Human early syncytiotrophoblasts are highly susceptible to SARS-CoV-2 infection.

Direct in vivo investigation of human placenta trophoblast's susceptibility to SARS-CoV-2 is challenging. Here we report that human trophoblast stem cells (hTSCs) and their derivatives are susceptible to SARS-CoV-2 infection, which reveals heterogeneity in hTSC cultures. Early syncytiotrophoblasts (eSTBs) generated from hTSCs have enriched transcriptomic features of peri-implantation trophoblasts, express high levels of angiotensin-converting enzyme 2 (ACE2), and are productively infected by SARS-CoV-2 and its Delta and Omicron variants to produce virions. Antiviral drugs suppress SARS-CoV-2 replication in eSTBs and antagonize the virus-induced blockage of STB maturation. Although less susceptible to SARS-CoV-2 infection, trophoblast organoids originating from hTSCs show detectable viral replication reminiscent of the uncommon placental infection. These findings implicate possible risk of COVID-19 infection in peri-implantation embryos, which may go unnoticed. Stem cell-derived human trophoblasts such as eSTBs can potentially provide unlimited amounts of normal and genome-edited cells and facilitate coronavirus research and antiviral discovery.

Comparative analysis of SARS-CoV-2 Omicron BA.2.12.1 and BA.5.2 variants.

The initial severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron subvariants, BA.1 and BA.2, are being progressively displaced by BA.5 in many countries. To provide insight on the replacement of BA.2 by BA.5 as the dominant SARS-CoV-2 variant, we performed a comparative analysis of Omicron BA.2.12.1 and BA.5.2 variants in cell culture and hamster models. We found that BA.5.2 exhibited enhanced replicative kinetics over BA.2.12.1 in vitro and in vivo, which is evidenced by the dominant BA.5.2 viral genome detected at different time points, regardless of immune selection pressure with vaccine-induced serum antibodies. Utilizing reverse genetics, we constructed a mutant SARS-CoV-2 carrying spike F486V substitution, which is an uncharacterized mutation that concurrently discriminates Omicron BA.5.2 from BA.2.12.1 variant. We noticed that the 486th residue does not confer viral replication advantage to the virus. We also found that 486V displayed generally reduced immune evasion capacity when compared with its predecessor, 486F. However, the surge of fitness in BA.5.2 over BA.2.12.1 was not due to stand-alone F486V substitution but as a result of the combination of multiple mutations. Our study upholds the urgency for continuous monitoring of SARS-CoV-2 Omicron variants with enhanced replication fitness.

Control of SARS-CoV-2 infection by MT1-MMP-mediated shedding of ACE2.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global pandemic. Angiotensin-converting enzyme 2 (ACE2) is an entry receptor for SARS-CoV-2. The full-length membrane form of ACE2 (memACE2) undergoes ectodomain shedding to generate a shed soluble form (solACE2) that mediates SARS-CoV-2 entry via receptor-mediated endocytosis. Currently, it is not known how the physiological regulation of ACE2 shedding contributes to the etiology of COVID-19 in vivo. The present study identifies Membrane-type 1 Matrix Metalloproteinase (MT1-MMP) as a critical host protease for solACE2-mediated SARS-CoV-2 infection. SARS-CoV-2 infection leads to increased activation of MT1-MMP that is colocalized with ACE2 in human lung epithelium. Mechanistically, MT1-MMP directly cleaves memACE2 at M706-S to release solACE218-706 that binds to the SARS-CoV-2 spike proteins (S), thus facilitating cell entry of SARS-CoV-2. Human solACE218-706 enables SARS-CoV-2 infection in both non-permissive cells and naturally insusceptible C57BL/6 mice. Inhibition of MT1-MMP activities suppresses solACE2-directed entry of SARS-CoV-2 in human organoids and aged mice. Both solACE2 and circulating MT1-MMP are positively correlated in plasma of aged mice and humans. Our findings provide in vivo evidence demonstrating the contribution of ACE2 shedding to the etiology of COVID-19.

A molecularly engineered, broad-spectrum anti-coronavirus lectin inhibits SARS-CoV-2 and MERS-CoV infection in vivo.

