Coagulopathy and the humoral response against viral proteins in patients at different stages of COVID-19.

BACKGROUND: Patients with severe coronavirus disease 2019 (COVID-19) often present with coagulopathies and have high titres of circulating antibodies against viral proteins. OBJECTIVES: Herein, we evaluated the association between D-dimer and circulating immunoglobulin levels against viral proteins in patients at different clinical stages of COVID-19. METHODS: For this, we performed a cross-sectional study involving patients of the first wave of COVID-19 clinically classified as oligosymptomatic (n = 22), severe (n = 30), cured (n = 27) and non-infected (n = 9). Next, we measured in the plasma samples the total and fraction of immunoglobulins against the nucleoprotein (NP) and the receptor-binding domain (RBD) of the spike proteins by enzyme-linked immunosorbent assay (ELISA) assays. FINDINGS: Patients with severe disease had a coagulation disorder with high levels of D-dimer as well as circulating IgG against the NP but not the RBD compared to other groups of patients. In addition, high levels of D-dimer and IgG against the NP and RBD were associated with disease severity among the patients in this study. MAIN CONCLUSIONS: Our data suggest that IgG against NP and RBD participates in the worsening of COVID-19. Although the humoral response against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is partially understood, and more efforts are needed to clarify gaps in the knowledge of this process.

Identifying Inconclusive Data in the SARS-CoV-2 Molecular Diagnostic Using Nucleocapsid Phosphoprotein Gene as a Target.

CONTEXT.­: The gold standard test to identify the presence of SARS-CoV-2 in COVID-19 patients is the real-time reverse transcription-quantitative polymerase chain reaction (RT-qPCR), but inconclusive data and false-positive diagnosis remain the major problem of this approach. OBJECTIVE.­: To compare the fitness of 2 primer sets to the SARS-CoV-2 nucleocapsid phosphoprotein gene (NP) in the molecular diagnosis of COVID-19, we verified the inconclusive data and confidence of high cycle threshold (Ct) values in SARS-CoV-2 detection. DESIGN.­: The 970 patient samples were tested by using United States Centers for Disease Control and Prevention protocol. We compared the fitness of 2 primer sets to 2 different regions of the NP gene. In addition, we checked the consistency of positive samples with high Ct values by retesting extracted SARS-CoV-2 RNA or by second testing of patients. RESULTS.­: N1 and N2 displayed similar fitness during testing, with no differences between Ct values. Then, we verified security range Ct values related to positive diagnostics, with Ct values above 34 failing in 21 of 32 cases (65.6%) after retesting of samples. The patient samples with Ct values above 34.89 that were doubly positive revealed a low sensitivity (52.4%) and specificity (63.6%) of the test in samples with Ct values above 34. CONCLUSIONS.­: It is safe to use 1 primer set for the NP gene to identify SARS-CoV-2 in samples. However, samples with high Ct values may be considered inconclusive and retested to avoid false-positive diagnosis.