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      Pooled testing conserves SARS-CoV-2 laboratory resources and improves test turn-around time: experience on the Kenyan Coast

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      Wellcome Open Research
      F1000 Research Limited
      COVID-19, Pooled testing, SARS-CoV-2, Kilifi, Kenya

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          Abstract

          Background. International recommendations for the control of the coronavirus disease 2019 (COVID-19) pandemic emphasize the central role of laboratory testing for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the etiological agent, at scale. The availability of testing reagents, laboratory equipment and qualified staff are important bottlenecks to achieving this. Elsewhere, pooled testing (i.e. combining multiple samples in the same reaction) has been suggested to increase testing capacities in the pandemic period.

          Methods. We discuss our experience with SARS-CoV-2 pooled testing using real-time reverse transcription polymerase chain reaction (RT-PCR) on the Kenyan Coast.

          Results. In mid-May, 2020, our RT-PCR testing capacity for SARS-CoV-2 was improved by ~100% as a result of adoption of a six-sample pooled testing strategy. This was accompanied with a concomitant saving of ~50% of SARS-CoV-2 laboratory test kits at both the RNA extraction and RT-PCR stages. However, pooled testing came with a slight decline of test sensitivity. The RT-PCR cycle threshold value (ΔCt) was ~1.59 higher for samples tested in pools compared to samples tested singly.

          Conclusions. Pooled testing is a useful strategy to increase SARS-CoV-2 laboratory testing capacity especially in low-income settings.

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          Detection of 2019 novel coronavirus (2019-nCoV) by real-time RT-PCR

          Background The ongoing outbreak of the recently emerged novel coronavirus (2019-nCoV) poses a challenge for public health laboratories as virus isolates are unavailable while there is growing evidence that the outbreak is more widespread than initially thought, and international spread through travellers does already occur. Aim We aimed to develop and deploy robust diagnostic methodology for use in public health laboratory settings without having virus material available. Methods Here we present a validated diagnostic workflow for 2019-nCoV, its design relying on close genetic relatedness of 2019-nCoV with SARS coronavirus, making use of synthetic nucleic acid technology. Results The workflow reliably detects 2019-nCoV, and further discriminates 2019-nCoV from SARS-CoV. Through coordination between academic and public laboratories, we confirmed assay exclusivity based on 297 original clinical specimens containing a full spectrum of human respiratory viruses. Control material is made available through European Virus Archive – Global (EVAg), a European Union infrastructure project. Conclusion The present study demonstrates the enormous response capacity achieved through coordination of academic and public laboratories in national and European research networks.
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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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              Laboratory Diagnosis of COVID-19: Current Issues and Challenges

              The COVID-19 outbreak has had a major impact on clinical microbiology laboratories in the past several months. This commentary covers current issues and challenges for the laboratory diagnosis of infections caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In the preanalytical stage, collecting the proper respiratory tract specimen at the right time from the right anatomic site is essential for a prompt and accurate molecular diagnosis of COVID-19. Appropriate measures are required to keep laboratory staff safe while producing reliable test results.
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                Author and article information

