Ratiometric fluorescent Si-FITC nanoprobe for immunoassay of SARS-CoV-2 nucleocapsid protein.

Coronavirus disease 2019 (COVID-19) highlights the importance of rapid and reliable diagnostic assays for the management of virus transmission. Here, we developed a one-pot hydrothermal method to prepare Si-FITC nanoparticles (NPs) for the fluorescent immunoassay of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid protein (N protein). The synthesis of Si-FITC NPs did not need post-modification, which addressed the issue of quantum yield reduction during the coupling reaction. Si-FITC NPs showed two distinct peaks, Si fluorescence at λ em = 385 nm and FITC fluorescence at λ em = 490 nm. In the presence of KMnO4, Si fluorescence was decreased and FITC fluorescence was enhanced. Briefly, in the presence of N protein, catalase (CAT)-linked secondary antibody/reporter antibody/N protein/capture antibody immunocomplexes were formed on microplates. Subsequently, hydrogen peroxide (H2O2) and Si-FITC NPs/KMnO4 were injected into the microplate together. The decomposition of H2O2 by CAT resulted in remaining of KMnO4, which changed the fluorescence intensity ratio of Si-FITC NPs. The fluorescence intensity ratio correlated significantly with the N protein concentration ranging from 0.02 to 50.00 ng/mL, and the detection limit was 0.003 ng/mL, which was more sensitive than the commercial ELISA kit with a detection limit of 0.057 ng/mL. The N protein concentration can be accurately determined in human serum. Furthermore, the COVID-19 and non-COVID-19 patients were distinguishable by this method. Therefore, the ratiometric fluorescent immunoassay can be used for SARS-CoV-2 infection diagnosis with a high sensitivity and selectivity. Electronic Supplementary Material: Supplementary material (characterization of Si-FITC NPs (FTIR, HRXPS); stability investigation of Si-FITC NPs (photostability, pH stability, anti-interference ability); stability investigation of free FITC (pH value, KMnO4); quenching mechanism of KMnO4 (UV-vis absorption spectra, fluorescence lifetime decay curves); reaction condition optimization of biotin-CAT with H2O2 (pH value, temperature, time); detection of N protein using commercial ELISA Kit; selectivity investigation of assays for SARS-CoV-2 N protein detection; determination results of SARS-CoV-2 N protein in human serum) is available in the online version of this article at 10.1007/s12274-022-5005-z.

Association of individual resilience with organizational resilience, perceived social support, and job performance among healthcare professionals in township health centers of China during the COVID-19 pandemic.

Background: Primary healthcare professionals were overworked and psychologically overwhelmed during the COVID-19 pandemic. Resilience is an important shield for individuals to cope with psychological stress and improve performance in crises. This study aims to explore the association of individual resilience with organizational resilience, perceived social support and job performance among healthcare professionals in township health centers of China during the COVID-19 pandemic. Methods: Data from 1,266 questionnaires were collected through a cross-sectional survey conducted in December 2021 in Shandong Province, China. Descriptive analysis of individual resilience, organizational resilience, perceived social support, and job performance was conducted. Pearson correlation analysis was used to examine the correlations among these variables, and structural equation modeling was performed to verify the relationships between these variables. Results: The score of individual resilience was 101.67 ± 14.29, ranging from 24 to 120. Organizational resilience (ß = 0.409, p < 0.01) and perceived social support (ß = 0.410, p < 0.01) had significant direct effects on individual resilience. Individual resilience (ß = 0.709, p < 0.01) had a significant direct effect on job performance. Organizational resilience (ß = 0.290, p < 0.01) and perceived social support (ß = 0.291, p < 0.01) had significant indirect effects on job performance. Conclusion: During the COVID-19 pandemic, the individual resilience of healthcare professionals in township health centers was at a moderate level. Organizational resilience and perceived social support positively affected individual resilience, and individual resilience positively affected job performance. Furthermore, individual resilience mediated the effect of organizational resilience and perceived social support on job performance. It is recommended that multiple stakeholders work together to improve the individual resilience of primary healthcare professionals.

