Detection, sequencing, and tissue distribution of piscine orthoreovirus 2-like virus in diseased coho salmon in Alaska.

We performed a diagnostic disease investigation on a cohort of coho salmon (Oncorhynchus kisutch) fingerlings in Alaska exhibiting anorexia, gaping mouths, anemia, and increased mortality. Histologic examination revealed mild-to-severe myocardial degeneration and lymphohistiocytic and neutrophilic myocarditis, moderate splenic histiocytosis, and mild renal histiocytosis. Piscine orthoreoviruses 1 and 3 were not detected by molecular methods, and no other viruses could be cultured on 3 common diagnostic fish cell lines. De novo assembly produced a viral genome of 10 linear segments with >80% homology to piscine orthoreovirus 2 (PRV2) encoding all 11 PRV2 proteins. An in situ hybridization probe using RNAscope was developed against 697 viral nucleotides identified by sequencing, which revealed viral genome in heart, spleen, gill, kidney, liver, blood, and the lamina propria of the intestines. Our findings are supportive of a novel piscine orthoreovirus most closely related to PRV2 associated with morbidity and mortality of coho salmon in the northeastern Pacific.

Genomic characterization of antimicrobial resistance in 61 aquatic bacterial isolates.

Antimicrobial resistance (AMR) in pathogens important to aquatic animal health is of increasing concern but vastly understudied. Antimicrobial therapy is used to both treat and prevent bacterial disease in fish and is critical for a viable aquaculture industry and for maintenance of wild fish populations. Unfortunately, phenotypic antimicrobial susceptibility testing is technically difficult for bacteria recovered from aquatic animal hosts resulting in challenges in resistance monitoring using traditional methods. Whole-genome sequencing provides an appealing methodology for investigation of putative resistance. As part of the ongoing efforts of the FDA CVM Vet-LIRN to monitor AMR, source laboratories cultured and preliminarily identified pathogenic bacteria isolated from various fish species collected in 2019 from across the United States. Sixty-one bacterial isolates were evaluated using whole-genome sequencing. We present here the assembled draft genomes, AMR genes, predicted resistance phenotypes, and virulence factors of the 61 isolates and discuss concurrence of the identifications made by source laboratories using matrix-assisted laser desorption/time-of-flight mass spectrometry.

Facial and systemic mucormycosis caused by Lichtheimia corymbifera in a goat: case report and literature review of fungal infections in goats.

An 8-y-old Pygora doe was presented to the University of California-Davis, Veterinary Medical Teaching Hospital because of non-healing facial swelling of 2-wk duration. The lesion grew despite medical treatment, causing discomfort masticating, little-to-no airflow from the right nasal passage, and led to euthanasia. On gross examination, a large facial mass with a draining tract through the skin and hard palate was identified. On section, the mass was brown-pink, homogeneous, and friable. Abscess-like masses were identified in the lungs and kidney. Histopathology of the face, including oral and nasal cavities, salivary glands, and lymph nodes, as well as the lung and kidney lesions, revealed large areas of necrosis with numerous wide ribbon-like, mostly aseptate, fungal hyphae consistent with zygomycetes. PCR for fungal organisms performed on formalin-fixed, paraffin-embedded tissue from the face identified Lichtheimia corymbifera (formerly Absidia corymbifera) of the order Mucorales and an Aspergillus sp. The lesion was suspected to have started either as a fungal rhinitis or dental feed impaction, subsequently spreading to the face and systemically to the lungs and kidney. We describe here the lesions associated with facial mucormycosis in a goat and present a literature review of L. corymbifera infection in veterinary species and fungal infections in goats.

Investigation of laboratory methods for characterization of aquatic viruses in fish infected experimentally with infectious salmon anemia virus.

