The predicting roles of carcinoembryonic antigen and its underlying mechanism in the progression of coronavirus disease 2019 (preprint)

Background The coronavirus disease 2019 (COVID-19) has induced a worldwide pneumonia with a high infectivity and mortality. However, the predicting biomarkers and their potential mechanism in the progression of COVID-19 are not well known.Objective The aim of this study is to identify the candidate predictors of COVID-19 and investigate their underlying mechanism.Methods The retrospective study was conducted to identify the potential laboratory indicators with prognostic values of COVID-19 disease. Then, the prognostic nomogram was constructed to predict the overall survival of COVID-19 patients. Additionally, the scRNA-seq data of BALF and PBMCs from COVID-19 patients were downloaded to investigate the underlying mechanism of the most important prognostic indicators in lungs and peripherals, respectively.Results 304 hospitalized adult COVID-19 patients in Wuhan Jinyintan Hospital were included in the retrospective study. CEA was the only laboratory indicator with significant difference in the univariate (P < 0.001) and multivariate analysis (P = 0.021). The scRNA-seq data of BALF and PBMCs from COVID-19 patients were downloaded to investigate the underlying mechanism of CEA in lungs and peripherals, respectively. The results revealed the potential roles of CEA were significantly distributed in Type II pneumocytes of BALF and developing neutrophils of PBMCs, participating in the progression of COVID-19 by regulating the cell-cell communication.Conclusion This study identifies the prognostic roles of CEA in COVID-19 patients and implies the potential roles of CEACAM8-CEACAM6 in the progression of COVID-19 by regulating the cell-cell communication of developing neutrophils and Type II pneumocyte.

Secondary Infection in Severe and Critical COVID-19 Patients in China: A Multicenter Retrospective Study (preprint)

Background. Since 2020 COVID-19 pandemic became an emergent public sanitary incident. The epidemiology data and the impact on prognosis of secondary infection in severe and critical COVID-19 patients in China remained largely unclear.Methods. We retrospectively reviewed medical records of all adult patients with laboratory-confirmed COVID-19 who were admitted to ICUs from January 18th 2020 to April 26th 2020 at two hospitals in Wuhan, China and one hospital in Guangzhou, China. We measured the frequency of bacteria and fungi cultured from respiratory tract, blood and other body fluid specimens. The risk factors for and impact of secondary infection on clinical outcomes were also assessed. Results. Secondary infections were very common (86.6%) when patients were admitted to ICU for >72 hours. The majority of infections were respiratory, with the most common organisms being Klebsiella pneumoniae (24.5%), Acinetobacter baumannii (21.8%), Stenotrophomonas maltophilia (9.9%), Candida albicans (6.8%), and Pseudomonas spp. (4.8%). Furthermore, the proportions of multidrug resistant (MDR) bacteria and carbapenem resistant Enterobacteriaceae (CRE) were high. We also found that age ≥60 years and mechanical ventilation ≥13days independently increased the likelihood of secondary infection. Finally, patients with positive cultures had reduced ventilator free days in 28 days and patients with CRE and/or MDR bacteria positivity showed lower 28 day survival rate.Conclusions. In a retrospective cohort of severe and critical COVID-19 patients admitted to ICUs in China, the prevalence of secondary infection was high, especially with CRE and MDR bacteria, resulting in poor clinical outcomes.

Dynamics of the Upper Respiratory Tract Microbiota and Its Association with Fatality in COVID-19 Patients (preprint)

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.

Association between Risk of Venous Thromboembolism and Mortality in Patients with COVID-19 (preprint)

