Mortality associated with SARS-CoV-2 in nondomestic felids.

Between September and November 2021, 5 snow leopards (Panthera uncia) and 1 lion (Panthera leo) were naturally infected with severe acute respiratory coronavirus 2 (SARS-CoV-2) and developed progressive respiratory disease that resulted in death. Severe acute respiratory syndrome coronavirus 2 sequencing identified the delta variant in all cases sequenced, which was the predominant human variant at that time. The time between initial clinical signs and death ranged from 3 to 45 days. Gross lesions in all 6 cats included nasal turbinate hyperemia with purulent discharge and marked pulmonary edema. Ulcerative tracheitis and bronchitis were noted in 4 cases. Histologically, there was necrotizing and ulcerative rhinotracheitis and bronchitis with fibrinocellular exudates and fibrinosuppurative to pyogranulomatous bronchopneumonia. The 4 cats that survived longer than 8 days had fungal abscesses. Concurrent bacteria were noted in 4 cases, including those with more acute disease courses. Severe acute respiratory syndrome coronavirus 2 was detected by in situ hybridization using probes against SARS-CoV-2 spike and nucleocapsid genes and by immunohistochemistry. Viral nucleic acid and protein were variably localized to mucosal and glandular epithelial cells, pneumocytes, macrophages, and fibrinocellular debris. Based on established criteria, SARS-CoV-2 was considered a contributing cause of death in all 6 cats. While mild clinical infections are more common, these findings suggest that some SARS-CoV-2 variants may cause more severe disease and that snow leopards may be more severely affected than other felids.

Loop-mediated isothermal amplification (LAMP) assay for specific and rapid detection of Dickeya fangzhongdai targeting a unique genomic region.

Dickeya fangzhongdai, a bacterial pathogen of taro (Colocasia esculenta), onion (Allium sp.), and several species in the orchid family (Orchidaceae) causes soft rot and bleeding canker diseases. No field-deployable diagnostic tool is available for specific detection of this pathogen in different plant tissues. Therefore, we developed a field-deployable loop-mediated isothermal amplification (LAMP) assay using a unique genomic region, present exclusively in D. fangzhongdai. Multiple genomes of D. fangzhongdai, and other species of Dickeya, Pectobacterium and unrelated genera were used for comparative genomic analyses to identify an exclusive and conserved target sequence from the major facilitator superfamily (MFS) transporter gene region. This gene region had broad detection capability for D. fangzhongdai and thus was used to design primers for endpoint PCR and LAMP assays. In-silico validation showed high specificity with D. fangzhongdai genome sequences available in the NCBI GenBank genome database as well as the in-house sequenced genome. The specificity of the LAMP assay was determined with 96 strains that included all Dickeya species and Pectobacterium species as well as other closely related genera and 5 hosts; no false positives or false negatives were detected. The detection limit of the assay was determined by performing four sensitivity assays with tenfold serially diluted purified genomic DNA of D. fangzhongdai with and without the presence of crude host extract (taro, orchid, and onion). The detection limit for all sensitivity assays was 100 fg (18-20 genome copies) with no negative interference by host crude extracts. The assays were performed by five independent operators (blind test) and on three instruments (Rotor-Gene, thermocycler and dry bath); the assay results were concordant. The assay consistently detected the target pathogen from artificially inoculated and naturally infected host samples. The developed assay is highly specific for D. fangzhongdai and has applications in routine diagnostics, phytosanitary and seed certification programs, and epidemiological studies.

RNA-Sequencing based analysis of bovine endometrium during the maternal recognition of pregnancy.

BACKGROUND: Maternal recognition is the crucial step for establishing pregnancy in cattle. This study aims to identify endometrial genes and biological pathways involved in the maternal recognition of pregnancy. Caruncular endometrial tissues were collected from Day 15-17 of gestation (pregnant), non-pregnant (absence of conceptus), and cyclic (non-bred) heifers. RESULTS: Total RNAs were isolated from the caruncular endometrial tissues of pregnant, non-pregnant, and cyclic heifers, and were subjected to high-throughput RNA-sequencing. The genes with at least two-fold change and Benjamini and Hochberg p-value ≤ 0.05 were considered differentially expressed genes and further confirmed with quantitative real-time PCR. A total of 107 genes (pregnant vs cyclic) and 98 genes (pregnant vs non-pregnant) were differentially expressed in the pregnant endometrium. The most highly up-regulated genes in the pregnant endometrium were MRS2, CST6, FOS, VLDLR, ISG15, IFI6, MX2, C15H11ORF34, EIF3M, PRSS22, MS4A8, and TINAGL1. Interferon signaling, immune response, nutrient transporter, synthesis, and secretion of proteins are crucial pathways during the maternal recognition of pregnancy. CONCLUSIONS: The study demonstrated that the presence of conceptus at Day 15-17 of gestation affects the endometrial gene expression related to endometrial remodeling, immune response, nutrients and ion transporters, and relevant signaling pathways in the caruncular region of bovine endometrium during the maternal recognition of pregnancy.