"Pan-coronavirus" antivirals targeting conserved viral components can be designed. Here, we show that the rationally engineered H84T-banana lectin (H84T-BanLec), which specifically recognizes high mannose found on viral proteins but seldom on healthy human cells, potently inhibits Middle East respiratory syndrome coronavirus (MERS-CoV), severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (including Omicron), and other human-pathogenic coronaviruses at nanomolar concentrations. H84T-BanLec protects against MERS-CoV and SARS-CoV-2 infection in vivo. Importantly, intranasally and intraperitoneally administered H84T-BanLec are comparably effective. Mechanistic assays show that H84T-BanLec targets virus entry. High-speed atomic force microscopy depicts real-time multimolecular associations of H84T-BanLec dimers with the SARS-CoV-2 spike trimer. Single-molecule force spectroscopy demonstrates binding of H84T-BanLec to multiple SARS-CoV-2 spike mannose sites with high affinity and that H84T-BanLec competes with SARS-CoV-2 spike for binding to cellular ACE2. Modeling experiments identify distinct high-mannose glycans in spike recognized by H84T-BanLec. The multiple H84T-BanLec binding sites on spike likely account for the drug compound's broad-spectrum antiviral activity and the lack of resistant mutants.

SARS-CoV-2 hijacks macropinocytosis to facilitate its entry and promote viral spike-mediated cell-to-cell fusion.

Revealing the mechanisms of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) entry and cell-to-cell spread might provide insights for understanding the underlying mechanisms of viral pathogenesis, tropism, and virulence. The signaling pathways involved in SARS-CoV-2 entry and viral spike-mediated cell-to-cell fusion remain elusive. In the current study, we found that macropinocytosis inhibitors significantly suppressed SARS-CoV-2 infection at both the entry and viral spike-mediated cell-to-cell fusion steps. We demonstrated that SARS-CoV-2 entry required the small GTPase Rac1 and its effector kinase p21-activated kinase 1 by dominant-negative and RNAi assays in human embryonic kidney 293T-angiotensin-converting enzyme 2 cells and that the serine protease transmembrane serine protease 2 reversed the decrease in SARS-CoV-2 entry caused by the macropinocytosis inhibitors. Moreover, in the cell-to-cell fusion assay, we confirmed that macropinocytosis inhibitors significantly decreased viral spike-mediated cell-to-cell fusion. Overall, we provided evidence that SARS-CoV-2 utilizes a macropinocytosis pathway to enter target cells and to efficiently promote viral spike-mediated cell-to-cell fusion.

The spike receptor-binding motif G496S substitution determines the replication fitness of SARS-CoV-2 Omicron sublineage.

The replication and pathogenicity of SARS-CoV-2 Omicron BA.2 are comparable to that of BA.1 in experimental animal models. However, BA.2 has rapidly emerged to overtake BA.1 to become the predominant circulating SARS-CoV-2 variant worldwide. Here, we compared the replication fitness of BA.1 and BA.2 in cell culture and in the Syrian hamster model of COVID-19. Using a reverse genetics approach, we found that the BA.1-specific spike mutation G496S compromises its replication fitness, which may contribute to BA.1 being outcompeted by BA.2 in the real world. Additionally, the BA.1-unique G496S substitution confers differentiated sensitivity to therapeutic monoclonal antibodies, which partially recapitulates the immunoevasive phenotype of BA.1 and BA.2. In summary, our study identified G496S as an important determinant during the evolutionary trajectory of SARS-CoV-2.

Production of single-cycle infectious SARS-CoV-2 through a trans-complemented replicon.

Single-cycle infectious virus can elicit close-to-natural immune response and memory. One approach to generate single-cycle severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is through deletion of structural genes such as spike (S) and nucleocapsid (N). Transcomplementation of the resulting ΔS or ΔN virus through enforced expression of S or N protein in the cells gives rise to a live but unproductive virus. In this study, ΔS and ΔN BAC clones were constructed and their live virions were rescued by transient expression of S and N proteins from the ancestral and the Omicron strains. ΔS and ΔN virions were visualized by transmission electron microscopy. Virion production of ΔS was more efficient than that of ΔN. The coated S protein from ΔS was delivered to infected cells in which the expression of N protein was also robust. In contrast, expression of neither S nor N was detected in ΔN-infected cells. ΔS underwent viral RNA replication, induced type I interferon (IFN) response, but did not form plaques. Despite RNA replication in cells, ΔS infection did not produce viral progeny in culture supernatant. Interestingly, viral RNA replication was not further enhanced upon overexpression of S protein. Taken together, our work provides a versatile platform for development of single-cycle vaccines for SARS-CoV-2.

How epidemic information and policy information impact anti-infection behaviors: a cross-cultural study under social influence framing.

