Retrospective study of laboratory-based surveillance of rabies in wild and domestic animals in the southern United States, 2010-2021.

We performed a retrospective study of all case submissions for the rabies virus (RABV) direct fluorescent antibody test (DFAT) requested of the Tifton Veterinary Diagnostic and Investigational Laboratory (Tifton, GA, USA) between July 2010 and June 2021. Submitted were 792 samples from 23 animal species from 89 counties in Georgia, and 4 neighboring counties in Florida, 1 in South Carolina, and 1 in Alabama. In 13 (1.6%) cases, the DFAT result was inconclusive; 779 (98.4%) cases had a conclusive (positive or negative) test result. Of these 779 cases, 79 (10.1%) tested positive across 10 species. The remaining 700 (89.9%) cases were negative. The main reason for submission for RABV testing was human exposure to a potentially rabid animal in 414 (52.3%) cases. Among the 79 positive cases, 74 (93.7%) involved wildlife; raccoons (51 cases; 68.9%) were the primary host confirmed with RABV infection, followed by skunk and fox (8 cases each; 10.8%), bobcat (5 cases; 6.8%), and bats (2 cases; 2.7%). Only 5 domestic animals (6.3% of the positive cases) tested positive during our study period; one from each of the bovine, canine, caprine, equine, and feline species. Hence, the sylvatic cycle plays the predominant role in circulating RABV infection in our study area.

Predominance of Canine Parainfluenza Virus and Mycoplasma in Canine Infectious Respiratory Disease Complex in Dogs.

Canine infectious respiratory disease complex (CIRDC) is caused by different viruses and bacteria. Viruses associated with CIRDC include canine adenovirus type 2 (CAV-2), canine distemper virus (CDV), canine influenza virus (CIV), canine herpesvirus type 1 (CHV-1), canine respiratory coronavirus (CRCoV), and canine parainfluenza virus (CPIV). Bacteria associated with CIRDC include Bordetella bronchiseptica, Streptococcus equi subspecies zooepidemicus (S. zooepidemicus), and Mycoplasma spp. The present study examined the prevalence of CIRDC pathogens in specimens received by a Veterinary Diagnostic Laboratory in Georgia, USA., from 2018 to 2022. Out of 459 cases, viral agents were detected in 34% of cases and bacterial agents were detected in 58% of cases. A single pathogen was detected in 31% of cases, while two or more pathogens were identified in 24% of cases. The percentages of viral agents identified were CAV-2 (4%), CDV (3%), CPIV (16%), CRCoV (7%), and CIV (2%). The percentages of bacterial agents were B. bronchiseptica (10%), Mycoplasma canis (24%), Mycoplasma cynos (21%), and S. zooepidemicus (2%). Over the five-year period, the positive cases ranged from 2-4% for CAV-2, 1-7% for CDV, 1-4% for CHV-1, 9-22% for CPIV, 4-13% for CRCoV, and 1-4% for CIV. Overall, the most prevalent pathogens associated with CIRDC were CPIV, M. canis, and M. cynos.

Rapid Detection of SARS-CoV-2 RNA in Human Nasopharyngeal Specimens Using Surface-Enhanced Raman Spectroscopy and Deep Learning Algorithms.

A rapid and cost-effective method to detect the infection of SARS-CoV-2 is fundamental to mitigating the current COVID-19 pandemic. Herein, a surface-enhanced Raman spectroscopy (SERS) sensor with a deep learning algorithm has been developed for the rapid detection of SARS-CoV-2 RNA in human nasopharyngeal swab (HNS) specimens. The SERS sensor was prepared using a silver nanorod array (AgNR) substrate by assembling DNA probes to capture SARS-CoV-2 RNA. The SERS spectra of HNS specimens were collected after RNA hybridization, and the corresponding SERS peaks were identified. The RNA detection range was determined to be 103-109 copies/mL in saline sodium citrate buffer. A recurrent neural network (RNN)-based deep learning model was developed to classify 40 positive and 120 negative specimens with an overall accuracy of 98.9%. For the blind test of 72 specimens, the RNN model gave a 97.2% accuracy prediction for positive specimens and a 100% accuracy for negative specimens. All the detections were performed in 25 min. These results suggest that the DNA-functionalized AgNR array SERS sensor combined with a deep learning algorithm could serve as a potential rapid point-of-care COVID-19 diagnostic platform.