Rapid growth in aquaculture has resulted in high-density production systems in ecologically and geographically novel conditions in which the emergence of diseases is inevitable. Well-characterized methods for detection and surveillance of infectious diseases are vital for rapid identification, response, and recovery to protect economic and food security. We implemented a proof-of-concept approach for virus detection using a known high-consequence fish pathogen, infectious salmon anemia virus (ISAV), as the archetypal pathogen. In fish infected with ISAV, we integrated histopathology, virus isolation, whole-genome sequencing (WGS), electron microscopy (EM), in situ hybridization (ISH), and reverse transcription real-time PCR (RT-rtPCR). Fresh-frozen and formalin-fixed tissues were collected from virus-infected, control, and sham-infected Atlantic salmon (Salmo salar). Microscopic differences were not evident between uninfected and infected fish. Viral cytopathic effect was observed in cell cultures inoculated with fresh-frozen tissue homogenates from 3 of 3 ISAV-infected and 0 of 4 uninfected or sham-infected fish. The ISAV genome was detected by shotgun metagenomics in RNA extracted from the medium from 3 of 3 inoculated cell cultures, 3 of 3 infected fish, and 0 of 4 uninfected or sham-infected fish, yielding sufficient coverage for de novo assembly. An ISH probe against ISAV revealed ISAV genome in multiple organs, with abundance in renal hematopoietic tissue. Virus was detected by RT-rtPCR in gill, heart, kidney, liver, and spleen. EM and metagenomic WGS from tissues were challenging and unsuccessful. Our proof-of-concept methodology has promise for detection and characterization of unknown aquatic pathogens and also highlights some associated methodology challenges that require additional investigation.

ABORTION AND NEONATAL MORTALITY DUE TO TOXOPLASMA GONDII IN BIGHORN SHEEP (OVIS CANADENSIS).

Low lamb recruitment can be an obstacle to bighorn sheep (Ovis canadensis) conservation and restoration. Causes of abortion and neonate loss in bighorn sheep, which may affect recruitment, are poorly understood. Toxoplasma gondii is a major cause of abortion and stillbirth in domestic small ruminants worldwide, but no reports exist documenting abortion or neonatal death in bighorn sheep attributable to toxoplasmosis. Between March 2019 and May 2021, eight fetal and neonatal bighorn lamb cadavers from four western US states (Idaho, Montana, Nebraska, and Washington) were submitted to the Washington Animal Disease Diagnostic Laboratory for postmortem examination, histologic examination, and ancillary testing to determine the cause of abortion or neonatal death. Necrotizing encephalitis characteristic of toxoplasmosis was identified histologically in six of eight cases, and T. gondii infection was confirmed by PCR in five cases with characteristic lesions. Other lesions attributable to toxoplasmosis were pneumonia (3/5 cases) and myocarditis (2/5 cases). Protozoal cysts were identified histologically within brain, lung, heart, skeletal muscle, adipose tissue, or a combination of samples in all five sheep with PCR-confirmed T. gondii infections. Seroprevalence of T. gondii ranged from 40-81% of adult females sampled in the Washington population in October and November 2018-2021, confirming high rates of exposure before detection of Toxoplasma abortions in this study. Of 1,149 bighorn sheep postmortem samples submitted to Washington Animal Disease Diagnostic Laboratory between January 2000 and May 2021, 21 of which were from fetuses or neonates, a single case of chronic toxoplasmosis was diagnosed in one adult ewe. Recent identification of Toxoplasma abortions in bighorn sheep suggests that toxoplasmosis is an underappreciated cause of reproductive loss. Abortions and neonatal mortalities should be investigated through postmortem and histologic examination, particularly in herds that are chronically small, demographically stagnant, or exhibit reproductive rates lower than expected.

Household Transmission of SARS-CoV-2 from Humans to Pets, Washington and Idaho, USA.

SARS-CoV-2 likely emerged from an animal reservoir. However, the frequency of and risk factors for interspecies transmission remain unclear. We conducted a community-based study in Idaho, USA, of pets in households that had >1 confirmed SARS-CoV-2 infections in humans. Among 119 dogs and 57 cats, clinical signs consistent with SARS-CoV-2 were reported for 20 dogs (21%) and 19 cats (39%). Of 81 dogs and 32 cats sampled, 40% of dogs and 43% of cats were seropositive, and 5% of dogs and 8% of cats were PCR positive. This discordance might be caused by delays in sampling. Respondents commonly reported close human‒animal contact and willingness to take measures to prevent transmission to their pets. Reported preventive measures showed a slightly protective but nonsignificant trend for both illness and seropositivity in pets. Sharing of beds and bowls had slight harmful effects, reaching statistical significance for sharing bowls and seropositivity.