Three preregistered experiments examined to what extent information about an epidemic situation provided by experts and information about anti-infection policies promoted by governments/media influenced anti-infection behaviors. The above effects were examined among populations from different countries (in Experiments 2 and 3) and across self-construals (in Experiment 3). In three experiments, participants (N =706) were presented with a scenario where experts provided (or did not provide) information about an epidemic situation and governments/media promoted (or did not promote) information about anti-infection policies. After that, participants indicated their willingness to adopt anti-infection behaviors. Results across three experiments showed that both types of information independently increased participants' anti-infection behaviors. In Experiments 2 and 3, we further found that the epidemic information had a larger impact on inducing anti-infection behaviors than the policy information, which was robust and consistent across countries and self-construals. Findings were discussed under the framework of social influence and in terms of practical implications for pandemic situations like the COVID-19.

Phosphatidic acid phosphatase 1 impairs SARS-CoV-2 replication by affecting the glycerophospholipid metabolism pathway.

Viruses exploit the host lipid metabolism machinery to achieve efficient replication. We herein characterize the lipids profile reprogramming in vitro and in vivo using liquid chromatography-mass spectrometry-based untargeted lipidomics. The lipidome of SARS-CoV-2-infected Caco-2 cells was markedly different from that of mock-infected samples, with most of the changes involving downregulation of ceramides. In COVID-19 patients' plasma samples, a total of 54 lipids belonging to 12 lipid classes that were significantly perturbed compared to non-infected control subjects' plasma samples were identified. Among these 12 lipid classes, ether-linked phosphatidylcholines, ether-linked phosphatidylethanolamines, phosphatidylcholines, and ceramides were the four most perturbed. Pathway analysis revealed that the glycerophospholipid, sphingolipid, and ether lipid metabolisms pathway were the most significantly perturbed host pathways. Phosphatidic acid phosphatases (PAP) were involved in all three pathways and PAP-1 deficiency significantly suppressed SARS-CoV-2 replication. siRNA knockdown of LPIN2 and LPIN3 resulted in significant reduction of SARS-CoV-2 load. In summary, these findings characterized the host lipidomic changes upon SARS-CoV-2 infection and identified PAP-1 as a potential target for intervention for COVID-19.

Cross-variant protection against SARS-CoV-2 infection in hamsters immunized with monovalent and bivalent inactivated vaccines.

Rapid development and successful use of vaccines against SARS-CoV-2 might hold the key to curb the ongoing pandemic of COVID-19. Emergence of vaccine-evasive SARS-CoV-2 variants of concern (VOCs) has posed a new challenge to vaccine design and development. One urgent need is to determine what types of variant-specific and bivalent vaccines should be developed. Here, we compared homotypic and heterotypic protection against SARS-CoV-2 infection of hamsters with monovalent and bivalent whole-virion inactivated vaccines derived from representative VOCs. In addition to the ancestral SARS-CoV-2 Wuhan strain, Delta (B.1.617.2; δ) and Theta (P.3; θ) variants were used in vaccine preparation. Additional VOCs including Omicron (B.1.1.529) and Alpha (B.1.1.7) variants were employed in the challenge experiment. Consistent with previous findings, Omicron variant exhibited the highest degree of immune evasion, rendering all different forms of inactivated vaccines substantially less efficacious. Notably, monovalent and bivalent Delta variant-specific inactivated vaccines provided optimal protection against challenge with Delta variant. Yet, some cross-variant protection against Omicron and Alpha variants was seen with all monovalent and bivalent inactivated vaccines tested. Taken together, our findings support the notion that an optimal next-generation inactivated vaccine against SARS-CoV-2 should contain the predominant VOC in circulation. Further investigations are underway to test whether a bivalent vaccine for Delta and Omicron variants can serve this purpose.

Pathogenicity, transmissibility, and fitness of SARS-CoV-2 Omicron in Syrian hamsters.

The in vivo pathogenicity, transmissibility, and fitness of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron (B.1.1.529) variant are not well understood. We compared these virological attributes of this new variant of concern (VOC) with those of the Delta (B.1.617.2) variant in a Syrian hamster model of COVID-19. Omicron-infected hamsters lost significantly less body weight and exhibited reduced clinical scores, respiratory tract viral burdens, cytokine and chemokine dysregulation, and lung damage than Delta-infected hamsters. Both variants were highly transmissible through contact transmission. In noncontact transmission studies Omicron demonstrated similar or higher transmissibility than Delta. Delta outcompeted Omicron without selection pressure, but this scenario changed once immune selection pressure with neutralizing antibodies-active against Delta but poorly active against Omicron-was introduced. Next-generation vaccines and antivirals effective against this new VOC are therefore urgently needed.

Intranasal administration of a single dose of a candidate live attenuated vaccine derived from an NSP16-deficient SARS-CoV-2 strain confers sterilizing immunity in animals.

