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• W4207074361 abstract "We read with interest the paper of Li et al. reporting serum amyloid A (SAA) as a possible biomarker to evaluate the severity of coronavirus disease and progression. (1Li H. Xiang X. Ren H. Xu L. Zhao L. Chen X. et al.Serum Amyloid A is a biomarker of severe Coronavirus Disease and poor prognosis.J Infect. Jun 2020; 80 (PubMed PMID: 32277967. Pubmed Central PMCID: PMC7141628. Epub 2020/04/12): 646-655Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar) They found that the ratio of SAA to lymphocyte (SAA/L) was significantly increased in critically severe patients. A recent meta-analysis also supports the use of this biomarker to assess the severity of the disease although a high heterogeneity among the studies was reported. (2Zinellu A. Paliogiannis P. Carru C. Mangoni A.A. Serum amyloid A concentrations, COVID-19 severity and mortality: an updated systematic review and meta-analysis.Int J Infect Dis. 2021; 105 (2021-04-01): 668-674Abstract Full Text Full Text PDF PubMed Scopus (10) Google Scholar) Other biomarkers have also been linked with disease severity and complications and we would like to share our data regarding the association between the level of SARS-CoV-2 nucleocapsid antigens and the severity of COVID-19. Eighty-one unvaccinated patients with documented nasopharyngeal (NP) RT-PCR positive for SARS-CoV-2 (eSwab tubes; Copan Italia, Brescia, Italy) for which a serum sample (BD SST II Advance®; Becton Dickinson, NJ, USA) was obtained within 12 h of NP sampling were included. The median delta collection time was 1 min (10th - 90th percentiles: −74 to 183 min). Thirty-six patients (44.4%) were women (median age = 77 years, min-max = 20-97) and 45 (55.6%) were men (median age = 76.5 years, min-max = 33–94). Eighteen patients were categorized as asymptomatics (WHO score of 1), 6 as symptomatic ambulatory mild disease (WHO score of 2 and 3), 43 as symptomatic hospitalized moderate disease (WHO scores of 4 and 5) and 14 as symptomatic hospitalized severe disease patients (WHO score of 6 to 10). Eleven patients died (WHO score of 10). The mean time since symptoms was 5.5 days (standard deviation (SD): 4.4) and 7.1 days (SD: 7.2) in non-severe (WHO scores of 2 to 5) and severe patients (WHO score of 6 to 10), respectively. The SARS-CoV-2 nucleocapsid (N) antigen was quantified automatically in patient sera by a Single Molecular Array (Simoa) immunoassay using the Simoa HD-X analyzer (Quanterix, Massachusetts, USA), as described elsewhere. (3Shan D. Johnson J.M. Fernandes S.C. Suib H. Hwang S. Wuelfing D. et al.N-protein presents early in blood, dried blood and saliva during asymptomatic and symptomatic SARS-CoV-2 infection.Nat Commun. 2021; 12 (Mar 26PubMed PMID: 33771993. Pubmed Central PMCID: PMC7997897. Epub 2021/03/28): 1931Crossref PubMed Scopus (19) Google Scholar) The results are quantitative and expressed as pg/mL. Serum samples that provided results upper the calibration range were retested using a 1000-fold dilution. The LOD of the assay is 0.099 pg/mL and was considered as positivity cut-off. The within-run coefficient of variation (CV) is less than 10% according to the information of the manufacturer. RT-PCR was performed on the GeneXpert instrument (Cepheid, California, USA) using the Xpress SARS-CoV-2 assay targeting N2 and E genes for the majority of the samples and on a the LightCycler 480 Instrument II (Roche Diagnostics, Rotkreuz, Switzerland) using the LightMix Modular SARS-CoV E-gene set for very few samples obtained during day-care. The