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1.
Lancet Microbe ; 2022 Mar 23.
Article in English | MEDLINE | ID: covidwho-1758021

ABSTRACT

Background: The memory immune response is crucial for preventing reinfection or reducing disease severity. However, the robustness and functionality of the humoral and T-cell response to SARS-CoV-2 remains unknown 12 months after initial infection. The aim of this study is to investigate the durability and functionality of the humoral and T-cell response to the original SARS-CoV-2 strain and variants in recovered patients 12 months after infection. Methods: In this longitudinal cohort study, we recruited participants who had recovered from COVID-19 and who were discharged from the Wuhan Research Center for Communicable Disease Diagnosis and Treatment at the Chinese Academy of Medical Sciences, Wuhan, China, between Jan 7 and May 29, 2020. Patients received a follow-up visit between Dec 16, 2020, and Jan 27, 2021. We evaluated the presence of IgM, IgA, and IgG antibodies against the SARS-CoV-2 nucleoprotein, Spike protein, and the receptor-binding domain 12 months after initial infection, using ELISA. Neutralising antibodies against the original SARS-CoV-2 strain, and the D614G, beta (B.1.351), and delta (B.1.617.2) variants were analysed using a microneutralisation assay in a subset of plasma samples. We analysed the magnitude and breadth of the SARS-CoV-2-specific memory T-cell responses using the interferon γ (IFNγ) enzyme-linked immune absorbent spot (ELISpot) assay and intracellular cytokine staining (ICS) assay. The antibody response and T-cell response (ie, IFN-γ, interleukin-2 [IL-2], and tumour necrosis factor α [TNFα]) were analysed by age and disease severity. Antibody titres were also analysed according to sequelae symptoms. Findings: We enrolled 1096 patients, including 289 (26·4%) patients with moderate initial disease, 734 (67·0%) with severe initial disease, and 73 (6·7%) with critical initial disease. Paired plasma samples were collected from 141 patients during the follow-up visits for the microneutralisation assay. PBMCs were collected from 92 of 141 individuals at the 12-month follow-up visit, of which 80 were analysed by ELISpot and 92 by ICS assay to detect the SARS-CoV-2-specific memory T-cell responses. N-IgG (899 [82·0%]), S-IgG (1043 [95·2%]), RBD-IgG (1032 [94·2%]), and neutralising (115 [81·6%] of 141) antibodies were detectable 12 months after initial infection in most individuals. Neutralising antibodies remained stable 6 and 12 months after initial infection in most individuals younger than 60 years. Multifunctional T-cell responses were detected for all SARS-CoV-2 viral proteins tested. There was no difference in the magnitude of T-cell responses or cytokine profiles in individuals with different symptom severity. Moreover, we evaluated both antibody and T-cell responses to the D614G, beta, and delta viral strains. The degree of reduced in-vitro neutralising antibody responses to the D614G and delta variants, but not to the beta variant, was associated with the neutralising antibody titres after SARS-CoV-2 infection. We also found poor neutralising antibody responses to the beta variant; 83 (72·2%) of 115 patients showed no response at all. Moreover, the neutralising antibody titre reduction of the recovered patient plasma against the delta variant was similar to that of the D614G variant and lower than that of the beta variant. By contrast, T-cell responses were cross-reactive to the beta variant in most individuals. Importantly, T-cell responses could be detected in all individuals who had lost the neutralising antibody response to SARS-CoV-2 12 months after the initial infection. Interpretation: SARS-CoV-2-specific neutralising antibody and T-cell responses were retained 12 months after initial infection. Neutralising antibodies to the D614G, beta, and delta viral strains were reduced compared with those for the original strain, and were diminished in general. Memory T-cell responses to the original strain were not disrupted by new variants. This study suggests that cross-reactive SARS-CoV-2-specific T-cell responses could be particularly important in the protection against severe disease caused by variants of concern whereas neutralising antibody responses seem to reduce over time. Funding: Chinese Academy of Medical Sciences, National Natural Science Foundation, and UK Medical Research Council.

2.
EuropePMC; 2020.
Preprint in English | EuropePMC | ID: ppcovidwho-319974

ABSTRACT

Coronavirus Disease 2019 (COVID-19) has caused global pandemic. Here we profiled the humoral response against SARS-CoV-2 by measuring immunoglobulin (Ig) A, IgM and IgG against nucleocapsid, spike proteins and IgM, IgG antibodies against receptor-binding domain (RBD) of the spike protein along with total neutralizing antibodies. We tested 279 plasma samples collected from 176 COVID-19 patients. We demonstrate more severe cases have a late onset in the humoral response compared to mild/moderate infections. All the antibody titers continue to rise in patients with COVID-19 over the disease course. However, these levels are mostly unrelated to the disease severity. The appearance time and titers of neutralizing antibodies showed significant positive correlation to the antibodies against spike protein. Our results suggest late onset of antibody response as a risk factor for disease severity, however there is a limited role of antibody titers in predicting disease severity of COVID-19.

