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      Just 2% of SARS-CoV-2−positive individuals carry 90% of the virus circulating in communities

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      Proceedings of the National Academy of Sciences of the United States of America
      National Academy of Sciences
      viral load, SARS-CoV-2, transmission

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          Significance

          We analyzed data from saliva-based COVID-19 screening deployed on the University of Colorado Boulder campus. Our dataset is unique in that all SARS-CoV-2−positive individuals reported no symptoms at the time of saliva collection, and therefore were infected but asymptomatic or presymptomatic. We found that 1) the distribution of viral loads observed in our asymptomatic college population was indistinguishable from what has been reported in hospitalized populations; 2) regardless of symptomatic status, approximately 50% of individuals who test positive for SARS-CoV-2 seem to be in noninfectious phases of the infection; and 3) just 2% of infected individuals carry 90% of the virions circulating within communities, serving as viral “supercarriers” and likely also superspreaders.

          Abstract

          We analyze data from the fall 2020 pandemic response efforts at the University of Colorado Boulder, where more than 72,500 saliva samples were tested for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) using qRT-PCR. All samples were collected from individuals who reported no symptoms associated with COVID-19 on the day of collection. From these, 1,405 positive cases were identified. The distribution of viral loads within these asymptomatic individuals was indistinguishable from what has been previously observed in symptomatic individuals. Regardless of symptomatic status, ∼50% of individuals who test positive for SARS-CoV-2 seem to be in noninfectious phases of the disease, based on having low viral loads in a range from which live virus has rarely been isolated. We find that, at any given time, just 2% of individuals carry 90% of the virions circulating within communities, serving as viral “supercarriers” and possibly also superspreaders.

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          A pneumonia outbreak associated with a new coronavirus of probable bat origin

          Since the outbreak of severe acute respiratory syndrome (SARS) 18 years ago, a large number of SARS-related coronaviruses (SARSr-CoVs) have been discovered in their natural reservoir host, bats 1–4 . Previous studies have shown that some bat SARSr-CoVs have the potential to infect humans 5–7 . Here we report the identification and characterization of a new coronavirus (2019-nCoV), which caused an epidemic of acute respiratory syndrome in humans in Wuhan, China. The epidemic, which started on 12 December 2019, had caused 2,794 laboratory-confirmed infections including 80 deaths by 26 January 2020. Full-length genome sequences were obtained from five patients at an early stage of the outbreak. The sequences are almost identical and share 79.6% sequence identity to SARS-CoV. Furthermore, we show that 2019-nCoV is 96% identical at the whole-genome level to a bat coronavirus. Pairwise protein sequence analysis of seven conserved non-structural proteins domains show that this virus belongs to the species of SARSr-CoV. In addition, 2019-nCoV virus isolated from the bronchoalveolar lavage fluid of a critically ill patient could be neutralized by sera from several patients. Notably, we confirmed that 2019-nCoV uses the same cell entry receptor—angiotensin converting enzyme II (ACE2)—as SARS-CoV.
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            Virological assessment of hospitalized patients with COVID-2019

            Coronavirus disease 2019 (COVID-19) is an acute infection of the respiratory tract that emerged in late 20191,2. Initial outbreaks in China involved 13.8% of cases with severe courses, and 6.1% of cases with critical courses3. This severe presentation may result from the virus using a virus receptor that is expressed predominantly in the lung2,4; the same receptor tropism is thought to have determined the pathogenicity-but also aided in the control-of severe acute respiratory syndrome (SARS) in 20035. However, there are reports of cases of COVID-19 in which the patient shows mild upper respiratory tract symptoms, which suggests the potential for pre- or oligosymptomatic transmission6-8. There is an urgent need for information on virus replication, immunity and infectivity in specific sites of the body. Here we report a detailed virological analysis of nine cases of COVID-19 that provides proof of active virus replication in tissues of the upper respiratory tract. Pharyngeal virus shedding was very high during the first week of symptoms, with a peak at 7.11 × 108 RNA copies per throat swab on day 4. Infectious virus was readily isolated from samples derived from the throat or lung, but not from stool samples-in spite of high concentrations of virus RNA. Blood and urine samples never yielded virus. Active replication in the throat was confirmed by the presence of viral replicative RNA intermediates in the throat samples. We consistently detected sequence-distinct virus populations in throat and lung samples from one patient, proving independent replication. The shedding of viral RNA from sputum outlasted the end of symptoms. Seroconversion occurred after 7 days in 50% of patients (and by day 14 in all patients), but was not followed by a rapid decline in viral load. COVID-19 can present as a mild illness of the upper respiratory tract. The confirmation of active virus replication in the upper respiratory tract has implications for the containment of COVID-19.
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              SARS-CoV-2 Viral Load in Upper Respiratory Specimens of Infected Patients