Background: Hospitalized patients with COVID-19 appeared high risk of venous thromboembolism (VTE) defined by some risk assessment models (RAMs), which exhibited the predictor of mortality in patients with non-COVID-19. We aimed to investigate the association between risk of VTE with 30-day mortality in COVID-19 patients.Methods: In this retrospective cohort study, 1030 consecutive hospitalized patients, between January 26, 2020 and March 29, 2020, aged 14–98 years with a confirmed diagnosis of COVID-19-related pneumonia were recruited from Jinyintan Hospital and Union Hosptial, Wuhan, China. We collected baseline data on demographics, SOFA parameters, and VTE RAMs including Padua Prediction Score (PPS), IMPROVE RAM and Caprini RAM. The primary outcome of the study was 30-day mortality. The secondary outcome was the length of stay in hospital. Findings: Thirty-day mortality increased progressively from 2% in patients at low risk of VTE to 63% in those at high risk of VTE defined by PPS ≥ 4. Similar findings were also observed for risk of VTE defined by IMPROVE score ³ 2 and Caprini score ³ 5. Progressive increases in VTE risk also were associated with higher SOFA score. Our findings showed that the presence of high risk of VTE was independently associated with 30-day mortality regardless of adjusted gender, smoking status and some comorbidities with hazard ratios of 29.19 (95% CI 15.76 - 54.05), 37.37 (95% CI 18.43 - 75.78), 20.60 (95% CI 11.41 - 37.19) for PPS, IMPROVE RAM and Caprini RAM, respectively (P < 0.001 for all comparisons). Predictive accuracy of PPS (AUC, 0.900; 95% CI 0.881 - 0.919), IMPROVE RAM (AUC, 0.917; 95% CI 0.898 - 0.936) or Caprini RAM (AUC, 0.861; 95% CI 0.840 - 0.882) as the risk of 30-day mortality was markedly well.Interpretation: The presence of high risk of VTE identifies a group of patients with COVID-19 at higher risk for 30-day mortality. Furthermore, there is higher accuracy of VTE RAMs to predict 30-day mortality in COVID-19 hospitalized patients.Funding Statement: This study is funded by the National Nature Science Foundation of China (81870062 to Jinjun Jiang, 81900038 to Shujing Chen).Declaration of Interests: The authors have no conflict of interest to disclose.Ethics Approval Statement: The study was approved by Jinyintan Hospital Ethics Committee (KY2020-06.01) and Union Hospital Ethics Committee (2020-0039). Written informed consent was waived by the Ethics Commission.

Dynamics of the upper respiratory tract microbiota and its association with fatality in COVID-19 patients (preprint)

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. 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. 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. parasanguinis, 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. The generalization of the results in other populations and underlying mechanisms needs further investigations.

Pathological and molecular examination findings in postmortem testis biopsies reveal spermatogenesis damages in COVID-19 patients (preprint)

Despite widespread interest in the pathophysiology of COVID-19 in respiratory and cardiovascular systems, little is known about the morphologic and molecular changes in the testis of patients with COVID-19 and the effects of SARS-CoV-2 infection on male fertility. We report here on the pathophysiology and molecular feature of testes obtained at autopsy from six men with COVID-19, as compared with those of testes from three men with age-matched, uninfected SARS-CoV-2. Our histopathological results showed that all COVID-19 patients had severe spermatogenesis damages compared with controls. Importantly, we detected the nuclear acid of the SARS-CoV-2 virus, viral particles, and SARS-CoV-2 spike S1 protein in COVID-19 patient testes, and we also found ACE2 and TMPRSS2 significantly elevated in the testes from COVID-19 patients. Furthermore, we observed a prominent leukocyte infiltration, including CD3+ T lymphocytes, CD20+ B lymphocytes, CD68+ macrophages, HLA-DR+ myeloid cells, and CD38+ plasma cells in the testes of COVID-19 patients. RNA-Seq analyses further revealed SARS-CoV-2 infection could lead to dysfunction of the genes that regulate the spermatogenesis and inflammation response-related pathways. Collectively, our pathological and molecular examination findings indicate that SARS-CoV-2 could directly attack testicular cells, thereby inducing the damage of testicular immune privilege and spermatogenesis defects.