3.
EuropePMC; 2020.
Preprint in English | EuropePMC | ID: ppcovidwho-319973

ABSTRACT

Background: The pandemic of Coronavirus disease 2019 (COVID-19) is ongoing globally, which is a big challenge for public health. Alteration of human microbiota had been observed in COVID-19. However, it is unknown how the microbiota is associated with the fatality in COVID-19.Methods: We conducted metatranscriptome sequencing on 588 longitudinal oropharyngeal swab specimens collected from 192 COVID-19 patients recruited in the LOTUS clinical trial (Registration number: ChiCTR2000029308) (including 39 deceased patients), and 95 healthy controls from the same geographic area.Findings: The upper respiratory tract (URT) microbiota in COVID-19 patients differed from that in healthy controls, while deceased patients possessed a more distinct microbiota. Streptococcus was enriched in recovered patients, whereas potential pathogens, including Candida and Enterococcus, were more abundant in deceased patients. Moreover, the microbiota dominated by Streptococcus was more stable than that dominated by other species. In contrast, the URT microbiota in deceased patients showed a more significant alteration and became more deviated from the norm after admission. The abundance of Streptococcus on admission, particularly that of S. parasanguis, was identified as a strong predictor of fatality by Cox and L1 regularized logistic regression analysis, thus could be used as a potential prognostic biomarker of COVID-19.Interpretation Alteration of the URT microbiota was observed in COVID-19 patients and was associated with the fatality rate. A higher abundance of Streptococcus, especially S. parasanguis, on admission in oropharyngeal swabs predicts a better outcome. The generalization of the results in other populations and underlying mechanisms need further investigations.Trial Registration: Participants were enrolled in ChiCTR2000029308.Funding: This study was funded in part by the National Major Science & Technology Project for Control and Prevention of Major Infectious Diseases in China (2017ZX10103004, 2018ZX10301401), the Chinese Academy of Medical Sciences (CAMS) Innovation Fund for Medical Sciences (2019-I2M-2-XX, 2016-I2M-1-014, 2018-I2M-1-003), The Non-profit Central Research Institute Fund of CAMS (2020HY320001, 2019PT310029), Beijing Advanced Innovation Center for Genomics (ICG), and Beijing Advanced Innovation Center for Structural Biology (ICSB).Declaration of Interests: All authors declare no competing interests.Ethics Approval Statement: The study was approved by the Institutional Review Board of Jin Yin-Tan Hospital (KY2020-02.01). Written informed consent was obtained from all patients or their legal representatives if they were too unwell to provide consent.

4.
EuropePMC; 2020.
Preprint in English | EuropePMC | ID: ppcovidwho-311704

ABSTRACT

Background: Seasonal human coronaviruses (HCoVs) including HCoV-229E, -OC43, -NL63 and -HKU1 are widely spreading in global human populations. However, the relevance of humoral response against seasonal HCoVs to COVID-19 pathogenesis is elusive.Methods: We profiled the temporal changes of IgG antibodies against spike (S;S-IgG) proteins of SARS-CoV-2 and seasonal HCoVs in 838 plasma samples collected from 344 COVID-19 patients. We tested the antigenic cross-reactivity of S protein between SARS-CoV-2 and seasonal HCoVs and evaluated the correlations between HCoV-OC43 S-IgG antibody and disease severity in COVID-19 patients.Findings: SARS-CoV-2 S-IgG titers mounted until days 22–28, whereas HCoV-OC43 antibody titers increased until days 15–21 and then plateaued until day 46. However, IgG antibody titers against HCoV-NL63, -229E, and -HKU1 showed no significant increasing. A two-way cross-reactivity was identified between SARS-CoV-2 and HCoV-OC43. Neutralizing antibodies against SARS-CoV-2 were not detected in healthy controls who were positive for HCoV-OC43 S-IgG. HCoV-OC43 S-IgG titers were significantly higher in patients with severe disease than those in mild/moderate patients at days 1–21 post symptom onset (PSO). Higher levels of HCoV-OC43 S-IgG were also observed in patients requiring mechanical ventilation and the elderly. At days 1–10 PSO, HCoV-OC43 S-IgG titers correlated to disease severity in all age groups, and to fatality in over 60-year group.Interpretation: Our data indicate that there exist a humoral cross-reactive response between HCoV-OC43 and SARS-CoV-2. The cross-reactive HCoV-OC43 S-IgG antibody is not protective against SARS-CoV-2, but may be a risk factor for the severity and adverse outcome of COVID-19.Funding Statement: This study was funded in part by the National Major Science & Technology Project for Control and Prevention of Major Infectious Diseases in China (2017ZX10204401, 2018ZX10734404), Chinese Academy of Medical Sciences (CAMS) Innovation Fund for Medical Sciences (2016-I2M-1–014, 2018-I2M-1-003, 2020-I2M-1-001, 2020-I2M-CoV19-005), Natural Science Foundation of China (82041011/H0104), and National Key R&D Program of China (2020YFA0707600). Declaration of Interests: The authors declare no competing interests.Ethics Approval Statement: This study was approved by the Ethical Review Board of Wuhan Jinyintan Hospital, Infectious Disease Hospital of Heilongjiang Province (Harbin), and Institute of Pathogen Biology, Chinese Academy of Medical Sciences. Written informed consent was obtained from each healthy volunteer and COVID-19 patients in cohort 4. Written informed consents from the remaining patients were waived in light of the emerging infectious disease of high public health relevance.

