ABSTRACT
The coronavirus disease 2019 (COVID-19) pandemic presents a continued public health challenge across the world. Veterinary diagnostic laboratories in the U.S. use real-time reverse transcriptase PCR (RT-PCR) for animal testing, and many are certified for testing human samples, so ensuring laboratories have sensitive and specific SARS-CoV-2 testing methods is a critical component of the pandemic response. In 2020, the FDA Veterinary Laboratory Investigation and Response Network (Vet-LIRN) led the first round of an Inter-Laboratory Comparison (ILC) Exercise to help laboratories evaluate their existing real-time RT-PCR methods for detecting SARS-CoV-2. The ILC1 results indicated that all participating laboratories were able to detect the viral RNA spiked in buffer and PrimeStore molecular transport medium (MTM). The current ILC (ILC2) aimed to extend ILC1 by evaluating analytical sensitivity and specificity of the methods used by participating laboratories to detect three SARS-CoV-2 variants (B.1, B.1.1.7 (Alpha) and B.1.351 (Beta)). ILC2 samples were prepared with RNA at levels between 10 to 10,000 copies per 50 μL MTM. Fifty-seven sets of results from 45 laboratories were qualitatively and quantitatively analyzed according to the principles of ISO 16140-2:2016. The results showed that over 95% of analysts detected the SARS-CoV-2 RNA in MTM at 500 copies or higher for all three variants. In addition, 81% and 92% of the analysts achieved a Level of Detection (LOD95 eff. vol. ) below 20 copies in the assays with nucleocapsid markers N1 and N2, respectively. The analytical specificity of the evaluated methods was over 99%. The study allowed participating laboratories 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 reliable diagnosis of COVID-19 in potentially infected animals and humans.
Subject(s)
COVID-19ABSTRACT
We describe the first cases of natural SARS-CoV-2 infection detected in animals in the United States. In March 2020, four tigers and three lions at the Bronx Zoo developed mild respiratory signs. SARS-CoV-2 RNA was detected by rRT-PCR in respiratory secretions and/or feces from all seven affected animals; viral RNA and/or antibodies were detected in their keepers. SARS-CoV-2 was isolated from respiratory secretions or feces from three affected animals; in situ hybridization co-localized viral RNA with cellular damage. Whole genome sequence and haplotype network analyses showed tigers and lions were infected with two different SARS-CoV-2 strains, suggesting independent viral introductions. The source of SARS-CoV-2 infection in the lions is unknown. Epidemiological data and genetic similarities between keeper and tiger viruses indicate human to animal transmission.