Understanding the risk of transmission of respiratory viral infections in childcare centres: protocol for the DISeases TrANsmission in ChildcarE (DISTANCE) multicentre cohort study.

INTRODUCTION: Childcare centre is considered a high-risk setting for transmission of respiratory viruses. Further evidence is needed to understand the risk of transmission in childcare centres. To this end, we established the DISeases TrANsmission in ChildcarE (DISTANCE) study to understand the interaction among contact patterns, detection of respiratory viruses from environment samples and transmission of viral infections in childcare centres. METHODS AND ANALYSIS: The DISTANCE study is a prospective cohort study in multiple childcare centres of Jiangsu Province, China. Study subjects will be childcare attendees and teaching staff of different grades. A range of information will be collected from the study subjects and participating childcare centres, including attendance, contact behaviours (collected by onsite observers), respiratory viral infection (weekly respiratory throat swabs tested by multiplex PCR), presence of respiratory viruses on touch surfaces of childcare centres and weekly follow-up survey on respiratory symptoms and healthcare seeking among subjects tested positive for any respiratory viruses. Detection patterns of respiratory viruses from study subjects and environment samples, contact patterns, and transmission risk will be analysed by developing statistical and mathematical models as appropriate. The study has been initiated in September 2022 in 1 childcare centre in Wuxi City, with a total of 104 children and 12 teaching staff included in the cohort; data collection and follow-up is ongoing. One more childcare centre in Nanjing City (anticipated to include 100 children and 10 teaching staff) will start recruitment in 2023. ETHICS AND DISSEMINATION: The study has received ethics approval from Nanjing Medical University Ethics Committee (No. 2022-936) and ethics approval from Wuxi Center for Disease Control and Prevention Ethics Committee (No. 2022-011). We plan to disseminate the study findings mainly through publications in peer-reviewed journals and presentations in academic conferences. Aggregated research data will be shared freely to researchers.

Real-time detection of SARS-CoV-2 in clinical samples via ultrafast ligation-dependent RNA transcription amplification.

RNA has been recognized as an important biomarker of many infectious pathogens; thus, sensitive, simple and rapid detection of RNA is urgently required for the control of epidemics. Herein, we report an ultrafast ligation-dependent RNA transcription amplification assay with high sensitivity and specificity for real-time detection of SARS-CoV-2 in real clinical samples, termed splint-based cascade transcription amplification (SCAN). Target RNA is first recognized by two DNA probes, which are then ligated together by SplintR, followed by the binding of the T7 promotor and T7 RNA polymerase to the ligated probe and the start of the transcription process. By introducing a vesicular stomatitis virus (VSV) terminator in the ligated probe, large amounts of RNA transcripts are rapidly produced within 10 min, which then directly hybridize with molecular beacons (MBs) and trigger the conformational switch of the MBs to generate a fluorescence signal that can be monitored in real time. The SCAN assay, which can be completed within 30-50 min, has a limit of detection of 104 copies per mL, while exhibiting high specificity to distinguish the target pathogen from those causing similar syndromes. More importantly, the results of SCAN for SARS-CoV-2 detection in clinical samples display great agreement with the most used qRT-PCR and qRT-LAMP, indicating great potential in the diagnosis of pathogens in clinical practice.

Expression and Immunogenicity of SARS-CoV-2 Virus-Like Particles based on Recombinant Truncated HEV-3 ORF2 Capsid Protein.

COVID-19 is an emerging disease that poses a severe threat to global public health. As such, there is an urgent demand for vaccines against SARS-CoV-2, the virus that causes COVID-19. Here, we describe a virus-like nanoparticle candidate vaccine against SARS-CoV-2 produced by an E. coli expression system. The fusion protein of a truncated ORF2-encoded protein of aa 439~608 (p170) from hepatitis E virus CCJD-517 and the receptor-binding domain of the spike protein from SARS-CoV-2 were expressed, purified and characterized. The antigenicity and immunogenicity of p170-RBD were evaluated in vitro and in Kunming mice. Our investigation revealed that p170-RBD self-assembled into approximately 24 nm virus-like particles, which could bind to serum from vaccinated people (p < 0.001) and receptors on cells. Immunization with p170-RBD induced the titer of IgG antibody vaccine increased from 14 days post-immunization and was significantly enhanced after a booster immunization at 28 dpi, ultimately reaching a peak level on 42 dpi with a titer of 4.97 log10. Pseudovirus neutralization tests showed that the candidate vaccine induced a strong neutralizing antibody response in mice. In this research, we demonstrated that p170-RBD possesses strong antigenicity and immunogenicity and could be a potential candidate for use in future SARS-CoV-2 vaccine development.

