key: cord-0814295-g5kbgymf
authors: Zhu, Hui; Fu, Liyun; Jin, Yinhua; Shao, Jiale; Zhang, Shun; Zheng, Nanhong; Fan, Lingyan; Yu, Zhe; Ying, Jun; Hu, Yaoren; Chen, Tongen; Chen, Yanglingzi; Chen, Min; Chen, Mingjue; Xiong, Zi; Kang, Junfei; Jin, Jiachang; Cai, Ting; Ye, Honghua
title: Clinical features of COVID‐19 convalescent patients with re‐positive nucleic acid detection
date: 2020-06-07
journal: J Clin Lab Anal
DOI: 10.1002/jcla.23392
sha: bbd0e43002baf8192c0f203b6d2b0b0210819857
doc_id: 814295
cord_uid: g5kbgymf

BACKGROUND: Coronavirus disease 2019 (COVID‐19) is a pandemic that has rapidly spread worldwide. Increasingly, confirmed patients being discharged according to the current diagnosis and treatment protocols, follow‐up of convalescent patients is important to knowing about the outcome. METHODS: A retrospective study was performed among 98 convalescent patients with COVID‐19 in a single medical center. The clinical features of patients during their hospitalization and 2‐week postdischarge quarantine were collected. RESULTS: Among the 98 COVID‐19 convalescent patients, 17 (17.3%) were detected positive severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) nucleic acid during 2‐week postdischarge quarantine. The median time from discharge to SARS‐CoV‐2 nucleic acid re‐positive was 4 days (IQR, 3‐8.5).The median time from symptoms onset to final respiratory SARS‐CoV‐2 detection of negative result was significantly longer in re‐positive group (34 days [IQR, 29.5‐42.5]) than in non‐re‐positive group (19 days [IQR, 16‐26]). On the other hand, the levels of CD3‐CD56 + NK cells during hospitalization and 2‐week postdischarge were higher in re‐positive group than in non‐re‐positive group (repeated measures ANOVA, P = .018). However, only one case in re‐positive group showed exudative lesion recurrence in pulmonary computed tomography (CT) with recurred symptoms. CONCLUSION: It is still possible for convalescent patients to show positive for SARS‐CoV‐2 nucleic acid detection, but most of the re‐positive patients showed no deterioration in pulmonary CT findings. Continuous quarantine and close follow‐up for convalescent patients are necessary to prevent possible relapse and spread of the disease to some extent.

A novel coronavirus, designated as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) by the International Committee on Taxonomy of Viruses on February 11, 2020. 1 Coronavirus disease 2019 (COVID-19) caused by SARS-CoV-2 was declared a pandemic by World Health Organization. 2 As of April 10, 2020, China had reported 83 307 cases with 3346 deaths. 3 Internationally, by the same time, 1 439 516 confirmed cases of COVID-19 had been reported in approximately 212 countries and territories with more than 85 711 deaths. 3 Despite the large number of infected cases and the wide geographical spread of the disease, the COVID-19 has been gradually and effectively controlled by quarantine of suspected cases, analysis of epidemiology, and improvement of diagnosis and treatment in China. 4 In particular, the current use of broad-spectrum antiviral drugs such as lopinavir-ritonavir, arbidol hydrochloride, and chloroquine has achieved some desirable outcomes. [5] [6] [7] Recently, over ninety percent (77862/83307, 93.5%) of confirmed COVID-19 patients had met the criteria for hospital discharge or discontinuation of quarantine in China according to the latest diagnosis and treatment protocols from the National Health Commission of the People's Republic of China. 8 While previous reports on COVID-19 primarily focused on epidemiological and clinical characteristics of confirmed cases, 9, 10 this study was conducted to retrospectively investigate the clinical features and inflammation and immune biomarkers of COVID-19 convalescent patients with re-positive SARS-CoV-2 nucleic acid detection. Patients were diagnosed based on the interim guidance from the World Health Organization. 11 COVID-19 was confirmed by real-time reverse-transcriptase polymerase-chain-reaction (RT-PCR) assay for sputum or nasopharyngeal swab specimens. 12 Primers targeting open reading frame (ORF) 1a/1b and nuclear gene were used. Patient with cycle threshold (Ct) values less than or equal to 40 was considered positive.