Genome-informed loop-mediated isothermal amplification assay for specific detection of Pectobacterium parmentieri in infected potato tissues and soil.

Pectobacterium parmentieri (formerly Pectobacterium wasabiae), which causes soft rot disease in potatoes, is a newly established species of pectinolytic bacteria within the family Pectobacteriaceae. Despite serious damage caused to the potato industry worldwide, no field-deployable diagnostic tests are available to detect the pathogen in plant samples. In this study, we aimed to develop a reliable, rapid, field-deployable loop-mediated isothermal amplification (LAMP) assay for the specific detection of P. parmentieri. Specific LAMP primers targeting the petF1 gene region, found in P. parmentieri but no other Pectobacterium spp., were designed and validated in silico and in vitro using extensive inclusivity (15 strains of P. parmentieri) and exclusivity (94 strains including all other species in the genus Pectobacterium and host DNA) panels. No false positives or negatives were detected when the assay was tested directly with bacterial colonies, and with infected plant and soil samples. Sensitivity (analytical) assays using serially diluted bacterial cell lysate and purified genomic DNA established the detection limit at 10 CFU/mL and 100 fg (18-20 genome copies), respectively, even in the presence of host crude DNA. Consistent results obtained by multiple users/operators and field tests suggest the assay's applicability to routine diagnostics, seed certification programs, biosecurity, and epidemiological studies.

First Documented Cases of Canine Neuroangiostrongyliasis Due to Angiostrongylus cantonensis in Hawaii.

Two young dogs domiciled in Honolulu, Hawaii, were presented in November and December 2018 (respectively) for spinal hyperesthesia, hindlimb weakness, and proprioceptive ataxia. Both dogs had neurologic findings referable to spinal cord disease. Both dogs had a combination of lower motor neuron signs (reduced muscle mass, decreased withdrawal reflexes, low tail carriage) and long tract signs (conscious proprioceptive deficits, crossed extensor response, increased myotatic reflexes). Peripheral eosinophilia was present in the second case, but hematology and serum biochemistries were otherwise unremarkable. Plain radiographs and computed tomography scans ± contrast were unremarkable. Cerebrospinal fluid (CSF) from both patients demonstrated eosinophilic pleocytosis, and real-time polymerase chain reaction testing demonstrated Angiostrongylus cantonensis deoxyribonucleic acid in CSF, confirming a diagnosis of neuroangiostrongyliasis. Treatment included glucocorticoid therapy, ± anthelmintic (fenbendazole). Both dogs made a complete recovery. These are the first confirmed cases of autochthonous neuroangiostrongyliasis in canine patients in the United States and the first dogs anywhere to be diagnosed definitively with A cantonensis infection based on real-time polymerase chain reaction testing of CSF. A clinician examining a patient with severe spinal hyperesthesia and a combination of upper and lower motor signs should consider A cantonensis as a differential, especially in endemic areas.

Causes of mortality in backyard poultry in eight states in the United States.

A comprehensive understanding of common diseases of backyard poultry flocks is important to providing poultry health information to flock owners, veterinarians, and animal health officials. We collected autopsy reports over a 3-y period (2015-2017) from diagnostic laboratories in 8 states in the United States; 2,509 reports were collected, involving autopsies of 2,687 birds. The primary cause of mortality was categorized as infectious, noninfectious, neoplasia or lymphoproliferative disease, or undetermined. Neoplasia or lymphoproliferative disease was the most common primary diagnosis and involved 42% of the total birds autopsied; 63% of these cases were diagnosed as Marek's disease or leukosis/sarcoma. Bacterial, parasitic, and viral organisms were commonly detected, involving 42%, 28%, and 7% of the birds autopsied, respectively, with 2 or more organisms detected in 69% of birds. Our findings demonstrate the importance of educating flock owners about disease prevention and biosecurity practices. The detection of zoonotic bacteria including paratyphoid salmonellae, Campylobacter spp., Listeria monocytogenes, and Mycobacterium avium, and the detection of lead and other heavy metals, indicate public health risks to flock owners and consumers of backyard flock egg and meat products.

Malignant edema in postpartum dairy cattle.

