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1.
Emerg Infect Dis ; 29(7): 1386-1396, 2023 07.
Article in English | MEDLINE | ID: covidwho-20237258

ABSTRACT

Isolating and characterizing emerging SARS-CoV-2 variants is key to understanding virus pathogenesis. In this study, we isolated samples of the SARS-CoV-2 R.1 lineage, categorized as a variant under monitoring by the World Health Organization, and evaluated their sensitivity to neutralizing antibodies and type I interferons. We used convalescent serum samples from persons in Canada infected either with ancestral virus (wave 1) or the B.1.1.7 (Alpha) variant of concern (wave 3) for testing neutralization sensitivity. The R.1 isolates were potently neutralized by both the wave 1 and wave 3 convalescent serum samples, unlike the B.1.351 (Beta) variant of concern. Of note, the R.1 variant was significantly more resistant to type I interferons (IFN-α/ß) than was the ancestral isolate. Our study demonstrates that the R.1 variant retained sensitivity to neutralizing antibodies but evolved resistance to type I interferons. This critical driving force will influence the trajectory of the pandemic.


Subject(s)
COVID-19 , Interferon Type I , Humans , SARS-CoV-2/genetics , Interferon Type I/genetics , Antibodies, Neutralizing , COVID-19 Serotherapy , Canada/epidemiology , Antibodies, Viral , Spike Glycoprotein, Coronavirus
2.
Microbiol Spectr ; 11(3): e0190022, 2023 Jun 15.
Article in English | MEDLINE | ID: covidwho-2304932

ABSTRACT

Genomic epidemiology can facilitate an understanding of evolutionary history and transmission dynamics of a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) outbreak. We used next-generation sequencing techniques to study SARS-CoV-2 genomes isolated from patients and health care workers (HCWs) across five wards of a Canadian hospital with an ongoing SARS-CoV-2 outbreak. Using traditional contact tracing methods, we show transmission events between patients and HCWs, which were also supported by the SARS-CoV-2 lineage assignments. The outbreak predominantly involved SARS-CoV-2 B.1.564.1 across all five wards, but we also show evidence of community introductions of lineages B.1, B.1.1.32, and B.1.231, falsely assumed to be outbreak related. Altogether, our study exemplifies the value of using contact tracing in combination with genomic epidemiology to understand the transmission dynamics and genetic underpinnings of a SARS-CoV-2 outbreak. IMPORTANCE Our manuscript describes a SARS-CoV-2 outbreak investigation in an Ontario tertiary care hospital. We use traditional contract tracing paired with whole-genome sequencing to facilitate an understanding of the evolutionary history and transmission dynamics of this SARS-CoV-2 outbreak in a clinical setting. These advancements have enabled the incorporation of phylogenetics and genomic epidemiology into the understanding of clinical outbreaks. We show that genomic epidemiology can help to explore the genetic evolution of a pathogen in real time, enabling the identification of the index case and helping understand its transmission dynamics to develop better strategies to prevent future spread of SARS-CoV-2 in congregate, clinical settings such as hospitals.


Subject(s)
COVID-19 , SARS-CoV-2 , Humans , SARS-CoV-2/genetics , Contact Tracing , COVID-19/epidemiology , Ontario/epidemiology , Tertiary Care Centers , Disease Outbreaks
3.
J Infect Dis ; 225(5): 768-776, 2022 03 02.
Article in English | MEDLINE | ID: covidwho-1722480

ABSTRACT

BACKGROUND: We determined the burden of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in air and on surfaces in rooms of patients hospitalized with coronavirus disease 2019 (COVID-19) and investigated patient characteristics associated with SARS-CoV-2 environmental contamination. METHODS: Nasopharyngeal swabs, surface, and air samples were collected from the rooms of 78 inpatients with COVID-19 at 6 acute care hospitals in Toronto from March to May 2020. Samples were tested for SARS-CoV-2 ribonucleic acid (RNA), cultured to determine potential infectivity, and whole viral genomes were sequenced. Association between patient factors and detection of SARS-CoV-2 RNA in surface samples were investigated. RESULTS: Severe acute respiratory syndrome coronavirus 2 RNA was detected from surfaces (125 of 474 samples; 42 of 78 patients) and air (3 of 146 samples; 3 of 45 patients); 17% (6 of 36) of surface samples from 3 patients yielded viable virus. Viral sequences from nasopharyngeal and surface samples clustered by patient. Multivariable analysis indicated hypoxia at admission, polymerase chain reaction-positive nasopharyngeal swab (cycle threshold of ≤30) on or after surface sampling date, higher Charlson comorbidity score, and shorter time from onset of illness to sampling date were significantly associated with detection of SARS-CoV-2 RNA in surface samples. CONCLUSIONS: The infrequent recovery of infectious SARS-CoV-2 virus from the environment suggests that the risk to healthcare workers from air and near-patient surfaces in acute care hospital wards is likely limited.