5.
Am J Respir Crit Care Med ; 204(12): 1379-1390, 2021 12 15.
Article in English | MEDLINE | ID: covidwho-1430274

ABSTRACT

Rationale: Alteration of human respiratory microbiota had been observed in coronavirus disease (COVID-19). How the microbiota is associated with the prognosis in COVID-19 is unclear. Objectives: To characterize the feature and dynamics of the respiratory microbiota and its associations with clinical features in patients with COVID-19. Methods: We conducted metatranscriptome sequencing on 588 longitudinal oropharyngeal swab specimens collected from 192 patients with COVID-19 (including 39 deceased patients) and 95 healthy controls from the same geographic area. Meanwhile, the concentration of 27 cytokines and chemokines in plasma was measured for patients with COVID-19. Measurements and Main Results: The upper respiratory tract (URT) microbiota in patients with COVID-19 differed from that in healthy controls, whereas deceased patients possessed a more distinct microbiota, both on admission and before discharge/death. The alteration of URT microbiota showed a significant correlation with the concentration of proinflammatory cytokines and mortality. Specifically, Streptococcus-dominated microbiota was enriched in recovered patients, and showed high temporal stability and resistance against pathogens. In contrast, the microbiota in deceased patients was more susceptible to secondary infections and became more deviated from the norm after admission. Moreover, the abundance of S. parasanguinis on admission was significantly correlated with prognosis in nonsevere patients (lower vs. higher abundance, odds ratio, 7.80; 95% CI, 1.70-42.05). Conclusions: URT microbiota dysbiosis is a remarkable manifestation of COVID-19; its association with mortality suggests it may reflect the interplay between pathogens, symbionts, and the host immune status. Whether URT microbiota could be used as a biomarker for diagnosis and prognosis of respiratory diseases merits further investigation.


Subject(s)
COVID-19/microbiology , COVID-19/mortality , Microbiota , Respiratory Tract Infections/microbiology , Respiratory Tract Infections/mortality , Adult , Aged , COVID-19/epidemiology , Female , Humans , Male , Middle Aged , Prognosis , SARS-CoV-2
6.
Biosaf Health ; 3(5): 238-243, 2021 Oct.
Article in English | MEDLINE | ID: covidwho-1401267

ABSTRACT

Many factors have been identified as having the ability to affect the sensitivity of rapid antigen detection (RAD) tests for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). This study aimed to identify the impact of sample processing on the sensitivity of the RAD tests. We explored the effect of different inactivation methods, viral transport media (VTM) solutions, and sample preservation on the sensitivity of four RAD kits based on two SARS-CoV-2 strains. Compared with non-inactivation, heat inactivation significantly impacted the sensitivity of most RAD kits; however, ß-propiolactone inactivation only had a minor effect. Some of the VTM solutions (VTM2, MANTACC) had a significant influence on the sensitivity of the RAD kits, especially for low viral-loads samples. The detection value of RAD kits was slightly decreased, while most of them were still in the detection range with the extension of preservation time and the increase of freeze-thaw cycles. Our results showed that selecting the appropriate inactivation methods and VTM solutions is necessary during reagent development, performance evaluation, and clinical application.

8.
Clin Infect Dis ; 72(10): e545-e551, 2021 05 18.
Article in English | MEDLINE | ID: covidwho-1232187

ABSTRACT

BACKGROUND: The characteristics of neutralizing antibodies (NAbs) and antibody against major antigen proteins related to clinical outcomes in severe coronavirus disease 2019 (COVID-19) patients were still less known. METHODS: NAbs and antibodies targeting nucleocapsid (N), spike protein (S), and the receptor-binding domain (RBD) in longitudinal plasma samples from the LOTUS China trial were measured by microneutralization assay and enzyme-linked immunosorbent assay (ELISA). Viral load was determined by real-time reverse transcription polymerase chain reaction (RT-PCR). A total of 576 plasma and 576 throat swabs were collected from 191 COVID-19 patients. Antibody titers related to adverse outcome and clinical improvement were analyzed. Multivariable adjusted generalized linear mixed model for random effects were developed. RESULTS: After day 28 post symptoms onset, the rate of antibody positivity reached 100% for RBD-immunoglobulin M (IgM), 97.8% for S-IgM, 100% for N-immunoglobulin G (IgG), 100% for RBD-IgG, 91.1% for N-IgM, and 91.1% for NAbs. The NAbs titers increased over time in both survivors and nonsurvivors and correlated to IgG antibodies against N, S, and RBD, whereas its presence showed no statistical correlation with death. N-IgG (slope -2.11, 95% confidence interval [CI] -3.04 to -1.18, P < .0001), S-IgG (slope -2.44, 95% CI -3.35 to -1.54, P < .0001), and RBD-IgG (slope -1.43, 95% CI -1.98 to -.88, P < .0001) were negatively correlated with viral load. S-IgG titers were lower in nonsurvivors than survivors (P = .020) at week 4 after symptoms onset. CONCLUSIONS: IgM and IgG against N, S, and RBD and NAbs developed in most severe COVID-19 patients and do not correlate clearly with clinical outcomes. The levels of IgG antibodies against N, S, and RBD were related to viral clearance.