mean Ct values obtained by RT-PCR were significantly lower in case of serum antigen positivity (p < 0.0001, unpaired t-test) (Fig. 1A). Using a Ct threshold of 33 as a surrogate of contagiousness(4Basile K. McPhie K. Carter I. Alderson S. Rahman H. Donovan L. et al.Cell-based Culture Informs Infectivity and Safe De-Isolation Assessments in Patients with Coronavirus Disease 2019.Clin Infect Dis. 2021; 73 (Nov 2PubMed PMID: 33098412. Pubmed Central PMCID: PMC7665383. Epub 2020/10/25): e2952-e29e9Crossref PubMed Scopus (35) Google Scholar), the clinical sensitivity was 100% and the clinical specificity was 92.3%. Indeed, patients with low viral load (i.e. Ct > 33) have a very low risk of being infective and capable of transmitting the virus. Therefore, the NP RT-PCR positivity in these subjects may be attributable to residual low viral load(4Basile K. McPhie K. Carter I. Alderson S. Rahman H. Donovan L. et al.Cell-based Culture Informs Infectivity and Safe De-Isolation Assessments in Patients with Coronavirus Disease 2019.Clin Infect Dis. 2021; 73 (Nov 2PubMed PMID: 33098412. Pubmed Central PMCID: PMC7665383. Epub 2020/10/25): e2952-e29e9Crossref PubMed Scopus (35) Google Scholar). Wang et al. also found that negative plasma N antigen in COVID-19 patients corresponded to higher Ct values (38.2 = median; IQR = 37.4–39.2). (5Wang H. Hogan C.A. Verghese M. Solis D. Sibai M. Huang C. et al.SARS-CoV-2 Nucleocapsid Plasma Antigen for Diagnosis and Monitoring of COVID-19.Clin. Chem. 2021; (Oct 4PubMed PMID: 34605900. Pubmed Central PMCID: PMC8522398. Epub 2021/10/05)Crossref Scopus (2) Google Scholar) Highly significant correlations were found between RT-PCR results and antigen levels (Pearson r = −0.61; p < 0.0001) (Fig. 1B). Other studies found a clinical sensitivity ranging from 85.2% to 93.0% (5Wang H. Hogan C.A. Verghese M. Solis D. Sibai M. Huang C. et al.SARS-CoV-2 Nucleocapsid Plasma Antigen for Diagnosis and Monitoring of COVID-19.Clin. Chem. 2021; (Oct 4PubMed PMID: 34605900. Pubmed Central PMCID: PMC8522398. Epub 2021/10/05)Crossref Scopus (2) Google Scholar, 6Hingrat Q.L. Visseaux B. Laouenan C. Tubiana S. Bouadma L. Yazdanpanah Y. et al.Detection of SARS-CoV-2N-antigen in blood during acute COVID-19 provides a sensitive new marker and new testing alternatives.Clin Microbiol Infect. 2020 Dec 8; (PubMed PMID: 33307227. Pubmed Central PMCID: PMC7724284. Epub 2020/12/12)Google Scholar, 7Zhang Y., Ong C.M., Yun C., Mo W., Whitman J.D., Lynch K.L., et al. Diagnostic Value of Nucleocapsid Protein in Blood for SARS-CoV-2 Infection. Clinical chemistry. 2021 Aug 6. PubMed PMID: 34358289. Pubmed Central PMCID: PMC8436384. Epub 2021/08/07.Google Scholar) in patients that developed symptoms up to maximum two weeks. The clinical sensitivity increased to 94.2% to 100% considering samples collected within the first days since the onset of symptoms. After 2 weeks, the clinical sensitivity progressively decreased to values comprised between 43.2% to 74.5%. (3Shan D. Johnson J.M. Fernandes S.C. Suib H. Hwang S. Wuelfing D. et al.N-protein presents early in blood, dried blood and saliva during asymptomatic and symptomatic SARS-CoV-2 infection.Nat Commun. 2021; 12 (Mar 26PubMed PMID: 33771993. Pubmed Central PMCID: PMC7997897. Epub 2021/03/28): 1931Crossref PubMed Scopus (19) Google Scholar, 5Wang H. Hogan C.A. Verghese M. Solis D. Sibai M. Huang C. et al.SARS-CoV-2 Nucleocapsid Plasma Antigen for Diagnosis and Monitoring of COVID-19.Clin. Chem. 2021; (Oct 4PubMed PMID: 34605900. Pubmed Central PMCID: PMC8522398. Epub 2021/10/05)Crossref Scopus (2) Google Scholar, 6Hingrat Q.L. Visseaux B. Laouenan C. Tubiana S. Bouadma L. Yazdanpanah Y. et al.Detection of SARS-CoV-2N-antigen in blood during acute COVID-19 provides a sensitive new marker and new testing alternatives.Clin Microbiol Infect. 