid hormones in cetacean blubber, a significant gap persists in the comprehensive analysis of synthetic steroids within these samples. In this work, a QuEChERS (Quick, Easy, Cheap, Effective, Rugged, and Safe) extraction combined with UHPLC-MS/MS (ultra-high-performance liquid chromatography with tandem mass spectrometry) analysis was performed to determine six synthetic steroid hormones (nandrolone, prednisolone, prednisone, megestrol acetate, norethisterone, and norgestrel) in 11 stranded cetacean blubber samples. Despite the complex nature of blubber samples, the developed methodology showed promising results, with recoveries ranging from 70 % to 120 % for most target compounds at low concentration levels (150 ng·g-1). The method demonstrated high precision with relative standard deviations under 20 % in both intra- and inter-day experiments. Regarding the matrix effect, ion suppressions of about 50 % were also calculated for those samples spiked at concentration levels of 150 ng·g-1, nevertheless, all compounds were still able to be detected, in most cases below quantification limits. Notably, one sample presented a quantified concentration of megestrol acetate, a steroid hormone used in contraceptives and cancer treatments, at 26.98 ± 2.62 ng·g-1. The results affirm the methodology's effectiveness for analyzing synthetic steroid hormone levels in cetacean blubber, providing a valuable tool for assessing environmental concentrations of these anthropogenic endocrine disruptors and their impact on cetacean welfare and conservation.

Bone lesions, particularly osteonecrosis, have been rarely described in cetaceans, with limited studies correlating imaging, gross and microscopic findings. Aseptic osteonecrosis is especially relevant as a chronic form of decompression sickness, well-documented in human divers but controversially reported in cetaceans. This study examined the humeral heads of five stranded short-finned pilot whales (Globicephala macrorhynchus) using computed tomography (CT) and plastic embedding histologic techniques. Three specimens exhibited CT-detectable lesions, while two did not. CT imaging revealed distinct bone lesion patterns in three animals, including one case with a hypoattenuated triangular lesion constitutes histologically by well reorganized bone tissue. This lesion is highly consistent with aseptic dysbaric osteonecrosis (DON). Circular, focal, hyperattenuated necrotic bone areas most likely related to septic lesions were detected in two affected pilot whales. The two unaffected humoral heads exhibited normal bone histology, reinforcing the pathological significance of the osteonecrotic lesions of the three affected animals. As this was a retrospective study on fixed samples, microbiological analysis was not possible. Although not conclusive, this study provides the first histological evidence of chronic humeral head lesions in cetaceans consistent with DON, underscoring the need for further research using CT imaging, histology, and microbiology.