Augmentation of anti-MDA5 antibody implies severe disease in COVID-19 patients (preprint)

Recent studies have provided insights into the autoinflammation triggered by severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) infection, which is associated with high mortality of coronavirus disease 2019 (COVID-19). Striking similarities has been noted between COVID-19 and anti-melanoma differentiation-associated gene 5 (MDA5) antibody (Ab)-related dermatomyositis (DM), implying a shared autoinflammatory aberrance. However, it is unclear whether anti-MDA5 Ab is present in COVID-19 and correlates with the severity and adverse outcome of COVID-19 patients. Here, we found that the positive rate of anti-MDA5 Ab in patients with COVID-19 was 48.2% and the anti-MDA5 Ab positive patients tended to develop severe disease (88.6% vs 66.9%, P<0.0001). In particular, the titer of anti-MDA5 Ab was increased in the non-survivals (5.95{+/-}5.16 vs 8.22{+/-}6.64, P=0.030) and the positive rate was also higher than that in the survivals (23.5% vs 12.0%, P=0.012). Regarding to severe COVID-19 patients, we found that high titer of anti-MDA5 Ab ([≥]10.0 U/mL) was more prevalent in the non-survivals (31.2% vs 14.0%, P=0.006). Moreover, early profiling of anti-MDA5 Ab could distinguish severe patients from those with non-severe ones. Overall, our data reveal that anti-MDA5 Ab is prevalent in the COVID-19 patients and high titer of this antibody is correlated with severe disease and unfavorable outcomes.

The incidence, risk factors and prognosis of acute kidney injury in severe and critically ill patients with COVID-19 in mainland China: a retrospective study (preprint)

Background: Since the clinical correlates, prognosis and determinants of AKI in patients with Covid-19 remain largely unclear, we perform a retrospective study to evaluate the incidence, risk factors and prognosis of AKI in severe and critically ill patients with Covid-19.Methods: We reviewed medical records of all adult patients (>18 years) with laboratory-confirmed Covid-19 who were admitted to the intensive care unit (ICU) between January 23rd 2020 and April 6th 2020 at Wuhan JinYinTan Hospital and The First Affiliated Hospital of Guangzhou Medical University. The clinical data, including patient demographics, clinical symptoms and signs, laboratory findings, treatment [including respiratory supports, use of medications and continuous renal replacement therapy (CRRT)] and clinical outcomes, were extracted from the electronic records, and we access the incidence of AKI and the use of CRRT, risk factors for AKI, the outcomes of renal diseases, and the impact of AKI on the clinical outcomes.Results: Among 210 subjects, 131 were males (62.4%). The median age was 64 years (IQR: 56-71). Of 92 (43.8%) patients who developed AKI during hospitalization, 13 (14.1%), 15 (16.3%) and 64 (69.6%) patients were classified as stage 1, 2 and 3, respectively. 54 cases (58.7%) received CRRT. Age, sepsis, Nephrotoxic drug, IMV and elevated baseline Scr were associated with AKI occurrence. The renal recover during hospitalization among 16 AKI patients (17.4%), who had a significantly shorter time from admission to AKI diagnosis, lower incidence of right heart failure and higher P/F ratio. Of 210 patients, 93 patients deceased within 28 days of ICU admission. AKI stage 3, critical disease, greater age and minimum P/F <150mmHg independently associated with it.Conclusions: Among patients with Covid-19, the incidence of AKI was high. age , sepsis, nephrotoxic drug, IMV and baseline Scr were strongly associated with the development of AKI. Time from admission to AKI diagnosis, right heart failure and P/F ratio were independently associated with the potential of renal recovery. Finally, AKI KIDGO stage 3 independently predicted the risk of death within 28 days of ICU admission.

Impaired T Cell Functions Along with Elevated Activated Tregs at the Early Stage of Asymptomatic SARS-CoV-2 Infection (preprint)

Background: Limited data are available on the T cell responses for the asymptomatic SARS-CoV-2 infection case. Methods: The first imported SARS-CoV-2 infection case in Wuhan was admitted in hospital for quarantine and observation. The T cell responses were followed up by flow cytometry analysis of the peripheral blood nonnuclear cells (PBMCs) at days 7, 13, 22, and 28 after admission. Findings: We found the first imported SARS-CoV-2 infection in Wuhan is an asymptomatic case. His T cell differentiation, proliferation and activation matched the classical kinetics of T cell responses induced by viral infection, but the activation maintained at a relatively low level. Function analysis indicated frequencies of IFN-γ producing CD4+ and CD8+ T cells were notably lower than that of the healthy controls (HC) at day 7, and then rebound gradually. But IFN-γ+ CD8+ T cells were detained at a significant lower level even at day 28, when the SARS-CoV-2 virus had already become undetectable for 3 weeks. Moreover, percentage of IL-17 producing CD4+ T cells was also detained constantly at a much lower level compared to HC. At day 7, although percentage of Tregs was in normal range, the frequency of activated Treg (aTreg) was remarkably as high as 4·4-fold of that in HC. Interpretation: The T cell activation in the asymptomatic SARS-CoV-2 infection experienced a significant suppression and presented impairment of Th1/Th17 and CD8+ T cell functions. Early elevation of the aTregs might play role in the activation and function of T cells in the asymptomatic SARS-CoV-2 infection. Funding Statement: None. Declaration of Interests: All authors declare no competing interests.Ethics Approval Statement: This study was reviewed and approved by the Medical Ethical Committee of Wuhan Jinyintan hospital (approval number KY-2020-47·01). Written informed consent was obtained from the patient and the healthy controls.

