key: cord-0751607-qznr6e37 authors: Yuan, Jing; Kou, Shanglong; Liang, Yanhua; Zeng, JianFeng; Pan, Yanchao; Liu, Lei title: PCR Assays Turned Positive in 25 Discharged COVID-19 Patients date: 2020-04-08 journal: Clin Infect Dis DOI: 10.1093/cid/ciaa398 sha: 364889728c0e7d8f40b45b3ab094bc1ab6b9f7cf doc_id: 751607 cord_uid: qznr6e37 We report the observation that 14.5% of COVID-19 patients had positive RT-PCR testing again after discharge. We describe correlations between laboratory parameters and treatment duration (r= -0.637; p=0.002) and time to virus recrudescence (r= 0.52; p=0.008) respectively, suggesting the need for additional measures to confirm illness resolution in COVID-19 patients. Since Dec 8 th 2019, many cases has been reported by investigators who described the clinical characteristics of hospitalized patients with COVID-19 infection 1-3 . We noticed the recent report 4 from Lan et al. that 4 medical staff were still virus carriers after recovery from COVID-19 infection. Here we gave a report on 25 discharged patients with their medical record review and further analysis. Enrolled from Jan 23th 2020 to Feb 21th 2020, total 172 COVID-19 infected patients were discharged from Shenzhen Third People's Hospital. They all met the following criteria of hospital discharge in China: (1) Normal body temperature for more than 3 consecutive days. (2) Significant reduction of respiratory symptoms evaluated by following indicators: cough and expectoration disappeared, normal ranges for inflammatory markers IL-6 and CRP, as well as oxygenation index ≥350. (3) Substantial improvement over conventional chest radiography detection. (4) At least two consecutively negative results of RT-PCR testing separated by at least 24-hour interval. Considering the risk of reinfection, all discharged patients were required another 14 days of self-segregating at home for further observation. The cloacal swab and nasopharyngeal swab samples were both collected from these discharged patients each three days for RT-PCR detection of COVID-19 in the same way as they were in hospital. Among them, 25 discharged patients were identified with positive results again. COVID-19 RNA Detection Kits (Real-time fluorescent PCR method) approved by the National Medical Products Administration were used for virus testing as described previously 5 . Conditions for the amplifications include reverse transcription at 50°C for 15min, pre-denaturation at 95°C for 15 min, followed by 45 cycles of 94°C for 15 s and 55°C for 45 s for fluoresce detection. The receiver operating characteristic (ROC) curve analysis was used to determine the optimal threshold cut-off value. And a cycle threshold (Ct) value ≤40 was defined as a positive test. Other demographic, clinical, radiologic and laboratory findings were extracted from the electronic medical records of the patients. The anti-viral compound ritonavir/lopinavir (Kaletra ®) combined with interferon alpha (IFN-α) was used as the potential antiviral therapy for all these patients during their first hospitalization. Ritonavir/lopinavir table was administrated with 500 mg once daily while 50 μg IFN-α was aerosolized 2 times a day. Besides, Chinese herbal medicines were widely recommended by the Chinese Clinical Guideline for the Diagnosis and Treatment of Novel Coronavirus Pneumonia (On Trials) issued by National Health Commission of China. The herbal formula of Lung Cleansing and Detoxifying Decoction was used for these 25 patients during their second hospital stay. The study was approved by Shenzhen Third People's Hospital Ethics Committee and the informed consent was waived. The study population included 172 discharged COVID-19 patients from Jan 23th 2020 to Feb 21th On the other hand, previous clinical and laboratory outcomes were compared between these 25 patients (with conversion of RT-PCR results from negative to positive after being discharged from the hospital) and rest 147 patients (continuously represented negative for the virus after discharge). They showed no distinguished differences on the levels of laboratory parameters before leaving the hospital. Subsequently, correlation analysis indicated that there was a significant inverse correlation existed between serum D-Dimer level before discharging and the duration of treatment in these 25 patients (r=-0.637, p=0.002), instead of the rest 147 patients. Furthermore, lymphocyte concentrations before these 25 patient leaving the hospital were significantly positively correlated (r=0.52, p=0.008) with the time interval for virus reappearing. These 25 patients with COVID-19 infection all met the criteria for hospital release from quarantine, while the RT-PCR testing then conversed to positive without aggravation on symptoms after 2 to 13 days. It seems that there was a fluctuating period between of seeming improvement in clinics and full recovery from virus. These less symptomatic carriers brought more challenges to the management and control of COVID-19 epidemic in China and any other affected countries. According to our study, it is probably that two negative RT-PCR tests 24 hours apart may not be sufficient for viral clearance evaluation. Repeated viral RT-PCR testing separated by prolonged duration like 48 hours is essential to assure that virus has actually cleared and the discharged patients no longer transmitted the virus. On the other hand, we suggested that some immunological parameters such as D-dimer and absolute lymphocyte count, and even antibody test should be combined with RT-PCR negative test as additional measures to assure that infected patients have completely recovered and can be released from quarantine. Besides, the RT-PCR results then turning to negative within an average of 2.73 days of hospital stay. Even the Chinese herbal medicine was used for these readmitted patients, it is hard to assess the effect on virus clearance. Further case-control study and cohort study will be needed to pursue that. Data are given as mean ± standard deviation (SD). Determined using one-way analysis of variance. Abbreviations: LDH, lactate dehydrogenase; CRP, C-reactive protein; IL-6, interleukin-6. Figure. Correlation analysis for serum D-Dimer level and duration of treatment, as well as lymphocyte count and time interval for virus reappearing. Correlation analysis on serum D-Dimer level and the duration of treatment (r=-0.637, p=0.002) (A), as well as correlation between lymphocyte concentration and the time interval for virus reappearing (r=0.52, p=0.008) (B). The D-Dimer values and lymphocyte counts were collected from the last measurements before their first hospital discharge. The duration of treatment was derived from their previous records. The time interval for virus reappearing was calculated from last negative before previously leaving the hospital to turning positive again of RT-PCR testing on COVID-19. Clinical Characteristics of 138 Hospitalized Patients With 2019 Novel Coronavirus-Infected Pneumonia in Wuhan, China Early Transmission Dynamics in Wuhan, China, of Novel Coronavirus-Infected Pneumonia A familial cluster of pneumonia associated with the 2019 novel coronavirus indicating person-to-person transmission: a study of a family cluster Positive RT-PCR Test Results in Patients Recovered From COVID-19 Clinical features of patients infected with 2019 novel coronavirus in Wuhan, China All authors declare that they have no conflict of interest exists.