Thought and prospect of safety clinical transfusion under major public health emergency

As of the end of January 2020, 31 provinces, municipalities and autonomous regions across the country had activated their Level 1 responses to major public health emergencies due to the outbreak of the novel coronavirus. Under the state of emergency prevention and control, work and production were suspended, cities and roads were closed, people were isolated at home, blood donations were cancelled, and blood stock management was severely tested. If the epidemic continues to develop or an unknown public health event occurs, it is imperative that blood collection agencies and hospital transfusion departments at all levels respond in a safe and effective manner to safeguard the clinical blood supply. This article summarises the measures that can be taken to ensure adequate blood stocks and the safety of blood collection and supply in the event of a major public health emergency, with a view to providing new ideas for clinical blood collection and supply and safe clinical blood use.

Ratiometric fluorescent Si-FITC nanoprobe for immunoassay of SARS-CoV-2 nucleocapsid protein

Coronavirus disease 2019 (COVID-19) highlights the importance of rapid and reliable diagnostic assays for the management of virus transmission. Here, we developed a one-pot hydrothermal method to prepare Si-FITC nanoparticles (NPs) for the fluorescent immunoassay of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid protein (N protein). The synthesis of Si-FITC NPs did not need post-modification, which addressed the issue of quantum yield reduction during the coupling reaction. Si-FITC NPs showed two distinct peaks, Si fluorescence at λem = 385 nm and FITC fluorescence at λem = 490 nm. In the presence of KMnO4, Si fluorescence was decreased and FITC fluorescence was enhanced. Briefly, in the presence of N protein, catalase (CAT)-linked secondary antibody/reporter antibody/N protein/capture antibody immunocomplexes were formed on microplates. Subsequently, hydrogen peroxide (H2O2) and Si-FITC NPs/KMnO4 were injected into the microplate together. The decomposition of H2O2 by CAT resulted in remaining of KMnO4, which changed the fluorescence intensity ratio of Si-FITC NPs. The fluorescence intensity ratio correlated significantly with the N protein concentration ranging from 0.02 to 50.00 ng/mL, and the detection limit was 0.003 ng/mL, which was more sensitive than the commercial ELISA kit with a detection limit of 0.057 ng/mL. The N protein concentration can be accurately determined in human serum. Furthermore, the COVID-19 and non-COVID-19 patients were distinguishable by this method. Therefore, the ratiometric fluorescent immunoassay can be used for SARS-CoV-2 infection diagnosis with a high sensitivity and selectivity. Electronic Supplementary Material Supplementary material (characterization of Si-FITC NPs (FTIR, HRXPS);stability investigation of Si-FITC NPs (photostability, pH stability, anti-interference ability);stability investigation of free FITC (pH value, KMnO4);quenching mechanism of KMnO4 (UV-vis absorption spectra, fluorescence lifetime decay curves);reaction condition optimization of biotin-CAT with H2O2 (pH value, temperature, time);detection of N protein using commercial ELISA Kit;selectivity investigation of assays for SARS-CoV-2 N protein detection;determination results of SARS-CoV-2 N protein in human serum) is available in the online version of this article at 10.1007/s12274-022-5005-z.

Total infectome characterization of respiratory infections in pre-COVID-19 Wuhan, China.

