key: cord-0328827-std4jddn
authors: Ma, Kun-Long; Liu, Zhi-Heng; Cao, Chun-feng; Liu, Ming-Ke; Liao, Juan; Zou, Jing-Bo; Kong, Ling-Xi; Wan, Ke-Qiang; Zhang, Jun; Wang, Qun-Bo; Tian, Wen-Guang; Qin, Guang-Mei; Zhang, Lei; Luan, Fun-Jun; Li, Shi-Ling; Hu, Liang-Bo; Li, Qian-Lu; Wang, Hai-Qiang
title: COVID-19 Myocarditis and Severity Factors： An Adult Cohort Study
date: 2020-03-23
journal: nan
DOI: 10.1101/2020.03.19.20034124
sha: 9cbb982f243a17d8cb9f47bbf9eaa186cb24a001
doc_id: 328827
cord_uid: std4jddn

Background Notwithstanding the clinical hallmarks of COVID-19 patients were reported, several critical issues still remain mysterious, i.e., prognostic factors for COVID-19 including extrinsic factors as viral load of SARS-CoV-2 and intrinsic factors as individual's health conditions; myocarditis incidence rate and hallmarks. Methods Demographic, epidemiologic, radiologic and laboratory data were collected by medical record reviews of adult hospitalized patients diagnosed as COVID-19. Cycle threshold (Ct) value data of real-time PCR (RT-PCR) were collected. The time duration was from 21 January to 2 March, 2020. Pulmonary inflammation index (PII) values were used for chest CT findings. Multivariate logistic regression analysis was used to identify independent severity risk factors. RESULTS In total, 84 hospitalized adult patients diagnosed as COVID-19 were included, including 20 severe and 64 nonsevere cases. The viral load of the severe group was significantly higher than that of the non-severe group, regardless of the Ct values for N or ORF1ab gene of virus (all p<0.05).Typical CT abnormalities was more likely existing in the severe group than in the nonsevere group in patchy shadows or ground glass opacities, consolidation, and interlobular septal thickening (all p<0.05). In addition, the PII values in the severe group was significantly higher than that in the nonsevere group (52.5 [42.5-62.5] vs 20 [5.0-31.6]; p<0.001). Amongst 84 patients, 13 patients (15.48%) were noted with abnormal electrocardiograms (ECGs) and serum myocardial enzyme levels; whereas 4 (4.8%) were clinically diagnosed as SARS-CoV-2 myocarditis. Multivariable logistic regress analysis distinguished three key independent risk factors for the severity of COVID-19, including age [OR 2.350; 95% CI (1.206 to 4.580); p=0.012], Ct value [OR 0.158; 95% CI (0.025 to 0.987); p=0.048] and PII [OR 1.912; 95% CI (1.187 to 3.079); p=0.008]. Interpretation Three key-independent risk factors of COVID-19 were identified, including age, PII, and Ct value. The Ct value is closely correlated with the severity of COVID-19, and may act as a predictor of clinical severity of COVID-19 in the early stage. SARS-CoV-2 myocarditis should be highlighted despite a relatively low incidence rate (4.8%). The oxygen pressure and blood oxygen saturation should not be neglected as closely linked with the altitude of epidemic regions.

the severe group was significantly higher than that in the nonsevere group (52.5

[42. .5] vs 20 [5.0-31.6 ]; p<0.001). Amongst 84 patients, 13 patients (15.48%) were noted with abnormal electrocardiograms (ECGs) and serum myocardial enzyme levels;

whereas 4 (4.8%) were clinically diagnosed as SARS-CoV-2 myocarditis. Multivariable logistic regress analysis distinguished three key independent risk factors for the severity of COVID- 19 

Three key-independent risk factors of COVID-19 were identified, including age, PII, and Ct value. The Ct value is closely correlated with the severity of COVID-19, and may act as a predictor of clinical severity of COVID-19 in the early stage. SARS-CoV-2 myocarditis should be highlighted despite a relatively low incidence rate (4.8%). The oxygen pressure and blood oxygen saturation should not be neglected as closely linked with the altitude of epidemic regions.

. CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. ICU-free groups (73.9%). The median age was 56 years with 54.3% male patients.

