Targeting Multiple Conserved T-Cell Epitopes for Protection against COVID-19 Moderate-Severe Disease by a Pan-Sarbecovirus Vaccine (preprint)

Background Most of current approved vaccines, based on a Spike-only as single immunogen, fall short of producing a full-blown T-cell immunity. SARS-CoV-2 continues to evolve with ever-emergent higher-contagious mutants that may take a turn going beyond Omicron to bring about a new pandemic outbreak. New recombinant SARS-CoV-2 species could be man-made through genetic manipulation to infect systemically. Development of composition-innovated, pan-variant COVID-19 vaccines to prevent from hospitalization and severe disease, and to forestall the next pandemic catastrophe, is an urgent global objective. Methods and findings In a retrospective, e-questionnaire Observational Study, extended from a clinical Phase-2 trial conducted in Taiwan, during the prime time of Omicron outbreak dominated by BA.2 and BA.5 variants, we investigated the preventive effects against COVID-19 moderate-severe disease (hospitalization and ICU admission) by a pan-Sarbecovirus vaccine UB-612 that targets monomeric S1-RBD-focused subunit protein and five designer peptides comprising sequence-conserved, non-mutable helper and cytotoxic T lymphocyte (Th/CTL) epitopes derived from Spike (S2), Membrane (M) and Nucleocapsid (N) proteins. Per UB-612 vaccination, there were no hospitalization and ICU admission cases (0% rate, 6 months after Omicron outbreak) reported [≥]14 months post-2nd dose of primary series, and [≥]10 months post-booster (3rd dose), to which the potent memory cytotoxic CD8 T cell immunity may be the pivotal in control of the infection disease severity. Six months post-booster, the infection rate (asymptomatic and symptomatic mild) was only 1.2%, which increased to 27.8% observed [≥]10 months post-booster. The notable protection effects are in good alignment with a preliminary Phase-3 heterologous booster trial report showing that UB-612 can serve as a competent booster substitute for other EUA-approved vaccine platforms to enhance their seroconversion rate and viral-neutralizing titer against Omicron BA.5. Conclusions UB-612, a universal multitope vaccine promoting full-blown T cell immunity, may work as a competent primer and booster for persons vulnerable to Sarbecovirus infection. Trial Registration. Clinical Trials.gov ID: NCT04773067.

Development of a multi-recombinase polymerase amplification assay for rapid identification of COVID-19, influenza A and B.

The coronavirus disease 2019 (COVID-19) pandemic caused extensive loss of life worldwide. Further, the COVID-19 and influenza mix-infection had caused great distress to the diagnosis of the disease. To control illness progression and limit viral spread within the population, a real-time reverse-transcription PCR (RT-PCR) assay for early diagnosis of COVID-19 was developed, but detection was time-consuming (4-6 h). To improve the diagnosis of COVID-19 and influenza, we herein developed a recombinase polymerase amplification (RPA) method for simple and rapid amplification of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19 and Influenza A (H1N1, H3N2) and B (influenza B). Genes encoding the matrix protein (M) for H1N1, and the hemagglutinin (HA) for H3N2, and the polymerase A (PA) for Influenza B, and the nucleocapsid protein (N), the RNA-dependent-RNA polymerase (RdRP) in the open reading frame 1ab (ORF1ab) region, and the envelope protein (E) for SARS-CoV-2 were selected, and specific primers were designed. We validated our method using SARS-CoV-2, H1N1, H3N2 and influenza B plasmid standards and RNA samples extracted from COVID-19 and Influenza A/B (RT-PCR-verified) positive patients. The method could detect SARS-CoV-2 plasmid standard DNA quantitatively between 102 and 105 copies/ml with a log linearity of 0.99 in 22 min. And this method also be very effective in simultaneous detection of H1N1, H3N2 and influenza B. Clinical validation of 100 cases revealed a sensitivity of 100% for differentiating COVID-19 patients from healthy controls when the specificity was set at 90%. These results demonstrate that this nucleic acid testing method is advantageous compared with traditional PCR and other isothermal nucleic acid amplification methods in terms of time and portability. This method could potentially be used for detection of SARS-CoV-2, H1N1, H3N2 and influenza B, and adapted for point-of-care (POC) detection of a broad range of infectious pathogens in resource-limited settings.

