Orf virus DNA prime-protein boost strategy is superior to adenovirus-based vaccination in mice and sheep.

Contagious ecthyma (Orf), an acute and highly contagious zoonosis, is prevalent worldwide. Orf is caused by Orf virus (ORFV), which mainly infects sheep/goats and humans. Therefore, effective and safe vaccination strategies for Orf prevention are needed. Although immunization with single-type Orf vaccines has been tested, heterologous prime-boost strategies still need to be studied. In the present study, ORFV B2L and F1L were selected as immunogens, based on which DNA, subunit and adenovirus vaccine candidates were generated. Of note, heterologous immunization strategies using DNA prime-protein boost and DNA prime-adenovirus boost in mice were performed, with single-type vaccines as controls. We have found that the DNA prime-protein boost strategy induces stronger humoral and cellular immune responses than DNA prime-adenovirus boost strategy in mice, which was confirmed by the changes in specific antibodies, lymphocyte proliferation and cytokine expression. Importantly, this observation was also confirmed when these heterologous immunization strategies were performed in sheep. In summary, by comparing the two immune strategies, we found that DNA prime-protein boost strategy can induce a better immune response, which provides a new attempt for exploring Orf immunization strategy.

Preparation of equine anti-SARS-CoV-2 immunoglobulin special IgG and special F (ab')2

After immunizing healthy horses with SARS-CoV-2 virus-like particles (VLPs) as immunogens, immunized horse serum was collected. The total IgG in the serum was separated by affinity chromatography, and then digested with pepsin to obtain immunoglobulin F(ab')2, the IgG and F(ab')2 using an immunochro-matographic column that binds to the RBD protein to obtain a highly specific horse Anti-SARS-CoV-2 IgG and F(ab')2. It's concentration of IgG and F(ab')2 is 2.36 mg/mL and 1.05 mg/mL, whi le the recovery rates were 11% and 4.89%, and the purities of prepared IgG and F(ab')2 were 91% and 96%. Semi-inhibited concentrations of pseudovirus (IC50) were 1.406 g/mL and 0.862 g/mL. These results show that a high purity, specificity, activity of specific IgG and F(ab')2 against SARS-CoV-2 was prepared successfully, which laid a foundation for preparing safe and efficient anti-SARS-CoV-2 therapeutic antibody drugs.

Long-Acting Protective Ocular Surface by Instilling Adhesive Dual-Antiviral Nanoparticles.

The eye is susceptible to viral infections, causing severe ocular symptoms or even respiratory diseases. Methods capable of protecting the eye from external viral invasion in a long-term and highly effective way are urgently needed but have been proved to be extremely challenging. Here, a strategy of forming a long-acting protective ocular surface is described by instilling adhesive dual-antiviral nanoparticles. Taking pseudotyped severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) as a model virus, antiviral agent-loaded nanoparticles are coated with a "double-lock" hybrid cell membrane abundant with integrin-ß1 and angiotensin converting enzyme II (ACE2). After instillation, the presence of integrin-ß1 endows coated nanoparticles with steady adhesion via specific binding to Arg-Gly-Asp sequence on the fibronectin of ocular epithelium, achieving durable retention on the ocular surface. In addition to loaded inhibitors, the exposure of ACE2 can trap SARS-CoV-2 and subsequently neutralize the associated spike protein, playing a dual antiviral effect of the resulting nanoparticles. Adhesive dual-antiviral nanoparticles enabled by coating with a "double-lock" hybrid cell membrane could be a versatile platform for topical long-acting protection against viral infection of the eye.

Dressing Bacteria With a Hybrid Immunoactive Nanosurface to Elicit Dual Anticancer and Antiviral Immunity.

