Heparan sulfate promotes ACE2 super-cluster assembly to enhance SARS-CoV-2-associated syncytium formation.

The mechanism of syncytium formation, caused by spike-induced cell-cell fusion in severe COVID-19, is largely unclear. Here we combine chemical genetics with 4D confocal imaging to establish the cell surface heparan sulfate (HS) as a critical host factor exploited by SARS-CoV-2 to enhance spike’s fusogenic activity. HS binds spike to facilitate ACE2 clustering, generating synapse-like cell-cell contacts to promote fusion pore formation. ACE2 clustering, and thus, syncytium formation is significantly mitigated by chemical or genetic elimination of cell surface HS, while in a cell-free system consisting of purified HS, spike, and lipid-anchored ACE2, HS directly induces ACE2 clustering. Importantly, the interaction of HS with spike allosterically enables a conserved ACE2 linker in receptor clustering, which concentrates spike at the fusion site to overcome fusion-associated activity loss. This fusion-boosting mechanism can be effectively targeted by an investigational HS-binding drug, which reduces syncytium formation in vitro and viral infection in mice.

Research on a Sustainable Teaching Model Based on the OBE Concept and the TSEM Framework

This paper reports the results of a study on the implementation of a sustainable teaching model based on the OBE (Outcome-Based Education) concept and the TSEM (Teach, Study, Evaluate, and Manage) framework in computer science and technology at NingboTech University, China. In the context of digital education, the OBE concept and the TSEM framework are integrated to explore sustainable teaching and learning models based on "artificial intelligence and education”. Based on the core concept of engineering professional education accreditation, the course is designed by using the PCCM (Professional Competency Correlation Matrix) method to build a model based on big data analysis, deepen the classroom teaching reform of "artificial intelligence and education”, and explore the integrated digital sustainable teaching mode of "teaching, learning, evaluation, and management”. The aim of this study is to explore the effectiveness of the teaching model based on OBE and the TSEM framework on students' sustainable development. The results show that students deepen their learning in computer science while enhancing their own learning initiative, teamwork skills, innovation skills, and awareness of sustainable development. Research shows that our teaching model plays an important role in the development of student sustainable education, enhancing student engineering practice and innovation capabilities and cultivating applied innovative talents. The efficacy of the teaching model based on the OBE concept and the TSEM framework for improving students' competence in sustainable education warrants further investigation.

Abandoned disposable masks become hot substrates for plastisphere, whether in soil, atmosphere or water.

A large number of surgical masks (SMs) to be discarded indiscriminately during the spread of COVID-19. The relationship between the changes of masks entering the environment and the succession of the microorganisms on them is not yet clear. The natural aging process of SMs in different environments (water, soil, and atmosphere) was simulated, the changes and succession of the microbial community on SMs with aging time were explored. The results showed that the SMs in water environment had the highest aging degree, followed by atmospheric environment, and SMs in soil had the lowest aging degree. The results of high-throughput sequencing demonstrated the load capacity of SMs for microorganisms, showed the important role of environment in determining microbial species on SMs. According to the relative abundance of microorganisms, it is found that compared with the water environment, the microbial community on SMs in water is dominated by rare species. While in soil, in addition to rare species, there are a lot of swinging strains on the SMs. Uncovering the ageing of SMs in the environment and its association with the colonization of microorganisms will help us understand the potential of microorganisms, especially pathogenic bacteria, to survive and migrate on SMs.

PCCT: Progressive Class-Center Triplet Loss for Imbalanced Medical Image Classification.

Imbalanced training data in medical image diagnosis is a significant challenge for diagnosing rare diseases. For this purpose, we propose a novel two-stage Progressive Class-Center Triplet (PCCT) framework to overcome the class imbalance issue. In the first stage, PCCT designs a class-balanced triplet loss to coarsely separate distributions of different classes. Triplets are sampled equally for each class at each training iteration, which alleviates the imbalanced data issue and lays solid foundation for the successive stage. In the second stage, PCCT further designs a class-center involved triplet strategy to enable a more compact distribution for each class. The positive and negative samples in each triplet are replaced by their corresponding class centers, which prompts compact class representations and benefits training stability. The idea of class-center involved loss can be extended to the pair-wise ranking loss and the quadruplet loss, which demonstrates the generalization of the proposed framework. Extensive experiments support that the PCCT framework works effectively for medical image classification with imbalanced training images. On four challenging class-imbalanced datasets (two skin datasets Skin7 and Skin 198, one chest X-ray dataset ChestXray-COVID, and one eye dataset Kaggle EyePACs), the proposed approach respectively obtains the mean F1 score 86.20, 65.20, 91.32, and 87.18 over all classes and 81.40, 63.87, 82.62, and 79.09 for rare classes, achieving state-of-the-art performance and outperforming the widely used methods for the class imbalance issue.