A novel gene signature based on the hub genes of COVID-19 predicts the prognosis of idiopathic pulmonary fibrosis.

Background: Increasing evidence has demonstrated that there was a strong correlation between COVID-19 and idiopathic pulmonary fibrosis (IPF). However, the studies are limited, and the real biological mechanisms behind the IPF progression were still uncleared. Methods: GSE70866 and GSE 157103 datasets were downloaded. The weight gene co-expression network analysis (WGCNA) algorithms were conducted to identify the most correlated gene module with COVID-19. Then the genes were extracted to construct a risk signature in IPF patients by performing Univariate and Lasso Cox Regression analysis. Univariate and Multivariate Cox Regression analyses were used to identify the independent value for predicting the prognosis of IPF patients. What's more, the Kyoto Encyclopedia of Genes and Genomes (KEGG), Gene Ontology (GO), and gene set enrichment analysis (GSEA) were conducted to unveil the potential biological pathways. CIBERSORT algorithms were performed to calculate the correlation between the risk score and immune cells infiltrating levels. Results: Two hundred thirty three differentially expressed genes were calculated as the hub genes in COVID-19. Fourteen of these genes were identified as the prognostic differentially expressed genes in IPF. Three (MET, UCHL1, and IGF1) of the fourteen genes were chosen to construct the risk signature. The risk signature can greatly predict the prognosis of high-risk and low-risk groups based on the calculated risk score. The functional pathway enrichment analysis and immune infiltrating analysis showed that the risk signature may regulate the immune-related pathways and immune cells. Conclusion: We identified prognostic differentially expressed hub genes related to COVID-19 in IPF. A risk signature was constructed based on those genes and showed great value for predicting the prognosis in IPF patients. What's more, three genes in the risk signature may be clinically valuable as potential targets for treating IPF patients and IPF patients with COVID-19.

Role of air pollutants in dengue fever incidence: evidence from two southern cities in Taiwan.

Air pollution may be involved in spreading dengue fever (DF) besides rainfalls and warmer temperatures. While particulate matter (PM), especially those with diameter of 10 µm (PM10) or 2.5 µm or less (PM25), and NO2 increase the risk of coronavirus 2 infection, their roles in triggering DF remain unclear. We explored if air pollution factors predict DF incidence in addition to the classic climate factors. Public databases and DF records of two southern cities in Taiwan were used in regression analyses. Month order, PM10 minimum, PM2.5 minimum, and precipitation days were retained in the enter mode model, and SO2 minimum, O3 maximum, and CO minimum were retained in the stepwise forward mode model in addition to month order, PM10 minimum, PM2.5 minimum, and precipitation days. While PM2.5 minimum showed a negative contribution to the monthly DF incidence, other variables showed the opposite effects. The sustain of month order, PM10 minimum, PM2.5 minimum, and precipitation days in both regression models confirms the role of classic climate factors and illustrates a potential biological role of the air pollutants in the life cycle of mosquito vectors and dengue virus and possibly human immune status. Future DF prevention should concern the contribution of air pollution besides the classic climate factors.

Targeting COVID-19 vaccine hesitancy among nurses in Shanghai: A latent profile analysis

