Activation of T-bet, FOXP3, and EOMES in Target Organs From Piglets Infected With the Virulent PRRSV-1 Lena Strain

Transcription factors (TFs) modulate genes involved in cell-type-specific proliferative and migratory properties, metabolic features, and effector functions. Porcine reproductive and respiratory syndrome virus (PRRSV) is one of the most important pathogen agents in the porcine industry; however, TFs have been poorly studied during the course of this disease. Therefore, we aimed to evaluate the expressions of the TFs T-bet, GATA3, FOXP3, and Eomesodermin (EOMES) in target organs (the lung, tracheobronchial lymph node, and thymus) and those of different effector cytokines (IFNG, TNFA, and IL10) and the Fas ligand (FASL) during the early phase of infection with PRRSV-1 strains of different virulence. Target organs from mock-, virulent Lena-, and low virulent 3249-infected animals humanely euthanized at 1, 3, 6, 8, and 13 days post-infection (dpi) were collected to analyze the PRRSV viral load, histopathological lesions, and relative quantification through reverse transcription quantitative PCR (RT-qPCR) of the TFs and cytokines. Animals belonging to both infected groups, but mainly those infected with the virulent Lena strain, showed upregulation of the TFs T-bet, EOMES, and FOXP3, together with an increase of the cytokine IFN-γ in target organs at the end of the study (approximately 2 weeks post-infection). These results are suggestive of a stronger polarization to Th1 cells and regulatory T cells (Tregs), but also CD4+ cytotoxic T lymphocytes (CTLs), effector CD8+ T cells, and γδT cells in virulent PRRSV-1-infected animals; however, their biological functionality should be the object of further studies.

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.

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 in multiple mink farms in Utah and was associated with high mink mortality and rapid viral transmission between animals. The outbreak's epidemiology, pathology, molecular characterization, and tissue distribution of virus within infected mink is provided. Infection of mink was likely by reverse zoonosis. Once established, infection spread rapidly between independently housed animals and farms, and caused severe respiratory disease and death. Clinical signs 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. Whole genome sequencing from multiple mink was consistent with published SARS-CoV-2 genomes with few polymorphisms. The Utah mink SARS-CoV-2 strain fell into Clade GH, which is unique among mink and other animal strains sequenced to date and did not share other spike RBD mutations Y453F and F486L found in mink. Localization of viral RNA by in situ hybridization revealed a more localized infection, particularly of the upper respiratory tract. Mink in the outbreak reported herein had high levels of virus in the upper respiratory tract associated with mink-to-mink transmission in a confined housing environment and were particularly susceptible to disease and death due to SARS-CoV-2 infection.

Experimental Inoculation of Young Calves with SARS-CoV-2

The host range of SARS-CoV-2 and the susceptibility of animal species to the virus are topics of great interest to the international scientific community. The angiotensin I converting enzyme 2 (ACE2) protein is the major receptor for the virus, and sequence and structural analysis of the protein has been performed to determine its cross-species conservation. Based on these analyses, cattle have been implicated as a potential susceptible species to SARS-CoV-2 and have been reported to have increased ACE2 receptor distribution in the liver and kidney, and lower levels in the lungs. The goal of the current study was to determine the susceptibility of cattle to SARS-CoV-2 utilizing inoculation routes that facilitated exposure to tissues with increased ACE2 receptor distribution. For this, colostrum-deprived calves approximately 6 weeks of age were inoculated via the intratracheal or intravenous routes. Nasal and rectal swab samples, as well as blood and urine samples, were collected over the course of the study to evaluate viral shedding, viremia, and seroconversion. Pyrexia was used as the primary criteria for euthanasia and tissue samples were collected during necropsy. Importantly, SARS-CoV-2 RNA was detected in only two nasal swab samples collected on days 3 and 10 post-inoculation (pi) in two calves; one calf in the intratracheal group and the other calf in the intravenous group, respectively. Additionally, the calf in the intratracheal group that was positive on the nasal swab on day 3 pi also had a positive tracheobronchial lymph node on day 9 pi. Viral nucleic acid load on these samples, based on PCR cycle threshold values, were low and infectious virus was not recovered from the samples. These results suggest that there was no productive replication of SARS-CoV-2 in calves following intratracheal and intravenous inoculation.

Metastatic renal interstitial cell tumor in a dog

Renal interstitial cell tumor (RICT) is a rare renal sarcoma of dogs that arises from renal interstitial cells. Herein we describe a RICT in an 8-y-old female Labrador Retriever dog that died after a 2-d history of lethargy and disorientation. Grossly, soft white nodules of 1–10 mm diameter were present in the renal cortex and corticomedullary junction of both kidneys, left cardiac ventricular wall, and right cerebral hemisphere. A pale-white to yellow, firm, irregular mass effaced 80% of the right pulmonary parenchyma, involving mainly the cranial and middle lobes, and the adjacent tracheobronchial lymph nodes. Histologically, the renal, myocardial, and cerebral neoplasm consisted of interlacing bundles of stellate-to-spindle cells with eosinophilic vacuolated cytoplasm and round-to-oval nuclei with finely stippled chromatin. The mitotic count was 28 per 2.37 mm2. Alcian blue stain revealed an extracellular myxomatous matrix throughout the neoplasm. Neoplastic cells had cytoplasmic immunolabeling for vimentin and cyclooxygenase 2. The pulmonary and tracheobronchial neoplasm consisted of infiltrative nodules of cuboidal epithelial cells that had a moderate amount of eosinophilic cytoplasm and round nuclei with coarsely stippled chromatin. There were 5 mitoses per 2.37 mm2. Neoplastic cells had cytoplasmic and nuclear immunolabeling for cytokeratin AE1/AE3 and thyroid transcription factor 1, respectively. Morphologic and immunohistochemical findings were consistent with a RICT with cardiac and cerebral metastases, and a pulmonary carcinoma with tracheobronchial lymph node metastasis.

Effects of Pseudorabies Virus Infection on the Tracheobronchial Lymph Node Transcriptome

This study represents the first swine transcriptome hive plots created from gene set enrichment analysis (GSEA) data and provides a novel insight into the global transcriptome changes occurring in tracheobronchial lymph nodes (TBLN) and spanning the swine genome. RNA isolated from draining TBLN from 5-week-old pigs, either clinically infected with a feral isolate of Pseudorabies virus or uninfected, was interrogated using Illumina Digital Gene Expression Tag Profiling. More than 100 million tag sequences were observed, representing 4,064,189 unique 21-base sequences collected from TBLN at time points 1, 3, 6, and 14 days post-inoculation (dpi). Multidimensional statistical tests were applied to determine the significant changes in tag abundance, and then the tags were annotated. Hive plots were created to visualize the differential expression within the swine transcriptome defined by the Broad Institute's GSEA reference datasets between infected and uninfected animals, allowing us to directly compare different conditions.