              To the Editor: The 2019 novel coronavirus (SARS-CoV-2) epidemic, which was first reported in December 2019 in Wuhan, China, and has been declared a public health emergency of international concern by the World Health Organization, may progress to a pandemic associated with substantial morbidity and mortality. SARS-CoV-2 is genetically related to SARS-CoV, which caused a global epidemic with 8096 confirmed cases in more than 25 countries in 2002–2003. 1 The epidemic of SARS-CoV was successfully contained through public health interventions, including case detection and isolation. Transmission of SARS-CoV occurred mainly after days of illness 2 and was associated with modest viral loads in the respiratory tract early in the illness, with viral loads peaking approximately 10 days after symptom onset. 3 We monitored SARS-CoV-2 viral loads in upper respiratory specimens obtained from 18 patients (9 men and 9 women; median age, 59 years; range, 26 to 76) in Zhuhai, Guangdong, China, including 4 patients with secondary infections (1 of whom never had symptoms) within two family clusters (Table S1 in the Supplementary Appendix, available with the full text of this letter at NEJM.org). The patient who never had symptoms was a close contact of a patient with a known case and was therefore monitored. A total of 72 nasal swabs (sampled from the mid-turbinate and nasopharynx) (Figure 1A) and 72 throat swabs (Figure 1B) were analyzed, with 1 to 9 sequential samples obtained from each patient. Polyester flock swabs were used for all the patients. From January 7 through January 26, 2020, a total of 14 patients who had recently returned from Wuhan and had fever (≥37.3°C) received a diagnosis of Covid-19 (the illness caused by SARS-CoV-2) by means of reverse-transcriptase–polymerase-chain-reaction assay with primers and probes targeting the N and Orf1b genes of SARS-CoV-2; the assay was developed by the Chinese Center for Disease Control and Prevention. Samples were tested at the Guangdong Provincial Center for Disease Control and Prevention. Thirteen of 14 patients with imported cases had evidence of pneumonia on computed tomography (CT). None of them had visited the Huanan Seafood Wholesale Market in Wuhan within 14 days before symptom onset. Patients E, I, and P required admission to intensive care units, whereas the others had mild-to-moderate illness. Secondary infections were detected in close contacts of Patients E, I, and P. Patient E worked in Wuhan and visited his wife (Patient L), mother (Patient D), and a friend (Patient Z) in Zhuhai on January 17. Symptoms developed in Patients L and D on January 20 and January 22, respectively, with viral RNA detected in their nasal and throat swabs soon after symptom onset. Patient Z reported no clinical symptoms, but his nasal swabs (cycle threshold [Ct] values, 22 to 28) and throat swabs (Ct values, 30 to 32) tested positive on days 7, 10, and 11 after contact. A CT scan of Patient Z that was obtained on February 6 was unremarkable. Patients I and P lived in Wuhan and visited their daughter (Patient H) in Zhuhai on January 11 when their symptoms first developed. Fever developed in Patient H on January 17, with viral RNA detected in nasal and throat swabs on day 1 after symptom onset. We analyzed the viral load in nasal and throat swabs obtained from the 17 symptomatic patients in relation to day of onset of any symptoms (Figure 1C). Higher viral loads (inversely related to Ct value) were detected soon after symptom onset, with higher viral loads detected in the nose than in the throat. Our analysis suggests that the viral nucleic acid shedding pattern of patients infected with SARS-CoV-2 resembles that of patients with influenza 4 and appears different from that seen in patients infected with SARS-CoV. 3 The viral load that was detected in the asymptomatic patient was similar to that in the symptomatic patients, which suggests the transmission potential of asymptomatic or minimally symptomatic patients. These findings are in concordance with reports that transmission may occur early in the course of infection 5 and suggest that case detection and isolation may require strategies different from those required for the control of SARS-CoV. How SARS-CoV-2 viral load correlates with culturable virus needs to be determined. Identification of patients with few or no symptoms and with modest levels of detectable viral RNA in the oropharynx for at least 5 days suggests that we need better data to determine transmission dynamics and inform our screening practices.
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                Author and article information