Approaches capable of simultaneously treating cancer and protecting susceptible patients from lethal infections such as coronavirus disease 2019, are highly desirable but prove to be difficult. Here, dressing bacteria with a hybrid immunoactive nanosurface is reported to elicit dual anticancer and antiviral immunity. A combination of a checkpoint blocking antibody and a virus-specific antigen is covalently conjugated to polydopamine nanoparticles, which can be anchored onto bacterial surface, by a one-step in situ polymerization of dopamine under a cell-friendly condition. By virtue of the ability to colonize and penetrate deep tumor tissue, dressed bacteria enable sustained release and expanded exposure of carried immunoactivators to stimulate immune cells. In addition to a carrier role, bacteria are able to further provoke innate immunity due to the native immunogenicity of the pathogen-associated molecular patterns. Immunization with dressed bacteria promotes the maturation, and activation of antigen-presenting cells, which induces robust humoral and cellular immune responses in tumor-bearing mice. As evidenced by efficient production of viral-antigen-specific immunoglobulin G antibody in serum and significantly suppressed tumor growth in different models, dressing bacteria with a hybrid immunoactive nanosurface paves an avenue to prepare next-generation therapeutics for synergistic treatment and prevention.

PEGylation Prolongs the Half-Life of Equine Anti-SARS-CoV-2 Specific F(ab')2.

Therapeutic antibodies-F(ab')2 obtained from hyperimmune equine plasma could treat emerging infectious diseases rapidly because of their high neutralization activity and high output. However, the small-sized F(ab')2 is rapidly eliminated by blood circulation. This study explored PEGylation strategies to maximize the half-life of equine anti-SARS-CoV-2 specific F(ab')2. Equine anti-SARS-CoV-2 specific F(ab')2 were combined with 10 KDa MAL-PEG-MAL in optimum conditions. Specifically, there were two strategies: Fab-PEG and Fab-PEG-Fab, F(ab')2 bind to a PEG or two PEG, respectively. A single ion exchange chromatography step accomplished the purification of the products. Finally, the affinity and neutralizing activity was evaluated by ELISA and pseudovirus neutralization assay, and ELISA detected the pharmacokinetic parameters. The results displayed that equine anti-SARS-CoV-2 specific F(ab')2 has high specificity. Furthermore, PEGylation F(ab')2-Fab-PEG-Fab had a longer half-life than specific F(ab')2. The serum half-life of Fab-PEG-Fab, Fab-PEG, and specific F(ab')2 were 71.41 h, 26.73 h, and 38.32 h, respectively. The half-life of Fab-PEG-Fab was approximately two times as long as the specific F(ab')2. Thus far, PEGylated F(ab')2 has been prepared with high safety, high specificity, and a longer half-life, which could be used as a potential treatment for COVID-19.

Chiral-Selective Antigen-Presentation by Supramolecular Chiral Polymer Micelles.

Chirality is ubiquitous in biological systems, which is closely related to biological functions, life process, and even pathogenesis of diseases. However, the interface between the chirality of synthetic materials and organisms, particularly the immune system, remains poorly understood. Here, supramolecular chiral polymer micelles (SCPMs) are prepared by complexing antigenic proteins with chiral amino acid modified polyethyleneimine. The introduction of chirality not only reduces the toxicity of cationic polymer, but also benefits cell uptake and antigen presentation. Especially, D-chirality presents the lowest cytotoxicity, while promotes the highest expression level of costimulatory molecules on dendritic cells compared to L-chirality and achirality. The superiority of D-chirality to stimulate dendritic cell maturation is supported by immunization with D-SCPMs, which achieves significant antigen-specific proliferation of T cells in the spleen, lymph nodes and tumor of mice. Chirality-mediated antigen processing and presentation is demonstrated by D-SCPMs self-assembled from chiral alkaline histidine or neutral phenylalanine modified polyethyleneimine and tumor associated ovalbumin or severe acute respiratory syndrome coronavirus 2 spike 1 antigenic protein. Immunoactivation enabled by D-chirality opens a window to prepare potent nanotherapeutics for disease prevention and treatment. This article is protected by copyright. All rights reserved.

Correction: Effect of the "Art Coloring" Online Coloring Game on Subjective Well-Being Increase and Anxiety Reduction During the COVID-19 Pandemic: Development and Evaluation.