Objectives This study aims to clarify the profiles of the psychological antecedents of vaccine hesitancy among Shanghai nurses with a person-centered approach. Methods A population-based cross-sectional online survey was conducted on Shanghai nurses from July to August 2021 (N = 1,928). In the online survey, participants were asked to report their sociodemographic, the 5C vaccine hesitancy components, their knowledge level of COVID-19 vaccine and vaccination, and the COVID-19 vaccination uptake intention and attention to vaccine news. Latent profile analysis was used to reveal distinct profiles of vaccine hesitancy. Results The results revealed four profiles, including “believers” (68.9%;high confidence and collective responsibility), “free riders” (12.7%;similar characteristics to believers, except for a low collective responsibility), “middlemen” (14.6%;middle in all 5C constructs), and “contradictors” (3.7%;high in all 5C constructs). Compared to believers, middlemen were younger, more likely to be female, childless, less educated, held lower professional titles, had fewer years of nursing service, sometimes or never complied with recommended vaccinations, had satisfactory or poor self-assessed health status, had no work experience during the COVID-19 epidemic, and possessed greater levels of knowledge. Free riders were more likely to work in community health centers and have a lower degree than believers. Contradictors were more likely to work in community health centers, had junior college degrees or lower, and had no work experience during the COVID-19 epidemic than believers. From the highest to the lowest on vaccination intention and attention to vaccine news were believers, then free riders, contradictors, and finally middlemen. Conclusion This study could aid in the development of personalized vaccination strategies based on nurses' vaccine hesitancy profiles and predictors. In addition to vaccine believers, we identified other three profiles based on their 5C psychological antecedents, emphasizing the significance of establishing tailored vaccination campaigns. Further research into the prevalence of profile structure in other groups of healthcare workers is required.

A novel gene signature based on the hub genes of COVID-19 predicts the prognosis of idiopathic pulmonary fibrosis

Background: Increasing evidence has demonstrated that there was a strong correlation between COVID-19 and idiopathic pulmonary fibrosis (IPF). However, the studies are limited, and the real biological mechanisms behind the IPF progression were still uncleared. Methods: GSE70866 and GSE 157103 datasets were downloaded. The weight gene co-expression network analysis (WGCNA) algorithms were conducted to identify the most correlated gene module with COVID-19. Then the genes were extracted to construct a risk signature in IPF patients by performing Univariate and Lasso Cox Regression analysis. Univariate and Multivariate Cox Regression analyses were used to identify the independent value for predicting the prognosis of IPF patients. What’s more, the Kyoto Encyclopedia of Genes and Genomes (KEGG), Gene Ontology (GO), and gene set enrichment analysis (GSEA) were conducted to unveil the potential biological pathways. CIBERSORT algorithms were performed to calculate the correlation between the risk score and immune cells infiltrating levels. Results: Two hundred thirty three differentially expressed genes were calculated as the hub genes in COVID-19. Fourteen of these genes were identified as the prognostic differentially expressed genes in IPF. Three (MET, UCHL1, and IGF1) of the fourteen genes were chosen to construct the risk signature. The risk signature can greatly predict the prognosis of high-risk and low-risk groups based on the calculated risk score. The functional pathway enrichment analysis and immune infiltrating analysis showed that the risk signature may regulate the immune-related pathways and immune cells. Conclusion: We identified prognostic differentially expressed hub genes related to COVID-19 in IPF. A risk signature was constructed based on those genes and showed great value for predicting the prognosis in IPF patients. What’s more, three genes in the risk signature may be clinically valuable as potential targets for treating IPF patients and IPF patients with COVID-19.

Effectiveness of the talent cultivation training program for industry transformation in Taiwan during the COVID-19 pandemic

In response to the impact of the COVID-19 pandemic on the Taiwanese tourism and hospitality industry, the Taiwanese government launched an industry revival and revitalization project. Subsidies were given for cultivating talent and industry transformation. The industry implemented talent cultivation courses to improve the skills and knowledge of workers and facilitate financial subsidization. This study reviewed the talent cultivation subsidy and training results for employees participating in training programs within Taiwan’s lodging industry. The results showed that the government promoted large-scale industry transformation and talent cultivation, and industry employees who received the training responded to it positively.

The Moderating Effect of Compassionate Mindfulness on the Psychological Needs and Emotions of Generation Y in the 21st Century in Taiwan.