Subject(s)
COVID-19 , Adult , Antibodies, Viral , Antibody Formation , China/epidemiology , Humans , Immunoglobulin M , SARS-CoV-2
9.
Lancet ; 397(10279): 1075-1084, 2021 03 20.
Article in English | MEDLINE | ID: covidwho-1142326

ABSTRACT

BACKGROUND: Wuhan was the epicentre of the COVID-19 outbreak in China. We aimed to determine the seroprevalence and kinetics of anti-SARS-CoV-2 antibodies at population level in Wuhan to inform the development of vaccination strategies. METHODS: In this longitudinal cross-sectional study, we used a multistage, population-stratified, cluster random sampling method to systematically select 100 communities from the 13 districts of Wuhan. Households were systematically selected from each community and all family members were invited to community health-care centres to participate. Eligible individuals were those who had lived in Wuhan for at least 14 days since Dec 1, 2019. All eligible participants who consented to participate completed a standardised electronic questionnaire of demographic and clinical questions and self-reported any symptoms associated with COVID-19 or previous diagnosis of COVID-19. A venous blood sample was taken for immunological testing on April 14-15, 2020. Blood samples were tested for the presence of pan-immunoglobulins, IgM, IgA, and IgG antibodies against SARS-CoV-2 nucleocapsid protein and neutralising antibodies were assessed. We did two successive follow-ups between June 11 and June 13, and between Oct 9 and Dec 5, 2020, at which blood samples were taken. FINDINGS: Of 4600 households randomly selected, 3599 families (78·2%) with 9702 individuals attended the baseline visit. 9542 individuals from 3556 families had sufficient samples for analyses. 532 (5·6%) of 9542 participants were positive for pan-immunoglobulins against SARS-CoV-2, with a baseline adjusted seroprevalence of 6·92% (95% CI 6·41-7·43) in the population. 437 (82·1%) of 532 participants who were positive for pan-immunoglobulins were asymptomatic. 69 (13·0%) of 532 individuals were positive for IgM antibodies, 84 (15·8%) were positive for IgA antibodies, 532 (100%) were positive for IgG antibodies, and 212 (39·8%) were positive for neutralising antibodies at baseline. The proportion of individuals who were positive for pan-immunoglobulins who had neutralising antibodies in April remained stable for the two follow-up visits (162 [44·6%] of 363 in June, 2020, and 187 [41·2%] of 454 in October-December, 2020). On the basis of data from 335 individuals who attended all three follow-up visits and who were positive for pan-immunoglobulins, neutralising antibody levels did not significantly decrease over the study period (median 1/5·6 [IQR 1/2·0 to 1/14·0] at baseline vs 1/5·6 [1/4·0 to 1/11·2] at first follow-up [p=1·0] and 1/6·3 [1/2·0 to 1/12·6] at second follow-up [p=0·29]). However, neutralising antibody titres were lower in asymptomatic individuals than in confirmed cases and symptomatic individuals. Although titres of IgG decreased over time, the proportion of individuals who had IgG antibodies did not decrease substantially (from 30 [100%] of 30 at baseline to 26 [89·7%] of 29 at second follow-up among confirmed cases, 65 [100%] of 65 at baseline to 58 [92·1%] of 63 at second follow-up among symptomatic individuals, and 437 [100%] of 437 at baseline to 329 [90·9%] of 362 at second follow-up among asymptomatic individuals). INTERPRETATION: 6·92% of a cross-sectional sample of the population of Wuhan developed antibodies against SARS-CoV-2, with 39·8% of this population seroconverting to have neutralising antibodies. Our durability data on humoral responses indicate that mass vaccination is needed to effect herd protection to prevent the resurgence of the epidemic. FUNDING: Chinese Academy of Medical Sciences & Peking Union Medical College, National Natural Science Foundation, and Chinese Ministry of Science and Technology. TRANSLATION: For the Chinese translation of the abstract see Supplementary Materials section.