Patients treated in ICU were older, with underlying comorbidities, complaining of dyspnea and anorexia, in comparison with ICU-free patients. Laboratory indicators with statistical significance between two groups included higher levels of white blood cell and neutrophils . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted March 23, 2020. . https://doi.org/10.1101/2020.03. 19.20034124 doi: medRxiv preprint counts, D-dimer, creatine kinase, and creatine. Notably, higher level of lactate dehydrogenase (LDH) was linked with ICU-treated group. Xu et al 7 is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted March 23, 2020. . https://doi.org/10.1101/2020.03. 19.20034124 doi: medRxiv preprint of RT-PCR for SARS-CoV-2. 5 Despite high viral load reflected by low Ct values (13.73 for ORF1ab gene, 15 .57 for N gene), the infant was asymptomatic. Notably, scarce evidence is available on viral load and clinical outcome, notwithstanding RT-PCR has been widely used in clinical practice.

Whereas these frontier studies presented critical lines of evidence on the emerging infectious disease as COVID-19, most patients were undergoing treatment with unclear outcome when data were summarized. Therefore, several critical issues still remain mysterious, i.e., prognostic factors for COVID-19 including extrinsic factors as viral load of SARS-CoV-2 and intrinsic factors as individual's health conditions, including those identified as candidate risk factors (older age, underlying comorbidities, higher levels of white blood cell and neutrophils counts, D-dimer, creatine kinase, and creatine). 6 Nevertheless, the clinical significance of abnormal serum myocardial enzymes identified by previous studies [6] [7] [8] has not been well documented, in particular viral potentiality of myocardial damage and underlying mechanisms. Accordingly, the study aimed for addressing these important issues based on a retrospective observational study of 84 adult cases in Chongqing municipality, China (adjacent to Hubei Province).

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The copyright holder for this preprint this version posted March 23, 2020. Demographic, epidemiologic, radiologic imaging and laboratory data were collected by intensive medical record reviews of included cases, with double checks and additional inquiries when appropriate. The diagnosis and treatment were conducted strictly conforming to the COVID-19 national program provided by the National Health Commission of China (5 th edition). 9

Each patient underwent double tests for SARS-CoV-2 nucleic acid detection every 24 hours on his/her nasopharyngeal swab samples. In brief, viral nucleic acid from samples was conducted using viral isolation kits (Daangene, Guangzhou, China). Fluorescence . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)

The copyright holder for this preprint this version posted March 23, 2020. 

Full spectrum laboratory tests were performed at admission and successive appropriate times, including blood count, serum biochemistry, serum inflammatory indicators, myocardial enzymes. Additional tests were performed according to patients' detailed conditions of background diseases. Lymphocytes sorting analyses were conducted using flow cytometre (Bricyte e6, Mindray, Shenzhen, China) and pertaining assay kits (Mindray, Shenzhen, China).

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The copyright holder for this preprint this version posted March 23, 2020. Association of China, 10 with distribution and size of lesions estimated. As for distribution, one lung segment equals one score with highest score as 20, representing left and right lung lesions. For lesion size, 1 score indicates over 50% lung segment volume invasive, whereas 0 score less than 50%. Accordingly, PII= (Distribution score+ Size score)/40*100%. PII values were calculated twice at three week intervals by two experienced radiologists. Interclass coefficient was used to assess inter-rater reliability.

At admission, each patient underwent ECG examinations routinely. Each electrocardiogram was evaluated and reported by senior technicians. Abnormal ECGs were collected and analyzed in combination of clinical manifestations and serum myocardial enzyme tests. Repeated ECGs were ordered for patients suspected with myocardial damage.