Point-of-care testing in companion and food animal disease diagnostics.

Laboratory diagnoses of animal diseases has advanced tremendously in recent decades with the advent of cutting-edge technologies such as real-time polymerase chain reaction, next generation sequencing (NGS), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and others However, most of these technologies need sophisticated equipment, laboratory space and highly skilled workforce. Therefore, there is an increasing market demand for point-of-care testing (POCT) in animal health and disease diagnostics. A wide variety of assays based on antibodies, antigens, nucleic acid, and nanopore sequencing are currently available. Each one of these tests have their own advantages and disadvantages. However, a number of research and developmental activities are underway in both academia and industry to improve the existing tests and develop newer and better tests in terms of sensitivity, specificity, turnaround time and affordability. In both companion and food animal disease diagnostics, POCT has an increasing role to play, especially in resource-limited settings. It plays a critical role in improving animal health and wellbeing in rural communities in low- and middle-income countries. At the same time, ensuring high standard of quality through proper validation, quality assurance and regulation of these assays are very important for accurate diagnosis, surveillance, control and management of animal diseases. This review addresses the different types of POCTs currently available for companion and food animal disease diagnostics, tests in the pipeline and their advantages and disadvantages.

Isothermal amplification using sequence-specific fluorescence detection of SARS coronavirus 2 and variants in nasal swabs.

Coronavirus disease 2019 is a public health challenge requiring rapid testing for the detection of infections and transmission. Nucleic acid amplification tests targeting SARS coronavirus 2 (CoV2) are used to detect CoV2 in clinical samples. Real-time reverse transcription quantitative PCR is the standard nucleic acid amplification test for CoV2, although reverse transcription loop-mediated isothermal amplification is used in diagnostics. The authors demonstrate a sequence-specific reverse transcription loop-mediated isothermal amplification-based nucleic acid amplification assay that is finished within 30 min using minimally processed clinical nasal swab samples and describe a fluorescence-quenched reverse transcription loop-mediated isothermal amplification assay using labeled primers and a quencher oligonucleotide. This assay can achieve rapid (30 min) and sensitive (1000 plaque-forming units/ml) fluorescence detection of CoV2 (WA1/2020), B.1.1.7 (Alpha) and variants of concern Delta (B.1.617.2) and Omicron (B.1.1.529) in nasal samples.

Determining the role of natural SARS-CoV-2 infection in the death of domestic pets: 10 cases (2020-2021).

OBJECTIVE: To establish a pathoepidemiological model to evaluate the role of SARS-CoV-2 infection in the first 10 companion animals that died while infected with SARS-CoV-2 in the US. ANIMALS: 10 cats and dogs that tested positive for SARS-CoV-2 and died or were euthanized in the US between March 2020 and January 2021. PROCEDURES: A standardized algorithm was developed to direct case investigations, determine the necessity of certain diagnostic procedures, and evaluate the role, if any, that SARS-CoV-2 infection played in the animals' course of disease and death. Using clinical and diagnostic information collected by state animal health officials, state public health veterinarians, and other state and local partners, this algorithm was applied to each animal case. RESULTS: SARS-CoV-2 was an incidental finding in 8 animals, was suspected to have contributed to the severity of clinical signs leading to euthanasia in 1 dog, and was the primary reason for death for 1 cat. CONCLUSIONS AND CLINICAL RELEVANCE: This report provides the global community with a standardized process for directing case investigations, determining the necessity of certain diagnostic procedures, and determining the clinical significance of SARS-CoV-2 infections in animals with fatal outcomes and provides evidence that SARS-CoV-2 can, in rare circumstances, cause or contribute to death in pets.