Non-carbapenemase producing carbapenem-resistant Klebsiella pneumoniae isolated from the urinary tract of a dog.

Objective: Carbapenems are broad-spectrum ß-lactams with excellent activity against multidrug-resistant (MDR) Enterobacterales. Unfortunately, resistance to carbapenems within this bacterial family, known as carbapenem-resistant Enterobacterales (CRE), occurs and challenges the ability to treat difficult MDR infections. Although the impact of carbapenem-resistance has been greatest in human medicine, reports in the veterinary literature are increasing especially as national veterinary antimicrobial resistance surveillance programs are now in place. In this brief communication, we report the isolation of a non-carbapenemase-producing, carbapenem-resistant Klebsiella pneumoniae from the urine of a dog, discuss the likely mechanism of resistance, and wider implications. Animal: Canine. Procedure: Whole genome sequencing and phenotypic antimicrobial susceptibility testing was performed on a K. pneumoniae isolated from the urine of a dog. Results: Antimicrobial susceptibility testing identified phenotypic resistance to imipenem and meropenem. Phenotypic detection of carbapenemase production was negative. Whole genome sequencing identified efflux pump genes associated with carbapenem resistance and point mutations in membrane porin genes. No carbapenemase gene was identified. Conclusion: Phenotypic antimicrobial susceptibility testing identified the K. pneumoniae as a non-carbapenemase producing carbapenem-resistant organism with the proposed genotypic mechanism including alteration of efflux pumps and membrane porin activity and/or expression. Clinical significance: Currently, there is limited use of carbapenem antimicrobial drugs in veterinary medicine, and practitioners may be unfamiliar or unaware of this type of resistance, its significance on routine antimicrobial susceptibility test reports, and implications for antimicrobial therapy and public health. Carbapenem-resistant Enterobacterales are infrequently isolated from companion animals; however, due to increasing adoption of advanced medical and surgical interventions, they may become more prevalent.

Objectif: Les carbapénèmes sont des ß-lactamines à large spectre avec une excellente activité contre les Enterobacterales multirésistantes (MDR). Malheureusement, la résistance aux carbapénèmes au sein de cette famille bactérienne, connue sous le nom d'Enterobacterales résistantes aux carbapénèmes (CRE), se produit et remet en question la capacité de traiter les infections MDR difficiles. Bien que l'impact de la résistance aux carbapénèmes ait été plus important en médecine humaine, les rapports dans la littérature vétérinaire se multiplient, d'autant plus que des programmes nationaux de surveillance de la résistance aux antimicrobiens vétérinaires sont désormais en place. Dans cette brève communication, nous rapportons l'isolement d'une Klebsiella pneumoniae non-productrice de carbapénémase et résistante aux carbapénèmes à partir de l'urine d'un chien, discutons du mécanisme probable de résistance et des implications plus larges. Animal: Canin. Procédure: Le séquençage du génome entier et les tests de sensibilité phénotypique aux antimicrobiens ont été effectués sur un isolat de K. pneumoniae provenant de l'urine d'un chien. Résultats: Les tests de sensibilité aux antimicrobiens ont identifié une résistance phénotypique à l'imipénème et au méropénème. La détection phénotypique de production de carbapénèmase était négative. Le séquençage du génome entier a identifié des gènes de pompe à efflux associés à la résistance aux carbapénèmes et à des mutations ponctuelles dans les gènes des porines membranaires. Aucun gène de carbapénémase n'a été identifié. Conclusion: Les tests de sensibilité phénotypique aux antimicrobiens ont identifié cet isolat de K. pneumoniae comme un organisme résistant aux carbapénèmes ne produisant pas de carbapénémase avec le mécanisme génotypique proposé, y compris l'altération des pompes à efflux et l'activité et/ou l'expression de porines membranaires. Signification clinique: Actuellement, l'utilisation des médicaments antimicrobiens à base de carbapénème en médecine vétérinaire est limitée, et les praticiens peuvent ne pas être familiers ou ne pas être au fait de ce type de résistance, de son importance dans les rapports de routine sur les tests de sensibilité aux antimicrobiens et de ses implications pour la thérapie antimicrobienne et la santé publique. Les Enterobacterales résistantes aux carbapénèmes sont rarement isolées des animaux de compagnie; cependant, en raison de l'adoption croissante d'interventions médicales et chirurgicales avancées, elles peuvent devenir plus répandues.(Traduit par Dr Serge Messier).