2020 Dec 8; (PubMed PMID: 33307227. Pubmed Central PMCID: PMC7724284. Epub 2020/12/12)Google Scholar, 7Zhang Y., Ong C.M., Yun C., Mo W., Whitman J.D., Lynch K.L., et al. Diagnostic Value of Nucleocapsid Protein in Blood for SARS-CoV-2 Infection. Clinical chemistry. 2021 Aug 6. PubMed PMID: 34358289. Pubmed Central PMCID: PMC8436384. Epub 2021/08/07.Google Scholar) Given that the peak of N antigens is reached after 7 days, as for the viral load in NP samples, (8Lippi G. Simundic A.M. Plebani M. Potential preanalytical and analytical vulnerabilities in the laboratory diagnosis of coronavirus disease 2019 (COVID-19).Clin Chem Lab Med. 2020 Jun 25; 58 (PubMed PMID: 32172228. Epub 2020/03/17): 1070-1076Crossref PubMed Scopus (311) Google Scholar) and that a continuous decline is observed afterwards, the timing since symptoms is therefore essential for the evaluation of antigenemia. Interestingly, we found that N antigen levels were significantly increased in severe patients (median = 7673 pg/mL) compared to moderately affected (351.6 pg/mL, p = 0.0174) and asymptomatic patients (median = 4.6 pg/mL, p < 0.0001). Moderate patients also had significantly higher antigen levels compared to asymptomatic subjects (p = 0.0030) (Fig. 2A). Using ROC curve analysis for the classification of hospitalized (WHO score of 4 to 10) versus non-hospitalized patients (WHO score of 1 to 3), a cut-off of 1631.9 pg/mL was identified (sensitivity = 45.6%, specificity = 95.8%; AUC = 0.76, p < 0.0001) (Fig. 2B – green line). This AUC was similar to the one found by Li et al. using SAA/L for clinical classification (i.e. AUC = 0.75)(1Li H. Xiang X. Ren H. Xu L. Zhao L. Chen X. et al.Serum Amyloid A is a biomarker of severe Coronavirus Disease and poor prognosis.J Infect. Jun 2020; 80 (PubMed PMID: 32277967. Pubmed Central PMCID: PMC7141628. Epub 2020/04/12): 646-655Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar). For the classification of severe (WHO score of 6 to 10) versus non-severe patients (WHO score of 1 to 5), a cut-off of 4039.4 pg/mL was identified with a sensitivity of 64.0% and a specificity of 89.0%. The calculated AUC was 0.84 (p < 0.0001) (Fig. 2C – green line). Higher concentrations of N antigens were also observed in other studies in more severe patients (3Shan D. Johnson J.M. Fernandes S.C. Suib H. Hwang S. Wuelfing D. et al.N-protein presents early in blood, dried blood and saliva during asymptomatic and symptomatic SARS-CoV-2 infection.Nat Commun. 2021; 12 (Mar 26PubMed PMID: 33771993. Pubmed Central PMCID: PMC7997897. Epub 2021/03/28): 1931Crossref PubMed Scopus (19) Google Scholar, 5Wang H. Hogan C.A. Verghese M. Solis D. Sibai M. Huang C. et al.SARS-CoV-2 Nucleocapsid Plasma Antigen for Diagnosis and Monitoring of COVID-19.Clin. Chem. 2021; (Oct 4PubMed PMID: 34605900. Pubmed Central PMCID: PMC8522398. Epub 2021/10/05)Crossref Scopus (2) Google Scholar, 7Zhang Y., Ong C.M., Yun C., Mo W., Whitman J.D., Lynch K.L., et al. Diagnostic Value of Nucleocapsid Protein in Blood for SARS-CoV-2 Infection. Clinical chemistry. 