Identification of conserved linear epitopes in the SARS-CoV-2 receptor-binding region using monoclonal antibodies.

The ongoing coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has caused millions of cases of infections, leading to a global health emergency. The SARS-CoV-2 spike (S) protein plays the most important role in viral infection, and S1 subunit and its receptor-binding domain (RBD) are widely considered the most attractive vaccine targets. The RBD is highly immunogenic and its linear epitopes are important for vaccine development and therapy, but linear epitopes on the RBD have rarely been reported. In this study, 151 mouse monoclonal antibodies (mAbs) against the SARS-CoV-2 S1 protein were characterized and used to identify epitopes. Fifty-one mAbs reacted with eukaryotic SARS-CoV-2 RBD. Sixty-nine mAbs reacted with the S proteins of Omicron variants B.1.1.529 and BA.5, indicating their potential as rapid diagnostic materials. Three novel linear epitopes of RBD, R6 (391CFTNVYADSFVIRGD405), R12 (463PFERDISTEIYQAGS477), and R16 (510VVVLSFELLHAPAT523), were identified; these were highly conserved in SARS-CoV-2 variants of concern and could be detected in the convalescent serum of COVID-19 patients. From pseudovirus neutralization assays, some mAbs including one detecting R12 were found to possess neutralizing activity. Together, from the reaction of mAbs with eukaryotic RBD (N501Y), RBD (E484K), and S1 (D614G), we found that a single amino acid mutation in the SARS-CoV-2 S protein may cause a structural alteration, exerting substantial impact on mAb recognition. Our results could, therefore, help us better understand the function of the SARS-CoV-2 S protein and develop diagnostic tools for COVID-19.

Comparative analysis of prevention and control measures toward COVID-19 epidemic between Shanghai and Beijing.

Purpose: By serving and providing a guide for other regional places, this study aims to advance and guide the epidemic prevention and control methods, and practices and strengthen people's ability to respond to COVID-19 and other future potential public health risks. Design/methodology/approach: A comparative analysis was conducted that the COVID-19 epidemic development trend and prevention and control effects both in Beijing and Shanghai. In fact, regarding the COVID-19 policy and strategic areas, the differences between governmental, social, and professional management were discussed and explored. To prevent and be ready for potential pandemics, experience and knowledge were used and summarized. Findings: The strong attack of the Omicron variant in early 2022 has posed challenges to epidemic prevention and control practices in many Chinese cities. Shanghai, which had achieved relatively good performance in the fight against the epidemic, has exposed limitations in its epidemic prevention and control system in the face of Omicron. In fact, the city of Beijing has undertaken prompt and severe lockdown measures and achieved rather good results in epidemic prevention and control because of learning from Shanghai's experience and lessons; adhering to the overall concept of "dynamic clearing," implementing precise prevention and monitoring, enhancing community control, and making emergency plans and preparations. All these actions and measures are still essential in the shift from pandemic response to pandemic control. Research limitations/implications: Different places have introduced different urgent policies to control the spread of the pandemic. Strategies to control COVID-19 have often been based on preliminary and limited data and have tended to be slow to evolve as new evidence emerges. Hence, the effects of these anti-epidemic policies need to be further tested.

Characterization and application of a series of monoclonal antibodies against SARS-CoV-2 nucleocapsid protein.

The ongoing coronavirus disease 2019 (COVID-19) pandemic has a significant global social and economic impact, and the emergence of new and more destructive mutant strains highlights the need for accurate virus detection. Here, 90 monoclonal antibodies (MAbs) that exclusively reacted with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid protein (NP) were generated. These MAbs did not cross-react with NPs of common human coronaviruses (HCoVs, i.e., 229E, OC43, HKU1, and NL63) and Middle East Respiratory Syndrome Coronavirus. Subsequently, overlapped peptides in individual fragments (N1-N4) of NP were synthesized. N1-3 (25-GSNQNGERSGARSKQ-39), N3-1 (217-AALALLLLDRLNQL-230), and N4-8 (393-TLLPAADLDDFSKQL-407) were identified as major epitopes using enzyme-linked immunoassay (ELISA) and recognized by 47, 1, and 18 MAbs, respectively. The 24 remaining MAbs exhibited no reactivity with all synthetic peptides. Among MAb-epitope pairs, only MAbs targeting epitope N1-3 displayed no cross-reaction with NPs of SARS-CoV-1 and other SARS-related CoVs. All Omicron variants contained a three-amino acid deletion (31ERS33) in the N1-3 region. Thus, MAbs targeting N1-3 failed to recognize these variants. Furthermore, a double-antibody sandwich ELISA for antigen detection was established using the optimal MAbs. Overall, a series of MAbs targeting SARS-CoV-2 NP was prepared, characterized with epitope mapping, and applied for the detection of SARS-CoV-2 antigens, and some novel B-cell epitopes of the viral NP were identified.