Subject(s)
Antibodies, Neutralizing/blood , Antibodies, Viral/blood , COVID-19/immunology , SARS-CoV-2/immunology , Adolescent , Adult , Aged , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19/blood , COVID-19/epidemiology , COVID-19/prevention & control , Child , Child, Preschool , China/epidemiology , Coronavirus Nucleocapsid Proteins/immunology , Cross-Sectional Studies , Female , Follow-Up Studies , Humans , Immunity, Herd/immunology , Immunity, Humoral , Infant , Infant, Newborn , Longitudinal Studies , Male , Mass Vaccination/organization & administration , Middle Aged , Seroepidemiologic Studies , Young Adult
10.
Emerg Microbes Infect ; 10(1): 664-676, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1139855

ABSTRACT

Seasonal human coronaviruses (HCoVs) including HCoV-229E, -OC43, -NL63, and -HKU1 widely spread in global human populations. However, the relevance of humoral response against seasonal HCoVs to COVID-19 pathogenesis is elusive. In this study, we profiled the temporal changes of IgG antibody against spike proteins (S-IgG) of SARS-CoV-2 and seasonal HCoVs in 838 plasma samples collected from 344 COVID-19 patients. We tested the antigenic cross-reactivities of S protein between SARS-CoV-2 and seasonal HCoVs and evaluated the correlations between the levels of HCoV-OC43 S-IgG and the disease severity in COVID-19 patients. We found that SARS-CoV-2 S-IgG titres mounted until days 22-28, whereas HCoV-OC43 antibody titres increased until days 15-21 and then plateaued until day 46. However, IgG titres against HCoV-NL63, -229E, and -HKU1 showed no significant increase. A two-way cross-reactivity was identified between SARS-CoV-2 and HCoV-OC43. Neutralizing antibodies against SARS-CoV-2 were not detectable in healthy controls who were positive for HCoV-OC43 S-IgG. HCoV-OC43 S-IgG titres were significantly higher in patients with severe disease than those in mild patients at days 1-21 post symptom onset (PSO). Higher levels of HCoV-OC43 S-IgG were also observed in patients requiring mechanical ventilation. At days 1-10 PSO, HCoV-OC43 S-IgG titres correlated to disease severity in the age group over 60. Our data indicate that there is a correlation between cross-reactive antibody against HCoV-OC43 spike protein and disease severity in COVID-19 patients.


Subject(s)
Antibodies, Viral/blood , COVID-19/immunology , Coronavirus OC43, Human/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Adolescent , Adult , Aged , Aged, 80 and over , Antibodies, Neutralizing/blood , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19/pathology , Cross Reactions/immunology , Enzyme-Linked Immunosorbent Assay , Female , Humans , Immunoglobulin G/blood , Immunoglobulin G/immunology , Male , Middle Aged , Severity of Illness Index , Young Adult
11.
Cell Mol Immunol ; 18(4): 945-953, 2021 04.
Article in English | MEDLINE | ID: covidwho-1104474

ABSTRACT

SARS-CoV-2 is the pathogenic agent of COVID-19, which has evolved into a global pandemic. Compared with some other respiratory RNA viruses, SARS-CoV-2 is a poor inducer of type I interferon (IFN). Here, we report that SARS-CoV-2 nsp12, the viral RNA-dependent RNA polymerase (RdRp), suppresses host antiviral responses. SARS-CoV-2 nsp12 attenuated Sendai virus (SeV)- or poly(I:C)-induced IFN-ß promoter activation in a dose-dependent manner. It also inhibited IFN promoter activation triggered by RIG-I, MDA5, MAVS, and IRF3 overexpression. Nsp12 did not impair IRF3 phosphorylation but suppressed the nuclear translocation of IRF3. Mutational analyses suggested that this suppression was not dependent on the polymerase activity of nsp12. Given these findings, our study reveals that SARS-CoV-2 RdRp can antagonize host antiviral innate immunity and thus provides insights into viral pathogenesis.


Subject(s)
COVID-19/metabolism , Coronavirus RNA-Dependent RNA Polymerase/metabolism , Interferon Regulatory Factor-3/metabolism , Interferon Type I/metabolism , SARS-CoV-2/metabolism , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/metabolism , Cell Nucleus/metabolism , DEAD Box Protein 58/genetics , DEAD Box Protein 58/metabolism , Host-Pathogen Interactions/immunology , Humans , Immunity, Innate , Interferon Regulatory Factor-3/genetics , Interferon Type I/genetics , Interferon-Induced Helicase, IFIH1/genetics , Interferon-Induced Helicase, IFIH1/metabolism , Interferon-beta/genetics , Interferon-beta/metabolism , Mutation , Phosphorylation , Promoter Regions, Genetic , Receptors, Immunologic/genetics , Receptors, Immunologic/metabolism , SARS-CoV-2/enzymology , Sendai virus/metabolism
12.
Front Immunol ; 11: 586572, 2020.
Article in English | MEDLINE | ID: covidwho-979019