Antidepressants for COVID-19: A systematic review.

OBJECTIVE: To systematically examine the efficacy and safety of antidepressants for the treatment of coronavirus disease 2019 (COVID-19). METHODS: A systematic search was performed independently by two researchers based on Chinese Journal Net, WanFang, PsycINFO, Cochrane Library, PubMed, and EMBASE. RESULTS: Seven studies (n = 92,947) including three retrospective studies (n = 91,083), two randomized clinical trials (RCTs, n = 1649), two prospective cohort study (n = 215) involving (n = 92,947) patients with COVID-19 were examined. For RCTs, fluvoxamine outperformed placebo in reducing clinical deterioration and hospitalisation for COVID-19 patients. For retrospective studies, antidepressants (2 studies) and fluoxetine (1 study) possibly reduced the risk of mortality in patients with COVID-19. Results from two remaining studies supported the superiority of fluvoxamine in reducing risk of mortality in COVID-19 patients. The two RCTs that examined the safety of fluvoxamine for COVID-19 patients found inconsistent results but no significant group differences in the dropout rate. CONCLUSION: This systematic review found emerging evidence for fluvoxamine in reducing the risk of mortality and hospitalisation in COVID-19 patients, but inconsistent evidence for the safety of fluvoxamine in COVID-19 patients. More studies are needed to determine the efficacy and safety of antidepressants for the treatment of COVID-19.

Targeting the Fusion Process of SARS-CoV-2 Infection by Small Molecule Inhibitors.

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has become a serious threat to global public health, underscoring the urgency of developing effective therapies. Therapeutics and, more specifically, direct-acting antiviral development are still very much in their infancy. Here, we report that two hepatitis C virus (HCV) fusion inhibitors identified in our previous study, dichlorcyclizine and fluoxazolevir, broadly block human coronavirus entry into various cell types. Both compounds were effective against various human-pathogenic CoVs in multiple assays based on vesicular stomatitis virus (VSV) pseudotyped with the spike protein and spike-mediated syncytium formation. The antiviral effects were confirmed in SARS-CoV-2 infection systems. These compounds were equally effective against recently emerged variants, including the delta variant. Cross-linking experiments and structural modeling suggest that the compounds bind to a hydrophobic pocket near the fusion peptide of S protein, consistent with their potential mechanism of action as fusion inhibitors. In summary, these fusion inhibitors have broad-spectrum antiviral activities and may be promising leads for treatment of SARS-CoV-2, its variants, and other pathogenic CoVs. IMPORTANCE SARS-CoV-2 is an enveloped virus that requires membrane fusion for entry into host cells. Since the fusion process is relatively conserved among enveloped viruses, we tested our HCV fusion inhibitors, dichlorcyclizine and fluoxazolevir, against SARS-CoV-2. We performed in vitro assays and demonstrated their effective antiviral activity against SARS-CoV-2 and its variants. Cross-linking experiments and structural modeling suggest that the compounds bind to a hydrophobic pocket in spike protein to exert their inhibitory effect on the fusion step. These data suggest that both dichlorcyclizine and fluoxazolevir are promising candidates for further development as treatment for SARS-CoV-2.

The impact of moral leadership on physical education teachers' innovation behavior: The role of identification with leader and psychological safety.

