A T cell-based SARS-CoV-2 spike protein vaccine provides protection without antibodies.

SARS-CoV-2 spike-based vaccines are used to control the COVID-19 pandemic. However, emerging variants have become resistant to antibody neutralization and further mutations may lead to full resistance. We tested whether T cells alone could provide protection without antibodies. We designed a T cell-based vaccine in which SARS-CoV-2 spike sequences were rearranged and attached to ubiquitin. Immunization of mice with the vaccine induced no specific antibodies, but strong specific T cell responses. We challenged mice with SARS-CoV-2 wild-type strain or an Omicron variant after the immunization and monitored survival or viral titers in the lungs. The mice were significantly protected against death and weight loss caused by the SARS-CoV-2 wild-type strain, and the viral titers in the lungs of mice challenged with the SARS-CoV-2 wild-type strain or the Omicron variant were significantly reduced. Importantly, depletion of CD4+ or CD8+ T cells led to significant loss of the protection. Our analyses of spike protein sequences of the variants indicated that fewer than one-third presented by dominant HLA alleles were mutated and that most of the mutated epitopes were in the subunit 1 region. As the subunit 2 region is conservative, the vaccines targeting spike protein are expected to protect against future variants due to the T cell responses.

Integrated Transcriptomics and Metabolomics Analysis Promotes the Understanding of Adventitious Root Formation in Eucommia ulmoides Oliver.

As a primary approach to nutrient propagation for many woody plants, cutting roots is essential for the breeding and production of Eucommia ulmoides Oliver. In this study, hormone level, transcriptomics, and metabolomics analyses were performed on two E. ulmoides varieties with different adventitious root (AR) formation abilities. The higher JA level on the 0th day and the lower JA level on the 18th day promoted superior AR development. Several hub genes executed crucial roles in the crosstalk regulation of JA and other hormones, including F-box protein (EU012075), SAUR-like protein (EU0125382), LOB protein (EU0124232), AP2/ERF transcription factor (EU0128499), and CYP450 protein (EU0127354). Differentially expressed genes (DEGs) and metabolites of AR formation were enriched in phenylpropanoid biosynthesis, flavonoid biosynthesis, and isoflavonoid biosynthesis pathways. The up-regulated expression of PAL, CCR, CAD, DFR, and HIDH genes on the 18th day could contribute to AR formation. The 130 cis-acting lncRNAs had potential regulatory functions on hub genes in the module that significantly correlated with JA and DEGs in three metabolism pathways. These revealed key molecules, and vital pathways provided more comprehensive insight for the AR formation mechanism of E. ulmoides and other plants.

Discovery of Nanosota-2, -3, and -4 as super potent and broad-spectrum therapeutic nanobody candidates against COVID-19.

IMPORTANCE: The COVID-19 pandemic exposed limitations of conventional antibodies as therapeutics, including high cost, limited potency, ineffectiveness against new viral variants, and primary reliance on injection-only delivery. Nanobodies are single-domain antibodies with therapeutic potentials. We discovered three anti-SARS-CoV-2 nanobodies, named Nanosota-2, -3, and -4, from an immunized alpaca. Nanosota-2 is super potent against prototypic SARS-CoV-2, Nanosota-3 is highly potent against the omicron variant, and Nanosota-4 is effective against both SARS-CoV-1 and SARS-CoV-2. In addition to their super potency and combined broad antiviral spectrum, these nanobodies are cost-effective, can be easily adapted to new viral variants through phage display, and can potentially be administered as inhalers. The Nanosota series are powerful therapeutic candidates to combat circulating SARS-CoV-2 and prepare for possible future coronavirus pandemics.

Glycosylated Delta-receptor-binding domain mucosal vaccine elicits broadly neutralizing antibodies with protection against SARS-CoV-2 challenge.