Epidemiological characteristics, clinical symptoms and signs, laboratory findings, chest computed tomography (CT) assessments, antiviral treatment, and outcome data were extracted from electronic medical records. Epidemiological exposure was defined as a history of traveling to or residing in Wuhan city or the areas surrounding Wuhan, or a history of having contact with confirmed COVID-19 patients or patients with fever or respiratory symptoms from Wuhan city and its surrounding areas within 14 days before onset of the disease. The cluster events, such as the one when people collectively prayed in Tiantong temple on January 19, 2020, were also investigated. 13 Pulmonary CT scanning was conducted in all patients on admission. Additionally, repeat CT scans were performed every 5 days or in case of deterioration during hospitalization. The CT imaging was analyzed by two experienced radiologists. Primary parameters for CT imaging include lesion distribution, single or multiple lesions within each lobe, lesion density and interstitial pulmonary fibrosis.

Sputum and nasopharyngeal swab specimens were collected from all patients on admission, and confirmation testing for SARS-CoV-2 RNA was performed at Ningbo Municipal Centers for Disease Control (CDC) of Zhejiang Province or HwaMei Hospital, University of Chinese Academy of Sciences following the standard protocol. 12 Laboratory testing includes blood count, liver and kidney function, high-sensitivity C-reactive protein (hs-CRP), procalcitonin, creatine kinase, lactic dehydrogenase (LDH), D-dimer, electrolytes, and so on. We analyzed the temporal changes in total serum IgM and IgG specific for SARS-CoV-2 using COVID-19 Antibody (IgM/IgG) Combined Test Kit (Medical System Biotechnology Co., Ltd) and 2019-nCoV Antibody Test Kit (Innovita Biotechnology Co., Ltd). Above two kits were used for qualitative determination of specific antibodies by the latex agglutination ethod and the colloidal gold method, respectively.

Patients have to meet the following criteria for hospital discharge:

(a) temperature returned to normal for more than 3 days, (b) respiratory symptoms are relieved or resolved, (c) pulmonary computed tomography (CT) images show significant improvement in acute exudative lesions, and (d)two consecutive negative detections of respiratory SARS-CoV-2 (sample collection interval of at least 1 day). 14

This study was approved by the Ethics Committee of HwaMei Hospital, University of Chinese Academy of Sciences. Written informed consents were obtained from all subjects. The procedures followed were in accordance with the ethical standards of the Helsinki Declaration.

Quantitative variables were presented as means ± standard deviation (SD) or median and interquartile range (IQR), and the differences between groups were evaluated with unpaired Student's t test or the Mann-Whitney U test. Categorical variables were presented as absolute frequencies (n) and relative frequencies (%), and chi-square tests or Fisher's exact test was used for categorical variables. The ANOVA test with Greenhouse-Geisser correction was conducted to analyze repeated measures. All analyses were made by IBM SPSS statistics version 24.0.

The epidemiological and clinical data were presented in Table 1 In the re-positive group, the median age was 54 years (IQR, 44-63), and 70.6% were females (Table 1) . Less than 20% of re-positive group patients had underlying disease including hypertension (2 [11.8%]) and diabetes (1 [5.9%]). There was no significant difference of initial clinical symptoms between re-positive group and non-re-positive group.

Chest computed tomography (CT) scan on admission showed that the lesions were more apparent in the peripheral zone of lungs (73/98, 74.5%) ( Seventy-seven (77/98, 78.6%) patients had more than or equal to three pulmonary lesions. There was no statistically significant difference of imaging signs on admission between the two groups.

Patients who met criteria for hospital discharge or discontinuation of quarantine showed improvement in acute exudative lesions on chest CT images ( Table 2) presented the absorption rate for acute exudative lesions greater than or equal to 50%. Non-re-positive group showed a better absorption for acute exudative lesions than positive group at discharge.

It is worth mentioning that the chest CT of one case in re-positive group presented recurrent symptoms with blurred image in the upper lobe of both lungs, more prominent on the left side during the convalescent period, but the severity of image is less than that of late period of hospitalization ( Figure 1 ).

