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1.
J Vet Diagn Invest ; 34(5): 825-834, 2022 Sep.
Article in English | MEDLINE | ID: covidwho-2002060

ABSTRACT

The COVID-19 pandemic presents a continued public health challenge. Veterinary diagnostic laboratories in the United States use RT-rtPCR for animal testing, and many laboratories are certified for testing human samples; hence, ensuring that laboratories have sensitive and specific SARS-CoV2 testing methods is a critical component of the pandemic response. In 2020, the FDA Veterinary Laboratory Investigation and Response Network (Vet-LIRN) led an interlaboratory comparison (ILC1) to help laboratories evaluate their existing RT-rtPCR methods for detecting SARS-CoV2. All participating laboratories were able to detect the viral RNA spiked in buffer and PrimeStore molecular transport medium (MTM). With ILC2, Vet-LIRN extended ILC1 by evaluating analytical sensitivity and specificity of the methods used by participating laboratories to detect 3 SARS-CoV2 variants (B.1; B.1.1.7 [Alpha]; B.1.351 [Beta]) at various copy levels. We analyzed 57 sets of results from 45 laboratories qualitatively and quantitatively according to the principles of ISO 16140-2:2016. More than 95% of analysts detected the SARS-CoV2 RNA in MTM at ≥500 copies for all 3 variants. In addition, for nucleocapsid markers N1 and N2, 81% and 92% of the analysts detected ≤20 copies in the assays, respectively. The analytical specificity of the evaluated methods was >99%. Participating laboratories were able to assess their current method performance, identify possible limitations, and recognize method strengths as part of a continuous learning environment to support the critical need for the reliable diagnosis of COVID-19 in potentially infected animals and humans.


Subject(s)
COVID-19 , SARS-CoV-2 , Animals , COVID-19/diagnosis , COVID-19/veterinary , COVID-19 Testing , Humans , Immunity, Innate , Laboratories , Lymphocytes , Pandemics/veterinary , RNA, Viral/analysis , SARS-CoV-2/genetics , Sensitivity and Specificity , United States/epidemiology
2.
Viruses ; 14(5)2022 04 21.
Article in English | MEDLINE | ID: covidwho-1822442

ABSTRACT

A canine coronavirus (CCoV) has now been reported from two independent human samples from Malaysia (respiratory, collected in 2017-2018; CCoV-HuPn-2018) and Haiti (urine, collected in 2017); these two viruses were nearly genetically identical. In an effort to identify any novel adaptations associated with this apparent shift in tropism we carried out detailed evolutionary analyses of the spike gene of this virus in the context of related Alphacoronavirus 1 species. The spike 0-domain retains homology to CCoV2b (enteric infections) and Transmissible Gastroenteritis Virus (TGEV; enteric and respiratory). This domain is subject to relaxed selection pressure and an increased rate of molecular evolution. It contains unique amino acid substitutions, including within a region important for sialic acid binding and pathogenesis in TGEV. Overall, the spike gene is extensively recombinant, with a feline coronavirus type II strain serving a prominent role in the recombinant history of the virus. Molecular divergence time for a segment of the gene where temporal signal could be determined, was estimated at around 60 years ago. We hypothesize that the virus had an enteric origin, but that it may be losing that particular tropism, possibly because of mutations in the sialic acid binding region of the spike 0-domain.


Subject(s)
Coronavirus, Canine , Animals , Cats , Dogs , N-Acetylneuraminic Acid , Spike Glycoprotein, Coronavirus/genetics , Tropism , Zoonoses
3.
J Vet Diagn Invest ; 33(6): 1039-1051, 2021 Nov.
Article in English | MEDLINE | ID: covidwho-1322903

ABSTRACT

The continued search for intermediate hosts and potential reservoirs for SARS-CoV2 makes it clear that animal surveillance is critical in outbreak response and prevention. Real-time RT-PCR assays for SARS-CoV2 detection can easily be adapted to different host species. U.S. veterinary diagnostic laboratories have used the CDC assays or other national reference laboratory methods to test animal samples. However, these methods have only been evaluated using internal validation protocols. To help the laboratories evaluate their SARS-CoV2 test methods, an interlaboratory comparison (ILC) was performed in collaboration with multiple organizations. Forty-four sets of 19 blind-coded RNA samples in Tris-EDTA (TE) buffer or PrimeStore transport medium were shipped to 42 laboratories. Results were analyzed according to the principles of the International Organization for Standardization (ISO) 16140-2:2016 standard. Qualitative assessment of PrimeStore samples revealed that, in approximately two-thirds of the laboratories, the limit of detection with a probability of 0.95 (LOD95) for detecting the RNA was ≤20 copies per PCR reaction, close to the theoretical LOD of 3 copies per reaction. This level of sensitivity is not expected in clinical samples because of additional factors, such as sample collection, transport, and extraction of RNA from the clinical matrix. Quantitative assessment of Ct values indicated that reproducibility standard deviations for testing the RNA with assays reported as N1 were slightly lower than those for N2, and they were higher for the RNA in PrimeStore medium than those in TE buffer. Analyst experience and the use of either a singleplex or multiplex PCR also affected the quantitative ILC test results.


Subject(s)
COVID-19 , RNA, Viral , Animals , COVID-19/veterinary , Laboratories , RNA, Viral/genetics , Reproducibility of Results , SARS-CoV-2 , Sensitivity and Specificity
5.
J Vet Diagn Invest ; 33(1): 80-86, 2021 Jan.
Article in English | MEDLINE | ID: covidwho-920981

ABSTRACT

In the United States, horses are used for a variety of purposes including recreation, exhibition, and racing. As farm, performance, and companion animals, horses are a unique species from a zoonotic disease risk perspective, and the risks of subclinical infections spreading among horses can pose challenges. Using a nanoscale real-time PCR platform, we investigated the prevalence of 14 enteric pathogens, 11 Escherichia coli genes, and 9 respiratory pathogens in fecal samples from 97 apparently healthy horses at a multi-day horse event. In addition, sugar flotation test was performed for fecal parasites. E. coli f17 was commonly detected, prevalent in 59% of horses, followed closely by Streptococcus equi subsp. zooepidemicus (55%). Additional pathogens recognized included betacoronavirus, Campylobacter jejuni, Cryptosporidium sp., E. coli O157, equine adenovirus 1, equine rhinitis B virus, and others. The use of PCR data may overestimate the true prevalence of these pathogens but provides a sensitive overview of common pathogens present in healthy horses. Our results prompt the continued need for practical biosecurity measures at horse shows, both to protect individuals interacting with these horses and to minimize transmission among horses.


Subject(s)
Animal Husbandry , Cryptosporidiosis/epidemiology , Cryptosporidium/isolation & purification , Escherichia coli Infections/veterinary , Escherichia coli/isolation & purification , Horse Diseases/epidemiology , Animals , Cryptosporidium/genetics , Escherichia coli/genetics , Escherichia coli Infections/epidemiology , Female , Horse Diseases/diagnosis , Horses , Male , New York/epidemiology , Population Surveillance , Real-Time Polymerase Chain Reaction/veterinary
6.
Microbiol Resour Announc ; 9(22)2020 May 28.
Article in English | MEDLINE | ID: covidwho-401519

ABSTRACT

This report describes the identification and characterization of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in a Malayan tiger in a U.S. zoo.

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