Household transmission of SARS-CoV-2 from humans to pets in Washington and Idaho: burden and risk factors.

SARS-CoV-2 is believed to have emerged from an animal reservoir; however, the frequency of and risk factors for inter-species transmission remain unclear. We carried out a community-based study of pets in households with one or more confirmed SARS-CoV-2 infection in humans. Among 119 dogs and 57 cats with completed surveys, clinical signs consistent with SARS-CoV-2 were reported in 20 dogs (21%) and 19 cats (39%). Out of 81 dogs and 32 cats sampled for testing, 40% of dogs and 43% of cats were seropositive, and 5% of dogs and 8% of cats were PCR positive; this discordance may be due to delays in sampling. Respondents commonly reported close human-animal contact and willingness to take measures to prevent transmission to their pets. Reported preventative measures showed a slightly protective trend for both illness and seropositivity in pets, while sharing of beds and bowls had slight harmful effects.

An outbreak of SARS-CoV-2 with high mortality in mink (Neovison vison) on multiple Utah farms.

The breadth of animal hosts that are susceptible to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and may serve as reservoirs for continued viral transmission are not known entirely. In August 2020, an outbreak of SARS-CoV-2 occurred on five mink farms in Utah and was associated with high mink mortality (35-55% of adult mink) and rapid viral transmission between animals. The premise and clinical disease information, pathology, molecular characterization, and tissue distribution of virus within infected mink during the early phase of the outbreak are provided. Infection spread rapidly between independently housed animals and farms, and caused severe respiratory disease and death. Disease indicators were most notably sudden death, anorexia, and increased respiratory effort. Gross pathology examination revealed severe pulmonary congestion and edema. Microscopically there was pulmonary edema with moderate vasculitis, perivasculitis, and fibrinous interstitial pneumonia. Reverse transcriptase polymerase chain reaction (RT-PCR) of tissues collected at necropsy demonstrated the presence of SARS-CoV-2 viral RNA in multiple organs including nasal turbinates, lung, tracheobronchial lymph node, epithelial surfaces, and others. Localization of viral RNA by in situ hybridization revealed a more localized infection, particularly of the upper respiratory tract. Whole genome sequencing from multiple mink was consistent with published SARS-CoV-2 genomes with few polymorphisms. The Utah mink SARS-CoV-2 strains fell into Clade GH, which is unique among mink and other animal strains sequenced to date. While sharing the N501T mutation which is common in mink, the Utah strains did not share other spike RBD mutations Y453F and F486L found in nearly all mink from the United States. Mink in the outbreak reported herein had high levels of SARS-CoV-2 in the upper respiratory tract associated with symptomatic respiratory disease and death.

Detection of Yersinia ruckeri in Pacific Lamprey (Entosphenus tridentatus) on the Olympic Peninsula in Washington, USA.

Pacific lamprey (Entosphenus tridentatus) are important anadromous fish throughout their range in western North America. As conservation programs for lamprey expand, disease surveillance is becoming more prevalent. During routine surveillance, Yersinia ruckeri biotype II was isolated from Pacific lamprey. This is the first documented Y. ruckeri detection in Pacific lamprey.

Fatal Meningoencephalitis From a Rhizomucor pusillus Infection in a Juvenile Magellanic Penguin (Spheniscus magellanicus).

A 7-month-old, male Magellanic penguin (Spheniscus magellanicus), housed in an outdoor exhibit, developed acute neurologic signs that progressed to death over 2 days. On gross examination, the bird had congested, edematous lungs, and cerebellar hemorrhage. Histologic examination identified granulomatous pneumonia and encephalitis, with thrombosis and eosinophilic, branching fungal hyphae that had invaded the meningeal vessel walls. Polymerase chain reaction identified the fungus in the brain as Rhizomucor pusillus, an uncommon cause of mucormycosis. This organism has previously been reported in respiratory, skeletal, and sino-orbital lesions of avian species. This clinical report describes meningoencephalitis associated with Rhizomucor pusillus in a penguin.