Understanding the condition of disease prevention and control workforce by disciplines, duties, and work stress during the COVID-19 pandemic: A case from Beijing disease prevention and control system.

Objective: The duties, discipline cross-complementation, and work stress of professional staff during the COVID-19 pandemic are analyzed and summarized to provide a scientific basis for workforce allocation and reserve in respect of infectious disease prevention and control in the disease prevention and control (DPC) system. Method: The cross-sectional survey was made in April-May 2021 on professional staff in the Beijing DPC system by way of typical + cluster sampling. A total of 1,086 staff were surveyed via electronic questionnaire, which was independently designed by the Study Group and involves three dimensions, i.e., General Information, Working Intensity & Satisfaction, and Need for Key Capacity Building. This paper focuses on the former two dimensions: General Information, Working Intensity, and Satisfaction. The information collected is stored in a database built with Microsoft Excel 2010 and analyzed statistically with SPSS 22.0. The results are expressed in absolute quantities and proportions. Assuming that the overload of work stress is brought by incremental duties and cross-discipline tasks, a binary logistic regression model is constructed. Results: Among the 1086 staff surveyed, 1032 staff were engaged in COVID-19 prevention and control works, and they can be roughly divided into two groups by their disciplines: Public Health and Preventive Medicine (hereinafter referred to P, 637 staff, as 61.72%) and Non-Public Health and Preventive Medicine (hereinafter referred to N-P, 395 staff, as 38.28%). During the COVID-19 pandemic, the 1,032 staff assumed a total of 2239 duties, that is, 2.17 per person (PP), or 2.45 PP for the P group and 1.72 PP for the N-P group. As to four categories of duties, i.e., Spot Epidemiological Investigation and Sampling, Information Management and Analysis, On-site Disposal, Prevention, Control Guidance, and Publicity, the P group accounts for 76.14, 78.50, 74.74, and 57.66%, respectively, while the N-P group accounts for 23.86, 21.50, 25.26, and 42.34%, respectively. Obviously, the former proportions are higher than the latter proportions. The situation is the opposite of the Sample Detection and Other Works, where the P group accounts for 25.00 and 31.33%, respectively, while the N-P group accounts for 75.00 and 68.67%, respectively. The analysis of work stress reveals that the P group and N-P group have similar proportions in view of full load work stress, being 48.67 and 50.13%, respectively, and the P group shows a proportion of 34.38% in view of overload work stress, apparently higher than the N-P group (24.05%). Moreover, both groups indicate their work stresses are higher than the pre-COVID-19 period levels. According to the analysis of work stress factors, the duty quantity and cross-discipline tasks are statistically positively correlated with the probability of overload work stress. Conclusion: The front-line staff in the DPC system involved in the COVID-19 prevention and control primarily fall in the category of Public Health and Preventive Medicine discipline. The P group assumes the most duties, and the N-P group serves as an important cross-complement. The study results indicate that the prevention and control of same-scale epidemic require the duty post setting at least twice than usual. As to workforce recruitment, allocation, and reserve in respect of the DPC system, two solutions are optional: less addition of P staff, or more addition of N-P staff. A balance between P and N-P staff that enables the personnel composition to accommodate both routine DPC and unexpected epidemic needs to be further discussed.

Establishment and evaluation of a 30-minute detection method for SARS-CoV-2 nucleic acid using a novel ultra-fast real-time PCR instrument.