Mucosal COVID-19 vaccines are needed to block SARS-CoV-2 infection at the mucosal site. Intranasal delivery of a glycosylated Delta variant receptor-binding domain (Delta-RBD) mucosal vaccine elicited potent and balanced systemic antibody titers comparable to those induced by the intramuscular injection of the same vaccine or Omicron-S subunit vaccine, as well as high mucosal IgA antibody responses. It elicited broadly neutralizing antibodies against the original SARS-CoV-2 strain, Delta and Omicron BA1/BA2 variants, completely protecting transgenic mice from lethal challenge with a Delta variant, including complete absence of weight loss. Of note, intramuscular priming with the Omicron-S protein followed by intranasal boosting with the Delta-RBD protein improved the vaccine's ability to generate broad-spectrum neutralizing antibodies against recent BA5 and XBB Omicron variants. Overall, this vaccine has the potential to prevent the SARS-CoV-2 infection of the respiratory mucosa, while the i.m. priming and i.n. boosting vaccination strategy may offer protection against known and emerging SARS-CoV-2 variants.

MERS-CoV RBD-mRNA vaccine induces potent and broadly neutralizing antibodies with protection against MERS-CoV infection.

Middle East respiratory syndrome coronavirus (MERS-CoV), a highly pathogenic coronavirus in the same Betacoronavirus genus and Coronaviridae family as SARS-CoV-2, continues to post a threat to human health. Mortality remains high; therefore, there is a need to develop effective vaccines to prevent MERS-CoV infection. The receptor-binding domain (RBD) within the MERS-CoV spike (S) protein is a critical vaccine target. The latest mRNA technology has enabled rapid development of much-needed vaccines with high efficiency and scalable manufacturing capacity. Here, we designed a mRNA vaccine encoding the RBD of MERS-CoV S protein (RBD-mRNA) and evaluated its immunogenicity and protective efficacy in a mouse model. The data showed that nucleoside-modified RBD-mRNA, but not RBD-mRNA lacking the nucleoside modification, was stable and elicited broadly and durable neutralizing antibody and cellular immune responses, which neutralized the original strain and multiple MERS-CoV variants. Among all immunization routes tested, the intradermal route was appropriate for this RBD-mRNA to induce strong B-cell responses and the highest neutralizing antibody titers. Importantly, injection of nucleoside-modified RBD-mRNA through the intradermal route protected immunized mice against challenge with MERS-CoV. This protection correlated with serum neutralizing antibody titers. Overall, we have developed an effective MERS-CoV RBD-based mRNA vaccine (with potential for further development) that prevents infection by divergent strains of MERS-CoV.

Allocation strategy of medical supplies during a public health emergency: a tripartite evolutionary game perspective.

Ensuring the rational and orderly circulation of medical supplies during a public health emergency is crucial to quickly containing the further spread of the epidemic and restoring the order of rescue and treatment. However, due to the shortage of medical supplies, there are challenges to rationalizing the allocation of critical medical supplies among multiple parties with conflicting interests. In this paper, a tripartite evolutionary game model is constructed to study the allocation of medical supplies in the rescue environment of public health emergencies under conditions of incomplete information. The game's players include Government-owned Nonprofit Organizations (GNPOs), hospitals, and the government. By analyzing the equilibrium of the tripartite evolutionary game, this paper makes an in-depth study on the optimal allocation strategy of medical supplies. The findings indicate that: (1) the hospital should reasonably increase its willingness to accept the allocation plan of medical supplies, which can help medical supplies allocate more scientifically. (2) The government should design a reasonable reward and punishment mechanism to ensure the rational and orderly circulation of medical supplies, which can reduce the interference of GNPOs and hospitals in the allocation process of medical supplies. (3) Higher authorities should strengthen the supervision of the government and the accountability for loose supervision. The findings of this research can guide the government in promoting better circulation of medical supplies during public health emergencies by formulating more reasonable allocation schemes of emergency medical supplies, as well as incentives and penalties. At the same time, for GNPOs with limited emergency medical supplies, the equal allocation of emergency supplies is not the optimal solution to improve the efficiency of emergency relief, and it is simpler to achieve the goal of maximizing social benefits by allocating limited emergency resources to the demand points that match the degree of urgency. For example, in Corona Virus Disease 2019, emergency medical supplies should be prioritized for allocation to government-designated fever hospitals that are have a greater need for medical supplies and greater treatment capacity.