During the 2020 COVID-19 pandemic in Taiwan, 6.5% of Generation Y required medical treatment for emotional and stress-related mental disorders. This study explores the moderating effect of mindfulness training on psychological needs and emotions to propose effective measures to promote the mental health of Generation Y. This study was carried out by questionnaire, using the data of respondents born in 1980-1999, collected in three different periods for quantitative analysis with compassionate mindfulness as the main variable. The results show that the compassionate mindfulness effect on emotion regulation varies greatly among different educational levels. However, it still plays a positive role in the psychological needs of Generation Y. Most members of Generation Y who receive compassionate mindfulness training have fewer basic needs and more interpersonal trust. They pay more attention to individual-oriented self-realization. Compassionate mindfulness has a greater positive moderating effect on the mental health of women aged 30-39 and those who are highly educated. Compassionate mindfulness has a more positive moderating effect on the psychological needs of members of Generation Y who were born more recently. During the COVID-19 pandemic, providing compassionate mindfulness has a significant positive effect on the prevention of mental disorders of Generation Y in Taiwan.

Evaluation of multi-component interventions for prevention of nosocomial pneumonia in older adults: a randomized, controlled trial.

AIMS: To evaluate the efficacy of multi-component interventions for prevention of hospital-acquired pneumonia in older patients hospitalized in geriatric wards. METHODS: A randomized, parallel-group, controlled trial was undertaken in patients aged 65 and above who were admitted to a tertiary hospital geriatric unit from January 1, 2016 to June 30, 2018 for an acute non-respiratory illness. Participants were randomized by to receive either a multi-component intervention (consisting of reverse Trendelenburg position, dysphagia screening, oral care and vaccinations), or usual care. The outcome measures were the proportion of patients who developed hospital-acquired pneumonia during hospitalisation, and mean time from randomization to the next hospitalisation due to respiratory infections in 1 year. RESULTS: A total of 123 participants (median age, 85; 43.1% male) were randomized, (n = 59) to intervention group and (n = 64) to control group. The multi-component interventions did not significantly reduce the incidence of hospital-acquired pneumonia but did increase the mean time to next hospitalisation due to respiratory infection (11.5 months vs. 9.5 months; P = 0.049), and reduced the risk of hospitalisation in 1 year (18.6% vs. 34.4%; P = 0.049). Implementation of multi-component interventions increased diagnoses of oropharyngeal dysphagia (35.6% vs. 20.3%; P < 0.001) and improved the influenza (54.5% vs 17.2%; P < 0.001) and pneumococcal vaccination rates (52.5% vs. 20.3%; P < 0.001). CONCLUSIONS: The nosocomial pneumonia multi-component intervention did not significantly reduce the incidence of hospital-acquired pneumonia during hospitalisation but reduce subsequent hospitalisations for respiratory infections. CLINICAL TRIAL REGISTRATION: ClinicalTrial.gov, NCT04347395.

Long Noncoding RNAs as Emerging Regulators of COVID-19.

Coronavirus disease 2019 (COVID-19), which has high incidence rates with rapid rate of transmission, is a pandemic that spread across the world, resulting in more than 3,000,000 deaths globally. Currently, several drugs have been used for the clinical treatment of COVID-19, such as antivirals (radecivir, baritinib), monoclonal antibodies (tocilizumab), and glucocorticoids (dexamethasone). Accumulating evidence indicates that long noncoding RNAs (lncRNAs) are essential regulators of virus infections and antiviral immune responses including biological processes that are involved in the regulation of COVID-19 and subsequent disease states. Upon viral infections, cellular lncRNAs directly regulate viral genes and influence viral replication and pathology through virus-mediated changes in the host transcriptome. Additionally, several host lncRNAs could help the occurrence of viral immune escape by inhibiting type I interferons (IFN-1), while others could up-regulate IFN-1 production to play an antiviral role. Consequently, understanding the expression and function of lncRNAs during severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection will provide insights into the development of lncRNA-based methods. In this review, we summarized the current findings of lncRNAs in the regulation of the strong inflammatory response, immune dysfunction and thrombosis induced by SARS-CoV-2 infection, discussed the underlying mechanisms, and highlighted the therapeutic challenges of COVID-19 treatment and its future research directions.

Inhibitors of Coronavirus 3CL Proteases Protect Cells from Protease-Mediated Cytotoxicity.