The Kinetics of Humoral Response and its Relationship with the Disease Severity in COVID-19 (preprint)

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.

Impaired T cell functions along with elevated activated Tregs at the early stage of asymptomatic SARS-CoV-2 infection (preprint)

Background Limited data are available on the T cell responses for the asymptomatic SARS-CoV-2 infection case. Methods The first imported SARS-CoV-2 infection case in Wuhan was admitted in hospital for quarantine and observation. The T cell responses were followed up by flow cytometry analysis of the peripheral blood nonnuclear cells (PBMCs) at days 7, 13, 22, and 28 after admission. Findings We found the first imported SARS-CoV-2 infection in Wuhan is an asymptomatic case. His T cell differentiation, proliferation and activation matched the classical kinetics of T cell responses induced by viral infection, but the activation maintained at a relatively low level. Function analysis indicated frequencies of IFN-{gamma} producing CD4+ and CD8+ T cells were notably lower than that of the healthy controls (HC) at day 7, and then rebound gradually. But IFN-{gamma}+CD8+ T cells were detained at a significant lower level even at day 28, when the SARS-CoV-2 virus had already become undetectable for 3 weeks. Moreover, percentage of IL-17 producing CD4+ T cells was also detained constantly at a much lower level compared to HC. At day 7, although percentage of Tregs was in normal range, the frequency of activated Treg (aTreg) was remarkably as high as 4.4-fold of that in HC. Interpretation The T cell activation in the asymptomatic SARS-CoV-2 infection experienced a significant suppression and presented impairment of Th1/Th17 and CD8+ T cell functions. Early elevation of the aTregs might play role in the activation and function of T cells in the asymptomatic SARS-CoV-2 infection.

Derivation and validation of a prognostic model for predicting in-hospital mortality in patients admitted with COVID-19 in Wuhan, China: the PLANS (Platelet Lymphocyte Age Neutrophil Sex) model (preprint)

OBJECTIVETo develop and validate a prognostic model for in-hospital mortality in COVID-19 patients using routinely collected demographic and clinical characteristics. DESIGNMulticenter, retrospective cohort study. SETTINGJinyintan Hospital, Union Hospital, and Tongji Hosptial in Wuhan, China. PARTICIPANTSA pooled derivation cohort of 1008 COVID-19 patients from Jinyintan Hospital, Union Hospital in Wuhan and an external validation cohort of 1031 patients from Tongji Hospital in Wuhan, China. MAIN OUTCOME MEASURESOutcome of interest was in-hospital mortality, treating discharged alive from hospital as the competing event. Fine-Gray models, using backward elimination for inclusion of predictor variables and allowing non-linear effects of continuous variables, were used to derive a prognostic model for predicting in-hospital mortality among COVID-19 patients. Internal validation was implemented to check model overfitting using bootstrap approach. External validation to a separate hospital was implemented to evaluate the generalizability of the model. RESULTSThe derivation cohort was a case-mix of mild-to-severe hospitalized COVID-19 patients (n=1008, 43.6% females, median age 55). The final model (PLANS), including five predictor variables of platelet count, lymphocyte count, age, neutrophil count, and sex, had an excellent predictive performance (optimism-adjusted C-index: 0.85, 95% CI: 0.83 to 0.87; averaged calibration slope: 0.95, 95% CI: 0.82 to 1.08). Internal validation showed little overfitting. External validation using an independent cohort (n=1031, 47.8% female, median age 63) demonstrated excellent predictive performance (C-index: 0.87, 95% CI: 0.85 to 0.89; calibration slope: 1.02, 95% CI: 0.92 to 1.12). The averaged predicted survival curves were close to the observed survival curves across patients with different risk profiles. CONCLUSIONSThe PLANS model based on the five routinely collected demographic and clinical characteristics (platelet count, lymphocyte count, age, neutrophil count, and sex) showed excellent discriminative and calibration accuracy in predicting in-hospital mortality in COVID-19 patients. This prognostic model would assist clinicians in better triaging patients and allocating healthcare resources to reduce COVID-19 fatality.