With the growth of people's health needs and the impact of the COVID-19 pandemic, it is an inevitable trend to promote innovation behaviors of physical education (PE) teachers to innovate traditional physical education and adapt to national needs of sustainable development in the sports industry. Considering that moral leadership can promote innovation behavior of individuals through psychological factors, this study defines the types of innovation behavior, and from the perspective of psychological safety and identifying with leaders, discusses the impact of moral leadership on individuals' innovation behavior by using hierarchical multivariate regression analysis, which provides inspiration for schools to strengthen the innovation behavior of physical education teachers. In this study, 327 questionnaires were distributed to PE teachers in Chinese provinces and 287 valid questionnaires were collected. The analysis of the collected data was performed with the help of the SPSSAU data analysis platform. The following conclusions were drawn: First, moral leadership has a significant positive impact on the psychological safety and internal and external innovation of physical education teachers. Secondly, moral leadership influences employees' innovation behavior through psychological safety, and plays a part of intermediary role between moral leadership and internal and external innovation behavior; Third, by comparing the two impact mechanisms of innovation behavior, we found that moral leadership encourages employees to produce more external innovation behavior through psychological safety; Finally, strong leadership identity plays a positive role in regulating the relationship between moral leadership and innovation behavior.

Maternal and infant outcomes in women with and without gestational diabetes mellitus in the COVID-19 era in China: Lessons learned

Aims The global COVID-19 pandemic has required a drastic transformation of prenatal care services. Whether the reformulation of the antenatal care systems affects maternal and infant outcomes remains unknown. Particularly, women with gestational diabetes mellitus (GDM) are among those who bear the greatest brunt. Thus, this study aimed to evaluate the impact of COVID-19 lockdown during late pregnancy on maternal and infant outcomes in women stratified by the GDM status in China. Study design The participants were women who experienced the COVID-19 lockdown during late pregnancy (3185 in the 2020 cohort) or not (2540 in the 2019 cohort) that were derived from the Beijing Birth Cohort Study. Maternal metabolic indicators, neonatal outcomes, and infant anthropometrics at 12 months of age were compared between the two cohorts, stratified by the GDM status. Results Participants who experienced COVID-19 lockdown in late pregnancy showed lower gestational weight gain than those in the control cohort. Nevertheless, they displayed a worse metabolic profile. COVID-19 lockdown during pregnancy was associated with higher glycosylated hemoglobin (HbA1c) (β= 0.11, 95% CI = 0.05–0.16, q-value = 0.002) and lower high density lipoprotein cholesterol level (HDL-C) level (β=–0.09, 95% CI = –0.14 to –0.04, q-value = 0.004) in women with GDM, adjusted for potential confounders. In normoglycemic women, COVID-19 lockdown in late pregnancy was associated with higher fasting glucose level (β= 0.10, 95% CI = 0.08–0.12, q-value <0.0001), lower HDL-C level (β=–0.07, 95% CI = –0.08 to –0.04, q-value <0.0001), and increased risk of pregnancy-induced hypertension (adjusted OR=1.80, 95%CI=1.30–2.50, q-value=0.001). The fasting glucose level decreased less from early to late pregnancy in women who experienced COVID-19 lockdown than in the controls, regardless of the GDM status. The HDL-C has risen less with COVID-19 lockdown in the normoglycemic subgroup. In contrast, no significant differences regarding neonatal outcomes or infant weight were found between the two cohorts. Conclusion Experiencing the COVID-19 lockdown in pregnancy was associated with worse maternal metabolic status but similar neonatal outcomes and infant weight.

The SARS-CoV-2 spike protein binds and modulates estrogen receptors.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) protein binds angiotensin-converting enzyme 2 as its primary infection mechanism. Interactions between S and endogenous proteins occur after infection but are not well understood. We profiled binding of S against >9000 human proteins and found an interaction between S and human estrogen receptor α (ERα). Using bioinformatics, supercomputing, and experimental assays, we identified a highly conserved and functional nuclear receptor coregulator (NRC) LXD-like motif on the S2 subunit. In cultured cells, S DNA transfection increased ERα cytoplasmic accumulation, and S treatment induced ER-dependent biological effects. Non-invasive imaging in SARS-CoV-2-infected hamsters localized lung pathology with increased ERα lung levels. Postmortem lung experiments from infected hamsters and humans confirmed an increase in cytoplasmic ERα and its colocalization with S in alveolar macrophages. These findings describe the discovery of a S-ERα interaction, imply a role for S as an NRC, and advance knowledge of SARS-CoV-2 biology and coronavirus disease 2019 pathology.

