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SARS-CoV-2, the causative agent of COVID-19 1 , features a receptor-binding domain (RBD) for binding to the host cell ACE2 protein 1 – 6 . Neutralizing antibodies that block RBD-ACE2 interaction are candidates for the development of targeted therapeutics 7 – 17 . Llama-derived single-domain antibodies (nanobodies, ~15 kDa) offer advantages in bioavailability, amenability, and production and storage owing to their small sizes and high stability. Here, we report the rapid selection of 99 synthetic nanobodies (sybodies) against RBD by in vitro selection using three libraries. The best sybody, MR3 binds to RBD with high affinity ( K D  = 1.0 nM) and displays high neutralization activity against SARS-CoV-2 pseudoviruses (IC 50  = 0.42 μg mL −1 ). Structural, biochemical, and biological characterization suggests a common neutralizing mechanism, in which the RBD-ACE2 interaction is competitively inhibited by sybodies. Various forms of sybodies with improved potency have been generated by structure-based design, biparatopic construction, and divalent engineering. Two divalent forms of MR3 protect hamsters from clinical signs after live virus challenge and a single dose of the Fc-fusion construct of MR3 reduces viral RNA load by 6 Log 10 . Our results pave the way for the development of therapeutic nanobodies against COVID-19 and present a strategy for rapid development of targeted medical interventions during an outbreak. Here, the authors report the engineering, structural and biological characterization of synthetic nanobodies (sybodies) that display potent therapeutic activity against SARS-CoV-2 infection in animal models via targeting the virus receptor-binding domain.

The COVID pandemic fueled by emerging SARS-CoV-2 new variants of concern remains a major global health concern, and the constantly emerging mutations present challenges to current therapeutics. The spike glycoprotein is not only essential for the initial viral entry, but is also responsible for the transmission of SARS-CoV-2 components via syncytia formation. Spike-mediated cell-cell transmission is strongly resistant to extracellular therapeutic and convalescent antibodies via an unknown mechanism. Here, we describe the antibody-mediated spike activation and syncytia formation on cells displaying the viral spike. We found that soluble antibodies against receptor binding motif (RBM) are capable of inducing the proteolytic processing of spike at both the S1/S2 and S2’ cleavage sites, hence triggering ACE2-independent cell-cell fusion. Mechanistically, antibody-induced cell-cell fusion requires the shedding of S1 and exposure of the fusion peptide at the cell surface. By inhibiting S1/S2 proteolysis, we demonstrated that cell-cell fusion mediated by spike can be re-sensitized towards antibody neutralization in vitro . Lastly, we showed that cytopathic effect mediated by authentic SARS-CoV-2 infection remain unaffected by the addition of extracellular neutralization antibodies. Hence, these results unveil a novel mode of antibody evasion and provide insights for antibody selection and drug design strategies targeting the SARS-CoV-2 infected cells.

An essential step for SARS‐CoV‐2 infection is the attachment to the host cell receptor by its Spike receptor‐binding domain (RBD). Most of the existing RBD‐targeting neutralizing antibodies block the receptor‐binding motif (RBM), a mutable region with the potential to generate neutralization escape mutants. Here, we isolated and structurally characterized a non‐RBM‐targeting monoclonal antibody (FD20) from convalescent patients. FD20 engages the RBD at an epitope distal to the RBM with a K D of 5.6 nM, neutralizes SARS‐CoV‐2 including the current Variants of Concern such as B.1.1.7, B.1.351, P.1, and B.1.617.2 (Delta), displays modest cross‐reactivity against SARS‐CoV, and reduces viral replication in hamsters. The epitope coincides with a predicted “ideal” vulnerability site with high functional and structural constraints. Mutation of the residues of the conserved epitope variably affects FD20‐binding but confers little or no resistance to neutralization. Finally, in vitro mode‐of‐action characterization and negative‐stain electron microscopy suggest a neutralization mechanism by which FD20 destructs the Spike. Our results reveal a conserved vulnerability site in the SARS‐CoV‐2 Spike for the development of potential antiviral drugs. SYNOPSIS A monoclonal antibody (FD20) from convalescent COVID‐19 patients has been isolated and structurally and biologically characterized. Various SARS‐CoV‐2 strains, including the Alpha, Beta, Gamma, and Delta variants, and naturally occurring epitope mutants, can be neutralized by FD20 with similar potency. A broadly active mAb is identified with consistent neutralizing activity against 14 SARS‐CoV‐2 strains/mutants and weak activity against SARS‐CoV. The conservation of FD20's epitope residues is supported by their low mutation frequencies both in nature and in laboratory experiments. A neutralizing mechanism through which the surface glycoprotein is destructed by FD20 is proposed based on electron microscopy evidence. Graphical Abstract A monoclonal antibody (FD20) from convalescent COVID‐19 patients has been isolated and structurally and biologically characterized. Various SARS‐CoV‐2 strains, including the Alpha, Beta, Gamma, and Delta variants, and naturally occurring epitope mutants, can be neutralized by FD20 with similar potency.

