COVID-19 immune features revealed by a large-scale single-cell transcriptome atlas.

A dysfunctional immune response in coronavirus disease 2019 (COVID-19) patients is a recurrent theme impacting symptoms and mortality, yet a detailed understanding of pertinent immune cells is not complete. We applied single-cell RNA sequencing to 284 samples from 196 COVID-19 patients and controls and created a comprehensive immune landscape with 1.46 million cells. The large dataset enabled us to identify that different peripheral immune subtype changes are associated with distinct clinical features, including age, sex, severity, and disease stages of COVID-19. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA was found in diverse epithelial and immune cell types, accompanied by dramatic transcriptomic changes within virus-positive cells. Systemic upregulation of S100A8/A9, mainly by megakaryocytes and monocytes in the peripheral blood, may contribute to the cytokine storms frequently observed in severe patients. Our data provide a rich resource for understanding the pathogenesis of and developing effective therapeutic strategies for COVID-19.

Potent Neutralizing Antibodies against SARS-CoV-2 Identified by High-Throughput Single-Cell Sequencing of Convalescent Patients' B Cells.

The COVID-19 pandemic urgently needs therapeutic and prophylactic interventions. Here, we report the rapid identification of SARS-CoV-2-neutralizing antibodies by high-throughput single-cell RNA and VDJ sequencing of antigen-enriched B cells from 60 convalescent patients. From 8,558 antigen-binding IgG1+ clonotypes, 14 potent neutralizing antibodies were identified, with the most potent one, BD-368-2, exhibiting an IC50 of 1.2 and 15 ng/mL against pseudotyped and authentic SARS-CoV-2, respectively. BD-368-2 also displayed strong therapeutic and prophylactic efficacy in SARS-CoV-2-infected hACE2-transgenic mice. Additionally, the 3.8 Å cryo-EM structure of a neutralizing antibody in complex with the spike-ectodomain trimer revealed the antibody's epitope overlaps with the ACE2 binding site. Moreover, we demonstrated that SARS-CoV-2-neutralizing antibodies could be directly selected based on similarities of their predicted CDR3H structures to those of SARS-CoV-neutralizing antibodies. Altogether, we showed that human neutralizing antibodies could be efficiently discovered by high-throughput single B cell sequencing in response to pandemic infectious diseases.

Analysis of SARS-CoV-2 variant mutations reveals neutralization escape mechanisms and the ability to use ACE2 receptors from additional species.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants continue to emerge during the global pandemic and may facilitate escape from current antibody therapies and vaccine protection. Here we showed that the South African variant B.1.351 was the most resistant to current monoclonal antibodies and convalescent plasma from coronavirus disease 2019 (COVID-19)-infected individuals, followed by the Brazilian variant P.1 and the United Kingdom variant B.1.1.7. This resistance hierarchy corresponded with Y144del and 242-244del mutations in the N-terminal domain and K417N/T, E484K, and N501Y mutations in the receptor-binding domain (RBD) of SARS-CoV-2. Crystal structure analysis of the B.1.351 triple mutant (417N-484K-501Y) RBD complexed with the monoclonal antibody P2C-1F11 revealed the molecular basis for antibody neutralization and escape. B.1.351 and P.1 also acquired the ability to use mouse and mink ACE2 receptors for entry. Our results demonstrate major antigenic shifts and potential broadening of the host range for B.1.351 and P.1 variants, which poses serious challenges to current antibody therapies and vaccine protection.

The N-Degron Pathway Mediates ER-phagy.