Single-walled carbon nanotubes-based RNA protection and extraction improves RT-qPCR sensitivity for SARS-CoV-2 detection.

The false-negative result of nucleic acid testing is an important cause of continued spread of COVID-19, while SARS-CoV-2 RNA degradation during transportation and nucleic acid extraction can lead to false-negative results. Here, we investigated that single-walled carbon nanotubes (SCNTs) could protect RNA from degradation for at least 4 days at room temperature. By constructing magnetism-functionalized SCNTs (MSCNTs), we developed a method that enabled protection and simple extraction of SARS-CoV-2 RNA, and the RNA-bound MSCNTs can be directly used for reverse transcription polymerase chain reaction (RT-qPCR) detection. The experimental results showed that 1 µg of MSCNTs adsorbed up to 24 ng of RNA. Notably, the MSCNTs-based method for extracting SARS-CoV-2 RNA from simulated nasopharyngeal swabs and saliva samples with mean recovery rates of 103% and 106% improved the sensitivity of RT-qPCR detection by 8-32 fold in comparison to current common methods. This improvement was largely attributable to the protection of RNA, enabling increased RNA load for downstream assays.

Real-time monitoring of isothermal nucleic acid amplification on a smartphone by using a portable electrochemical device for home-testing of SARS-CoV-2.

Home-testing of SARS-CoV-2 is an ideal approach for controlling the pandemic of COVID-19 and alleviating the shortage of medical resource caused by this acute infectious disease. Herein, a portable device that enables real-time monitoring of isothermal nucleic acid amplification tests (INAATs) through the electrochemistry method was fabricated for home-testing of SARS-CoV-2. First, a disposable plug-and-play pH-sensitive potentiometric sensor that matches this electrochemical INAATs (E-INAATs) device was designed to allow the label-free pH sensing detection of nucleic acid. By applying Nafion film on the polyaniline-based working electrode, this sensor exhibited an excellent linear potentiometric response to pH value in the range of 6.0-8.5 with a slope of -37.45 ± 1.96 mV/pH unit. A Bluetooth module was integrated into this device to enable the users real-time monitoring INAATs on their smartphones at home. Moreover, by presetting criteria, the detection results could be automatically judged by the device to avoid human errors. Finally, the utility of this E-INAATs device was demonstrated by detecting the presence of SARS-CoV-2 nucleocapsid protein gene in artificial samples with a sensitivity of 2 × 102 copies/test within 25 min, which was comparable with fluorescence and colorimetric assay. This portable, easy-operated, sensitive, and affordable device is particularly desirable for the full integration of household SARS-CoV-2 detection products and will open a new prospect for the control of infectious diseases via electrochemical NAATs.

Reliability and validity of the Pittsburgh Sleep Quality Index among frontline COVID-19 health care workers using classical test theory and item response theory.

STUDY OBJECTIVES: The applicability of sleep-related scales to frontline medical staff for the COVID-19 pandemic has not been fully proved, so sleep survey results lack credibility and accuracy, creating difficulties for the guidance and treatment of frontline medical staff with sleep disorders, which is not conducive to the prevention and control of COVID-19. This study sought to analyze the reliability and validity of the Pittsburgh Sleep Quality Index (PSQI) among frontline medical staff fighting the COVID-19 pandemic. METHODS: A network questionnaire survey was used to investigate the PSQI among frontline medical staff who fought COVID-19 in Wuhan, China from March 19 to April 15, 2020. Combined with classical test theory and item response theory, the content validity, internal consistency, construct validity, and other aspects of the PSQI were evaluated. RESULTS: According to classical test theory, content validity, criterion validity, and construct validity of the PSQI were good. But the internal consistency was better after the deletion of the "daytime dysfunction" subscale. With regard to item response theory, difficulty, the differential item function, and the Wright map performed well. CONCLUSIONS: The original PSQI showed acceptable applicability in frontline COVID-19 medical staff, and its characteristics moderately improved after the "daytime dysfunction" subscale was removed. CITATION: Wang L, Wu Y-X, Lin Y-Q, et al. Reliability and validity of the Pittsburgh Sleep Quality Index among frontline COVID-19 health care workers using classical test theory and item response theory. J Clin Sleep Med. 2022;18(2):541-551.

Nucleic acid extraction without electrical equipment via magnetic nanoparticles in Pasteur pipettes for pathogen detection.