Advances in SARS-CoV-2 receptor-binding domain-based COVID-19 vaccines.

INTRODUCTION: The Coronavirus Disease 2019 (COVID-19) pandemic has caused devastating human and economic costs. Vaccination is an important step in controlling the pandemic. Severe acute respiratory coronavirus-2 (SARS-CoV-2), the causative agent of COVID-19, infects cells by binding a cellular receptor through the receptor-binding domain (RBD) within the S1 subunit of the spike (S) protein. Viral entry and membrane fusion are mediated by the S2 subunit. AREAS COVERED: SARS-CoV-2 S protein, particularly RBD, serves as an important target for vaccines. Here we review the structure and function of SARS-CoV-2 S protein and its RBD, summarize current COVID-19 vaccines targeting the RBD, and outline potential strategies for improving RBD-based vaccines. Overall, this review provides important information that will facilitate rational design and development of safer and more effective COVID-19 vaccines. EXPERT OPINION: The S protein of SARS-CoV-2 harbors numerous mutations, mostly in the RBD, resulting in multiple variant strains. Although many COVID-19 vaccines targeting the RBD of original virus strain (and previous variants) can prevent infection of these strains, their ability against recent dominant variants, particularly Omicron and its offspring, is significantly reduced. Collective efforts are needed to develop effective broad-spectrum vaccines to control current and future variants that have pandemic potential.

The chromosome-level genome of Eucommia ulmoides provides insights into sex differentiation and α-linolenic acid biosynthesis.

Eucommia ulmoides Oliver is a typical dioecious plant endemic to China that has great medicinal and economic value. Here, we report a high-quality chromosome-level female genome of E. ulmoides obtained by PacBio and Hi-C technologies. The size of the female genome assembly was 1.01 Gb with 17 pseudochromosomes and 31,665 protein coding genes. In addition, Hi-C technology was used to reassemble the male genome released in 2018. The reassembled male genome was 1.24 Gb with the superscaffold N50 (48.30 Mb), which was increased 25.69 times, and the number of predicted genes increased by 11,266. Genome evolution analysis indicated that E. ulmoides has undergone two whole-genome duplication events before the divergence of female and male, including core eudicot γ whole-genome triplication event (γ-WGT) and a recent whole genome duplication (WGD) at approximately 27.3 million years ago (Mya). Based on transcriptome analysis, EuAP3 and EuAG may be the key genes involved in regulating the sex differentiation of E. ulmoides. Pathway analysis showed that the high expression of ω-3 fatty acid desaturase coding gene EU0103017 was an important reason for the high α-linolenic acid content in E. ulmoides. The genome of female and male E. ulmoides presented here is a valuable resource for the molecular biological study of sex differentiation of E. ulmoides and also will provide assistance for the breeding of superior varieties.

Effective vaccination strategy using SARS-CoV-2 spike cocktail against Omicron and other variants of concern.

The SARS-CoV-2 Omicron variant harbors more than 30 mutations in its spike (S) protein. Circulating Omicron subvariants, particularly BA5 and other variants of concern (VOCs), show increased resistance to COVID-19 vaccines that target the original S protein, calling for an urgent need for effective vaccines to prevent multiple SARS-CoV-2 VOCs. Here, we evaluated the neutralizing activity and protection conferred by a BA1-S subunit vaccine when combined with or used as booster doses after, administration of wild-type S protein (WT-S). A WT-S/BA1-S cocktail, or WT-S prime and BA1-S boost, induced significantly higher neutralizing antibodies against pseudotyped Omicron BA1, BA2, BA2.12.1, and BA5 subvariants, and similar or higher neutralizing antibodies against the original SARS-CoV-2, than the WT-S protein alone. The WT-S/BA1-S cocktail also elicited higher or significantly higher neutralizing antibodies than the WT-S-prime-BA1-S boost, WT-S alone, or BA1-S alone against pseudotyped SARS-CoV-2 Alpha, Beta, Gamma, and Delta VOCs, and SARS-CoV, a closely related beta-coronavirus using the same receptor as SARS-CoV-2 for viral entry. By contrast, WT-S or BA1-S alone failed to induce potent neutralizing antibodies against all these viruses. Similar to the WT-S-prime-BA1-S boost, the WT-S/BA1-S cocktail completely protected mice against the lethal challenge of a Delta variant with negligible weight loss. Thus, we have identified an effective vaccination strategy that elicits potent, broadly, and durable neutralizing antibodies against circulating SARS-CoV-2 Omicron subvariants, other VOCs, original SARS-CoV-2, and SARS-CoV. These results will provide useful guidance for developing efficacious vaccines that inhibit current and future SARS-CoV-2 variants to control the COVID-19 pandemic.