[This corrects the article DOI: 10.2196/37026.].

Discovery and characterization of SARS-CoV-2 reactive and neutralizing antibodies from humanized CAMouseHG mice through rapid hybridoma screening and high-throughput single-cell V(D)J sequencing.

The coronavirus disease 2019 pandemic has caused more than 532 million infections and 6.3 million deaths to date. The reactive and neutralizing fully human antibodies of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are effective detection tools and therapeutic measures. During SARS-CoV-2 infection, a large number of SARS-CoV-2 reactive and neutralizing antibodies will be produced. Most SARS-CoV-2 reactive and neutralizing fully human antibodies are isolated from human and frequently encoded by convergent heavy-chain variable genes. However, SARS-CoV-2 viruses can mutate rapidly during replication and the resistant variants of neutralizing antibodies easily survive and evade the immune response, especially in the face of such focused antibody responses in humans. Therefore, additional tools are needed to develop different kinds of fully human antibodies to compensate for current deficiency. In this study, we utilized antibody humanized CAMouseHG mice to develop a rapid antibody discovery method and examine the antibody repertoire of SARS-CoV-2 RBD-reactive hybridoma cells derived from CAMouseHG mice by using high-throughput single-cell V(D)J sequencing analysis. CAMouseHG mice were immunized by 28-day rapid immunization method. After electrofusion and semi-solid medium screening on day 12 post-electrofusion, 171 hybridoma clones were generated based on the results of SARS-CoV-2 RBD binding activity assay. A rather obvious preferential usage of IGHV6-1 family was found in these hybridoma clones derived from CAMouseHG mice, which was significantly different from the antibodies found in patients with COVID-19. After further virus neutralization screening and antibody competition assays, we generated a noncompeting two-antibody cocktail, which showed a potent prophylactic protective efficacy against SARS-CoV-2 in cynomolgus macaques. These results indicate that humanized CAMouseHG mice not only provide a valuable platform to obtain fully human reactive and neutralizing antibodies but also have a different antibody repertoire from humans. Thus, humanized CAMouseHG mice can be used as a good complementary tool in discovery of fully human therapeutic and diagnostic antibodies.

Proteomic analysis and identification reveal the anti-inflammatory mechanism of clofazimine on lipopolysaccharide-induced acute lung injury in mice.

BACKGROUND AND OBJECTIVE: Acute lung injury (ALI)/ acute respiratory distress syndrome (ARDS) was increasingly recognized as one of the most severe acute hyperimmune response of coronavirus disease 2019 (COVID-19). Clofazimine (CFZ) has attracted attention due to its anti-inflammatory property in immune diseases as well as infectious diseases. However, the role and potential molecular mechanism of CFZ in anti-inflammatory responses remain unclear. METHODS: We analyze the protein expression profiles of CFZ and LPS from Raw264.7 macrophages using quantitative proteomics. Next, the protective effect of CFZ on LPS-induced inflammatory model is assessed, and its underlying mechanism is validated by molecular biology analysis. RESULTS: LC-MS/MS-based shotgun proteomics analysis identified 4746 (LPS) and 4766 (CFZ) proteins with quantitative information. The key proteins and their critical signal transduction pathways including TLR4/NF-κB/HIF-1α signaling was highlighted, which was involved in multiple inflammatory processes. A further analysis of molecular biology revealed that CFZ could significantly inhibit the proliferation of Raw264.7 macrophages, decrease the levels of TNF-α and IL-1ß, alleviate lung histological changes and pulmonary edema, improve the survival rate, and down-regulate TLR4/NF-κB/HIF-1α signaling in LPS model. CONCLUSION: This study can provide significant insight into the proteomics-guided pharmacological mechanism study of CFZ and suggest potential therapeutic strategies for infectious disease.

A new circular RNA-encoded protein BIRC6-236aa inhibits transmissible gastroenteritis virus (TGEV)-induced mitochondrial dysfunction.