The outbreak of COVID-19 makes epidemic prevention and control become a growing global concern. Nucleic acid amplification testing (NAAT) can realize early and rapid detection of targets, thus it is considered as an ideal approach for detecting pathogens of severe acute infectious diseases. Rapid acquisition of high-quality target nucleic acid is the prerequisite to ensure the efficiency and accuracy of NAAT. Herein, we proposed a simple system in which magnetic nanoparticles (MNPs) based nucleic acid extraction was carried out in a plastic Pasteur pipette. Different from traditional approaches, this proposed system could be finished in 15 min without the supports of any electrical instruments. Furthermore, this system was superior to traditional MNPs based extraction methods in the aspects of rapid extraction and enhancing the sensitivity of a NAAT method, accelerated denaturation bubbles mediated strand exchange amplification (ASEA), to the pathogens from various artificial samples. Finally, this Pasteur pipette system was utilized for pathogen detection in actual samples of throat swabs, cervical swabs and gastric mucosa, the diagnosis results of which were identical with that provided by hospital. This rapid, easy-performing and efficiency extraction method ensures the applications of the NAAT in pathogen detection in regions with restricted resources.

Persistence of Antibody and Cellular Immune Responses in Coronavirus Disease 2019 Patients Over Nine Months After Infection.

BACKGROUND: The duration of humoral and T and B cell response after the infection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remains unclear. METHODS: We performed a cross-sectional study to assess the virus-specific antibody and memory T and B cell responses in coronavirus disease 2019 (COVID-19) patients up to 343 days after infection. Neutralizing antibodies and antibodies against the receptor-binding domain, spike, and nucleoprotein of SARS-CoV-2 were measured. Virus-specific memory T and B cell responses were analyzed. RESULTS: We enrolled 59 patients with COVID-19, including 38 moderate, 16 mild, and 5 asymptomatic patients; 31 (52.5%) were men and 28 (47.5%) were women. The median age was 41 years (interquartile range, 30-55). The median day from symptom onset to enrollment was 317 days (range 257 to 343 days). We found that approximately 90% of patients still have detectable immunoglobulin (Ig)G antibodies against spike and nucleocapsid proteins and neutralizing antibodies against pseudovirus, whereas ~60% of patients had detectable IgG antibodies against receptor-binding domain and surrogate virus-neutralizing antibodies. The SARS-CoV-2-specific IgG+ memory B cell and interferon-γ-secreting T cell responses were detectable in more than 70% of patients. CONCLUSIONS: Severe acute respiratory syndrome coronavirus 2-specific immune memory response persists in most patients approximately 1 year after infection, which provides a promising sign for prevention from reinfection and vaccination strategy.

Dense GAN and multi-layer attention based lesion segmentation method for COVID-19 CT images.

As the COVID-19 virus spreads around the world, testing and screening of patients have become a headache for governments. With the accumulation of clinical diagnostic data, the imaging big data features of COVID-19 are gradually clear, and CT imaging diagnosis results become more important. To obtain clear lesion information from the CT images of patients' lungs is helpful for doctors to adopt effective medical methods, and at the same time, is helpful to screen the patients with real infection. Deep learning image segmentation is widely used in the field of medical image segmentation. However, there are some challenges in using deep learning to segment the lung lesions of COVID-19 patients. Since image segmentation requires the labeling of lesion information on a pixel by pixel basis, most professional radiologists need to screen and diagnose patients on the front line, and they do not have enough energy to label a large amount of image data. In this paper, an improved Dense GAN to expand data set is developed, and a multi-layer attention mechanism method, combined with U-Net's COVID-19 pulmonary CT image segmentation, is proposed. The experimental results showed that the segmentation method proposed in this paper improved the segmentation accuracy of COVID-19 pulmonary medical CT image by comparing with other image segmentation methods.

Reduced Sleep Duration and Sleep Efficiency Were Independently Associated With Frequent Nightmares in Chinese Frontline Medical Workers During the Coronavirus Disease 2019 Outbreak.