BACKGROUND: The coronavirus disease 2019 (COVID-19) pandemic is still raging worldwide. Efficient, fast and low-cost severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleic acid detection methods are urgently needed. METHODS: A rapid PCR temperature change mode was explored by moving the reaction tube between the independent temperature modules with large temperature differences and a portable ultra-fast real-time PCR instrument were developed. We established a rapid SARS-CoV-2 test method using the ultra-fast real-time PCR instrument, a China Food and Drug Administration-certified SARS-CoV-2 reagent and optimized reaction condition. The analytical and clinical performances of the rapid tests were evaluated by comparing with the standard SARS-CoV-2 tests. RESULTS: The new temperature change mode can effectively shorten the amplification reaction time and be successfully used in the development of the ultra-fast real-time PCR instrument. The rapid SARS-CoV-2 test method was established and the time to yield results were greatly shortened from 81 min of the standard test to 31 min. Specificity of the rapid test was assessed and no non-specific amplification (0/63) was observed. The limits of detection of the rapid and standard tests were similar. Clinical performance was evaluated using 184 respiratory specimens from patients with suspected SARS-CoV-2 infection. The positive agreement between the rapid and standard tests was 100% (67/67), the negative agreement was 97.4% (114/117), and the kappa statistic was 0.965 (P<0.001). No significant differences in the Ct values for each target gene were observed between the rapid test and the standard test (P>0.05). CONCLUSIONS: We had developed a 30-minute detection method for SARS-CoV-2 nucleic acid using a novel ultra-fast real-time PCR instrument. The rapid test method may impact on patient management.

The hotspots and trends of coronavirus disease 2019 (COVID-19) and physical therapy: a bibliometric and visual analysis.

[Purpose] With the COVID-19 pandemic, more and more articles have been published to explore the role of physical therapy on COVID-19. In order to analyze the research hotspots and the trends of physical therapy and COVID-19, we conducted bibliometric and visual analysis. [Methods] Data were collected from the Science Citation Index Expanded (SCI-EXPANDED) and Social Sciences Citation Index (SSCI) of the Web of Science Core Collect (WoSCC) from 2019 to 2021. CiteSpace and VOSviewer were used to perform the visual analysis of keywords and references to help quickly get key information. [Results] A total of 466 publications were retrieved. Exercise, sedentary behavior, and mental health were research hotspots. The relationship between exercise and immunity, as well as the management of COVID-19 patients after discharge were the research trends. [Conclusion] This study provided relevant information for future research. Findings suggested that physical therapy is beneficial for suspected or confirmed COVID-19 patients during isolation. It is hoped that academic exchanges can be quickly established in the face of infectious diseases. And in the future, we should focus on the rehabilitation of discharged patients.

The Running Track of Government Responsibility During the Pandemic, Characteristic Analysis of Policy Documents Relevant to the COVID-19 Pandemic Released by Beijing Municipal Government in 2020.

Objective: Provide a reference point for the division of labor during the collaboration of multiple departments and the planning for the prevention and control of the Covid-19 epidemic of departments of the Beijing Municipal Government, from the perspective of policy documents. Methods: Policy documents and daily updates on COVID-19 cases published in 2020 are taken from the official website of Beijing Municipal Government and Beijing Municipal Health Commission. The characteristics of the pandemic situation and the content of relevant documents issued by different departments are described in five stages. Results: There were 988 confirmed cases of COVID-19 in Beijing in 2020, and policy analysis covered 444 documents (257 policy documents and 187 explanations of policy). A total of 153 policy documents were directly issued by the Beijing Municipal Government and its 45 subordinate commissions and bureaus, while others were policy forwarding from the central government and its relevant departments, county-level governments of Beijing and other organizations. Most cases and documents emerged during the initial stage of the pandemic (Level-I of the Emergency Response, which is the most serious). It was found that as many as 109 documents published by Beijing Municipal Government during the Level I emergency response period were relevant to economic and social development, 83 documents were related to disease control and medical services, and the rest were in close relation to the production and daily life of the people. Overall, major policy measures taken were relevant to 7 fields: finance, transportation, economic activities, employment people's lives, epidemic prevention and control and medical insurance. Policy implementation objectives were centered on promoting epidemic prevention and control and maintaining the stability of social production and residents' life. However, there are different emphases in different stages of the epidemic. Conclusion: Beijing municipality realized an effective mode of collaboration among multiple departments and organizations in the prevention and control of the COVID-19 epidemic, which was an example of the practice of "Health in All Policies."