The endoplasmic reticulum (ER) is susceptible to wear-and-tear and proteotoxic stress, necessitating its turnover. Here, we show that the N-degron pathway mediates ER-phagy. This autophagic degradation initiates when the transmembrane E3 ligase TRIM13 (also known as RFP2) is ubiquitinated via the lysine 63 (K63) linkage. K63-ubiquitinated TRIM13 recruits p62 (also known as sequestosome-1), whose complex undergoes oligomerization. The oligomerization is induced when the ZZ domain of p62 is bound by the N-terminal arginine (Nt-Arg) of arginylated substrates. Upon activation by the Nt-Arg, oligomerized TRIM13-p62 complexes are separated along with the ER compartments and targeted to autophagosomes, leading to lysosomal degradation. When protein aggregates accumulate within the ER lumen, degradation-resistant autophagic cargoes are co-segregated by ER membranes for lysosomal degradation. We developed synthetic ligands to the p62 ZZ domain that enhance ER-phagy for ER protein quality control and alleviate ER stresses. Our results elucidate the biochemical mechanisms and pharmaceutical means that regulate ER homeostasis.

Highly stabilized and efficient thermoelectric copper selenide.

The liquid-like feature of thermoelectric superionic conductors is a double-edged sword: the long-range migration of ions hinders the phonon transport, but their directional segregation greatly impairs the service stability. We report the synergetic enhancement in figure of merit (ZT) and stability in Cu1.99Se-based superionic conductors enabled by ion confinement effects. Guided by density functional theory and nudged elastic band simulations, we elevated the activation energy to restrict ion migrations through a cation-anion co-doping strategy. We reduced the carrier concentration without sacrificing the low thermal conductivity, obtaining a ZT of ∼3.0 at 1,050 K. Notably, the fabricated device module maintained a high conversion efficiency of up to ∼13.4% for a temperature difference of 518 K without obvious degradation after 120 cycles. Our work could be generalized to develop electrically and thermally robust functional materials with ionic migration characteristics.

Key gp120 Glycans Pose Roadblocks to the Rapid Development of VRC01-Class Antibodies in an HIV-1-Infected Chinese Donor.

VRC01-class antibodies neutralize diverse HIV-1 strains by targeting the conserved CD4-binding site. Despite extensive investigations, crucial events in the early stage of VRC01 development remain elusive. We demonstrated how VRC01-class antibodies emerged in a Chinese donor by antigen-specific single B cell sorting, structural and functional studies, and longitudinal antibody and virus repertoire analyses. A monoclonal antibody DRVIA7 with modest neutralizing breadth was isolated that displayed a subset of VRC01 signatures. X-ray and EM structures revealed a VRC01-like angle of approach, but less favorable interactions between the DRVIA7 light-chain CDR1 and the N terminus with N276 and V5 glycans of gp120. Although the DRVIA7 lineage was unable to acquire broad neutralization, longitudinal analysis revealed a repertoire-encoded VRC01 light-chain CDR3 signature and VRC01-like neutralizing heavy-chain precursors that rapidly matured within 2 years. Thus, light chain accommodation of the glycan shield should be taken into account in vaccine design targeting this conserved site of vulnerability.

Efficacy of Voluntary Medical Male Circumcision to Prevent HIV Infection Among Men Who Have Sex With Men : A Randomized Controlled Trial.

BACKGROUND: Observational studies suggest that voluntary medical male circumcision (VMMC) may lower HIV risk among men who have sex with men (MSM). A randomized controlled trial (RCT) is needed to confirm this. OBJECTIVE: To assess the efficacy of VMMC in preventing incident HIV infection among MSM. DESIGN: An RCT with up to 12 months of follow-up. (Chinese Clinical Trial Registry: ChiCTR2000039436). SETTING: 8 cities in China. PARTICIPANTS: Uncircumcised, HIV-seronegative men aged 18 to 49 years who self-reported predominantly practicing insertive anal intercourse and had 2 or more male sex partners in the past 6 months. INTERVENTION: VMMC. MEASUREMENTS: Rapid testing for HIV was done at baseline and at 3, 6, 9, and 12 months. Behavioral questionnaires and other tests for sexually transmitted infections were done at baseline, 6 months, and 12 months. The primary outcome was HIV seroconversion using an intention-to-treat analysis. RESULTS: The study enrolled 124 men in the intervention group and 123 in the control group, who contributed 120.7 and 123.1 person-years of observation, respectively. There were 0 seroconversions in the intervention group (0 infections [95% CI, 0.0 to 3.1 infections] per 100 person-years) and 5 seroconversions in the control group (4.1 infections [CI, 1.3 to 9.5 infections] per 100 person-years). The HIV hazard ratio was 0.09 (CI, 0.00 to 0.81; P = 0.029), and the HIV incidence was lower in the intervention group (log-rank P = 0.025). The incidence rates of syphilis, herpes simplex virus type 2, and penile human papillomavirus were not statistically significantly different between the 2 groups. There was no evidence of HIV risk compensation. LIMITATION: Few HIV seroconversions and limited follow-up period. CONCLUSION: Among MSM who predominantly practice insertive anal intercourse, VMMC is efficacious in preventing incident HIV infection; MSM should be included in VMMC guidelines. PRIMARY FUNDING SOURCE: The National Science and Technology Major Project of China.