We describe a mammalian cell-based assay to identify coronavirus 3CL protease (3CLpro) inhibitors. This assay is based on rescuing protease-mediated cytotoxicity and does not require live virus. By enabling the facile testing of compounds across a range of 15 distantly related coronavirus 3CLpro enzymes, we identified compounds with broad 3CLpro-inhibitory activity. We also adapted the assay for use in compound screening and in doing so uncovered additional severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) 3CLpro inhibitors. We observed strong concordance between data emerging from this assay and those obtained from live-virus testing. The reported approach democratizes the testing of 3CLpro inhibitors by developing a simplified method for identifying coronavirus 3CLpro inhibitors that can be used by the majority of laboratories, rather than the few with extensive biosafety infrastructure. We identified two lead compounds, GC376 and compound 4, with broad activity against all 3CL proteases tested, including 3CLpro enzymes from understudied zoonotic coronaviruses. IMPORTANCE Multiple coronavirus pandemics have occurred over the last 2 decades. This has highlighted a need to be proactive in the development of therapeutics that can be readily deployed in the case of future coronavirus pandemics. We developed and validated a simplified cell-based assay for the identification of chemical inhibitors of 3CL proteases encoded by a wide range of coronaviruses. This assay is reporter free, does not require specialized biocontainment, and is optimized for performance in high-throughput screening. By testing reported 3CL protease inhibitors against a large collection of 3CL proteases with variable sequence similarity, we identified compounds with broad activity against 3CL proteases and uncovered structural insights into features that contribute to their broad activity. Furthermore, we demonstrated that this assay is suitable for identifying chemical inhibitors of proteases from families other than 3CL proteases.

Lead compounds for the development of SARS-CoV-2 3CL protease inhibitors.

We report the identification of three structurally diverse compounds - compound 4, GC376, and MAC-5576 - as inhibitors of the SARS-CoV-2 3CL protease. Structures of each of these compounds in complex with the protease revealed strategies for further development, as well as general principles for designing SARS-CoV-2 3CL protease inhibitors. These compounds may therefore serve as leads for the basis of building effective SARS-CoV-2 3CL protease inhibitors.

Author Correction: Cobrotoxin could be an effective therapeutic for COVID-19.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Clinical characteristics and corticosteroids application of different clinical types in patients with corona virus disease 2019.

To describe the epidemiological and clinical characteristics of patients with Corona Virus Disease 2019 (COVID-19) in Beijing. To analyze the application of corticosteroids in patients with severe pneumonia. We collected information on demographic characteristics, exposure history, clinical characteristics, corticosteroids use, and outcomes of the 65 confirmed cases of COVID-19 at Fifth Medical Center of PLA General Hospital from Jan 20 to Feb 23, 2020. The final follow-up date observed was April 15th, 2020. The number of patients with mild, general, severe, and critical type were 10 (15.38%), 32 (49.23%), 8 (12.31%), and 15 (23.08%), respectively. The median incubation period was 6 days. Notable outliers were 1 patient at 16 days and 1 patient at 21 days. In lymphocyte subgroup analysis, decreases in total, T, CD4, and CD8 lymphocytes were more common as the disease worsened (All P < 0.05). Methylprednisolone (mPSL) was applied to 31 (47.69%) patients with pneumonia, including 10 (31.25%) general, 8 (100%) severe, and 13 (86.67%) critical patients, respectively. Corticosteroids inhibited Interleukin-6(IL-6) production (P = 0.0215) but did not affect T lymphocyte (P = 0.0796). There was no significant difference between patients using lower dose (≤ 2 mg/kg day) and higher dose (> 2 mg/kg day) mPSL in inhibiting IL-6 production (P = 0.5856). Thirty of 31 patients (96.77%) had stopped mPSL due to improvement of pneumonia. Virus RNA clearance time lengthened with disease progression (P = 0.0001). In general type, there was no significant difference in virus clearance time between patients with (15, 12-19 days) and without (14.5, 11-18 days) (P = 0.7372) mPSL use. Lymphocyte, especially T lymphocyte, in severe and critical patients showed a dramatic decrease. Application of lower dose corticosteroids (≤ 2 mg/kg day) could inhibit IL-6 production (a representative of cytokines) as effectively as a higher dose. Proper use corticosteroids in general type patients did not delay virus clearance.