Five cases of postparturient vulvovaginitis and metritis in cattle caused by Clostridium septicum (malignant edema) are described in the current report. The diagnosis was established based on detection of C. septicum by culture and fluorescent antibody test. All animals were Holsteins, and 4 were primiparous (the parity of 1 animal was not reported). All animals developed clinical signs 1-3 days after calving, consisting of swelling of perineal and perivulvar areas, fever, and depression. Perineal, perivulvar, and perivaginal gelatinous and often hemorrhagic edema was consistently observed on gross examination. Longitudinal vulvar, vaginal, cervical, and uterine body tears, covered by fibrinous exudates, were also present. Microscopically, vulvar, vaginal, and uterine mucosae were multifocally necrotic and ulcerated. Large Gram-positive rods, some with subterminal spores, were present within the edematous subcutaneous and submucosal tissues. Clostridium septicum was demonstrated by culture and/or fluorescent antibody test in tissues of most animals. These cases of malignant edema were considered to be produced by C. septicum and predisposed by the trauma occurring during parturition.

Notoedric mange in two free-ranging mountain lions (Puma concolor).

Two mountain lions (Puma con-color) were found dead in the Simi Hills area of southern California (Ventura County). Postmortem examination and toxicological analyses indicated that the cause of death was anticoagulant rodenticide intoxication. In addition, both lions had marked alopecia and skin crusts, caused by Notoedres cati. The diagnosis of notoedric mange was confirmed by histology and direct examination of mites obtained from skin scrapings of the two animals. Histologically, the affected skin showed acanthotic epidermis with parakeratosis and parasitic tunnels in the stratum corneum. This is the first report of the pathological changes associated with notoedric mange in free-ranging mountain lions.

Association between findings on palmarodorsal radiographic images and detection of a fracture in the proximal sesamoid bones of forelimbs obtained from cadavers of racing Thoroughbreds.

OBJECTIVE: To determine the distribution for limbs and bones in horses with fractures of the proximal sesamoid bones and relationships with findings on palmarodorsal radiographic images. SAMPLE POPULATION: Proximal sesamoid bones obtained from both forelimbs of cadavers of 328 racing Thoroughbreds. PROCEDURE: Osteophytes; large vascular channels; and fracture location, orientation, configuration, and margin distinctness were categorized by use of high-detail contact palmarodorsal radiographs. Distributions of findings were determined. Relationships between radiographic findings and fracture characteristics were examined by use of chi2 and logistic regression techniques. RESULTS: Fractures were detected in 136 (41.5%) horses. Biaxial fractures were evident in 109 (80%) horses with a fracture. Osteophytes and large vascular channels were evident in 266 (81%) and 325 (99%) horses, respectively. Medial bones typically had complete transverse or split transverse simple fractures, indistinct fracture margins, > 1 vascular channel that was > 1 mm in width, and osteophytes in abaxial wing and basilar middle or basilar abaxial locations. Lateral bones typically had an oblique fracture and distinct fracture margins. Odds of proximal sesamoid bone fracture were approximately 2 to 5 times higher in bones without radiographic evidence of osteophytes or large vascular channels, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: Biaxial fractures of proximal sesamoid bones were common in cadavers of racing Thoroughbreds. Differences between medial and lateral bones for characteristics associated with fracture may relate to differences in fracture pathogeneses for these bones. Osteophytes and vascular channels were common findings; however, fractures were less likely to occur in bones with these features.

Abortion and ulcerative posthitis associated with caprine herpesvirus-1 infection in goats in California.

Three outbreaks of late-gestation abortions in does and ulcerative posthitis in bucks, associated with caprine herpes virus-1 (CHV-1), in California are described. In herd A, 10 of 17 does aborted in a 7-day period, whereas in herd B, 4 of 130 does aborted in a 45-day period and in herd C, 100 of 300 does aborted in a 3-week period. Most fetuses had multifocal pinpoint depressed foci with a zone of hyperemia on external and cut surfaces of the kidneys, liver, lungs, and adrenal glands. Histologically, scattered multifocal areas of necrosis with mild neutrophilic infiltrate were observed in kidneys, brain, liver, adrenal glands, and lungs of most fetuses of the 3 herds. Large amphophilic intranuclear inclusion bodies, which displaced the chromatin, were observed in cells within and around the necrotic foci in kidneys and adrenal glands. Particles 85-113 nm in size with morphology compatible with herpes virus were observed in the nuclei of these cells when examined by electron microscopy. Irregular, shallow, red ulcers were observed in the prepuce of 1 buck from herd C. Prepuce biopsies from this animal had necrosis of the superficial mucosal epithelium and severe submucosal lymphoplasmocytic infiltrates. Large intranuclear amphophilic inclusion bodies were observed in most cells of the stratum spinosum of the preputial epithelium, but no viral particles were observed in these cells. Caprine herpes virus-1 was isolated from tissue pools of fetuses from the 3 herds but not from prepuce biopsies. Positive results were obtained when tissues of a fetus from herd C were processed by a polymerase chain reaction technique to amplify the amino terminus of the glycoprotein C gene of CHV-1. Sera from aborted does from herds B and C and from the 3 bucks from herd C had high antibody titers to CHV-1. The results presented here support the hypothesis that the male goat is involved in the transmission of CHV-1. However, other forms of transmission cannot be ruled out.