Complete Genome Sequences of Six Staphylococcus pseudintermedius Strains from Dogs with Superficial Pyoderma in Georgia, USA.

Staphylococcus pseudintermedius is a pathogen of veterinary importance, as it is the major causative agent of superficial pyoderma in dogs. We present the complete genome sequences of six strains of S. pseudintermedius derived from dogs affected with epidermal collarettes and superficial bacterial folliculitis, which are two variants of superficial pyoderma.

A milk-line sampling system to detect foodborne pathogens: A field case investigation from the United States and Argentina.

The objective of this short communication was to discuss two field case investigations to determine the usefulness of a milk-line sampling device to detect bacteria either coming from a group of cows suffering from mastitis or from the milking line potentially contaminated with environmental bacteria. In Case 1, the in-line sampling device was able to detect certain segments of the milk-line contaminated with environmental bacteria, but not coming from the cows. In Case 2, 19 out of 25 pooled in-line samples were in agreement with at least one of the individual sampled cows shedding either Staphylococcus or Streptococcus spp. or both, which accounted for 76% accordance between both methods. The in-line system, although not perfect, provided a reliable method to detect individual cows shedding mastitis-causing organisms. In conclusion, the milk-line sampling device system was able to help identify foodborne pathogens. Regular monitoring of the microbial quality of milk through a milk-line sampling device is recommended for groups of cows within the dairy herd to detect potential mastitis-causing microorganisms. Furthermore, the sampling device was an effective tool to screen the efficacy of cleaning and disinfecting mechanisms of the milk lines to identify and control potential foodborne pathogens that are collected in the bulk tank.

Systemic humoral immunity in beef bulls following therapeutic vaccination against Tritrichomonas foetus.

The utility of therapeutic vaccination of bulls against Tritrichomonas foetus has been advocated in previous studies, but anecdotal reports suggest this practice does not clear infections and may additionally confound diagnostic testing by reducing parasite burdens below detectable limits. The objective of this study was to characterize the systemic humoral immune response to therapeutic vaccination in T. foetus-infected bulls over a period of four months using an indirect ELISA and to compare the dynamics of this response to culture and PCR results to establish the existence of a relationship (or lack thereof) between immunization and infection status. A study population of 4- to 6-year-old T. foetus-infected beef bulls (n = 20) was divided equally into a treatment group and a control group. The treatment group received two doses of commercially prepared whole cell killed vaccine 2 weeks apart while the control group received injections of vaccine diluent. Blood samples were collected at each injection and at 4 subsequent dates every 4 weeks thereafter (i.e. 0, 2, 6, 10, 14, and 18 wks) to measure IgG1 and IgG2 antibody subisotype response via an indirect ELISA. Preputial smegma samples were collected at the four monthly intervals following vaccination for diagnosis of infection via InPouch™ culture, Modified Diamond's Medium (MDM) culture, and PCR. Humoral response for both IgG isotypes from week 2 through week 18 were significantly increased in vaccinates compared to controls. No significant decrease in infection prevalence was detected in the treatment group for any of the diagnostic methods used. The apparent lack of pathogen clearance during a stimulated immune response suggests that therapeutic vaccination may not be a useful T. foetus management practice.

Adenovirus and cryptosporidium co-infection in a corn snake (Elaphae guttata guttata).

A 1-yr-old albino male corn snake (Elaphae guttata guttata), which was part of a large breeding stock, was presented to the University of Florida, College of Veterinary Medicine, Zoo and Exotic Animal Clinic with a history of anorexia for 2 wk and progressively declining body condition. The animal was euthanized due to a poor prognosis. Histopathology, electron microscopy, and polymerase chain reaction analysis on tissues revealed concurrent infection with adenovirus and Cryptosporidium. Primary infection with adenovirus could have caused immunodeficiency in the snake, thus predisposing it to secondary infection with Cryptosporidium. To the authors' knowledge, this is the first report of co-infection of adenovirus and Cryptosporidium in a Colubrid species of snake.