On admission, 25.5%, 34.7%, and 23.5% of cases showed leucopenia (white blood cell count <4 × 10 9 /L), lymphopenia (lymphocyte count <1.0 × 10 9 /L), and platelet suppression (blood platelet count <150 × 10 9 /L), respectively (Table 3) 

The changes of T lymphocyte subsets and Th1/Th2 cytokines during hospitalization and after discharge in re-positive group and non-re-positive cases were compared (Table S1 and Table S2 ). The levels of CD3-CD56 + NK cells at different time during hospitalization and follow-up were overall higher in positive group than in non-positive group (repeated measures ANOVA, P = .018) ( Figure 2 and Table S3 ). The measures of cytokines at different times found that IL-4, IL-6, IL-10, and TNF-a increased on admission and declined during recovery. However, there was no statistically significant difference between re-positive group and non-re-positive group (Table S4) . 

All patients received antiviral therapy during hospitalization, and (Table 4) .

Overall, the length of hospital stay was 15 days at median (IQR, [13] [14] [15] [16] [17] [18] [19] , and no significant differences were discovered between Our study has several limitations. First, our study was conducted in convalescent patients with mild or moderate COVID-19. Second, further follow-up study needs be conducted among these patients.

Third, our study was limited to a small number of patients in Ningbo city, Zhejiang province. Multi-center research on a larger cohort is warranted in future study.

In conclusion, SARS-CoV-2 nucleic acid can still be detected in a small proportion of convalescent patients based on our study.

The repeated measures of CD3-CD56 + NK cell. Difference of CD3-CD56 + NK cell between the positive group and the non-positive group was tested using repeated measures two-way analysis of variance with Greenhouse-Geisser correction Note: Data are presented as median and interquartile range (IQR) or n (%).

P values comparing non-re-positive cases and re-positives cases are from Mann-Whitney U test and Fisher's exact test.

Percentages do not total 100% owing to missing data.

Taxonomy of Viruses. Naming the 2019 coronavirus

WHO Declares COVID-19 a Pandemic

World Health Organization. Coronavirus disease (COVID-19) outbreak situation

Novel coronavirus 2019-nCoV: prevalence, biological and clinical characteristics comparison with SARS-CoV and MERS-CoV

Case of the index patient who caused tertiary transmission of COVID-19 infection in korea: the application of lopinavir/ritonavir for the treatment of COVID-19 infected pneumonia monitored by quantitative RT-PCR

Towards optimization of hydroxychloroquine dosing in intensive care unit COVID-19 patients

Remdesivir and chloroquine effectively inhibit the recently emerged novel coronavirus (2019-nCoV) in vitro

The diagnosis and treatment protocols from the national health commission of the People's Republic of China

4856d 5b045 8141f a9f37 68532 24d41 d7.shtml. Accessed March

Clinical characteristics of 138 hospitalized patients with 2019 novel coronavirus-infected pneumonia in Wuhan, China

Novel coronavirus infection in hospitalized infants under 1 year of age in China

Clinical management of severe acute respiratory infection when Novel coronavirus (nCoV) infection is suspected: interim guidance

Clinical features of patients infected with 2019 novel coronavirus in Wuhan

Epidemiologic and clinical characteristics of 91 hospitalized patients with COVID-19 in Zhejiang, China: a retrospective, multi-centre case series. QJM. 2020. Epub ahead of print

Positive RT-PCR test results in patients recovered from COVID-19

CD56(+dim) and CD56(+bright) cell activation and apoptosis in hepatitis C virus infection

The human liver contains multiple populations of NK cells, T cells, and CD3+CD56+ natural T cells with distinct cytotoxic activities and Th1, Th2, and Th0 cytokine secretion patterns

CD1d-unrestricted human NKT cells release chemokines upon Fas engagement

Immunomodulation by mesenchymal stem cells in treating human autoimmune disease-associated lung fibrosis

CD1d-restricted IFN-gamma-secreting NKT cells promote immune complex-induced acute lung injury by regulating macrophage-inflammatory protein-1alpha production and activation of macrophages and dendritic cells

High frequencies of CD158b+ NK cells are associated with persistent hepatitis C virus infections

Relapse might occur in COVID-19 patients. Strict quantification of pulmonary CT evaluation of inflammation absorption may improve the discharge criteria. Continuous quarantine and follow-up for convalescent patients are necessary to control possible spread to some extent.

The authors declare that they have no competing interests.

Honghua Ye and Ting Cai designed the study. Zhu Hui and Liyun 

https://orcid.org/0000-0002-3159-8549