2021 Aug 6. PubMed PMID: 34358289. Pubmed Central PMCID: PMC8436384. Epub 2021/08/07.Google Scholar, 9Brasen C.L. Christensen H. Olsen D.A. Kahns S. Andersen R.F. Madsen J.B. et al.Daily monitoring of viral load measured as SARS-CoV-2 antigen and RNA in blood, IL-6, CRP and complement C3d predicts outcome in patients hospitalized with COVID-19.Clin Chem Lab Med. 2021 Nov 25; 59 (PubMed PMID: 34455731. Epub 2021/08/30): 1988-1997Crossref PubMed Scopus (4) Google Scholar, 10Perna F. Bruzzaniti S. Piemonte E. Maddaloni V. Atripaldi L. Sale S. et al.Serum levels of SARS-CoV-2 nucleocapsid antigen associate with inflammatory status and disease severity in COVID-19 patients.Clin Immunol. May 2021; 226 (PubMed PMID: 33819577. Pubmed Central PMCID: PMC8017913. Epub 2021/04/06)108720Crossref PubMed Scopus (6) Google Scholar) as well as positive correlations with inflammatory biomarker levels (i.e. CRP or IL-6). (9Brasen C.L. Christensen H. Olsen D.A. Kahns S. Andersen R.F. Madsen J.B. et al.Daily monitoring of viral load measured as SARS-CoV-2 antigen and RNA in blood, IL-6, CRP and complement C3d predicts outcome in patients hospitalized with COVID-19.Clin Chem Lab Med. 2021 Nov 25; 59 (PubMed PMID: 34455731. Epub 2021/08/30): 1988-1997Crossref PubMed Scopus (4) Google Scholar, 10Perna F. Bruzzaniti S. Piemonte E. Maddaloni V. Atripaldi L. Sale S. et al.Serum levels of SARS-CoV-2 nucleocapsid antigen associate with inflammatory status and disease severity in COVID-19 patients.Clin Immunol. May 2021; 226 (PubMed PMID: 33819577. Pubmed Central PMCID: PMC8017913. Epub 2021/04/06)108720Crossref PubMed Scopus (6) Google Scholar) Our study confirms that severe patients exhibit higher N antigen levels compared to non-severe at the time of diagnosis. This may help in patients’ triage and monitoring of those more prone to develop a severe form of the disease. Compared to N antigen levels in serum, Ct values of RT-PCR were less associated to severity, especially considering the hospitalization status (Fig. 2B and C – blue line). In our cohort, only asymptomatics had significantly higher Ct values compared to severe patients (28.4 versus 18.8, p = 0.037), an observation consistent with previous investigations. (5Wang H. Hogan C.A. Verghese M. Solis D. Sibai M. Huang C. et al.SARS-CoV-2 Nucleocapsid Plasma Antigen for Diagnosis and Monitoring of COVID-19.Clin. Chem. 2021; (Oct 4PubMed PMID: 34605900. Pubmed Central PMCID: PMC8522398. Epub 2021/10/05)Crossref Scopus (2) Google Scholar) In conclusion, sensitive N antigen determination in serum provides a valuable marker for COVID-19 diagnosis and evaluation of severity. Further studies with more patients are needed to complement our data. A better discrimination of N antigen levels based on the days since symptoms as well as correlation with antibody seropositive are also needed. None. Conceptualization: JFA – JBA – JDO; Data curation: JFA – JBA – CDA – JDO; Formal analysis: JFA – JBA – CDA – JDO; Funding acquisition: JDO; Investigation: JFA – JDO; Methodology: JFA – JDO; Project administration: JMD – JDO; Resources: JDM – JDO; Supervision: JDO; Validation: JFA – JDO; Visualization: JFA – JDO; Writing – original draft: JFA – JDO; Writing – review & editing: JFA – JBA – CDA – JMD – JDO. The data that support the findings of this study are available from the corresponding author JDO, upon reasonable request. The authors would like to thank the technical teams from the laboratories of QUALIblood, Clinique Saint-Luc Bouge and Clinique Saint-Pierre Ottignies for collecting the samples and performing the analyses." @default.
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• W4207074361 title "Nucleocapsid serum antigen determination in SARS-CoV-2 infected patients using the single molecule array technology and prediction of disease severity" @default.
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