Correction: A Truncated Receptor-Binding Domain of MERS-CoV Spike Protein Potently Inhibits MERS-CoV Infection and Induces Strong Neutralizing Antibody Responses: Implication for Developing Therapeutics and Vaccines.

[This corrects the article DOI: 10.1371/journal.pone.0081587.].

mRNA vaccines elicit potent neutralization against multiple SARS-CoV-2 omicron subvariants and other variants of concern.

SARS-CoV-2 variants of concern (VOCs) have shown resistance to vaccines targeting the original virus strain. An mRNA vaccine encoding the spike protein of Omicron BA1 (BA1-S-mRNA) was designed, and its neutralizing activity, with or without the original receptor-binding domain (RBD)-mRNA, was tested against SARS-CoV-2 VOCs. First-dose of BA1-S-mRNA followed by two-boosts of RBD-mRNA elicited potent neutralizing antibodies (nAbs) against pseudotyped and authentic original SARS-CoV-2; pseudotyped Omicron BA1, BA2, BA2.12.1 and BA5 subvariants, and Alpha, Beta, Gamma and Delta VOCs; authentic Omicron BA1 subvariant and Delta VOC. By contrast, other vaccination strategies, including RBD-mRNA first-dose plus BA1-S-mRNA two-boosts, RBD-mRNA or BA1-S-mRNA three-doses, or their combinations, failed to elicit high nAb titers against all of these viruses. Overall, this vaccination strategy was effective for inducing broadly and potent nAbs against multiple SARS-CoV-2 VOCs, particularly Omicron BA5, and may guide the rational design of next-generation mRNA vaccines with greater efficacy against future variants.

A Glycosylated RBD Protein Induces Enhanced Neutralizing Antibodies against Omicron and Other Variants with Improved Protection against SARS-CoV-2 Infection.