Transmissible gastroenteritis virus (TGEV), a member of the coronavirus family, is the pathogen responsible for transmissible gastroenteritis, which results in mitochondrial dysfunction in host cells. Previously, we identified 123 differentially expressed circular RNAs (cRNA)from the TGEV-infected porcine intestinal epithelial cell line jejunum 2 (IPEC-J2). Previous bioinformatics analysis suggested that, of these, circBIRC6 had the potential to regulate mitochondrial function. Furthermore, mitochondrial permeability transition, a key step in the process of mitochondrial dysfunction, is known to be caused by abnormal opening of mitochondrial permeability transition pores (mPTPs) regulated by the voltage-dependent anion-selective channel protein 1 (VDAC)-Cyclophilin D (CypD) complex. Therefore, in the present study, we investigated the effects of circBIRC6-2 on mitochondrial dysfunction and opening of mPTPs. We found that TGEV infection reduced circBIRC6-2 levels, which in turn reduced mitochondrial calcium (Ca2+) levels, the decrease of mitochondrial membrane potential, and opening of mPTPs. In addition, we also identified ORFs and internal ribosomal entrance sites within the circBIRC6-2 RNA. We demonstrate circBIRC6-2 encodes a novel protein, BIRC6-236aa, which we show inhibits TGEV-induced opening of mPTPs during TGEV infection. Mechanistically, we identified an interaction between BIRC6-236aa and VDAC1, suggesting that BIRC6-236aa destabilizes the VDAC1-CypD complex. Taken together, the results suggest that the novel protein BIRC6-236aa encoded by cRNA circBIRC6-2 inhibits mPTP opening and subsequent mitochondrial dysfunction by interacting with VDAC1.

Effect of the "Art Coloring" Online Coloring Game on Subjective Well-Being Increase and Anxiety Reduction During the COVID-19 Pandemic: Development and Evaluation.

BACKGROUND: COVID-19 has spread worldwide and generated tremendous stress on human beings. Unfortunately, it is often hard for distressed individuals to access mental health services under conditions of restricted movement or even lockdown. OBJECTIVE: The study first aims to develop an online digital intervention package based on a commercially released coloring game. The second aim is to test the effectiveness of difference intervention packages for players to increase subjective well-being (SWB) and reduce anxiety during the pandemic. METHODS: An evidence-based coloring intervention package was developed and uploaded to an online coloring game covering almost 1.5 million players worldwide in January 2021. Players worldwide participated to color either 4 rounds of images characterized by awe, pink, nature, and blue or 4 rounds of irrelevant images. Participants' SWB and anxiety and the perceived effectiveness of the game in reducing anxiety (subjective effectiveness [SE]) were assessed 1 week before the intervention (T1), after the participants completed pictures in each round (T2-T5), and after the intervention (T6). Independent 2-tailed t tests were conducted to examine the general intervention (GI) effect and the intervention effect of each round. Univariate analysis was used to examine whether these outcome variables were influenced by the number of rounds completed. RESULTS: In total, 1390 players worldwide responded and completed at least 1 assessment. Overall, the GI group showed a statistical significantly greater increase in SWB than the general control (GC) group (N=164, t162=3.59, Cohen d=0.59, 95% CI 0.36-1.24, P<.001). Compared to the control group, the best effectiveness of the intervention group was seen in the awe round, in which the increase in SWB was significant (N=171, t169=2.51, Cohen d=0.39, 95% CI 0.10-0.82, P=.01), and players who colored all 4 pictures had nearly significant improvements in SWB (N=171, F4,170=2.34, partial ŋ2=0.053, P=.06) and a significant decrease in anxiety (N=171, F4,170=3.39, partial ŋ2=0.075, P=.01). CONCLUSIONS: These data indicate the effectiveness of online psychological interventions, such as coloring games, for mental health in the specific period. They also show the feasibility of applying existing commercial games embedded with scientific psychological interventions that can fill the gap in mental crises and services for a wider group of people during the pandemic. The results would inspire innovations to prevent the psychological problems caused by public emergencies and encourage more games, especially the most popular ones, to take more positive action for the common crises of humankind.