At the end of 2019 Wuhan witnessed an outbreak of "atypical pneumonia" that later developed into a global pandemic. Metagenomic sequencing rapidly revealed the causative agent of this outbreak to be a novel coronavirus denoted SARS-CoV-2. To provide a snapshot of the pathogens in pneumonia-associated respiratory samples from Wuhan prior to the emergence of SARS-CoV-2, we collected bronchoalveolar lavage fluid samples from 408 patients presenting with pneumonia and acute respiratory infections at the Central Hospital of Wuhan between 2016 and 2017. Unbiased total RNA sequencing was performed to reveal their "total infectome", including viruses, bacteria and fungi. We identified 35 pathogen species, comprising 13 RNA viruses, 3 DNA viruses, 16 bacteria and 3 fungi, often at high abundance and including multiple co-infections (13.5%). SARS-CoV-2 was not present. These data depict a stable core infectome comprising common respiratory pathogens such as rhinoviruses and influenza viruses, an atypical respiratory virus (EV-D68), and a single case of a sporadic zoonotic pathogen-Chlamydia psittaci. Samples from patients experiencing respiratory disease on average had higher pathogen abundance than healthy controls. Phylogenetic analyses of individual pathogens revealed multiple origins and global transmission histories, highlighting the connectedness of the Wuhan population. This study provides a comprehensive overview of the pathogens associated with acute respiratory infections and pneumonia, which were more diverse and complex than obtained using targeted PCR or qPCR approaches. These data also suggest that SARS-CoV-2 or closely related viruses were absent from Wuhan in 2016-2017.

Depressive State in the Emergency Department During COVID-19: A National Cross-Sectional Survey in China.

Chinese emergency department (ED) staff encountered significant mental stress while fighting the coronavirus disease 2019 (COVID-19) pandemic. We sought to investigate the prevalence and associated factors for depressive symptoms among ED staff (including physicians, nurses, allied health, and auxiliary ED staff). A cross-sectional national survey of ED staff who were on duty and participated in combating the COVID-19 pandemic was conducted March 1-15, 2020. A total of 6,588 emergency medical personnel from 1,060 hospitals responded to this survey. A majority of respondents scored above 10 points on the PHQ-9 standardized test, which is associated with depressive symptoms. Those aged 31-45, those working in the COVID-19 isolation unit, and those with relatives ≤ 16 or ≥70 years old at home all had statistically significant associations with scoring >10 points. Depressive symptoms among Chinese emergency medical staff were likely quite common during the response to the COVID-19 pandemic and reinforce the importance of targeted ED staff support during future outbreaks.

Remdesivir in adults with severe COVID-19: a randomised, double-blind, placebo-controlled, multicentre trial.

BACKGROUND: No specific antiviral drug has been proven effective for treatment of patients with severe coronavirus disease 2019 (COVID-19). Remdesivir (GS-5734), a nucleoside analogue prodrug, has inhibitory effects on pathogenic animal and human coronaviruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in vitro, and inhibits Middle East respiratory syndrome coronavirus, SARS-CoV-1, and SARS-CoV-2 replication in animal models. METHODS: We did a randomised, double-blind, placebo-controlled, multicentre trial at ten hospitals in Hubei, China. Eligible patients were adults (aged ≥18 years) admitted to hospital with laboratory-confirmed SARS-CoV-2 infection, with an interval from symptom onset to enrolment of 12 days or less, oxygen saturation of 94% or less on room air or a ratio of arterial oxygen partial pressure to fractional inspired oxygen of 300 mm Hg or less, and radiologically confirmed pneumonia. Patients were randomly assigned in a 2:1 ratio to intravenous remdesivir (200 mg on day 1 followed by 100 mg on days 2-10 in single daily infusions) or the same volume of placebo infusions for 10 days. Patients were permitted concomitant use of lopinavir-ritonavir, interferons, and corticosteroids. The primary endpoint was time to clinical improvement up to day 28, defined as the time (in days) from randomisation to the point of a decline of two levels on a six-point ordinal scale of clinical status (from 1=discharged to 6=death) or discharged alive from hospital, whichever came first. Primary analysis was done in the intention-to-treat (ITT) population and safety analysis was done in all patients who started their assigned treatment. This trial is registered with ClinicalTrials.gov, NCT04257656. FINDINGS: Between Feb 6, 2020, and March 12, 2020, 237 patients were enrolled and randomly assigned to a treatment group (158 to remdesivir and 79 to placebo); one patient in the placebo group who withdrew after randomisation was not included in the ITT population. Remdesivir use was not associated with a difference in time to clinical improvement (hazard ratio 1·23 [95% CI 0·87-1·75]). Although not statistically significant, patients receiving remdesivir had a numerically faster time to clinical improvement than those receiving placebo among patients with symptom duration of 10 days or less (hazard ratio 1·52 [0·95-2·43]). Adverse events were reported in 102 (66%) of 155 remdesivir recipients versus 50 (64%) of 78 placebo recipients. Remdesivir was stopped early because of adverse events in 18 (12%) patients versus four (5%) patients who stopped placebo early. INTERPRETATION: In this study of adult patients admitted to hospital for severe COVID-19, remdesivir was not associated with statistically significant clinical benefits. However, the numerical reduction in time to clinical improvement in those treated earlier requires confirmation in larger studies. FUNDING: Chinese Academy of Medical Sciences Emergency Project of COVID-19, National Key Research and Development Program of China, the Beijing Science and Technology Project.