Retrospective Review of Mycobacterial Conjunctivitis in Cockatiels (Nymphicus hollandicus).

The etiologic disease organism responsible for causing mycobacteriosis in avian species is an acid-fast gram-positive bacterium. This bacterium causes granulomatous disease in various internal organs, but in cockatiels (Nymphicus hollandicus) it has been commonly identified within the conjunctival tissues. Twenty-six cases of mycobacterial conjunctivitis in cockatiels were diagnosed through histopathologic assessment of diseased tissue samples, Fite acid-fast staining, and polymerase chain reaction in this retrospective study. Clinicians who saw these cases were contacted, and information was obtained regarding recommended treatment protocols prescribed for the patients, the Mycobacterium species identified, and case outcomes. All patients in this retrospective study had a biopsy performed on the affected conjunctival tissue, and because of the small size of the patients, this excisional biopsy removed the affected tissue in its entirety or significantly debulked the lesion. Of the 26 cases, 10 were lost to follow-up, 4 were euthanatized, 7 died, and 5 were alive at the time this information was submitted for publication.

Algal Lymphadenitis in a Dog Caused by Scenedesmus Species.

A 6-year-old, spayed female Labrador/Weimaraner cross-breed dog that had previously lived in Arizona presented in Montana for an annual examination with an incidentally enlarged popliteal lymph node, which was subsequently biopsied. Histologically, the lymph node was expanded by eosinophil-rich granulomas with both extracellular and intrahistiocytic green algae. These algae had intracytoplasmic, birefringent, and refractile granules; readily formed 2 to 3 mm green colonies on Columbia blood agar medium; and ultrastructurally had a multilayered cell wall and intracytoplasmic chloroplasts. Amplified product from the internal transcribed spacer and D1/D2 regions of the 28S ribosomal RNA gene had high sequence identity to Scenedesmus sp. Despite similar infection in the retropharyngeal lymph node 1 year later, the animal remained otherwise healthy with no clinical signs. To the authors' knowledge, this is the first case of Scenedesmus species infection in a dog and is a differential diagnosis for Coccidioides immitis.

Sarcocystis calchasi Outbreak in Feral Rock Pigeons ( Columba livia) in California.

Twenty-two feral rock pigeons ( Columba livia) from 10 counties in California with ataxia, torticollis, and difficulty standing and flying were admitted to rehabilitation centers in late winter and spring of 2017 and died or were euthanized. Common necropsy findings included thin body condition, generalized deep red discoloration of organs, and hemorrhagic subcutaneous neck tissues. Meningoencephalitis was observed microscopically in 16 pigeons, and 3 also had protozoal schizonts in the brain. The most consistently affected regions of the brain were cerebellum and brainstem. Skeletal muscles, and less frequently the heart, contained large intrasarcoplasmic bradyzoites typically without inflammation. Fifteen of the 22 birds tested positive using pan- Sarcocystis polymerase chain reaction. The sequence of the amplicon was most closely related to S. calchasi, and the 8 subtyped sequences had 100% homology with S. calchasi. This investigation demonstrated the transcontinental and North American spread of S. calchasi causing a disease outbreak in free-ranging rock pigeons, thus warranting increased surveillance in susceptible native columbids.

Disseminated mycobacteriosis caused by Mycobacterium kansasii in a pot-bellied pig.

A 1.5-y-old spayed female Juliana pot-bellied pig was presented to the Purdue University Veterinary Teaching Hospital with a history of wasting and anorexia. Enlarged and partially mineralized lymph nodes were identified on radiographs and computed tomography scan. Generalized lymphadenomegaly and disseminated nodules in the lungs, liver, spleen, and kidneys were identified on postmortem examination. Histologic examination revealed caseonecrotic granulomas with numerous intracellular, acid-fast bacilli. Mycobacterium kansasii type II was identified as the etiologic agent by PCR amplification using universal Mycobacterium primers, direct sequencing of the PCR amplicon, and comparison to sequences in GenBank. We describe a case in a pot-bellied pig of mycobacteriosis caused by an atypical mycobacterial species and highlight the important role of laboratory testing in suspected cases of tuberculosis.