ABSTRACT

COVID-19 pandemic has infected millions of people with mortality exceeding >1 million. There is an urgent need to find therapeutic agents that can help clear the virus to prevent severe disease and death. Identifying effective and safer drugs can provide more options to treat COVID-19 infections either alone or in combination. Here, we performed a high throughput screening of approximately 1,700 US FDA-approved compounds to identify novel therapeutic agents that can effectively inhibit replication of coronaviruses including SARS-CoV-2. Our two-step screen first used a human coronavirus strain OC43 to identify compounds with anti-coronaviral activities. The effective compounds were then screened for their effectiveness in inhibiting SARS-CoV-2. These screens have identified 20 anti-SARS-CoV-2 drugs including previously reported compounds such as hydroxychloroquine, amlodipine besylate, arbidol hydrochloride, tilorone 2HCl, dronedarone hydrochloride, mefloquine, and thioridazine hydrochloride. Five of the newly identified drugs had a safety index (cytotoxic/effective concentration) of >600, indicating a wide therapeutic window compared to hydroxychloroquine which had a safety index of 22 in similar experiments. Mechanistically, five of the effective compounds (fendiline HCl, monensin sodium salt, vortioxetine, sertraline HCl, and salifungin) were found to block SARS-CoV-2 S protein-mediated cell fusion. These FDA-approved compounds can provide much needed therapeutic options that we urgently need during the midst of the pandemic.


Subject(s)
Antiviral Agents/therapeutic use , COVID-19/drug therapy , High-Throughput Screening Assays/methods , Pandemics/prevention & control , SARS-CoV-2/drug effects , Animals , COVID-19/epidemiology , COVID-19/virology , Cell Line , Drug Repositioning/methods , Fendiline/therapeutic use , HEK293 Cells , Humans , Monensin/therapeutic use , SARS-CoV-2/physiology , Salicylanilides/therapeutic use , Sertraline/therapeutic use , Vortioxetine/therapeutic use
13.
Commun Biol ; 3(1): 780, 2020 12 11.
Article in English | MEDLINE | ID: covidwho-975030

ABSTRACT

Coronavirus Disease 2019 (COVID-19) has caused a global pandemic. Here we profiled the humoral response against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) by measuring immunoglobulin (Ig) A, IgM, and IgG against nucleocapsid and spike proteins, along with IgM and IgG antibodies against receptor-binding domain (RBD) of the spike protein and total neutralizing antibodies (NAbs). We tested 279 plasma samples collected from 176 COVID-19 patients who presented and enrolled at different stages of their disease. Plasma dilutions were optimized and based on the data, a single dilution of plasma was used. The mean absorbance at 450 nm was measured for Ig levels and NAbs were measured using geometric mean titers. We demonstrate that more severe cases have a late-onset in the humoral response compared to mild/moderate infections. All the antibody titers continue to rise in patients with COVID-19 over the disease course. However, these levels are mostly unrelated to disease severity. The appearance time and titers of NAbs showed a significant positive correlation to the antibodies against spike protein. Our results suggest the late onset of antibody response as a risk factor for disease severity, however, there is a limited role of antibody titers in predicting disease severity of COVID-19.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19/immunology , Immunity, Humoral/immunology , SARS-CoV-2/immunology , Adolescent , Adult , Aged , Aged, 80 and over , Animals , Antibodies, Neutralizing/blood , Antibodies, Viral/blood , COVID-19/epidemiology , COVID-19/virology , China/epidemiology , Chlorocebus aethiops , Female , Humans , Kinetics , Male , Middle Aged , Pandemics , SARS-CoV-2/physiology , Severity of Illness Index , Spike Glycoprotein, Coronavirus/immunology , Vero Cells , Young Adult
14.
Emerg Microbes Infect ; 9(1): 2707-2714, 2020 Dec.
Article in English | MEDLINE | ID: covidwho-953975

ABSTRACT

To identify the association between the kinetics of viral load and clinical outcome in severe coronavirus disease 2019 (COVID-19) patients, a retrospective study was performed by involved 188 hospitalized severe COVID-19 patients in the LOTUS China trial. Among the collected 578 paired throat swab (TS) and anal swab (AS) samples, viral RNA was detected in 193 (33.4%) TS and 121 (20.9%) AS. A higher viral RNA load was found in TS than that of AS, with means of 1.0 × 106 and 2.3 × 105 copies/ml, respectively. In non-survivors, the viral RNA in AS was detected earlier than that in survivors (median of 14 days vs 19 days, P = 0.007). The positivity and viral load in AS were higher in non-survivors than that of survivors at week 2 post symptom onset (P = 0.006). A high initial viral load in AS was associated with death (OR 1.368, 95% CI 1.076-1.741, P = 0.011), admission to the intensive care unit (OR 1.237, 95% CI 1.001-1.528, P = 0.049) and need for invasive mechanical ventilation (OR 1.340, 95% CI 1.076-1.669, P = 0.009). Our findings indicated viral replication in extrapulmonary sites should be monitored intensively during antiviral therapy.