SARS-CoV-2 has mutated frequently since its first emergence in 2019. Numerous variants, including the currently emerging Omicron variant, have demonstrated high transmissibility or increased disease severity, posing serious threats to global public health. This study describes the identification of an immunodominant non-neutralizing epitope on SARS-CoV-2 receptor-binding domain (RBD). A subunit vaccine against this mutant RBD, constructed by masking this epitope with a glycan probe, did not significantly affect RBD's receptor-binding affinity or antibody-binding affinity, or its ability to induce antibody production. However, this vaccine enhanced the neutralizing activity of this RBD and its protective efficacy in immunized mice. Specifically, this vaccine elicited significantly higher-titer neutralizing antibodies than the prototypic RBD protein against Alpha (B.1.1.7 lineage), Beta (B.1.351 lineage), Gamma (P.1 lineage), and Epsilon (B.1.427 or B.1.429 lineage) variant pseudoviruses containing single or combined mutations in the spike (S) protein, albeit the neutralizing antibody titers against some variants were slightly lower than against original SARS-CoV-2. This vaccine also significantly improved the neutralizing activity of the prototypic RBD against pseudotyped and authentic Delta (B.1.617.2 lineage) and Omicron (B.1.1.529 lineage) variants, although the neutralizing antibody titers were lower than against original SARS-CoV-2. In contrast to the prototypic RBD, the mutant RBD completely protected human ACE2 (hACE2)-transgenic mice from lethal challenge with a prototype SARS-CoV-2 strain and a Delta variant without weight loss. Overall, these findings indicate that this RBD vaccine has broad-spectrum activity against multiple SARS-CoV-2 variants, as well as the potential to be effective and have improved efficacy against Omicron and other pandemic variants. IMPORTANCE Several SARS-CoV-2 variants have shown increased transmissibility, calling for a need to develop effective vaccines with broadly neutralizing activity against multiple variants. This study identified a non-neutralizing epitope on the receptor-binding domain (RBD) of SARS-CoV-2 spike protein, and further shielded it with a glycan probe. A subunit vaccine based on this mutant RBD significantly enhanced the ability of prototypic RBD against multiple SARS-CoV-2 variants, including the Delta and Omicron strains, although the neutralizing antibody titers against some of these variants were lower than those against original SARS-CoV-2. This mutant vaccine also enhanced the protective efficacy of the prototypic RBD vaccine against SARS-CoV-2 infection in immunized animals. In conclusion, this study identified an engineered RBD vaccine against Omicron and other SARS-CoV-2 variants that induced stronger neutralizing antibodies and protection than the original RBD vaccine. It also highlights the need to improve the effectiveness of current COVID-19 vaccines to prevent pandemic SARS-CoV-2 variants.

A gossypol derivative effectively protects against Zika and dengue virus infection without toxicity.

BACKGROUND: Zika virus (ZIKV) and dengue virus (DENV) cause microcephaly and dengue hemorrhagic fever, respectively, leading to severe problems. No effective antiviral agents are approved against infections of these flaviviruses, calling for the need to develop potent therapeutics. We previously identified gossypol as an effective inhibitor against ZIKV and DENV infections, but this compound is toxic and not suitable for in vivo treatment. RESULTS: In this study, we showed that gossypol derivative ST087010 exhibited potent and broad-spectrum in vitro inhibitory activity against infections of at least ten ZIKV strains isolated from different hosts, time periods, and countries, as well as DENV-1-4 serotypes, and significantly reduced cytotoxicity compared to gossypol. It presented broad-spectrum in vivo protective efficacy, protecting ZIKV-infected Ifnar1-/- mice from lethal challenge, with increased survival and reduced weight loss. Ifnar1-/- mice treated with this gossypol derivative decreased viral titers in various tissues, including the brain and testis, after infection with ZIKV at different human isolates. Moreover, ST087010 potently blocked ZIKV vertical transmission in pregnant Ifnar1-/- mice, preventing ZIKV-caused fetal death, and it was safe for pregnant mice and their pups. It also protected DENV-2-challenged Ifnar1-/- mice against viral replication by reducing the viral titers in the brain, kidney, heart, and sera. CONCLUSIONS: Overall, our data indicate the potential for further development of this gossypol derivative as an effective and safe broad-spectrum therapeutic agent to treat ZIKV and DENV diseases.

RBD-mRNA vaccine induces broadly neutralizing antibodies against Omicron and multiple other variants and protects mice from SARS-CoV-2 challenge.

