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1.
PLoS Pathog ; 18(9): e1010828, 2022 09.
Article in English | MEDLINE | ID: covidwho-2039447

ABSTRACT

Spillover of sarbecoviruses from animals to humans has resulted in outbreaks of severe acute respiratory syndrome SARS-CoVs and the ongoing COVID-19 pandemic. Efforts to identify the origins of SARS-CoV-1 and -2 has resulted in the discovery of numerous animal sarbecoviruses-the majority of which are only distantly related to known human pathogens and do not infect human cells. The receptor binding domain (RBD) on sarbecoviruses engages receptor molecules on the host cell and mediates cell invasion. Here, we tested the receptor tropism and serological cross reactivity for RBDs from two sarbecoviruses found in Russian horseshoe bats. While these two viruses are in a viral lineage distinct from SARS-CoV-1 and -2, the RBD from one virus, Khosta 2, was capable of using human ACE2 to facilitate cell entry. Viral pseudotypes with a recombinant, SARS-CoV-2 spike encoding for the Khosta 2 RBD were resistant to both SARS-CoV-2 monoclonal antibodies and serum from individuals vaccinated for SARS-CoV-2. Our findings further demonstrate that sarbecoviruses circulating in wildlife outside of Asia also pose a threat to global health and ongoing vaccine campaigns against SARS-CoV-2.


Subject(s)
COVID-19 , Chiroptera , Angiotensin-Converting Enzyme 2 , Animals , Antibodies, Monoclonal , Antibodies, Viral , COVID-19/prevention & control , COVID-19 Vaccines , Humans , Pandemics/prevention & control , SARS-CoV-2 , Spike Glycoprotein, Coronavirus
2.
J Clin Virol Plus ; 1(4): 100047, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1509983

ABSTRACT

Serologic testing of residual blood samples from 812 children from a hospital in New Orleans, LA, between March and May 2020, demonstrated a SARS-CoV-2 seroprevalence of 6.8% based on S and N protein IgG; Black and Hispanic children, and children living in zip codes with lower household incomes were over-represented.

3.
Nat Nanotechnol ; 16(9): 1039-1044, 2021 09.
Article in English | MEDLINE | ID: covidwho-1322483

ABSTRACT

Plasma SARS-CoV-2 RNA may represent a viable diagnostic alternative to respiratory RNA levels, which rapidly decline after infection. Quantitative PCR with reverse transcription (RT-qPCR) reference assays exhibit poor performance with plasma, probably reflecting the dilution and degradation of viral RNA released into the circulation, but these issues could be addressed by analysing viral RNA packaged into extracellular vesicles. Here we describe an assay approach in which extracellular vesicles directly captured from plasma are fused with reagent-loaded liposomes to sensitively amplify and detect a SARS-CoV-2 gene target. This approach accurately identified patients with COVID-19, including challenging cases missed by RT-qPCR. SARS-CoV-2-positive extracellular vesicles were detected at day 1 post-infection, and plateaued from day 6 to the day 28 endpoint in a non-human primate model, while signal durations for 20-60 days were observed in young children. This nanotechnology approach uses a non-infectious sample and extends virus detection windows, offering a tool to support COVID-19 diagnosis in patients without SARS-CoV-2 RNA detectable in the respiratory tract.


Subject(s)
COVID-19/diagnosis , Extracellular Vesicles/metabolism , Liposomes/therapeutic use , RNA, Viral/blood , SARS-CoV-2/isolation & purification , Animals , Biosensing Techniques , COVID-19/blood , COVID-19 Nucleic Acid Testing , Chlorocebus aethiops , Disease Models, Animal , HEK293 Cells , Humans , Kinetics , Liposomes/metabolism , RNA, Viral/genetics , SARS-CoV-2/genetics , Tetraspanin 28/immunology , Tetraspanin 28/metabolism
4.
J Clin Invest ; 131(7)2021 04 01.
Article in English | MEDLINE | ID: covidwho-1076050

ABSTRACT

BACKGROUNDCirculating severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA may represent a more reliable indicator of infection than nasal RNA, but quantitative reverse transcription PCR (RT-qPCR) lacks diagnostic sensitivity for blood samples.METHODSA CRISPR-augmented RT-PCR assay that sensitively detects SARS-CoV-2 RNA was employed to analyze viral RNA kinetics in longitudinal plasma samples from nonhuman primates (NHPs) after virus exposure; to evaluate the utility of blood SARS-CoV-2 RNA detection for coronavirus disease 2019 (COVID-19) diagnosis in adults cases confirmed by nasal/nasopharyngeal swab RT-PCR results; and to identify suspected COVID-19 cases in pediatric and at-risk adult populations with negative nasal swab RT-qPCR results. All blood samples were analyzed by RT-qPCR to allow direct comparisons.RESULTSCRISPR-augmented RT-PCR consistently detected SARS-CoV-2 RNA in the plasma of experimentally infected NHPs from 1 to 28 days after infection, and these increases preceded and correlated with rectal swab viral RNA increases. In a patient cohort (n = 159), this blood-based assay demonstrated 91.2% diagnostic sensitivity and 99.2% diagnostic specificity versus a comparator RT-qPCR nasal/nasopharyngeal test, whereas RT-qPCR exhibited 44.1% diagnostic sensitivity and 100% specificity for the same blood samples. This CRISPR-augmented RT-PCR assay also accurately identified patients with COVID-19 using one or more negative nasal swab RT-qPCR results.CONCLUSIONResults of this study indicate that sensitive detection of SARS-CoV-2 RNA in blood by CRISPR-augmented RT-PCR permits accurate COVID-19 diagnosis, and can detect COVID-19 cases with transient or negative nasal swab RT-qPCR results, suggesting that this approach could improve COVID-19 diagnosis and the evaluation of SARS-CoV-2 infection clearance, and predict the severity of infection.TRIAL REGISTRATIONClinicalTrials.gov. NCT04358211.FUNDINGDepartment of Defense, National Institute of Allergy and Infectious Diseases, National Institute of Child Health and Human Development, and the National Center for Research Resources.


Subject(s)
COVID-19/blood , COVID-19/virology , Cell-Free Nucleic Acids/blood , Cell-Free Nucleic Acids/genetics , RNA, Viral/blood , RNA, Viral/genetics , SARS-CoV-2 , Adolescent , Adult , Aged , Animals , COVID-19/diagnosis , COVID-19 Nucleic Acid Testing/methods , COVID-19 Nucleic Acid Testing/statistics & numerical data , CRISPR-Cas Systems , Child , Child, Preschool , Disease Models, Animal , Female , Humans , Infant , Longitudinal Studies , Macaca mulatta , Male , Middle Aged , Pandemics , SARS-CoV-2/genetics , Sensitivity and Specificity , Time Factors
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