Subject(s)
Anal Canal/virology , COVID-19/virology , SARS-CoV-2/isolation & purification , Viral Load , Adolescent , Adult , Aged , Aged, 80 and over , COVID-19/mortality , Female , Humans , Male , Middle Aged , Pharynx/virology , RNA, Viral/analysis , Retrospective Studies , Time Factors , Virus Replication , Young Adult
16.
Nat Commun ; 11(1): 4528, 2020 09 10.
Article in English | MEDLINE | ID: covidwho-759594

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spreads worldwide and leads to an unprecedented medical burden and lives lost. Neutralizing antibodies provide efficient blockade for viral infection and are a promising category of biological therapies. Here, using SARS-CoV-2 spike receptor-binding domain (RBD) as a bait, we generate a panel of humanized single domain antibodies (sdAbs) from a synthetic library. These sdAbs reveal binding kinetics with the equilibrium dissociation constant (KD) of 0.99-35.5 nM. The monomeric sdAbs show half maximal neutralization concentration (EC50) of 0.0009-0.07 µg/mL and 0.13-0.51 µg/mL against SARS-CoV-2 pseudotypes, and authentic SARS-CoV-2, respectively. Competitive ligand-binding experiments suggest that the sdAbs either completely block or significantly inhibit the association between SARS-CoV-2 RBD and viral entry receptor ACE2. Fusion of the human IgG1 Fc to sdAbs improve their neutralization activity by up to ten times. These results support neutralizing sdAbs as a potential alternative for antiviral therapies.


Subject(s)
Antibodies, Neutralizing/immunology , Coronavirus Infections/virology , Pneumonia, Viral/virology , Single-Domain Antibodies/immunology , Spike Glycoprotein, Coronavirus/immunology , Angiotensin-Converting Enzyme 2 , Animals , Antibodies, Neutralizing/pharmacology , COVID-19 , Chlorocebus aethiops , Coronavirus Infections/drug therapy , HEK293 Cells , Humans , Immunoglobulin G , Models, Molecular , Pandemics , Peptidyl-Dipeptidase A/metabolism , Pneumonia, Viral/drug therapy , Protein Binding , Receptors, Virus/immunology , Single-Domain Antibodies/pharmacology , Spike Glycoprotein, Coronavirus/metabolism , Vero Cells
17.
Nat Commun ; 11(1): 3810, 2020 07 30.
Article in English | MEDLINE | ID: covidwho-690732

ABSTRACT

The pandemic of COVID-19 has posed an unprecedented threat to global public health. However, the interplay between the viral pathogen of COVID-19, SARS-CoV-2, and host innate immunity is poorly understood. Here we show that SARS-CoV-2 induces overt but delayed type-I interferon (IFN) responses. By screening 23 viral proteins, we find that SARS-CoV-2 NSP1, NSP3, NSP12, NSP13, NSP14, ORF3, ORF6 and M protein inhibit Sendai virus-induced IFN-ß promoter activation, whereas NSP2 and S protein exert opposite effects. Further analyses suggest that ORF6 inhibits both type I IFN production and downstream signaling, and that the C-terminus region of ORF6 is critical for its antagonistic effect. Finally, we find that IFN-ß treatment effectively blocks SARS-CoV-2 replication. In summary, our study shows that SARS-CoV-2 perturbs host innate immune response via both its structural and nonstructural proteins, and thus provides insights into the pathogenesis of SARS-CoV-2.


Subject(s)
Betacoronavirus/physiology , Coronavirus Infections/virology , Immune Evasion , Interferon Type I/metabolism , Pneumonia, Viral/virology , Signal Transduction , Betacoronavirus/genetics , Betacoronavirus/immunology , Betacoronavirus/metabolism , COVID-19 , Cell Line , Coronavirus Infections/immunology , Humans , Immunity, Innate , Interferon-beta/genetics , Interferon-beta/metabolism , Interferon-beta/pharmacology , Mutation , Open Reading Frames , Pandemics , Pneumonia, Viral/immunology , Promoter Regions, Genetic , SARS-CoV-2 , Signal Transduction/drug effects , Viral Proteins/genetics , Viral Proteins/metabolism , Virus Replication/drug effects
18.
JAMA ; 324(5): 460-470, 2020 08 04.
Article in English | MEDLINE | ID: covidwho-506058