Preclinical Pharmacokinetics and In Vitro Properties of GS-441524, a Potential Oral Drug Candidate for COVID-19 Treatment.

Preclinical pharmacokinetics (PK) and In Vitro ADME properties of GS-441524, a potential oral agent for the treatment of Covid-19, were studied. GS-441524 was stable in vitro in liver microsomes, cytosols, and hepatocytes of mice, rats, monkeys, dogs, and humans. The plasma free fractions of GS-441524 were 62-78% across all studied species. The in vitro transporter study results showed that GS-441524 was a substrate of MDR1, BCRP, CNT3, ENT1, and ENT2; but not a substrate of CNT1, CNT2, and ENT4. GS-441524 had a low to moderate plasma clearance (CLp), ranging from 4.1 mL/min/kg in dogs to 26 mL/min/kg in mice; the steady state volume distribution (Vdss) ranged from 0.9 L/kg in dogs to 2.4 L/kg in mice after IV administration. Urinary excretion appeared to be the major elimination process for GS-441524. Following oral administration, the oral bioavailability was 8.3% in monkeys, 33% in rats, 39% in mice, and 85% in dogs. The PK and ADME properties of GS-441524 support its further development as an oral drug candidate.

Chlorination of isothiazolinone biocides: kinetics, reactive species, pathway, and toxicity evolution.

Due to the Covid-19 pandemic, the worldwide biocides application has been increased, which will eventually result in enhanced residuals in treated wastewater. At the same time, chlorine disinfection of secondary effluents and hospital wastewaters has been intensified. With respect to predicted elevated exposure in wastewater, the chlorination kinetics, transformation pathways and toxicity evolution were investigated in this study for two typical isothiazolinone biocides, methyl-isothiazolinone (MIT) and chloro-methyl-isothiazolinone (CMIT). Second-order rate constants of 0.13 M-1·s-1, 1.95 × 105 M-1·s-1 and 5.14 × 105 M-1·s-1 were determined for the reaction of MIT with HOCl, Cl2O and Cl2, respectively, while reactivity of CMIT was around 1-2 orders of magnitude lower. While chlorination of isothiazolinone biocides at pH 7.1 was dominated by Cl2O-oxidation, acidic pH and elevated Cl- concentration favored free active chlorine (FAC) speciation into Cl2 and increased overall isothiazolinone removal. Regardless of the dominant FAC species, the elimination of MIT and CMIT resulted in an immediate loss of acute toxicity under all experimental conditions, which was attributed to a preferential attack at the S-atom resulting in subsequent formation of sulfoxides and sulfones and eventually an S-elimination. However, chlorination of isothiazolinone biocides in secondary effluent only achieved <10% elimination at typical disinfection chlorine exposure 200 mg·L-1·min, but was predicted to be remarkably increased by acidizing solution to pH 5.5. Alternative measures might be needed to minimize the discharge of these toxic chemicals into the aquatic environment.

A humanized nanobody phage display library yields potent binders of SARS CoV-2 spike.

Neutralizing antibodies targeting the SARS-CoV-2 spike protein have shown a great preventative/therapeutic potential. Here, we report a rapid and efficient strategy for the development and design of SARS-CoV-2 neutralizing humanized nanobody constructs with sub-nanomolar affinities and nanomolar potencies. CryoEM-based structural analysis of the nanobodies in complex with spike revealed two distinct binding modes. The most potent nanobody, RBD-1-2G(NCATS-BL8125), tolerates the N501Y RBD mutation and remains capable of neutralizing the B.1.1.7 (Alpha) variant. Molecular dynamics simulations provide a structural basis for understanding the neutralization process of nanobodies exclusively focused on the spike-ACE2 interface with and without the N501Y mutation on RBD. A primary human airway air-lung interface (ALI) ex vivo model showed that RBD-1-2G-Fc antibody treatment was effective at reducing viral burden following WA1 and B.1.1.7 SARS-CoV-2 infections. Therefore, this presented strategy will serve as a tool to mitigate the threat of emerging SARS-CoV-2 variants.

Synthesis and anti-SARS-CoV-2 activity of deuterated GS-441524 analogs.