Live attenuated vaccines might elicit mucosal and sterilizing immunity against SARS-CoV-2 that the existing mRNA, adenoviral vector and inactivated vaccines fail to induce. Here, we describe a candidate live attenuated vaccine strain of SARS-CoV-2 in which the NSP16 gene, which encodes 2'-O-methyltransferase, is catalytically disrupted by a point mutation. This virus, designated d16, was severely attenuated in hamsters and transgenic mice, causing only asymptomatic and nonpathogenic infection. A single dose of d16 administered intranasally resulted in sterilizing immunity in both the upper and lower respiratory tracts of hamsters, thus preventing viral spread in a contact-based transmission model. It also robustly stimulated humoral and cell-mediated immune responses, thus conferring full protection against lethal challenge with SARS-CoV-2 in a transgenic mouse model. The neutralizing antibodies elicited by d16 effectively cross-reacted with several SARS-CoV-2 variants. Secretory immunoglobulin A was detected in the blood and nasal wash of vaccinated mice. Our work provides proof-of-principle evidence for harnessing NSP16-deficient SARS-CoV-2 for the development of live attenuated vaccines and paves the way for further preclinical studies of d16 as a prototypic vaccine strain, to which new features might be introduced to improve safety, transmissibility, immunogenicity and efficacy.

Targeting papain-like protease for broad-spectrum coronavirus inhibition.

The emergence of SARS-CoV-2 variants of concern and repeated outbreaks of coronavirus epidemics in the past two decades emphasize the need for next-generation pan-coronaviral therapeutics. Drugging the multi-functional papain-like protease (PLpro) domain of the viral nsp3 holds promise. However, none of the known coronavirus PLpro inhibitors has been shown to be in vivo active. Herein, we screened a structurally diverse library of 50,080 compounds for potential coronavirus PLpro inhibitors and identified a noncovalent lead inhibitor F0213 that has broad-spectrum anti-coronaviral activity, including against the Sarbecoviruses (SARS-CoV-1 and SARS-CoV-2), Merbecovirus (MERS-CoV), as well as the Alphacoronavirus (hCoV-229E and hCoV-OC43). Importantly, F0213 confers protection in both SARS-CoV-2-infected hamsters and MERS-CoV-infected human DPP4-knockin mice. F0213 possesses a dual therapeutic functionality that suppresses coronavirus replication via blocking viral polyprotein cleavage, as well as promoting antiviral immunity by antagonizing the PLpro deubiquitinase activity. Despite the significant difference of substrate recognition, mode of inhibition studies suggest that F0213 is a competitive inhibitor against SARS2-PLpro via binding with the 157K amino acid residue, whereas an allosteric inhibitor of MERS-PLpro interacting with its 271E position. Our proof-of-concept findings demonstrated that PLpro is a valid target for the development of broad-spectrum anti-coronavirus agents. The orally administered F0213 may serve as a promising lead compound for combating the ongoing COVID-19 pandemic and future coronavirus outbreaks.

Broad-Spectrum Host-Based Antivirals Targeting the Interferon and Lipogenesis Pathways as Potential Treatment Options for the Pandemic Coronavirus Disease 2019 (COVID-19).

The ongoing Coronavirus Disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) signals an urgent need for an expansion in treatment options. In this study, we investigated the anti-SARS-CoV-2 activities of 22 antiviral agents with known broad-spectrum antiviral activities against coronaviruses and/or other viruses. They were first evaluated in our primary screening in VeroE6 cells and then the most potent anti-SARS-CoV-2 antiviral agents were further evaluated using viral antigen expression, viral load reduction, and plaque reduction assays. In addition to remdesivir, lopinavir, and chloroquine, our primary screening additionally identified types I and II recombinant interferons, 25-hydroxycholesterol, and AM580 as the most potent anti-SARS-CoV-2 agents among the 22 antiviral agents. Betaferon (interferon-ß1b) exhibited the most potent anti-SARS-CoV-2 activity in viral antigen expression, viral load reduction, and plaque reduction assays among the recombinant interferons. The lipogenesis modulators 25-hydroxycholesterol and AM580 exhibited EC50 at low micromolar levels and selectivity indices of >10.0. Combinational use of these host-based antiviral agents with virus-based antivirals to target different processes of the SARS-CoV-2 replication cycle should be evaluated in animal models and/or clinical trials.

SARS-CoV-2 and COVID-19: revisiting the most important research questions.

In February 2020, we highlighted the top nine important research questions on SARS-CoV-2 and COVID-19 concerning virus transmission, asymptomatic and presymptomatic virus shedding, diagnosis, treatment, vaccine development, origin of virus and viral pathogenesis. These and related questions are revisited at the end of 2021 to shed light on the roadmap of bringing an end to the pandemic.