ABSTRACT

Importance: Convalescent plasma is a potential therapeutic option for patients with coronavirus disease 2019 (COVID-19), but further data from randomized clinical trials are needed. Objective: To evaluate the efficacy and adverse effects of convalescent plasma therapy for patients with COVID-19. Design, Setting, and Participants: Open-label, multicenter, randomized clinical trial performed in 7 medical centers in Wuhan, China, from February 14, 2020, to April 1, 2020, with final follow-up April 28, 2020. The trial included 103 participants with laboratory-confirmed COVID-19 that was severe (respiratory distress and/or hypoxemia) or life-threatening (shock, organ failure, or requiring mechanical ventilation). The trial was terminated early after 103 of a planned 200 patients were enrolled. Intervention: Convalescent plasma in addition to standard treatment (n = 52) vs standard treatment alone (control) (n = 51), stratified by disease severity. Main Outcomes and Measures: Primary outcome was time to clinical improvement within 28 days, defined as patient discharged alive or reduction of 2 points on a 6-point disease severity scale (ranging from 1 [discharge] to 6 [death]). Secondary outcomes included 28-day mortality, time to discharge, and the rate of viral polymerase chain reaction (PCR) results turned from positive at baseline to negative at up to 72 hours. Results: Of 103 patients who were randomized (median age, 70 years; 60 [58.3%] male), 101 (98.1%) completed the trial. Clinical improvement occurred within 28 days in 51.9% (27/52) of the convalescent plasma group vs 43.1% (22/51) in the control group (difference, 8.8% [95% CI, -10.4% to 28.0%]; hazard ratio [HR], 1.40 [95% CI, 0.79-2.49]; P = .26). Among those with severe disease, the primary outcome occurred in 91.3% (21/23) of the convalescent plasma group vs 68.2% (15/22) of the control group (HR, 2.15 [95% CI, 1.07-4.32]; P = .03); among those with life-threatening disease the primary outcome occurred in 20.7% (6/29) of the convalescent plasma group vs 24.1% (7/29) of the control group (HR, 0.88 [95% CI, 0.30-2.63]; P = .83) (P for interaction = .17). There was no significant difference in 28-day mortality (15.7% vs 24.0%; OR, 0.59 [95% CI, 0.22-1.59]; P = .30) or time from randomization to discharge (51.0% vs 36.0% discharged by day 28; HR, 1.61 [95% CI, 0.88-2.95]; P = .12). Convalescent plasma treatment was associated with a negative conversion rate of viral PCR at 72 hours in 87.2% of the convalescent plasma group vs 37.5% of the control group (OR, 11.39 [95% CI, 3.91-33.18]; P < .001). Two patients in the convalescent plasma group experienced adverse events within hours after transfusion that improved with supportive care. Conclusion and Relevance: Among patients with severe or life-threatening COVID-19, convalescent plasma therapy added to standard treatment, compared with standard treatment alone, did not result in a statistically significant improvement in time to clinical improvement within 28 days. Interpretation is limited by early termination of the trial, which may have been underpowered to detect a clinically important difference. Trial Registration: Chinese Clinical Trial Registry: ChiCTR2000029757.


Subject(s)
Betacoronavirus/immunology , Coronavirus Infections/therapy , Pneumonia, Viral/therapy , Aged , Aged, 80 and over , Blood Component Transfusion , COVID-19 , China , Combined Modality Therapy , Coronavirus Infections/mortality , Female , Humans , Immunization, Passive/adverse effects , Male , Middle Aged , Pandemics , Patient Acuity , Pneumonia, Viral/mortality , SARS-CoV-2 , Treatment Outcome
19.
Nature ; 583(7818): 830-833, 2020 07.
Article in English | MEDLINE | ID: covidwho-220333

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of coronavirus disease 2019 (COVID-19), which has become a public health emergency of international concern1. Angiotensin-converting enzyme 2 (ACE2) is the cell-entry receptor for severe acute respiratory syndrome coronavirus (SARS-CoV)2. Here we infected transgenic mice that express human ACE2 (hereafter, hACE2 mice) with SARS-CoV-2 and studied the pathogenicity of the virus. We observed weight loss as well as virus replication in the lungs of hACE2 mice infected with SARS-CoV-2. The typical histopathology was interstitial pneumonia with infiltration of considerable numbers of macrophages and lymphocytes into the alveolar interstitium, and the accumulation of macrophages in alveolar cavities. We observed viral antigens in bronchial epithelial cells, macrophages and alveolar epithelia. These phenomena were not found in wild-type mice infected with SARS-CoV-2. Notably, we have confirmed the pathogenicity of SARS-CoV-2 in hACE2 mice. This mouse model of SARS-CoV-2 infection will be valuable for evaluating antiviral therapeutic agents and vaccines, as well as understanding the pathogenesis of COVID-19.


Subject(s)
Betacoronavirus/pathogenicity , Coronavirus Infections/pathology , Coronavirus Infections/virology , Lung/pathology , Peptidyl-Dipeptidase A/genetics , Peptidyl-Dipeptidase A/metabolism , Pneumonia, Viral/pathology , Pneumonia, Viral/virology , Transgenes , Angiotensin-Converting Enzyme 2 , Animals , Antigens, Viral/immunology , Antigens, Viral/metabolism , Betacoronavirus/immunology , Betacoronavirus/metabolism , Bronchi/pathology , Bronchi/virology , COVID-19 , Coronavirus Infections/immunology , Disease Models, Animal , Epithelial Cells/pathology , Epithelial Cells/virology , Female , Humans , Immunoglobulin G/immunology , Lung/immunology , Lung/virology , Lymphocytes/immunology , Macrophages, Alveolar/immunology , Macrophages, Alveolar/virology , Male , Mice , Mice, Transgenic , Pandemics , Pneumonia, Viral/immunology , Receptors, Complement 3d/genetics , Receptors, Complement 3d/metabolism , SARS-CoV-2 , Virus Replication , Weight Loss
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