                Contributors
                Role: ConceptualizationRole: Data CurationRole: Formal AnalysisRole: InvestigationRole: MethodologyRole: Project AdministrationRole: SupervisionRole: ValidationRole: VisualizationRole: Writing – Original Draft PreparationRole: Writing – Review & Editing
                Role: InvestigationRole: MethodologyRole: Project AdministrationRole: Writing – Review & Editing
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                Role: Data CurationRole: Writing – Review & Editing
                Role: Data CurationRole: Writing – Review & Editing
                Role: Data CurationRole: Writing – Review & Editing
                Role: InvestigationRole: Writing – Review & Editing
                Role: InvestigationRole: Writing – Review & Editing
                Role: InvestigationRole: Project AdministrationRole: Writing – Review & Editing
                Role: InvestigationRole: Writing – Review & Editing
                Role: Project AdministrationRole: Writing – Review & Editing
                Role: InvestigationRole: Writing – Review & Editing
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                Role: InvestigationRole: Writing – Review & Editing
                Role: InvestigationRole: Writing – Review & Editing
                Role: ConceptualizationRole: Funding AcquisitionRole: Project AdministrationRole: ResourcesRole: SupervisionRole: Writing – Review & Editing
                Role: ConceptualizationRole: InvestigationRole: MethodologyRole: Project AdministrationRole: SupervisionRole: ValidationRole: Writing – Review & Editing
                Role: ConceptualizationRole: Project AdministrationRole: SupervisionRole: Writing – Review & Editing
                Journal
                Wellcome Open Res
                Wellcome Open Res
                Wellcome Open Res
                Wellcome Open Research
                F1000 Research Limited (London, UK )
                2398-502X
                6 August 2020
                2020
                6 August 2020
                : 5
                : 186
                Affiliations
                [1 ]Kenya Medical Research Institute-Wellcome Trust Research Programme, Centre for Geographic Medicine Research, Kilifi, Kenya
                [2 ]Department of Biomedical Sciences, Pwani University, Kilifi, Kenya
                [3 ]Nuffield Department of Medicine, Centre for Clinical Vaccinology and Tropical Medicine, Churchill Hospital, University of Oxford, Oxford, UK
                [1 ]School of Medicine, International Medical University, Kuala Lumpur, Malaysia
                [1 ]Kenya Research Station, Institute of Tropical Medicine, KEMRI/Nagasaki University, Nairobi, Kenya
                [2 ]Directorate of Research and Innovation, Mount Kenya University, Thika, Kenya
                [3 ]School of Medicine, Mount Kenya University, Thika, Kenya
                Author notes

                No competing interests were disclosed.

                Competing interests: No competing interests were disclosed.

                Competing interests: No competing interests were disclosed.

                Author information
                https://orcid.org/0000-0002-2160-567X
                https://orcid.org/0000-0001-5312-0960
                https://orcid.org/0000-0002-3257-5920
                https://orcid.org/0000-0001-7467-7290
                https://orcid.org/0000-0002-3670-7521
                https://orcid.org/0000-0002-8014-7306
                https://orcid.org/0000-0001-6814-4439
                https://orcid.org/0000-0002-4015-718X
                https://orcid.org/0000-0001-8766-7470
                https://orcid.org/0000-0001-6482-9107
                https://orcid.org/0000-0002-8945-0002
                https://orcid.org/0000-0003-1961-3205
                https://orcid.org/0000-0003-2398-6717
                https://orcid.org/0000-0002-2619-0856
                https://orcid.org/0000-0002-2135-7549
                Article
                10.12688/wellcomeopenres.16113.1
                7590893
                33134555
                1c8a5078-4e02-42db-b15b-01ee3d03a5eb
                Copyright: © 2020 Agoti CN et al.

                This is an open access article distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                : 31 July 2020
                Funding
                Funded by: New Partnership for Africa's Development
                Award ID: DEL-15-003
                Funded by: Department for International Development, UK Government
                Funded by: African Academy of Sciences
                Award ID: DEL-15-003
                Funded by: Wellcome Trust
                Award ID: 107568
                Award ID: 107769
                Award ID: 203077
                Funded by: National Institute for Health Research
                Award ID: 17/63/82
                Funded by: Alliance for Accelerating Excellence in Science in Africa
                Award ID: DEL-15-003
                This work was supported by the Wellcome Trust through a Wellcome Intermediate Fellowship to LIOO which also supports VO [107568]; Core Support for the East African Major Overseas Programme [203077]; and support to the Initiative to Develop African Research Leaders (IDeAL) [107769]. This work was also supported by the African Academy of Sciences (AAS) through support to CA and PK as part of the IDeAL programme. IDeAL is a programme of the The Developing Excellence in Leadership, Training and Science (DELTAS) Africa [DEL-15-003]. The DELTAS Africa Initiative is an independent funding scheme of the AAS’s Alliance for Accelerating Excellence in Science in Africa (AESA) and supported by the New Partnership for Africa’s Development Planning and Coordinating Agency (NEPAD Agency). Support was also provided by the United Kingdom Department for International Development (DFID) in conjunction with Wellcome [107769 and 203077] and the National Institute for Health Research (NIHR) [project reference 17/63/82] using UK aid from the UK Government to support global health research. The views expressed in this report are those of the authors and not necessarily those of AAS, NEPAD Agency, The Wellcome, NIHR or the UK government.
                The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
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                covid-19,pooled testing,sars-cov-2,kilifi,kenya
                covid-19, pooled testing, sars-cov-2, kilifi, kenya

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