SARS-CoV-2 exploits host DGAT and ADRP for efficient replication.

Coronavirus Disease 2019 (COVID-19) is predominantly a respiratory tract infection that significantly rewires the host metabolism. Here, we monitored a cohort of COVID-19 patients' plasma lipidome over the disease course and identified triacylglycerol (TG) as the dominant lipid class present in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-induced metabolic dysregulation. In particular, we pinpointed the lipid droplet (LD)-formation enzyme diacylglycerol acyltransferase (DGAT) and the LD stabilizer adipocyte differentiation-related protein (ADRP) to be essential host factors for SARS-CoV-2 replication. Mechanistically, viral nucleo capsid protein drives DGAT1/2 gene expression to facilitate LD formation and associates with ADRP on the LD surface to complete the viral replication cycle. DGAT gene depletion reduces SARS-CoV-2 protein synthesis without compromising viral genome replication/transcription. Importantly, a cheap and orally available DGAT inhibitor, xanthohumol, was found to suppress SARS-CoV-2 replication and the associated pulmonary inflammation in a hamster model. Our findings not only uncovered the mechanistic role of SARS-CoV-2 nucleocapsid protein to exploit LDs-oriented network for heightened metabolic demand, but also the potential to target the LDs-synthetase DGAT and LDs-stabilizer ADRP for COVID-19 treatment.

Spatiotemporal analysis of COVID-19 outbreaks in Wuhan, China.

Few study has revealed spatial transmission characteristics of COVID-19 in Wuhan, China. We aimed to analyze the spatiotemporal spread of COVID-19 in Wuhan and its influence factors. Information of 32,682 COVID-19 cases reported through March 18 were extracted from the national infectious disease surveillance system. Geographic information system methods were applied to analysis transmission of COVID-19 and its influence factors in different periods. We found decrease in effective reproduction number (Rt) and COVID-19 related indicators through taking a series of effective public health measures including restricting traffic, centralized quarantine and strict stay-at home policy. The distribution of COVID-19 cases number in Wuhan showed obvious global aggregation and local aggregation. In addition, the analysis at streets-level suggested population density and the number of hospitals were associated with COVID-19 cases number. The epidemic situation showed obvious global and local spatial aggregations. High population density with larger number of hospitals may account for the aggregations. The epidemic in Wuhan was under control in a short time after strong quarantine measures and restrictions on movement of residents were implanted.

Self-Interest Bias in the COVID-19 Pandemic: A Cross-Cultural Comparison between the United States and China

In the global crisis of the COVID-19 pandemic, many countries attempt to enforce new social norms to prevent the further spread of the coronavirus. A key to the success of these measures is the individual adherence to norms that are collectively beneficial to contain the spread of the pandemic. However, individuals? self-interest bias (i.e., the prevalent tendency to license own but not others? self-serving acts or norm violations) can pose a challenge to the success of such measures. The current research examines COVID-19-related self-interest bias from a cross-cultural perspective. Two studies (N?=?1,558) sampled from the United States and China consistently revealed that participants from the United States evaluated their own self-serving acts (exploiting test kits in Study 1;social gathering and sneezing without covering the mouth in public in Study 2) as more acceptable than identical deeds of others, while such self-interest bias did not emerge among Chinese participants. Cultural underpinnings of independent versus interdependent self-construal may influence the extent to which individuals apply self-interest bias to justifications of their own self-serving behaviors during the pandemic.

In silico structure-based discovery of a SARS-CoV-2 main protease inhibitor.

The Coronavirus Disease 2019 (COVID-19) pandemic caused by the novel lineage B betacoroanvirus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in significant mortality, morbidity, and socioeconomic disruptions worldwide. Effective antivirals are urgently needed for COVID-19. The main protease (Mpro) of SARS-CoV-2 is an attractive antiviral target because of its essential role in the cleavage of the viral polypeptide. In this study, we performed an in silico structure-based screening of a large chemical library to identify potential SARS-CoV-2 Mpro inhibitors. Among 8,820 compounds in the library, our screening identified trichostatin A, a histone deacetylase inhibitor and an antifungal compound, as an inhibitor of SARS-CoV-2 Mpro activity and replication. The half maximal effective concentration of trichostatin A against SARS-CoV-2 replication was 1.5 to 2.7µM, which was markedly below its 50% effective cytotoxic concentration (75.7µM) and peak serum concentration (132µM). Further drug compound optimization to develop more stable analogues with longer half-lives should be performed. This structure-based drug discovery platform should facilitate the identification of additional enzyme inhibitors of SARS-CoV-2.