The COVID-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is continuing to spread around the world. GS-441524 is the parent nucleoside of remdesivir which is the first drug approved for the treatment of COVID-19, and demonstrates strong activity against SARS-Cov-2 in vitro and in vivo. Herein, we reported the synthesis of a series of deuterated GS-441524 analogs, which had deuterium atoms up to five at the ribose and the nucleobase moieties. Compared to GS-441524, all the deuterated compounds showed similar inhibitory activities against SARS-CoV-2 in vitro.

Progressive assembly of multi-domain protein structures from cryo-EM density maps

Progress in cryo-electron microscopy has provided the potential for large-size protein structure determination. However, the success rate for solving multi-domain proteins remains low because of the difficulty in modelling inter-domain orientations. Here we developed domain enhanced modeling using cryo-electron microscopy (DEMO-EM), an automatic method to assemble multi-domain structures from cryo-electron microscopy maps through a progressive structural refinement procedure combining rigid-body domain fitting and flexible assembly simulations with deep-neural-network inter-domain distance profiles. The method was tested on a large-scale benchmark set of proteins containing up to 12 continuous and discontinuous domains with medium- to low-resolution density maps, where DEMO-EM produced models with correct inter-domain orientations (template modeling score (TM-score) >0.5) for 97% of cases and outperformed state-of-the-art methods. DEMO-EM was applied to the severe acute respiratory syndrome coronavirus 2 genome and generated models with average TM-score and root-mean-square deviation of 0.97 and 1.3 Å, respectively, with respect to the deposited structures. These results demonstrate an efficient pipeline that enables automated and reliable large-scale multi-domain protein structure modelling from cryo-electron microscopy maps.

Atypical pneumonia caused by Chlamydia psittaci during the COVID-19 pandemic.

OBJECTIVES: Here, we retrospectively described the diagnosis and treatment of 32 cases diagnosed with Chlamydia psittaci pneumonia during the COVID-19 pandemic. METHODS: Clinical information was collected from all the patients. Reverse transcription-PCR and ELISAs were conducted for the detection of COVID-19 using nasal swabs and bronchoalveolar lavage fluid (BALF) samples. Metagenomic next-generation sequencing (mNGS) was performed for the identification of causative pathogens using BALF, peripheral blood and sputum samples. End-point PCR was performed to confirm the mNGS results. RESULTS: All 32 patients showed atypical pneumonia and had infection-like symptoms that were similar to COVID-19. Results of reverse transcription-PCR and ELISAs ruled out COVID-19 infection. mNGS identified C. psittaci as the suspected pathogen in these patients within 48 hours, which was validated by PCR, except for three blood samples. The sequence reads that covered fragments of C. psittaci genome were detected more often in BALF than in sputum or blood samples. All patients received doxycycline-based treatment regimens and showed favorable outcomes. CONCLUSION: This retrospective study, with the highest number of C. psittaci pneumonia enrolled cases in China so far, suggests that human psittacosis may be underdiagnosed and misdiagnosed clinically, especially in the midst of the COVID-19 pandemic.

Suite of TMPRSS2 Assays for Screening Drug Repurposing Candidates as Potential Treatments of COVID-19.

SARS-CoV-2 is the causative viral pathogen driving the COVID-19 pandemic that prompted an immediate global response to the development of vaccines and antiviral therapeutics. For antiviral therapeutics, drug repurposing allows for rapid movement of the existing clinical candidates and therapies into human clinical trials to be tested as COVID-19 therapies. One effective antiviral treatment strategy used early in symptom onset is to prevent viral entry. SARS-CoV-2 enters ACE2-expressing cells when the receptor-binding domain of the spike protein on the surface of SARS-CoV-2 binds to ACE2 followed by cleavage at two cut sites by TMPRSS2. Therefore, a molecule capable of inhibiting the protease activity of TMPRSS2 could be a valuable antiviral therapy. Initially, we used a fluorogenic high-throughput screening assay for the biochemical screening of 6030 compounds in NCATS annotated libraries. Then, we developed an orthogonal biochemical assay that uses mass spectrometry detection of product formation to ensure that hits from the primary screen are not assay artifacts from the fluorescent detection of product formation. Finally, we assessed the hits from the biochemical screening in a cell-based SARS-CoV-2 pseudotyped particle entry assay. Of the six molecules advanced for further studies, two are approved drugs in Japan (camostat and nafamostat), two have entered clinical trials (PCI-27483 and otamixaban), while the other two molecules are peptidomimetic inhibitors of TMPRSS2 taken from the literature that have not advanced into clinical trials (compounds 92 and 114). This work demonstrates a suite of assays for the discovery and development of new inhibitors of TMPRSS2.