Predictive factors of severe coronavirus disease 2019 in previously healthy young adults: a single-center, retrospective study.

BACKGROUND: Several previously healthy young adults have developed Coronavirus Disease 2019 (COVID-19), and a few of them progressed to the severe stage. However, the factors are not yet determined. METHOD: We retrospectively analyzed 123 previously healthy young adults diagnosed with COVID-19 from January to March 2020 in a tertiary hospital in Wuhan. Patients were classified as having mild or severe COVID-19 based on their respiratory rate, SpO2, and PaO2/FiO2 levels. Patients' symptoms, computer tomography (CT) images, preadmission drugs received, and the serum biochemical examination on admission were compared between the mild and severe groups. Significant variables were enrolled into logistic regression model to predict the factors affecting disease severity. A receiver operating characteristic (ROC) curve was applied to validate the predictive value of predictors. RESULT: Age; temperature; anorexia; and white blood cell count, neutrophil percentage, platelet count, lymphocyte count, C-reactive protein, aspartate transaminase, creatine kinase, albumin, and fibrinogen values were significantly different between patients with mild and severe COVID-19 (P < 0.05). Logistic regression analysis confirmed that lymphopenia (P = 0.010) indicated severe prognosis in previously healthy young adults with COVID-19, with the area under the curve (AUC) was 0.791(95% Confidence Interval (CI) 0.704-0.877)(P < 0.001). CONCLUSION: For previously healthy young adults with COVID-19, lymphopenia on admission can predict severe prognosis.

The biological characteristics of SARS-CoV-2 spike protein Pro330-Leu650.

SARS-CoV-2 is the cause of the worldwide outbreak of COVID-19 that has been characterized as a pandemic by the WHO. Since the first report of COVID-19 on December 31, 2019, 179,111 cases were confirmed in 160 countries/regions with 7426 deaths as of March 17, 2020. However, there have been no vaccines approved in the world to date. In this study, we analyzed the biological characteristics of the SARS-CoV-2 Spike protein, Pro330-Leu650 (SARS-CoV-2-SPL), using biostatistical methods. SARS-CoV-2-SPL possesses a receptor-binding region (RBD) and important B (Ser438-Gln506, Thr553-Glu583, Gly404-Aps427, Thr345-Ala352, and Lys529-Lys535) and T (9 CD4 and 11 CD8 T cell antigenic determinants) cell epitopes. High homology in this region between SARS-CoV-2 and SARS-CoV amounted to 87.7%, after taking the biological similarity of the amino acids into account and eliminating the receptor-binding motif (RBM). The overall topology indicated that the complete structure of SARS-CoV-2-SPL was with RBM as the head, and RBD as the trunk and the tail region. SARS-CoV-2-SPL was found to have the potential to elicit effective B and T cell responses. Our findings may provide meaningful guidance for SARS-CoV-2 vaccine design.