Selaginella moellendorffii is a lycophyte, a member of an ancient vascular plant lineage. Two distinct types of terpene synthase (TPS) genes were identified from this species, including S. moellendorffii TPS genes (SmTPSs) and S. moellendorffii microbial TPS-like genes (SmMTPSLs). The goal of this study was to investigate the biochemical functions of SmMTPSLs. Here, eight full-length SmMTPSL genes (SmMTPSL5, -15, -19, -23, -33, -37, -46, and -47) were functionally characterized from S. moellendorffii. Escherichia coli-expressed recombinant SmMTPSLs were tested for monoterpenes synthase and sesquiterpenes synthase activities. These enzymatic products were typical monoterpenes and sesquiterpenes that have been previous shown to be generated by typical plant TPSs when provided with geranyl diphosphate (GPP) and farnesyl diphosphate (FPP) as the substrates. Meanwhile, SmMTPSL23, -33, and -37 were up-regulated when induced by alamethicin (ALA) and methyl jasmonate (MeJA), suggesting a role for these genes in plants response to abiotic stresses. Furthermore, this study pointed out that the terpenoids products of SmMTPSL23, -33, and -37 have an antibacterial effect on Pseudomonas syringae pv. tomato DC3000 and Staphylococcus aureus. Taken together, these results provide more information about the catalytic and biochemical function of SmMTPSLs in S. moellendorffii plants.

The stability of mold oscillation can directly influence on slab surface quality and operational safety in continuous casting. In recent years, the hydraulic oscillation is developed and applied as the driven equipment of mold oscillation. In the present work, based on the slab continuous caster of hydraulic oscillations, the displacement and other parameters of hydraulic oscillator are measured and the evaluation method of oscillation is studied. The displacement difference and phase difference of oscillation with sinusoidal waveform and non-sinusoidal waveform are analyzed. Especially, the dynamic characteristics of the driving force of left and right cylinders are evaluated. The results indicated that the hydraulic oscillator is quite a good device in terms of precision. The method proposed may be useful for evaluating oscillators.

ABSTRACT SARS-CoV-2, the causative agent of COVID-191, recognizes host cells by attaching its receptor-binding domain (RBD) to the host receptor ACE22–7. Neutralizing antibodies that block RBD-ACE2 interaction have been a major focus for therapeutic development8–18. Llama-derived single-domain antibodies (nanobodies, ∼15 kDa) offer advantages including ease of production and possibility for direct delivery to the lungs by nebulization19, which are attractive features for bio-drugs against the global respiratory disease. Here, we generated 99 synthetic nanobodies (sybodies) by in vitro selection using three libraries. The best sybody, MR3 bound to RBD with high affinity (KD = 1.0 nM) and showed high neutralization activity against SARS-CoV-2 pseudoviruses (IC50 = 0.40 μg mL−1). Structural, biochemical, and biological characterization of sybodies suggest a common neutralizing mechanism, in which the RBD-ACE2 interaction is competitively inhibited by sybodies. Various forms of sybodies with improved potency were generated by structure-based design, biparatopic construction, and divalent engineering. Among these, a divalent MR3 conjugated with the albumin-binding domain for prolonged half-life displayed highest potency (IC50 = 12 ng mL−1) and protected mice from live SARS-CoV-2 challenge. Our results pave the way to the development of therapeutic nanobodies against COVID-19 and present a strategy for rapid responses for future outbreaks. Competing Interest Statement The authors have declared no competing interest. Footnotes * In this revision, we include data to show that the engineered sybody MR3 can protect mice from live SARS-CoV-2 infection.