A trust model for consumer conversion in community-based group buying: the dual roles of group leaders

Purpose>In the presence of coronavirus disease 2019 (COVID-19), due to the social distance restriction, consumers' regular consumption behaviors and patterns have been changing fundamentally. Thereafter, an innovative group buying model has emerged and developed explosively with a specific focus on consumer's location, known as community-based group buying (CGB). The purpose of this paper is to investigate the transfer mechanism of user's trust in dyadic contexts of social and commercial role-playing in the CGB program.Design/methodology/approach>This study adopts an empirical research method, with an online and offline questionnaire survey, a total of 382 responses have been obtained. Then, both descriptive analysis and hierarchical regression analysis are conducted to explore the dual roles of group leader and its corresponding effects on consumers' trust (i.e. emotional trust and behavioral trust) and engagement actions (i.e. purchase and share) in the CGB program.Findings>Results indicate that resident's trust and their perception of group leader's friend role can positively enhance their engagement actions in the CGB programs. Meanwhile, for the purpose of profit maximization, the group leader is more willing to play a friend role in transactions no matter whether the role conflict exists.Originality/value>Research findings provide some managerial insights for CGB platform on the selection and training of group leaders and the incentive mechanism design.

Assessing the impact of insurance type on COVID-19 mortality in black and white patients in the largest healthcare system in the state of georgia.

IMPORTANCE: The increased COVID-19 mortality for Black individuals over White individuals may be explained by the known racial disparities in access to insurance. OBJECTIVE: To determine whether racial disparities in COVID-19 mortality still exist when Blacks and Whites are equally insured. DESIGN: Routinely collected data on race, mortality, type of insurance, known risk factors, and lab results from the EPIC Patient Management System were analyzed using a multivariable logistic regression model. SETTING: Piedmont Healthcare is the largest hospital system in Georgia. Due to its multiple locations across the state of Georgia, it receives a relatively equitably insured population. PARTICIPANTS: All patients hospitalized with a positive COVID-19 status between March 1 and November 30, 2020. MAIN OUTCOMES: We hypothesized that Black patients would not have higher odds of mortality than White patients, and that type of insurance would predict COVID-19 mortality. RESULTS: 6,881 (3,674 Black, 3,207 White; 48% male, mean age = 60) patients were included. Race was not a significant predictor of COVID-19 mortality (p>0.05). When controlling for age and insurance, the mortality rate for Black patients was not statistically significant from that for White patients (p>0.05). Compared to those relying on Medicare, patients with commercial (OR=0.68, 95% CI: 0.48-0.96) or out-of-pocket (self-pay) insurance (OR=0.22, 95% CI: 0.03-0.88) had lower odds of mortality. CONCLUSIONS: National trends of racial disparities in COVID-19 mortality may be partially explained by disparities in insurance.

Systemic antiviral immunization by virus-mimicking nanoparticles-decorated erythrocytes.

New vaccine technologies are urgently needed to produce safe and effective vaccines in a more timely manner to prevent future infectious disease pandemics. Here, we describe erythrocyte-mediated systemic antiviral immunization, a versatile vaccination strategy that boosts antiviral immune responses by using erythrocytes decorated with virus-mimetic nanoparticles carrying a viral antigen and a Toll-like receptor (TLR) agonist. As a proof of concept, polydopamine nanoparticles were synthesized via a simple in situ polymerization in which the nanoparticles were conjugated with the SARS-CoV-2 spike protein S1 subunit and the TLR7/8 agonist R848. The resulting SARS-CoV-2 virus-mimetic nanoparticles were attached to erythrocytes via catechol groups on the nanoparticle. Erythrocytes naturally home to the spleen and interact with the immune system. Injection of the nanoparticle-decorated erythrocytes into mice resulted in greater maturation and activation of antigen-presenting cells, humoral and cellular immune responses in the spleen, production of S1-specific immunoglobulin G (IgG) antibodies, and systemic antiviral T cell responses than a control group treated with the nanoparticles alone, with no significant negative side effects. These results show that erythrocyte-mediated systemic antiviral immunization using viral antigen- and TLR agonist-presenting polydopamine nanoparticles-a generalizable method applicable to many viral infections-is effective new approach to developing vaccines against severe infectious diseases.

