Antigenic cartography using hamster sera identifies SARS-CoV-2 JN.1 evasion seen in human XBB.1.5 booster sera (preprint)

Antigenic assessments of SARS-CoV-2 variants inform decisions to update COVID-19 vaccines. Primary infection sera are often used for assessments, but such sera are rare due to population immunity from SARS-CoV-2 infections and COVID-19 vaccinations. Here, we show that neutralization titers and breadth of matched human and hamster pre-Omicron variant primary infection sera correlate well and generate similar antigenic maps. The hamster antigenic map shows modest antigenic drift among XBB sub-lineage variants, with JN.1 and BA.4/BA.5 variants within the XBB cluster, but with five to six-fold antigenic differences between these variants and XBB.1.5. Compared to sera following only ancestral or bivalent COVID-19 vaccinations, or with post-vaccination infections, XBB.1.5 booster sera had the broadest neutralization against XBB sub-lineage variants, although a five-fold titer difference was still observed between JN.1 and XBB.1.5 variants. These findings suggest that antibody coverage of antigenically divergent JN.1 could be improved with a matched vaccine antigen.

The substrate selectivity of papain-like proteases from human-infecting coronaviruses correlates with innate immune suppression.

Coronaviruses that can infect humans can cause either common colds (HCoV-NL63, HCoV-229E, HCoV-HKU1, and HCoV-OC43) or severe respiratory symptoms (SARS-CoV-2, SARS-CoV, and MERS-CoV). The papain-like proteases (PLPs) of SARS-CoV, SARS-CoV-2, MERS-CoV, and HCoV-NL63 function in viral innate immune evasion and have deubiquitinating (DUB) and deISGylating activities. We identified the PLPs of HCoV-229E, HCoV-HKU1, and HCoV-OC43 and found that their enzymatic properties correlated with their ability to suppress innate immune responses. A conserved noncatalytic aspartic acid residue was critical for both DUB and deISGylating activities, but the PLPs had differing ubiquitin (Ub) chain cleavage selectivities and binding affinities for Ub, K48-linked diUb, and interferon-stimulated gene 15 (ISG15) substrates. The crystal structure of HKU1-PLP2 in complex with Ub revealed binding interfaces that accounted for the unusually high binding affinity between this PLP and Ub. In cellular assays, the PLPs from the severe disease-causing coronaviruses strongly suppressed innate immune IFN-I and NF-κB signaling and stimulated autophagy, whereas the PLPs from the mild disease-causing coronaviruses generally showed weaker effects on immune suppression and autophagy induction. In addition, a PLP from a SARS-CoV-2 variant of concern showed increased suppression of innate immune signaling pathways. Overall, these results demonstrated that the DUB and deISGylating activities and substrate selectivities of these PLPs differentially contribute to viral innate immune evasion and may affect viral pathogenicity.

Efficacy of high-flow nasal oxygenation compared with laryngeal mask airway in children undergoing ambulatory oral surgery under deep sedation: A randomized controlled non-inferiority trial.

Background: High-flow nasal oxygenation (HFNO) has been suggested as an alternative oxygenation method during procedural sedation. This randomized, non-inferiority trial evaluated the safety and efficacy of HFNO compared with laryngeal mask airway (LMA) in pediatric ambulatory oral surgery under deep sedation. Methods: In total, 120 children aged 2-7 years (weight: 10-30 kg) were equally assigned into two groups, namely, HFNO with propofol total intravenous anesthesia infusion (HFNO-IV) or LMA with propofol total intravenous anesthesia infusion (LMA-IV). The primary objective was to monitor carbon dioxide (CO2) accumulation during perioperative surgery. Secondary objectives included monitoring transcutaneous oxygen saturation, grade exposure to the surgical field, perioperative adverse events, or other events. The predefined non-inferiority margin was 7 mmHg. During the COVID-19 pandemic, a novel WeChat applet was implemented to gather follow-up data after discharge. Results: Non-inferiority could be declared for HFNO relative to LMA (mean difference in transcutaneous CO2 (TcCO2) = -1.4 mmHg, 95% CI: -2.9, 0.1 mmHg; P > 0.05). The pre-surgical TcCO2 of the HFNO-IV group (45.4 ± 4.5 mmHg) was similar to that of the LMA-IV group (44.0 ± 3.5 mmHg), within the clinically acceptable normal range. All the children maintained SpO2 levels of >97%. The surgical field exposure score of the HFNO group was significantly better than that of the LMA group. There was no significant difference between the two groups regarding risk or adverse events. Conclusion: HFNO was not inferior to LMA for maintaining oxygenation and ventilation in patients undergoing pediatric ambulatory oral surgery under deep sedation under strict isolation from the oral cavity to the upper airway.