Data were recorded and summarized using spreadsheets. Patients in our cohort were classified into severe and nonsevere groups at admission in combination of national . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)

The copyright holder for this preprint this version posted March 23, 2020. . https://doi.org/10.1101/2020.03. 19.20034124 doi: medRxiv preprint program and the American Thoracic Society guidelines. 11 In brief, patients were included in severe group if they meet with the following criteria: 1) Shortness of breath with respiratory rate (RR) >30 times /min; 2) Oxygen saturation <93% at rest; 3)Arterial partial oxygen pressure (PaO2)/oxygen absorption concentration (FiO2)<277mmHg (The result was calculated according to the altitude of the epidemic area, Yongchuan city is about 700 meters above sea level, 1mmHg=0.133 kPa); 3) Pulmonary imaging showed that the lesion had progressed to > 50% at 24 ~ 48 h; 4) Respiratory failure;5) Shock; 6)Combined with other organ failure. Continuous (means for normally distributed variables and/or interquartile ranges for abnormally distributed variables) and categorical variables (percentage, %) were analyzed using SPSS, version 29.0. Unpaired t test, Fisher exact test, Mann-Whitney U test and chi-square test were performed where appropriate. Multivariable logistic regress model was used to control for confounding factors and distinguish independent risk factors for clinical severity. Risk factors with a univariable p-value<0.05 were eligible for inclusion in the model. Severe group was coded as "1" and nonsevere group was coded as "0". Variables (p≤0.10) were in the final model, with significant variables identified (p<0.05), via a forward and stepwise protocol. Adjusted odds ratios (ORs, indicating pertaining digital times more likely to present in severe group) and 95% confidence intervals (CIs) were calculated as independent risk factors.

Spearman's correlations were used to assess the links between multiple factors.

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The copyright holder for this preprint this version posted March 23, 2020. . https://doi.org/10.1101/2020.03. 19.20034124 doi: medRxiv preprint in charge of all parts of the study and confirm that issues regarding the accuracy or integrity of the study had been properly scrutinized and settled. The final version of the manuscript was approved by all authors.

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There were significant differences in a number of laboratory indicators between patients in the severe and non-severe groups ( CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted March 23, 2020. In addition, in the detection of lymphocyte subsets, total T lymphocyte count, B lymphocyte count, and Inhibitive/Cytotoxic T Lymphocyte count were significantly reduced in the severe group compared with the nonsevere group.

The Ct values of RT-PCR for two target genes (ORF1ab and N) were recorded in 62 of all patients, with a total of 207 values at corresponding time points (table 3) and for ORF1ab gene was 38.6(IQR, 36.0-41.5). There was significant difference in Ct . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted March 23, 2020. . values either for N gene (p=0.004) or for ORF1ab gene (p=0.002) between the two groups.

Totally 84 patients diagnosed with COVID-19 underwent at least two or more CT examinations (table 3 ). An excellent inter-rater reliability was noted in the assessment of CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

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Complications during hospitalization were documented ( failure, and 6(7.1%) cases with renal abnormalities. Except for gastrointestinal disorders, the incidence of all complications in the severe group was significantly higher than that in the non-severe group (p<0.05). Of the 20 patients with respiratory failure, 9(45%) were treated with assisted mechanical ventilation, 7(35%) with nasal catheters and 4(20%) with masks. The median time from onset to respiratory failure was 7.75 days and to mechanical ventilation was 8 days.

Among all 84 patients, we found 13 patients with abnormal myocardial injury indicators combined with abnormal ECG, including 8 severe patients and 5 nonsevere patients (table 5). Four of the 13 patients had a history of cardiovascular diseases, while the other 9 patients had no such history. There were 3 cases with a history of hypertension, 1 patient with hypertension, atrial fibrillation and coronary heart disease . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted March 23, 2020. . https://doi.org/10.1101/2020.03. 19.20034124 doi: medRxiv preprint who was excluded from our further analysis. The 12 remaining patients had abnormal changes in the myocardial enzymes or cTnI, amongst which 10 cases with high levels of inflammatory markers (except for patient no.9 and 11), and eleven of them exhibiting clinical symptoms of myocardial injury (except for patient no. 9).