Effects of Corticosteroid Treatment for Non-Severe COVID-19 Pneumonia: A Propensity Score-Based Analysis.

BACKGROUND AND OBJECTIVE: The effects of corticosteroid treatment on non-severe COVID-19 pneumonia patients are unknown. To determine the impacts of adjuvant corticosteroid administrated to patients with non-severe COVID-19 pneumonia. METHOD: A retrospective cohort study based on propensity score analysis was designed to explore the effects of corticosteroid on several clinical outcomes. RESULTS: One hundred thirty-two patients satisfied the inclusion criteria and 35 pairs were generated according to propensity score matching. Compared to non-corticosteroid group, the CT score on day 7 was significantly higher in corticosteroid group (8.6 (interquartile range [IQR], 2.8-11.5) versus 12.0 (IQR, 5.0-19.3), P = 0.046). In corticosteroid group, more patients progressed to severe cases (11.4% versus 2.9%, P = 0.353), hospital stay (23.5 days (IQR, 19-29 d) versus 20.2 days (IQR, 14-25.3 d), P = 0.079) and duration of viral shedding (20.3 days (IQR, 15.2-24.8 d) versus 19.4 days (IQR, 11.5-28.3 d), P = 0.669) were prolonged, while fever time (9.5 days (IQR, 6.5-12.2 d) versus 10.2 days (IQR, 6.8-14 d), P = 0.28) was shortened; however, all these data revealed no statistically significant differences. CONCLUSION: Corticosteroid might have a negative effect on lung injury recovery in non-severe COVID-19 pneumonia patients; however, the results of this study must be interpreted with caution because of confounding factors.

Hospitalization and Critical Care of 109 Decedents with COVID-19 Pneumonia in Wuhan, China.

Rationale: The current outbreak of coronavirus disease (COVID-19) pneumonia caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in Wuhan, China, spreads across national and international borders. The overall death rate of COVID-19 pneumonia in the Chinese population was 4%.Objectives: To describe the process of hospitalization and critical care of patients who died of COVID-19 pneumonia.Methods: This was a multicenter observational study of 109 decedents with COVID-19 pneumonia from three hospitals in Wuhan. Demographic, clinical, laboratory, and treatment data were collected and analyzed, and the final date of follow-up was February 24, 2020.Results: The mean age of 109 decedents with COVID-19 pneumonia was 70.7 years, 35 patients (32.1%) were female, and 85 patients (78.0%) suffered from one or more underlying comorbidities. Multiple organ failure, especially respiratory failure and heart failure, appeared in all patients even at the early stage of disease. Overall, the mean time from onset of symptoms to death was 22.3 days. All 109 hospitalized patients needed admission to an intensive care unit (ICU); however, because of limited availability, only 51 (46.8%) could be admitted. The period from hospitalization to death in the ICU group and non-ICU group was 15.9 days (standard deviation = 8.8 d) and 12.5 days (8.6 d, P = 0.044), respectively.Conclusions: Mortality due to COVID-19 pneumonia was concentrated in patients above the age of 65 years, especially those with major comorbidities. Patients who were admitted to the ICU lived longer than those who were not. Our findings should aid in the recognition and clinical management of such infections, especially with regard to ICU resource allocation.

Association of radiologic findings with mortality of patients infected with 2019 novel coronavirus in Wuhan, China.