A Randomized, Single-blind, Group sequential, Active-controlled Study to evaluate the clinical efficacy and safety of α-Lipoic acid for critically ill patients with coronavirus disease 2019(COVID-19) (preprint)

Object: To evaluate the clinical efficacy and safety of -Lipoic acid (ALA) for critically ill patients with coronavirus disease 2019 (COVID-19). Methods: A randomized, single-blind, group sequential, active-controlled trial was performed at JinYinTan Hospital, Wuhan, China. Between February 2020 and March 2020, 17 patients with critically ill COVID-19 were enrolled in our study. Eligible patients were randomly assigned in a 1:1 ratio to receive either ALA (1200 mg/d, intravenous infusion) once daily plus standard care or standard care plus equal volume saline infusion (placebo) for 7 days. All patients were monitored within the 7 days therapy and followed up to day 30 after therapy. The primary outcome of this study was the Sequential Organ Failure Estimate (SOFA) score, and the secondary outcome was the all-cause mortality within 30 days. Result: Nine patients were randomized to placebo group and 8 patients were randomized to ALA group. SOFA score was similar at baseline, increased from 4.3 to 6.0 in the placebo group and increased from 3.8 to 4.0 in the ALA group (P=0.36) after 7 days. The 30-day all-cause mortality tended to be lower in the ALA group (3/8, 37.5%) compared to that in the placebo group (7/9, 77.8%, P=0.09). Conclusion: In our study, ALA use is associated with lower SOFA score increase and lower 30-day all-cause mortality as compared with the placebo group. Although the mortality rate was two-folds higher in placebo group than in ALA group, only borderline statistical difference was evidenced due to the limited patient number. Future studies with larger patient cohort are warranted to validate the role of ALA in critically ill patients with COVID-19. Keywords: Pneumonia; COVID-19; SARS-CoV-2 ; -Lipoic acid

Practice of Pharmaceutical Care in Designated Hospital under the Condition of Novel Coronavirus Pneumonia Epidemic / 中国药房

OBJECTIVE：To shar e the experienc e of pharmaceutical care in Wuhan Jinyintan Hospital （herein after refers to “our hospital ”）under the condition of novel coronavirus pneumonia （COVID-19）epidemic，and to provide reference for other hospitals to deal with public health emergencies. METHODS ：The situation of pharmaceutical care in our hospital under the condition of COVID- 19 epidemic was summarized and shared ，including the epidemic prevention and control management （regional division ，disinfection management ，pharmacy personnel training ），supply of drugs and disinfection products ，the monitoring and education of rational drug use by information technology. RESULTS ：The pharmacy department of our hospital divided the activity scope into clean area ，potential pollution risk area ，semi pollution area ，and implement different disinfection management. All pharmacists received training ，involving personal health protection ，prevention and control knowledge of COVID-19，health status monitoring ，etc. For supply and guarantee of drugs and disinfectants ，the epidemic drug list of our hospital was formulated ，drugs and disinfectants were purchased accurately and stored in a standardized way. 24 h response telephone was set up in the clinical pharmacy room to receive consultation from clinicians on drug use at any time. The drugs mentioned in the COVID- 19 diagnosis scheme were compared in terms of the mechanism of action and the medication of special populations to form a tablet ，so as to help clinical rational choice treatment drug. CONCLUSIONS ：The pharmaceutical care in the designated hospital of COVID- 19 is a professional and complicated work ，involving a wide range of aspects. Pharmacy department must respond actively and adjust the strategy in time so as to play an important role in improving the ability of medical treatment.