Experience of using a smartphone WeChat applet for dental anxiety assessment and preoperative evaluation: A nationwide multicenter study

Introduction Dental anxiety is a multivariate phenomenon that regularly occurs during a dental procedure. Although it may lead to patients' safety concerns and adverse events in routine treatment, it is often ignored. The purpose of this research is to develop a novel WeChat Applet for dental anxiety (WADA) with the following features and aims: (1) to help patients with dental anxiety management;(2) provide patient with a physical status self-evaluation;and(3) provide a platform for online assessment and tele-consultancy by dentists. We aimed to test and verify whether such an applet could play a beneficial role before and after a dental procedure and facilitate management of high-risk patients during the COVID-19 pandemic. Materials and methods During the 12-month survey period (August 2020 to July 2021), a total of 180 patients aged 3–74 years from eight different cities (n = 180 at the end of treatment, n = 25 for the System Usability Scale (SUS) and follow-up interview) and 20 medical staff from eight different cities (n =20 for follow-up interview) were evaluated by WADA. At the end of the survey period, the results of the interviews were analyzed thematically. Results WADA assessment results from 180 patients and follow-up interview results from 45 participants were analyzed. In this study with a male to female ratio of 2:3, 75% were found to be suffering from dental anxiety, 86% were found with postoperative complications, and 11 cases were found to have contraindications to surgery. The total SUS score for WADA is 72.25 above the mean score, proving that WADA is a relevant and useful tool before and after dental treatment. Based on the results of the interviews, the following themes were identified: patient satisfaction;dentists' effectiveness;multi-center data integration;and increase its frequency of usage. Conclusions The WADA was developed for dental procedures and is effective for reducing treatment risks, improving patients' satisfaction and dentists' convenience, especially in terms of facilitating management of high-risk patient during the COVID-19 pandemic.

Broadly neutralizing antibodies target the coronavirus fusion peptide (preprint)

The potential for future coronavirus outbreaks highlights the need to develop strategies and tools to broadly target this group of pathogens. Here, using an epitope-agnostic approach, we identified six monoclonal antibodies that bound to spike proteins from all seven human-infecting coronaviruses. Epitope mapping revealed that all six antibodies target the conserved fusion peptide region adjacent to the S2' cleavage site. Two antibodies, COV44-62 and COV44-79, broadly neutralize a range of alpha and beta coronaviruses, including SARS-CoV-2 Omicron subvariants BA.1 and BA.2, albeit with lower potency than RBD-specific antibodies. In crystal structures of Fabs COV44-62 and COV44-79 with the SARS-CoV-2 fusion peptide, the fusion peptide epitope adopts a helical structure and includes the arginine at the S2' cleavage site. Importantly, COV44-79 limited disease caused by SARS-CoV-2 in a Syrian hamster model. These findings identify the fusion peptide as the target of the broadest neutralizing antibodies in an epitope-agnostic screen, highlighting this site as a candidate for next-generation coronavirus vaccine development.