SARS-CoV-2 infection of the liver directly contributes to hepatic impairment in patients with COVID-19.

Background & Aims: Liver enzyme abnormalities are common in patients with coronavirus disease 2019 (COVID-19). Whether or not severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection can lead to liver damage per se remains unknown. Herein, we reported the clinical characteristics and liver pathological manifestations of COVID-19 patients with liver enzyme abnormalities. Methods: We analyzed 156 patients diagnosed with COVID-19 from 2 designated centers in China and compared clinical features between patients with or without elevated aminotransferases. Postmortem liver biopsies were obtained from 2 cases who had elevated aminotransferases. We investigated the patterns of liver impairment by electron microscopy, immunohistochemistry, TUNEL assay and pathological studies. Results: Sixty-four out of 156 (41.0%) patients with COVID-19 had elevated aminotransferases. The median levels of alanine aminotransferase were 50 U/L vs. 19 U/L, respectively, aspartate aminotransferase were 45.5 U/L vs. 24 U/L, respectively in abnormal and normal aminotransferase groups. Liver enzyme abnormalities were associated with disease severity, as well as a series of laboratory tests including higher alveolar-arterial oxygen partial pressure difference, higher gamma-glutamyltransferase, lower albumin, decreased CD4+ T cells and B lymphocytes. Ultrastructural examination identified typical coronavirus particles, characterized by spike structures, in the cytoplasm of hepatocytes in 2 COVID-19 cases. SARS-CoV-2-infected hepatocytes displayed conspicuous mitochondrial swelling, endoplasmic reticulum dilatation and glycogen granule decrease. Histologically, massive hepatic apoptosis and some binuclear hepatocytes were observed. Taken together, both ultrastructural and histological evidence indicated a typical lesion of viral infection. Immunohistochemical results showed scarce CD4+ and CD8+ lymphocytes. No obvious eosinophil infiltration, cholestasis, fibrin deposition, granuloma, massive central necrosis, or interface hepatitis were observed. Conclusions: SARS-CoV-2 infection in the liver directly contributes to hepatic impairment in patients with COVID-19. Hence, a surveillance of viral clearance in liver and long-term outcome of COVID-19 is required. Lay summary: Liver enzyme abnormalities are common in patients with coronavirus disease 2019 (COVID-19). We reported the clinical characteristics and liver pathological manifestations of COVID-19 patients with elevated liver enzymes. Our findings suggested that SARS-CoV-2 infection of the liver is a crucial factor contributing to hepatic impairment in patients with COVID-19.

Clinical and pathological investigation of severe COVID-19 patients

BACKGROUND: Severe acute respiratory coronavirus 2 (SARS-CoV-2) caused coronavirus disease 2019 (COVID-19) has become a pandemic. This study addressed the clinical and immunopathological characteristics of severe COVID-19. METHODS: Sixty-nine COVID-19 patients were classified into as severe and non-severe groups to analyze their clinical and laboratory characteristics. A panel of blood cytokines was quantified over time. Biopsy specimens from two deceased cases were obtained for immunopathological, ultrastructural, and in situ hybridization examinations. RESULTS: Circulating cytokines, including IL8, IL6, TNFα, IP10, MCP1, and RANTES, were significantly elevated in severe COVID-19 patients. Dynamic IL6 and IL8 were associated with disease progression. SARS-CoV-2 was demonstrated to infect type II, type I pneumocytes and endothelial cells, leading to severe lung damage through cell pyroptosis and apoptosis. In severe cases, lymphopenia, neutrophilia, depletion of CD4+ and CD8+ T lymphocytes, and massive macrophage and neutrophil infiltrates were observed in both blood and lung tissues. CONCLUSIONS: A panel of circulating cytokines could be used to predict disease deterioration and inform clinical interventions. Severe pulmonary damage was predominantly attributed to both SARS-CoV-2 caused cytopathy and immunopathologic damage. Strategies that encourage pulmonary recruitment and overactivation of inflammatory cells by suppressing cytokine storm might improve the outcomes of severe COVID-19 patients.