                Journal
                Proc Natl Acad Sci U S A
                Proc Natl Acad Sci U S A
                pnas
                pnas
                PNAS
                Proceedings of the National Academy of Sciences of the United States of America
                National Academy of Sciences
                0027-8424
                1091-6490
                25 May 2021
                10 May 2021
                10 May 2021
                : 118
                : 21
                : e2104547118
                Affiliations
                [1] aBioFrontiers Institute, University of Colorado Boulder , Boulder, CO 80303;
                [2] bDepartment of Molecular, Cellular, and Developmental Biology, University of Colorado Boulder , Boulder, CO 80303;
                [3] cDepartment of Biochemistry, University of Colorado Boulder , Boulder, CO 80303;
                [4] dHHMI , University of Colorado Boulder , Boulder, CO 80303;
                [5] eDarwin Biosciences Inc., Boulder, CO 80303;
                [6] fInterdisciplinary Quantitative Biology Program, University of Colorado Boulder , Boulder, CO 80303;
                [7] gWardenburg Health Center, University of Colorado Boulder , Boulder, CO 80303;
                [8] hDepartment of Integrative Physiology, University of Colorado Boulder , Boulder, CO 80303;
                [9] iDepartment of Computer Science, University of Colorado Boulder , Boulder, CO 80303
                Author notes
                1To whom correspondence may be addressed. Email: Roy.Parker@ 123456Colorado.EDU or ssawyer@ 123456colorado.EDU .

                Edited by Peter Palese, Icahn School of Medicine at Mount Sinai, New York, NY, and approved April 11, 2021 (received for review March 8, 2021)

                Author contributions: Q.Y., T.K.S., P.K.G., E.L., C.J.D., N.R.M., G.R.B., M.B.M., R.D.D., L.L., R.P., and S.L.S. designed research; Q.Y., T.K.S., P.K.G., E.L., C.J.D., K.L.T., M.R.F., C.R.H., J.C.D., C.D.O., D.M., S.K.C., W.T.F., C.L.P., A.R.G., A.B.-G., E.R.W.-S., S.S.W., and R.D.D. performed research; Q.Y., R.D.D., R.P., and S.L.S. analyzed data; and Q.Y., T.K.S., R.D.D., L.L., R.P., and S.L.S. wrote the paper.

                Author information
                https://orcid.org/0000-0001-9053-3158
                https://orcid.org/0000-0002-2672-8919
                https://orcid.org/0000-0002-0567-3234
                https://orcid.org/0000-0001-8476-6617
                https://orcid.org/0000-0002-7620-3977
                https://orcid.org/0000-0001-7483-839X
                https://orcid.org/0000-0003-1470-4810
                https://orcid.org/0000-0002-8412-4152
                https://orcid.org/0000-0002-6965-1085
                Article
                202104547
                10.1073/pnas.2104547118
                8166196
                33972412
                06294c8c-5f2a-4e1e-a1aa-b169a2393564
                Copyright © 2021 the Author(s). Published by PNAS.

                This open access article is distributed under Creative Commons Attribution License 4.0 (CC BY).

                History
                Page count
                Pages: 6
                Funding
                Funded by: Burroughs Wellcome Fund (BWF) 100000861
                Award ID: PATH
                Award Recipient : Nicholas R Meyerson Award Recipient : Sara L Sawyer
                Funded by: Burroughs Wellcome Fund (BWF) 100000861
                Award ID: PDEP
                Award Recipient : Nicholas R Meyerson Award Recipient : Sara L Sawyer
                Funded by: HHS | National Institutes of Health (NIH) 100000002
                Award ID: DP1-DA-046108
                Award Recipient : Sara L Sawyer
                Categories
                423
                530
                Biological Sciences
                Microbiology
                Custom metadata
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                viral load,sars-cov-2,transmission
                viral load, sars-cov-2, transmission

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