Using the Internet Big Data to Investigate the Epidemiological Characteristics of Allergic Rhinitis and Allergic Conjunctivitis.

BACKGROUND: To explore the epidemiological characteristics of allergic rhinitis (AR) and allergic conjunctivitis (AC) based on the Internet big data. METHODS: The Baidu index (BDI) of keywords "allergic rhinitis" and "allergic conjunctivitis" in Mandarin, the daily pollen concentration (PC) released by the Beijing Meteorological Bureau and the volumes of outpatient visits (OV) of the Beijing Tongren Hospital (Beijing) and the Third Affiliated Hospital of Sun Yat-sen University (Guangzhou) from 2017 to 2020 were obtained. The temporal and spatial changes of AR and AC were discussed. The correlations between BDI and PC/OV were analyzed by Spearman correlation analysis. RESULTS: The trends of BDI of "AR"/"AC" in Beijing showed obvious seasonal variations, but not in Guangzhou. The BDI of "AR" and "AC" was consistent with the OV in both cities (r1AR-BJ=0.580, P<0.001; r1AR-GZ=0.360, P=0.031; r1AC-BJ=0.885, P<0.001; r1AC-GZ=0.694, P<0.001). The BDI of "AR" and "AC" was highly consistent with the change of the PC in Beijing (r AR-Pollen=0.826, P<0.001; r AC-Pollen=0.564, P<0.001). The OV of AR in Beijing and Guangzhou decreased significantly in the first half of 2020, but there was no significant change in AC. In the first half of 2020, the OV of AC in Beijing was significantly higher than that of AR, while that of AC in Guangzhou was slightly higher than that of AR. CONCLUSION: The BDI could reflect the real-world situation to some extent and has the potential to predict the epidemiological characteristics of AR and AC. The BDI and OV of AR decreased significantly, but those of AC were still at a high level, during the COVID-19 pandemic, in the environment where most people in Beijing and Guangzhou wore masks without eye protection.

The Influence Mechanisms of Chinese College Students' Individual Cognitions on Their COVID-19 Pop-Science Transmission Behaviors on Social Media

This study explores what specific factors might influence Chinese college students' information transmission patterns on COVID‐19 pop‐science in China. Based on the risk perception theory and the situational theory of problem solving, an influence mechanism model of health information transmission behaviors online was constructed. This study investigated the public's information transmission behaviors of a particular type of health information on social medias during this global public health emergency, and on, specifically, why and how health information were transferred.

Molecular diagnosis of COVID-19: Current situation and trend in China (Review).

COVID-19 is caused by a novel coronavirus (2019-nCoV or SARS-CoV-2) and has become a global public health emergency. Rapid and accurate molecular diagnostic technologies are crucial for the screening, isolation, treatment, prevention and control of COVID-19. Currently, nucleic acid detection-based techniques and rapid diagnostic tests that detect antigens or antibodies specific to 2019-nCoV infections are the primary diagnostic tools. China National Medical Products Administration has opened a special channel for approval of new pharmaceuticals owing to urgent clinical needs, with 18 nucleic acid detection kits, 11 protein detection kits and 1 sequencing-related equipment and supporting software having been approved until April 23, 2020. The current review summarizes the application situation, advantages, disadvantages and associated technology improvement trends of molecular diagnostics for COVID-19 in China, identifies knowledge gaps and indicates future priorities for research in this field. The most effective way to prevent and control COVID-19 is early detection, diagnosis, isolation and treatment. In the clinical application of molecular diagnosis technology, it is necessary to combine pathogenic microbiology, immunology and other associated detection technologies, advocate the combination of multiple technologies, determine how they complement each other, enhance practicability and improve the ability of rapid and accurate diagnosis and differential diagnosis of COVID-19.