Unpredicted protective function of Fc-mediated inhibitory antibodies for HIV and SARS-CoV-2 vaccines.

Developing effective vaccines is necessary in combating new virus pandemics. For HIV and SARS-CoV-2, the induction of neutralizing antibodies (NAb) is important for vaccine protection; however, the exact mechanisms underlying protection require further study. Recent data emphasize that even Abs that do not exhibit neutralizing activity may contribute to immune defense. Abs exhibiting this function may counter virus mutations, which are acquired to escape from NAbs, and therefore, broaden the protective Ab response induced by vaccination. However, the steps leading to Ab Fc-mediated inhibition are complex. How can these functions be measured in vitro? What inhibitory assay is the most physiologically relevant at mimicking effective in vivo protection? This review provides a comprehensive update on the current knowledge gaps on the Ab Fc-mediated functions involved in HIV and SARS-CoV-2 protection. Understanding the inhibitory effects of these Abs is vital for designing the next generation of protective HIV and SARS-CoV-2 vaccines.

Controllable and Low-Loss Electrochemiluminescence Waveguide Supported by a Micropipette Electrode.

One-dimensional molecular crystal waveguide (MCW) can transmit self-generated electrochemiluminescence (ECL), but heavy optical loss occurs because of the small difference in the refractive index between the crystal and its surroundings. Herein, we report a micropipette electrode-supported MCW (MPE/MCW) for precisely controlling the far-field transmission of ECL in air with a low optical loss. ECL is generated from one terminal of the MCW positioned inside the MPE, which is transmitted along the MCW to the other terminal in air. In comparison with conventional waveguides on solid substrates or in solutions, the MPE/MCW is propitious to the total internal reflection of light at the MCW/air interface, thus confining the ECL efficiently in MCW and improving the waveguide performance with an extremely low-loss coefficient of 4.49 × 10-3 dB µm-1. Moreover, by regulation of the gas atmosphere, active and passive waveguides can be resolved simultaneously inside MPE and in air. This MPE/MCW offers a unique advantage of spatially controlling and separating ECL signal readout from its generation, thus holding great promise in biosensing without or with less electrical/chemical disturbance.

CRISPR-Cas12a-Mediated Hue-Recognition Lateral Flow Assay for Point-of-Need Detection of Salmonella.

Nucleic acid detection of pathogens in a point-of-need (PON) manner is of great significance yet remains challenging for sensitive and accurate visual discrimination. Here, we report a CRISPR-Cas12a-mediated lateral flow assay for PON detection of Salmonella typhimurium (S.ty) that is a prevailing pathogen disseminated through tainted food. The variation of the fluorescence color of the test line is exploited to interpret the results, enabling the discrimination between positive and negative samples on the basis of a hue-recognition mechanism. By leveraging the cleavage activity of Cas12a and hue-recognition readout, the assay facilitated by recombinase polymerase amplification can yield a visual detection limit of 1 copy µL-1 for S.ty genomic DNA within 1 h. The assay also displays a high specificity toward S.ty in fresh chicken samples, as well as a sensitivity 10-fold better than that of the commercial test strip. Moreover, a semiquantitative detection of S.ty ranging from 0 to 4 × 103 CFU/mL by the naked eye is made possible, thanks to the easily discernible color change of the test line. This approach provides an easy, rapid, accurate, and user-friendly solution for the PON detection of Salmonella and other pathogens.