Study of optical-based speech acquisition system using vibration signals from speakers' medical masks.

Medical masks have become necessary of late because of the COVID-19 outbreak; however, they tend to attenuate the energy of speech signals and affect speech quality. Therefore, this study proposes an optical-based microphone approach to obtain speech signals from speakers' medical masks. Experimental results showed that the optical-based microphone approach achieved better performance (85.61%) than the two baseline approaches, namely, omnidirectional (24.17%) and directional microphones (31.65%), in the case of long-distance speech and background noise. The results suggest that the optical-based microphone method is a promising approach for acquiring speech from a medical mask.

Mitoxantrone modulates a heparan sulfate-spike complex to inhibit SARS-CoV-2 infection.

Spike-mediated entry of SARS-CoV-2 into human airway epithelial cells is an attractive therapeutic target for COVID-19. In addition to protein receptors, the SARS-CoV-2 spike (S) protein also interacts with heparan sulfate, a negatively charged glycosaminoglycan (GAG) attached to certain membrane proteins on the cell surface. This interaction facilitates the engagement of spike with a downstream receptor to promote viral entry. Here, we show that Mitoxantrone, an FDA-approved topoisomerase inhibitor, targets a heparan sulfate-spike complex to compromise the fusogenic function of spike in viral entry. As a single agent, Mitoxantrone inhibits the infection of an authentic SARS-CoV-2 strain in a cell-based model and in human lung EpiAirway 3D tissues. Gene expression profiling supports the plasma membrane as a major target of Mitoxantrone but also underscores an undesired activity targeting nucleosome dynamics. We propose that Mitoxantrone analogs bearing similar heparan sulfate-binding activities but with reduced affinity for DNA topoisomerases may offer an alternative therapy to overcome breakthrough infections in the post-vaccine era.

Graph Convolutional Network-Based Screening Strategy for Rapid Identification of SARS-CoV-2 Cell-Entry Inhibitors.

The cell entry of SARS-CoV-2 has emerged as an attractive drug development target. We previously reported that the entry of SARS-CoV-2 depends on the cell surface heparan sulfate proteoglycan (HSPG) and the cortex actin, which can be targeted by therapeutic agents identified by conventional drug repurposing screens. However, this drug identification strategy requires laborious library screening, which is time consuming, and often limited number of compounds can be screened. As an alternative approach, we developed and trained a graph convolutional network (GCN)-based classification model using information extracted from experimentally identified HSPG and actin inhibitors. This method allowed us to virtually screen 170,000 compounds, resulting in ∼2000 potential hits. A hit confirmation assay with the uptake of a fluorescently labeled HSPG cargo further shortlisted 256 active compounds. Among them, 16 compounds had modest to strong inhibitory activities against the entry of SARS-CoV-2 pseudotyped particles into Vero E6 cells. These results establish a GCN-based virtual screen workflow for rapid identification of new small molecule inhibitors against validated drug targets.

Repurposing drugs as COVID-19 therapies: A toxicity evaluation.

Drug repurposing is an appealing method to address the Coronavirus 2019 (COVID-19) pandemic because of the low cost and efficiency. We analyzed our in-house database of approved drug screens and compared their activity profiles with results from a severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) cytopathic effect (CPE) assay. The activity profiles of the human ether-à-go-go-related gene (hERG), phospholipidosis (PLD), and many cytotoxicity screens were found significantly correlated with anti-SARS-CoV-2 activity. hERG inhibition is a nonspecific off-target effect that has contributed to promiscuous drug interactions, whereas drug-induced PLD is an undesirable effect linked to hERG blockers. Thus, this study identifies preferred drug candidates as well as chemical structures that should be avoided because of their potential to induce toxicity. Lastly, we highlight the hERG liability of anti-SARS-CoV-2 drugs currently enrolled in clinical trials.