Epitope mapping of severe acute respiratory syndrome-related coronavirus nucleocapsid protein with a rabbit monoclonal antibody.

The emergency SARS-CoV-2, a member of severe acute respiratory syndrome-related coronaviruses (SARSr-CoV), is still greatly harming the health of mankind. SARS-CoV-2-specific monoclonal antibodies (MAbs), which can identify SARS-CoV-2 from common human coronaviruses, are considered to extensively apply to developing rapid and reliable antigen assays. In this study we generated a rabbit MAb (RAb) detecting SARS-CoV-2 nucleocapsid protein (NP), which has cross-reaction with SARS-CoV-1 NP, but not with NPs of MERS and common human CoVs (OC43, NL63, 229E, and HKU1). With truncated NP fragments and synthesized peptides, the linear epitope detected by RAb was mapped in peptide N4-8, 393-407 amino acid residue (TLLPAADLDDFSKQL) of SARS-CoV-2 NP. This epitope N4-8 was highly conserved in SARSr-CoVs, including SARS-CoV-2, SARS-CoV-1, and bat CoV RaTG13 strain. However, the corresponding peptide of bat SARSr-CoV BtKY72 strain could not be recognized by RAb, which indicates amino acid D399 may be critical for N4-8 epitope detected by RAb. The present study will be conducive to developing reliable diagnosis for SARS-CoV-2 and gaining insights into the function of the SARS-CoV-2 N protein.

A more secure and effective method for throat swab collection: The importance of adequate exposure of oral cavity in COVID-19 specimen collection.

OBJECTIVES: This study aims to propose a novel and effective throat swab collection method for coronavirus disease 2019 (COVID-19). METHODS: The subjects were randomly divided into two groups. The subjects were asked to open their mouth to make "ah" sound (traditional method) or simulate yawn (improved method) for throat swab collection. The usage of tongue depressor, collection time, adverse reactions and subjective discomfort (VAS score) were compared. The collection time, comprehensive indicators of adverse reactions and VAS score were also compared among three collectors. RESULTS: The tongue depressor was less used in the improved group (χ2 = 40.186, P < 0.01). The average collection time of the traditional group was 5.44 ± 2.97 and that of the improved group was 4.00 ± 2.31 (P < 0.01). The subjects in the improved group had fewer and milder adverse reactions. The VAS score of subjects in the improved group was lower than that in the traditional group (P < 0.01). Among different collectors, the collection time, comprehensive indicators of adverse reactions and VAS were the same as the overall trend. CONCLUSION: Simulating yawn is a safer and faster throat swab collection method.

Influence of Different Inactivation Methods on Severe Acute Respiratory Syndrome Coronavirus 2 RNA Copy Number.

The outbreak of coronavirus disease 2019 (COVID-19) has spread across the world and was characterized as a pandemic. To protect medical laboratory personnel from infection, most laboratories inactivate the virus causing COVID-19, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), in clinical samples before testing. However, the effect of inactivation on the detection results remains unknown. Here, we used a digital PCR assay to determine the absolute SARS-CoV-2 RNA copy number in 63 nasopharyngeal swab samples and assess the effect of inactivation methods on viral RNA copy number. Viral inactivation was performed by three different methods: (i) incubation with the TRIzol LS reagent for 10 min at room temperature, (ii) heating in a water bath at 56°C for 30 min, and (iii) high-temperature treatment, including autoclaving at 121°C for 20 min, boiling at 100°C for 20 min, and heating at 80°C for 20 min. Compared to the amount of RNA in the original sample, TRIzol treatment destroyed 47.54% of the nucleocapsid protein (N) gene and 39.85% of open reading frame (ORF) 1ab. For samples treated at 56°C for 30 min, the copy number of the N gene and ORF 1ab was reduced by 48.55% and 56.40%, respectively. The viral RNA copy number dropped by 50 to 66% after heating at 80°C for 20 min. Nearly no viral RNA was detected after autoclaving at 121°C or boiling at 100°C for 20 min. These results indicate that inactivation reduced the quantity of detectable viral RNA and may cause false-negative results, especially in weakly positive cases. Thus, use of the TRIzol reagent rather than heat inactivation is recommended for sample inactivation, as the TRIzol reagent had the least effect on the RNA copy number among the tested methods.