Typical abnormal ECG findings were noted in ten patients (except for patient no.2, 6 and 12). Major ECG abnormalities included sinus tachycardia or bradycardia, t-wave changes, atrioventricular block, and anomaly Q wave. Four patients underwent repeated ECG examinations. Patient no. 3 was an elderly female patient in the severe group. The first ECG indicated t-wave changes and sinus bradycardia. However, repeated ECG restored normal after treatment with a decrease in inflammatory markers and cardiac enzyme levels. Patient no. 4 was an elderly male patient in the severe group. His first ECG indicated First-degree atrioventricular block, t-wave changes, left axis deviation and abnormal Q wave in the lower wall, while the abnormal Q and t-wave disappeared after treatments. Patient no. 9 and 10 were in the nonsevere group. Their initial ECGs showed sinus tachycardia and sinus tachycardia associated with t-wave changes respectively, with repeated normal ECGs after treatment. . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted March 23, 2020. . https://doi.org/10.1101/2020.03. 19.20034124 doi: medRxiv preprint In order to explore the intrinsic correlations between cardiac enzymes, ECG and cTn I with inflammatory cytokines and lymphocyte levels, we used Mann-Whitney U test to analyze the relevant data (table 6).The results showed that the level of cardiac enzymes was significantly positively correlated with the level of CRP (p=0.004) and PCT (p=0.012).

In addition, this study also found that there was a significant correlation between the abnormality of ECG and the level of PCT (p=0.004) and lymphocytes counts (p=0.041).

However, we did not find any correlation between cTn I and levels of inflammatory cytokines or lymphocytes.

In this study, multivariable logistic regress model was used to identify the independent risk factors for clinical severity of COVID-19 (figure 6 . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted March 23, 2020. . . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted March 23, 2020. 

To our knowledge, this is the first clinical cohort study to analyze the extrinsic (viral load) and intrinsic risk factors of patients with COVID-19. Moreover, SARS-CoV-2 myocarditis was diagnosed clinically with an incidence rate of 4.8%. Previous neglected regional altitude issue was considered for COVID-19 severity classification.

Previous studies mainly focused on the clinical characteristics, imaging findings and treatment measures of COVID-19, 2,3,5-8 however, few studies pay attention to its risk factors and complications. [14] [15] [16] There were significant differences between severe and nonsevere COVID-19 in age, clinical manifestations, imaging findings, levels of inflammatory cytokines and myocardial enzymes, and distribution of lymphocytes and their subsets, consistent with previous studies. 3,5-8 Furthermore, multivariable logistic regress analysis identified three key independent risk factors for the severity of COVID-19, i.e., age, PII, and Ct values.

Ct values from the RT-PCR can act as a proxy for viral load. 17, 18 In viral respiratory infectious diseases, viral load has been proved to be closely related to the severity of the diseases. [19] [20] [21] It is still unclear whether the virus load is closed related to the severity of COVID-19 patients, although sporadic case studies have been reported 18, 22 . We found that the patients' viral load was associated with severity of COVID-19 as an independent risk factor.

In addition, all the complications (except for gastrointestinal disorders) of COVID-2019 during hospitalization were found significantly higher in severe than . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. There are some limitations in our study. Firstly, the sample size in our study was relatively small, only 84 patients were included. The deadline of the collected data was March 2, 2020, with 16 patients still hospitalized. Secondly, we did not get exact viral load of patients as viral copy numbers due to the urgent outbreak of COVID-19 and limited conditions. We used Ct values as a substitute. Nevertheless, the results of multivariable logistic regress model demonstrated that the Ct value of SARS-CoV-2 is valid and reliable.

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The copyright holder for this preprint this version posted March 23, 2020. 

Three key-independent risk factors of COVID-19 were identified, including age, PII, and Ct value. The Ct value is closely correlated with the severity of COVID-19, and may act as a predictor of clinical severity of COVID-19 in the early stage. SARS-CoV-2 myocarditis should be highlighted despite a relatively low incidence rate (4.8%). The oxygen pressure and blood oxygen saturation should not be neglected as closely linked with the altitude of epidemic regions.

. CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted March 23, 2020. 

The datasets used to support the current study are available from the authors upon reasonable request.

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The copyright holder for this preprint this version posted March 23, 2020. . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted March 23, 2020. . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted March 23, 2020. is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint a P values indicate differences between severe and nonsevere types of patients. A P value less than 0.05 was considered statistically significant.

Variables were expressed as mean± standard deviation given that they were normally distributed; otherwise as median (Q1-Q3).

.

It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted March 23, 2020. .

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