Electrochemical Lateral Flow Immunoassay with Built-In Electrodes for Ultrasensitive and Wireless Detection of Inflammatory Biomarkers.

Paper-based lateral flow immunoassays (LFIAs) are cost-effective, portable, and simple methods for detection of diverse analytes, which however only provide qualitative or semiquantitative results and lack sufficient sensitivity. A combination of LFIA and electrochemical detection, namely, electrochemical lateral flow immunoassay (eLFIA), enables quantitative detection of analytes with high sensitivity, but the integration of external electrodes makes the system relatively expensive and unstable. Herein, the working, counter, and reference electrodes were prepared directly on the nitrocellulose membrane using screen printing, which remarkably simplified the structure of eLFIA and decreased the cost. Moreover, a horseradish peroxidase (HRP)-based electrochemical signal amplification strategy was used for further increasing the analytical sensitivity. HRP captured on the working electrode can catalyze the oxidation of tetramethylbenzidine (TMB) to form the TMB-TMBox precipitate on the electrode surface, which as an electrochemically active product can output an amplified current for quantification. We demonstrated that the eLFIA could detect low-abundant inflammatory biomarkers in human plasma samples with limits of detection of 0.17 and 0.54 pg mL-1 for interleukin-6 and C-reactive protein, respectively. Finally, a fully portable system was fabricated by integrating eLFIA with a flexible and wireless electrochemical workstation, realizing the point-of-care detection of interleukin-6.

Structural insights into BirA from Haemophilus influenzae, a bifunctional protein as a biotin protein ligase and a transcriptional repressor.

Biotin is an essential coenzyme involved in various metabolic processes across all known organisms, with biotinylation being crucial for the activity of carboxylases. BirA from Haemophilus influenzae is a bifunctional protein that acts as a biotin protein ligase and a transcriptional repressor. This study reveals the crystal structures of Hin BirA in both its apo- and holo-(biotinyl-5'-AMP bound) forms. As a class II BirA, it consists of three domains: N-terminal DNA binding domain, central catalytic domain, and C-terminal SH3-like domain. The structural analysis shows that the biotin-binding loop forms an ordered structure upon biotinyl-5'-AMP binding. This facilitates its interaction with the ligand and promotes protein dimerization. Comparative studies with other BirA homologs from different organisms indicate that the residues responsible for binding biotinyl-5'-AMP are highly conserved. This study also utilized AlphaFold2 to model the potential heterodimeric interaction between Hin BirA and biotin carboxyl carrier protein, thereby providing insights into the structural basis for biotinylation. These findings enhance our understanding of the structural and functional characteristics of Hin BirA, highlighting its potential as a target for novel antibiotics that disrupt the bacterial biotin synthesis pathways.

SIRT1 regulates the localization and stability of telomerase protein by direct interaction.

Telomerase reverse transcriptase (TERT) not only upholds telomeric equilibrium but also plays a pivotal role in multiple non-canonical cellular mechanisms, particularly in the context of aging, cancer, and genomic stability. Though depletion of SIRT1 in mouse embryonic fibroblasts has demonstrated telomere shortening, the impact of SIRT1 on enabling TERT to regulate telomeric homeostasis remains enigmatic. Here, we reveal that SIRT1 directly interacts with TERT, and promotes the nuclear localization and stability of TERT. Reverse transcriptase (RT) domain of TERT and N-terminus of SIRT1 mainly participated in their direct interaction. TERT, concomitantly expressed with intact SIRT1, exhibits nuclear localization, whereas TERT co-expressed with N-terminal-deleted SIRT1 remains in the cytosol. Furthermore, overexpression of SIRT1 enhances the nuclear localization and protein stability of TERT, akin to overexpression of deacetylase-inactive SIRT1, whereas N-terminal-deleted SIRT1 has no effect on TERT. These findings suggest a novel regulatory role of SIRT1 for TERT through direct interaction. This interaction provides new insights into the fields of aging, cancer, and genome stability governed by TERT and SIRT1.