Ultrapotent bispecific antibodies neutralize emerging SARS-CoV-2 variants (preprint)

The emergence of SARS-CoV-2 variants that threaten the efficacy of existing vaccines and therapeutic antibodies underscores the urgent need for new antibody-based tools that potently neutralize variants by targeting multiple sites of the spike protein. We isolated 216 monoclonal antibodies targeting SARS-CoV-2 from plasmablasts and memory B cells of COVID-19 patients. The three most potent antibodies targeted distinct regions of the RBD, and all three neutralized the SARS-CoV-2 variants B.1.1.7 and B.1.351. The crystal structure of the most potent antibody, CV503, revealed that it binds to the ridge region of SARS-CoV-2 RBD, competes with the ACE2 receptor, and has limited contact with key variant residues K417, E484 and N501. We designed bispecific antibodies by combining non-overlapping specificities and identified five ultrapotent bispecific antibodies that inhibit authentic SARS-CoV-2 infection at concentrations of <1 ng/mL. Through a novel mode of action three bispecific antibodies cross-linked adjacent spike proteins using dual NTD/RBD specificities. One bispecific antibody was >100-fold more potent than a cocktail of its parent monoclonals in vitro and prevented clinical disease in a hamster model at a 2.5 mg/kg dose. Notably, six of nine bispecific antibodies neutralized B.1.1.7, B.1.351 and the wild-type virus with comparable potency, despite partial or complete loss of activity of at least one parent monoclonal antibody against B.1.351. Furthermore, a bispecific antibody that neutralized B.1.351 protected against SARS-CoV-2 expressing the crucial E484K mutation in the hamster model. Thus, bispecific antibodies represent a promising next-generation countermeasure against SARS-CoV-2 variants of concern.

UL36 Encoded by Marek's Disease Virus Exhibits Linkage-Specific Deubiquitinase Activity.

(1) Background: Deubiquitinase (DUB) regulates various important cellular processes via reversing the protein ubiquitination. The N-terminal fragment of a giant tegument protein, UL36, encoded by the Marek's disease (MD) virus (MDV), encompasses a putative DUB (UL36-DUB) and shares no homology with any known DUBs. The N-terminus 75 kDa fragment of UL36 exists in MD T lymphoma cells at a high level and participates in MDV pathogenicity. (2) Methods: To characterize deubiquitinating activity and substrate specificity of UL36-DUB, the UL36 N-terminal fragments, UL36(323), UL36(480), and mutants were prepared using the Bac-to-Bac system. The deubiquitinating activity and substrate specificity of these recombinant UL36-DUBs were analyzed using various ubiquitin (Ub) or ubiquitin-like (UbL) substrates and activity-based deubiquitinating enzyme probes. (3) Results: The results indicated that wild type UL36-DUBs show a different hydrolysis ability against varied types of ubiquitin chains. These wild type UL36-DUBs presented the highest activity to K11, K48, and K63 linkage Ub chains, weak activity to K6, K29, and K33 Ub chains, and no activity to K27 linkage Ub chain. UL36 has higher cleavage efficiency for K48 and K63 poly-ubiquitin than linear ubiquitin chain (M1-Ub4), but no activity on various ubiquitin-like modifiers. The mutation of C98 and H234 residues eliminated the deubiquitinating activity of UL36-DUB. D232A mutation impacted, but did not eliminated UL36(480) activity. The Ub-Br probe can bind to wild type UL36-DUB and mutants UL36(480)H234A and UL36(480)D232A, but not C98 mutants. These in vitro results suggested that the C98 and H234 are essential catalytic residues of UL36-DUB. UL36-DUB exhibited a strict substrate specificity. Inhibition assay revealed that UL36-DUB exhibits resistance to the Roche protease inhibitor cocktail and serine protease inhibitor, but not to the Solarbio protease inhibitor cocktail. (4) Conclusions: UL36-DUB exhibited a strict substrate preference, and the protocol developed in the current study for obtaining active UL36-DUB protein should promote the high-throughput screening of UL36 inhibitors and the study on the function of MDV-encoded UL36.