Subject(s)
COVID-19 , Nasopharynx/virology , Respiratory Aerosols and Droplets , SARS-CoV-2/isolation & purification , Adult , Aged , Air Microbiology , COVID-19/epidemiology , COVID-19/prevention & control , COVID-19/transmission , COVID-19 Nucleic Acid Testing , Canada/epidemiology , Environmental Exposure , Health Personnel , Humans , Inpatients , Middle Aged , Pandemics/prevention & control , SARS-CoV-2/genetics
4.
Methods Mol Biol ; 2327: 119-137, 2021.
Article in English | MEDLINE | ID: covidwho-1363726

ABSTRACT

Outbreak analysis and transmission surveillance of viruses can be performed via whole-genome sequencing after viral isolation. Such techniques have recently been applied to characterize and monitor SARS-CoV-2 , the etiological agent of the COVID-19 pandemic. However, the isolation and culture of SARS-CoV-2 is time consuming and requires biosafety level 3 containment, which is not ideal for many resource-constrained settings. An alternate method, bait capture allows target enrichment and sequencing of the entire SARS-CoV-2 genome eliminating the need for viral culture. This method uses a set of hybridization probes known as "baits" that span the genome and provide sensitive, accurate, and minimal off-target hybridization. Baits can be designed to detect any known virus or bacteria in a wide variety of specimen types, including oral secretions. The bait capture method presented herein allows the whole genome of SARS-CoV-2 in saliva to be sequenced without the need to culture and provides an outline of bait design and bioinformatic analysis to guide a bioinformatician.


Subject(s)
Genome, Viral , SARS-CoV-2/genetics , Saliva/virology , Whole Genome Sequencing/methods , Computational Biology , DNA, Complementary/genetics , Humans , Molecular Probes/genetics , Polymerase Chain Reaction/methods , SARS-CoV-2/isolation & purification , Specimen Handling/methods , Streptavidin , Whole Genome Sequencing/instrumentation
5.
Viruses ; 12(8)2020 08 15.
Article in English | MEDLINE | ID: covidwho-717763

ABSTRACT

Genome sequencing of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is increasingly important to monitor the transmission and adaptive evolution of the virus. The accessibility of high-throughput methods and polymerase chain reaction (PCR) has facilitated a growing ecosystem of protocols. Two differing protocols are tiling multiplex PCR and bait capture enrichment. Each method has advantages and disadvantages but a direct comparison with different viral RNA concentrations has not been performed to assess the performance of these approaches. Here we compare Liverpool amplification, ARTIC amplification, and bait capture using clinical diagnostics samples. All libraries were sequenced using an Illumina MiniSeq with data analyzed using a standardized bioinformatics workflow (SARS-CoV-2 Illumina GeNome Assembly Line; SIGNAL). One sample showed poor SARS-CoV-2 genome coverage and consensus, reflective of low viral RNA concentration. In contrast, the second sample had a higher viral RNA concentration, which yielded good genome coverage and consensus. ARTIC amplification showed the highest depth of coverage results for both samples, suggesting this protocol is effective for low concentrations. Liverpool amplification provided a more even read coverage of the SARS-CoV-2 genome, but at a lower depth of coverage. Bait capture enrichment of SARS-CoV-2 cDNA provided results on par with amplification. While only two clinical samples were examined in this comparative analysis, both the Liverpool and ARTIC amplification methods showed differing efficacy for high and low concentration samples. In addition, amplification-free bait capture enriched sequencing of cDNA is a viable method for generating a SARS-CoV-2 genome sequence and for identification of amplification artifacts.


Subject(s)
Betacoronavirus/genetics , Coronavirus Infections/virology , Pneumonia, Viral/virology , RNA, Viral/genetics , Base Sequence , Betacoronavirus/isolation & purification , COVID-19 , COVID-19 Testing , Clinical Laboratory Techniques/methods , Coronavirus Infections/diagnosis , DNA, Complementary/genetics , Genome, Viral , Humans , Molecular Epidemiology , Multiplex Polymerase Chain Reaction/methods , Pandemics , SARS-CoV-2 , Whole Genome Sequencing/methods
6.
Emerg Infect Dis ; 26(9): 2054-2063, 2020 09.
Article in English | MEDLINE | ID: covidwho-607956

ABSTRACT

Since its emergence in Wuhan, China, in December 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has infected ≈6 million persons worldwide. As SARS-CoV-2 spreads across the planet, we explored the range of human cells that can be infected by this virus. We isolated SARS-CoV-2 from 2 infected patients in Toronto, Canada; determined the genomic sequences; and identified single-nucleotide changes in representative populations of our virus stocks. We also tested a wide range of human immune cells for productive infection with SARS-CoV-2. We confirm that human primary peripheral blood mononuclear cells are not permissive for SARS-CoV-2. As SARS-CoV-2 continues to spread globally, it is essential to monitor single-nucleotide polymorphisms in the virus and to continue to isolate circulating viruses to determine viral genotype and phenotype by using in vitro and in vivo infection models.


Subject(s)
Betacoronavirus , Coronavirus Infections/virology , Leukocytes, Mononuclear/virology , Pneumonia, Viral/virology , Virus Replication/genetics , Betacoronavirus/genetics , Betacoronavirus/isolation & purification , Betacoronavirus/physiology , COVID-19 , DNA, Viral/genetics , DNA, Viral/isolation & purification , Genotype , Humans , Kinetics , Pandemics , Polymorphism, Single Nucleotide , SARS-CoV-2 , Whole Genome Sequencing
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