Natural antibacterial agents have attracted increasing attention due to safety, ecological, and environmental protection concerns. In this study, hinokitiol-grafted-chitosan (HTCS) was prepared via the Mannich reaction and used as an antibacterial agent for the treatment of cotton fabric. The results showed that, compared with chitosan (CS) and hinokitiol (HT) alone, HTCS exhibited a lower value of minimum inhibitory concentration (MIC). Compared with the fabrics treated by CS and HT, the antibacterial rate of the treated fabric against Escherichia coli using HTCS as antibacterial agent increased by approximately 90% and 27%, the bacterial reduction rate against Staphylococcus aureus increased by about 58% and 39%. The HTCS-treated cotton fabric possessed good antibacterial properties even after 25 washing cycles. Moreover, the antibacterial cotton fabric retained original hydrophilicity, handle, and strength. Consequently, HTCS has great potential as a natural antibacterial agent for textiles.

Chemotherapy resistance is reported to correlate with up-regulation of anti-tumor agent transporter ABCB1 (p-gp) in epithelial ovarian cancer (EOC), but the results remain controversial. To reconcile the results, a systematic review followed by meta-analysis was performed to assess the association between high ABCB1 status or ABCB1 gene variants and overall survival (OS), progression free survival (PFS), and total response rate (TR) in patients with EOC. Electronic searches were performed using Pubmed, EMBASE, Web of Science and Chinese Wanfang databases from January 1990 to February 2016. Summary hazard ratio (HR), risk ratio (RR) and 95% confidence intervals (CIs) were combined using fixed or random-effects models as appropriate. Thirty-eight retrospective studies of 8607 cases qualified for meta-analysis were identified. Our results suggested that ABCB1 over-expression was significantly associated with unfavorable OS (HR = 1.54; 95% CI, 1.25-1.90), PFS (HR = 1.49; 95% CI, 1.22-1.82) and TR (RR = 0.63; 95% CI, 0.54-0.75). After adjustment for age, clinical stage, residual disease, histological type and tumor grade, high ABCB1 status remained to be a significant risk factor for adverse OS and PFS. Patients with recurrent ABCB1 positivity suffered from poorer OS than those with primary ABCB1 positivity. However, stratified by chemotherapy regimen, inverse correlation between high ABCB1 status and poor OS, PFS and TR were only found in patients underwent platinum-based chemotherapy but not in patients received standard platinum/paclitaxel-based chemotherapy. No evidence was found for any association between ABCB1 gene polymorphisms and OS, PFS or TR. High ABCB1 status is significantly associated with chemo-resistance and poor prognosis in patients with EOC. Large-scale, prospective studies are needed to assess the clinical value of ABCB1 expression in EOC more accurately.

To protect the forest ecological environment, China implemented the Comprehensive Commercial Logging Ban Policy (CCLBP), yet it has a major impact on forest residents. Therefore, this study aimed to evaluate the CCLBP from the perspective of the satisfaction of residents. In this study, we used 242 questionnaires from residents in national forest areas in Northeast China and Inner Mongolia to evaluate satisfaction and its influencing factors by factor analysis combined with the entropy method and multiple linear regression, respectively. The results show that: (1) the overall policy satisfaction of residents in national forest areas is 60.9, which is lower than the theoretical neutral value of policy satisfaction. Increasing employment opportunities, transferring surplus employees and developing forest economy are important to improve the satisfaction of residents. (2) Life satisfaction was higher in the high-income group than in the low-income group. Increasing wage income can improve the satisfaction of residents in national forest areas. (3) The satisfaction of the worker group was significantly higher than that of the forest farmer group. Improving job stability and policy equity are important to promote life satisfaction of residents in national forest areas. (4) The satisfaction of the Yichun Forest Industry Group was significantly higher than the Inner Mongolia Forest Group. Natural resource endowment and adaptability to the CCLBP are the main factors affecting the satisfaction of different forest industry groups. At the same time, focusing on the institutional supply of external support policies in order to provide them access to information, employment advice and other services is very significant. This research provides a new approach to studying the CCLBP, which is of great practical significance for raising the wellbeing index of national forest areas.