On-Demand 3D Spatial Distribution of Magnetic Permeability Based on Fe3 O4 Nanoparticle Liquid Toward Micro-Cavity Detectors.

The change of 3D spatial distribution of magnetic permeability can lead to the generation of introduced electrical signals. However, present studies can only achieve rough regulation by simple shape deformation of magnetic elastomers such as compression, bending, or stretching. Accurate control of the 3D spatial distribution of magnetic permeability is still an open question. In this study, an on-demand 3D spatial distribution of magnetic permeability by controlled flowing of Fe3 O4 nanoparticle liquid (FNL) is demonstrated. The flowing routes of FNL are tuned by a 3D-printed cage with pre-designed hollow structure, thus changing the 3D spatial distribution of magnetic permeability. Then, eight symmetrically distributed coils under cage are used to receive characteristic induction voltage signals. Maxwell numerical simulation reveals the working mechanism of signal generation. Notably, those eight coils can detect FNL flowing status in eight directions, allowing recognition of up to 255 different FNL flowing combinations. By introducing machine learning, the micro-cavity detector based on FNL can distinguish nine kinds of micro-cavity structures with an accuracy of 98.77%. This work provides a new strategy for the adjustment of the 3D spatial distribution of the magnetic permeability and expands the application of FNL in the field of space exploration.

Water-Triggered Self-Healing of Ti3C2Tx MXene Standalone Electrodes: Systematic Examination of Factors Affecting the Healing Process.

MXenes, with their remarkable attributes, stand at the forefront of diverse applications. However, the challenge remains in sustaining their performance, especially concerning Ti3C2Tx MXene electrodes. Current self-healing techniques, although promising, often rely heavily on adjacent organic materials. This study illuminates a pioneering water-initiated self-healing mechanism tailored specifically for standalone MXene electrodes. Here, both water and select organic solvents seamlessly mend impaired regions. Comprehensive evaluations around solvent types, thermal conditions, and substrate nuances underline water's unmatched healing efficacy, attributed to its innate ability to forge enduring hydrogen bonds with MXenes. Optimal healing environments range from ambient conditions to a modest 50 °C. Notably, on substrates rich in hydroxyl groups, the healing efficiency remains consistently high. The proposed healing mechanism encompasses hydrogen bonding formation, capillary action-induced expansion of interlayer spacing, solvent lubrication, Gibbs free energy minimizing MXene nanosheet rearrangement, and solvent evaporation-triggered MXene layer recombination. MXenes' resilience is further showcased by their electrical revival from profound damages, culminating in the crafting of Joule-heated circuits and heaters.

Recent Advances in Electrochemiluminescence Visual Biosensing and Bioimaging.

Electrochemiluminescence (ECL) is one of the most powerful techniques that meet the needs of analysis and detection in a variety of scenarios, because of its highly analytical sensitivity and excellent spatiotemporal controllability. ECL combined with microscopy (ECLM) offers a promising approach for quantifying and mapping a wide range of analytes. To date, ECLM has been widely used to image biological entities and processes, such as cells, subcellular structures, proteins and membrane transport properties. In this review, we first introduced the mechanisms of several classic ECL systems, then highlighted the progress of visual biosensing and bioimaging by ECLM in the last decade. Finally, the characteristics of ECLM were summarized, as well as some of the current challenges. The future research interests and potential directions for the application of ECLM were also outlooked.

Murine Coronavirus Disease 2019 Lethality Is Characterized by Lymphoid Depletion Associated with Suppressed Antigen-Presenting Cell Functionality.