Radiologic characteristics of 2019 novel coronavirus (2019-nCoV) infected pneumonia (NCIP) which had not been fully understood are especially important for diagnosing and predicting prognosis. We retrospective studied 27 consecutive patients who were confirmed NCIP, the clinical characteristics and CT image findings were collected, and the association of radiologic findings with mortality of patients was evaluated. 27 patients included 12 men and 15 women, with median age of 60 years (IQR 47-69). 17 patients discharged in recovered condition and 10 patients died in hospital. The median age of mortality group was higher compared to survival group (68 (IQR 63-73) vs 55 (IQR 35-60), P = 0.003). The comorbidity rate in mortality group was significantly higher than in survival group (80% vs 29%, P = 0.018). The predominant CT characteristics consisted of ground glass opacity (67%), bilateral sides involved (86%), both peripheral and central distribution (74%), and lower zone involvement (96%). The median CT score of mortality group was higher compared to survival group (30 (IQR 7-13) vs 12 (IQR 11-43), P = 0.021), with more frequency of consolidation (40% vs 6%, P = 0.047) and air bronchogram (60% vs 12%, P = 0.025). An optimal cutoff value of a CT score of 24.5 had a sensitivity of 85.6% and a specificity of 84.5% for the prediction of mortality. 2019-nCoV was more likely to infect elderly people with chronic comorbidities. CT findings of NCIP were featured by predominant ground glass opacities mixed with consolidations, mainly peripheral or combined peripheral and central distributions, bilateral and lower lung zones being mostly involved. A simple CT scoring method was capable to predict mortality.

Clinical features of patients infected with 2019 novel coronavirus in Wuhan, China.

BACKGROUND: A recent cluster of pneumonia cases in Wuhan, China, was caused by a novel betacoronavirus, the 2019 novel coronavirus (2019-nCoV). We report the epidemiological, clinical, laboratory, and radiological characteristics and treatment and clinical outcomes of these patients. METHODS: All patients with suspected 2019-nCoV were admitted to a designated hospital in Wuhan. We prospectively collected and analysed data on patients with laboratory-confirmed 2019-nCoV infection by real-time RT-PCR and next-generation sequencing. Data were obtained with standardised data collection forms shared by WHO and the International Severe Acute Respiratory and Emerging Infection Consortium from electronic medical records. Researchers also directly communicated with patients or their families to ascertain epidemiological and symptom data. Outcomes were also compared between patients who had been admitted to the intensive care unit (ICU) and those who had not. FINDINGS: By Jan 2, 2020, 41 admitted hospital patients had been identified as having laboratory-confirmed 2019-nCoV infection. Most of the infected patients were men (30 [73%] of 41); less than half had underlying diseases (13 [32%]), including diabetes (eight [20%]), hypertension (six [15%]), and cardiovascular disease (six [15%]). Median age was 49·0 years (IQR 41·0-58·0). 27 (66%) of 41 patients had been exposed to Huanan seafood market. One family cluster was found. Common symptoms at onset of illness were fever (40 [98%] of 41 patients), cough (31 [76%]), and myalgia or fatigue (18 [44%]); less common symptoms were sputum production (11 [28%] of 39), headache (three [8%] of 38), haemoptysis (two [5%] of 39), and diarrhoea (one [3%] of 38). Dyspnoea developed in 22 (55%) of 40 patients (median time from illness onset to dyspnoea 8·0 days [IQR 5·0-13·0]). 26 (63%) of 41 patients had lymphopenia. All 41 patients had pneumonia with abnormal findings on chest CT. Complications included acute respiratory distress syndrome (12 [29%]), RNAaemia (six [15%]), acute cardiac injury (five [12%]) and secondary infection (four [10%]). 13 (32%) patients were admitted to an ICU and six (15%) died. Compared with non-ICU patients, ICU patients had higher plasma levels of IL2, IL7, IL10, GSCF, IP10, MCP1, MIP1A, and TNFα. INTERPRETATION: The 2019-nCoV infection caused clusters of severe respiratory illness similar to severe acute respiratory syndrome coronavirus and was associated with ICU admission and high mortality. Major gaps in our knowledge of the origin, epidemiology, duration of human transmission, and clinical spectrum of disease need fulfilment by future studies. FUNDING: Ministry of Science and Technology, Chinese Academy of Medical Sciences, National Natural Science Foundation of China, and Beijing Municipal Science and Technology Commission.