OBJECTIVES: Nightmares were related to emotion and behavioral problems and also emerged as one of the core features of post-traumatic stress disorder (PTSD). Our study aimed to investigate the associations of frequent nightmares with sleep duration and sleep efficiency among frontline medical workers in Wuhan during the coronavirus disease 2019 (COVID-19) outbreak. METHODS: A total of 528 health-care workers from the province of Fujian providing medical aid in Wuhan completed the online questionnaires. There were 114 doctors and 414 nurses. The age, sex, marital status, and work situation were recorded. A battery of scales including the Pittsburgh Sleep Quality Index (PSQI) and the 12-item General Health Questionnaire (GHQ-12) were used to evaluate subjects' sleep and general mental health. Frequent nightmares were defined as the response of at least once a week in the item of "nightmare" of PSQI. RESULTS: Frequent nightmares were found in 27.3% of subjects. The frequent nightmare group had a higher score of PSQI-sleep duration and PSQI-habitual sleep efficiency (frequent nightmares vs. non-frequent nightmares: PSQI-sleep duration, 1.08 ± 0.97 vs. 0.74 ± 0.85, P < 0.001; PSQI-habitual sleep efficiency, 1.08 ± 1.10 vs. 0.62 ± 0.88, P < 0.001). Reduced sleep duration and reduced sleep efficiency were independently associated with frequent nightmares after adjustment for age, sex, poor mental health, and regular sleeping medication use (reduced sleep duration: OR = 1.96, 95% CI = 1.07-3.58, P = 0.029; reduced sleep efficiency: OR = 2.17, 95% CI = 1.09-4.32, P = 0.027). Subjects with both reduced sleep duration and sleep efficiency were also associated with frequent nightmares (OR = 2.70, 95% CI = 1.57-4.65, P < 0.001). CONCLUSION: The present study found that sleep duration and sleep efficiency were both independently associated with frequent nightmares among frontline medical workers in Wuhan during the COVID-19 pandemic. We should pay attention to nightmares and even the ensuing PTSD symptoms among subjects with reduced sleep duration or sleep efficiency facing potential traumatic exposure.

Characteristics and evaluation of the effectiveness of monitoring and control measures for the first 69 Patients with COVID-19 from 18 January 2020 to 2 March in Wuxi, China.

BACKGROUND: The Coronavirus disease (COVID-19) has caused 91,305 confirmed cases and 4746 deaths in China by 13:50 on October 11, 2020. We analyzed data on 69 infections in Wuxi to describe the disease's characteristics, to analyze factors of cases clinical outcome and to evaluate the prevention and control measures. METHODS: The demographic characteristics, exposure history, time indicators and propagation dynamics in Wuxi were collected. RESULTS: The clinical severity of cases was mostly mild and normal (75.36 %). Aging (relative risk [RR] = 1.04, 95 % confidence interval [CI]: 1.001-1.08) and fever (RR = 10.33, 95 %CI: 2.75-38.78) were risk factors for disease severity. The mean incubation period was estimated to be 4.77 days (95 % CI: 3.61-5.94), with a mean serial interval of 6.31 days (95 % CI: 5.12-7.50). The controlled reproduction number was estimated to be 1.12 (95 %CI: 0.71-1.69). CONCLUSIONS: The incidence of COVID-19 in Wuxi has turned into a lower level, suggesting the early prevention and control measures have achieved effectiveness. Aging and fever of initial symptom were risk factors for severe clinical outcome. The family clusters provided further clues of the risk factors for COVID-19 transmission.

A cluster of the Corona Virus Disease 2019 caused by incubation period transmission in Wuxi, China.

BACKGROUND: The infectivity and transmission capacity of COVID-2019 cases during the incubation period are not very clear. The manuscript described a cluster to provide information for research on incubation period infection. METHODS: We collected the required data from "Public Health Emergency Reporting Management Information System", epidemiological questionnaires for the cases, and laboratories. RESULTS: The cluster involved four generations, each of which was transmitted to the next generation during the incubation period. The time was 2-7 days, 6-7days, 3-8 days and 9 days prior to onset. As of March 11, the fourth-generation cases had no symptoms. Combined with the epidemiological data, we inferred that the source of the cluster was caused by the first-generation, who contacted with more than ten Wuhan people during the annual meeting from January 15 to 16. Two cases in this cluster were tested positive again during isolation and observation after discharge. CONCLUSIONS: We determined incubation period was infectious, and confirmed that it was contagious 9 days before the onset. The patients who were discharged might need to be observed for a period of time. This study was useful for the practical work, such as in the investigation of close contacts.

[Instructions for clinical research of oral diseases under the first level response to public health emergencies]

Corona virus disease 2019(COVID-19) has swept across our country and local governments have successively initiated first level response to public health emergencies. The outbreak of COVID-19 poses a huge challenge to the implementation of various clinical researches including oral diseases. In view of the specialty of oral diseases, it is particularly necessary to protect the health and rights of the participators and related practitioners, and to standardize the implementation of clinical trials. Based on the normative standards issued by the National Health Commission of China, consensus on clinical trial management formulated by the organization of clinical research, and regulations for the diagnosis and treatment of oral diseases during public health emergencies by Chinese Stomatological Association, we drafted the guideline for clinical research of oral diseases, with a view to provide reference for the clinical studies of oral diseases under the first level response to major public health emergencies.