The disease severity of coronavirus disease 2019 (COVID-19) varies considerably from asymptomatic to serious, with fatal complications associated with dysregulation of innate and adaptive immunity. Lymphoid depletion in lymphoid tissues and lymphocytopenia have both been associated with poor disease outcomes in patients with COVID-19, but the mechanisms involved remain elusive. In this study, human angiotensin-converting enzyme 2 (hACE2) transgenic mouse models susceptible to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection were used to investigate the characteristics and determinants of lethality associated with the lymphoid depletion observed in SARS-CoV-2 infection. The lethality of Wuhan SARS-CoV-2 infection in K18-hACE2 mice was characterized by severe lymphoid depletion and apoptosis in lymphoid tissues related to fatal neuroinvasion. The lymphoid depletion was associated with a decreased number of antigen-presenting cells (APCs) and their suppressed functionality below basal levels. Lymphoid depletion with reduced APC function was a specific feature observed in SARS-CoV-2 infection but not in influenza A infection and had the greatest prognostic value for disease severity in murine COVID-19. Comparison of transgenic mouse models resistant and susceptible to SARS-CoV-2 infection revealed that suppressed APC function could be determined by the hACE2 expression pattern and interferon-related signaling. Thus, we demonstrated that lymphoid depletion associated with suppressed APC function characterizes the lethality of COVID-19 mouse models. Our data also suggest a potential therapeutic approach to prevent the severe progression of COVID-19 by enhancing APC functionality.

Safety and immunogenicity of homologous prime-boost CoronaVac vaccine in people living with HIV in China: A multicenter prospective cohort study.

People living with HIV (PLWH) are particularly vulnerable to SARS-CoV-2. This multicentre prospective cohort study evaluated the long-term immunogenicity and safety of a third homologous dose of Sinovac CoronaVac in PLWH in China. A total of 228 PLWH and 127 HIV-negative controls were finally included and followed up for 6 months. Fewer participants reported mild or moderate adverse reactions, and no serious adverse events were observed. The median levels of neutralizing antibodies (nAbs) and immunoglobulin G against the receptor-binding domain of the spike protein (S-IgG) in PLWH (655.92 IU/mL, IQR: 175.76-1663.55; 206.83 IU/mL, IQR: 85.20-397.82) were comparable to those in control group (1067.16 IU/mL, IQR: 239.85-1670.83; 261.70 IU/mL, IQR: 77.13-400.75), and reached their peak at 4 weeks, exhibiting a delayed peak pattern compared to the 2-week peak in control group. After then, the immune titres gradually decreased over time, but most participants still maintained positive seroconversion at the 6-month mark. Multivariable generalized estimating equation analysis indicated that CD4+T cell count, HIV viral load, and antiretroviral therapy (ART) were independent factors strongly associated with immune response (each p < 0.05). We suggested that PLWH should maintain well-controlled HIV status through ART and receive timely administration of the second booster dose for optimal protection.

Multilayer track-etched membrane-based electroosmotic pump for drug delivery.

Herein, we report an electroosmotic pump (EOP) based on a multilayer track-etched polycarbonate (PC) membrane. A remarkable increase of maximum backpressure (198.2-2400 mmH2 O) of a fundamental pump unit was obtained at 0.8 mA, when the number of PC membranes was increased from 1 to 10. Meanwhile, the corresponding flow rate was increased from 80.3 to 111.7 µL/min. Furthermore, multiple pump units were assembled in series to obtain a multistage EOP. For a three-stage EOP (EOP-3), the operating voltage and power can be decreased significantly by 52%-72% under different driving currents, with a minimum power of 26.7 µW. Thus, EOP-3 can run stably over 35 h at a pulse current of 0.1 mA without the generation of gas bubbles. The pump was further integrated into a miniature device, which was successfully used to decrease the blood glucose level of diabetic rats by subcutaneous delivery of fast-acting insulin. This work brings a facile and efficient strategy to enhance the backpressure and lower the operating voltage and power of EOPs, which may find promising applications in drug delivery.