Multiple SARS-CoV-2 variants are identified with higher rates of transmissibility or greater disease severity. Particularly, recent emergence of Omicron variant with rapid human-to-human transmission posts new challenges to the current prevention strategies. In this study, following vaccination with an mRNA vaccine encoding SARS-CoV-2 receptor-binding domain (RBD-mRNA), we detected serum antibodies that neutralized pseudoviruses expressing spike (S) protein harboring single or multiple mutations, as well as authentic SARS-CoV-2 variants, and evaluated its protection against SARS-CoV-2 infection. The vaccine induced durable antibodies that potently neutralized prototypic strain and B.1.1.7 lineage variant pseudoviruses containing N501Y or D614G mutations alone or in combination with a N439K mutation (B.1.258 lineage), with a L452R mutation (B.1.427 or B.1.429 lineage), or a L452R-E484Q double mutation (B.1.617.1 variant), although neutralizing activity against B.1.1.7 lineage variant containing 10 amino acid changes in the S protein was slightly reduced. The RBD-mRNA-induced antibodies exerted moderate neutralization against authentic B.1.617.2 and B.1.1.529 variants, and pseudotyped B.1.351 and P.1 lineage variants containing K417N/T, E484K, and N501Y mutations, the B.1.617.2 lineage variant harboring L452R, T478K, and P681R mutations, and the B.1.1.529 lineage variant containing 38 mutations in the S protein. Particularly, RBD-mRNA vaccine completely protected mice from challenge with a virulent mouse-adapted SARS-CoV-2 variant. Among these lineages, B.1.1.7, B.1.351, P.1, B.1.617.2, and B.1.1.529 belong to Alpha, Beta, Gamma, Delta, and Omicron variants, respectively. Our observations reveal that RBD-mRNA vaccine is promising and highlights the need to design novel vaccines with improved neutralization against current and future pandemic SARS-CoV-2 variants.

Whole genome re-sequencing reveals the genetic diversity and evolutionary patterns of Eucommia ulmoides.

Eucommia ulmoides (E. ulmoides) is a deciduous perennial tree belonging to the order Garryales, and is known as "living fossil" plant, along with ginkgo (Ginkgo biloba), metaspaca (Metasequoia glyptostroboides) and dove tree (Davidia involucrata Baill). However, the genetic diversity and population structure of E. ulmoides are still  ambiguous nowdays. In this study, we re-sequenced the genomes of 12 E. ulmoides accessions from different major climatic geography regions in China to elucidate the genetic diversity, population structure and evolutionary pattern. By integration of phylogenetic analysis, principal component analysis and population structure analysis based on a number of high-quality SNPs, a total of 12 E. ulmoides accessions were clustered into four different groups. This result is consistent with their geographical location except for group samples from Shanghai and Hunan province. E. ulmoides accessions from Hunan province exhibited a closer genetic relationship with E. ulmoides accessions from Shanghai in China compared with other regions, which is also supported by the result of population structure analyses. Genetic diversity analysis further revealed that E. ulmoides samples in Shanghai and Hunan province were with higher genetic diversity than those in other regions in this study. In addition, we treated the E. ulmoides materials from Shanghai and Hunan province as group A, and the other materials from other places as group B, and then analyzed the evolutionary pattern of E. ulmoides. The result showed the significant differentiation (Fst = 0.1545) between group A and group B. Some candidate highly divergent genome regions were identified in group A by selective sweep analyses, and the function analysis of candidate genes in these regions showed that biological regulation processes could be correlated with the Eu-rubber biosynthesis. Notably, nine genes were identified from selective sweep regions. They were involved in the Eu-rubber biosynthesis and expressed in rubber containing tissues. The genetic diversity research and evolution model of E. ulmoides were preliminarily explored in this study, which laid the foundation for the protection of germplasm resources and the development and utilization of multipurpose germplasm resources in the future.

Recent advances in nanotechnology-based COVID-19 vaccines and therapeutic antibodies.

COVID-19 has caused a global pandemic and millions of deaths. It is imperative to develop effective countermeasures against the causative viral agent, SARS-CoV-2 and its many variants. Vaccines and therapeutic antibodies are the most effective approaches for preventing and treating COVID-19, respectively. SARS-CoV-2 enters host cells through the activities of the virus-surface spike (S) protein. Accordingly, the S protein is a prime target for vaccines and therapeutic antibodies. Dealing with particles with dimensions on the scale of nanometers, nanotechnology has emerged as a critical tool for rapidly designing and developing safe, effective, and urgently needed vaccines and therapeutics to control the COVID-19 pandemic. For example, nanotechnology was key to the fast-track approval of two mRNA vaccines for their wide use in human populations. In this review article, we first explore the roles of nanotechnology in battling COVID-19, including protein nanoparticles (for presentation of protein vaccines), lipid nanoparticles (for formulation with mRNAs), and nanobodies (as unique therapeutic antibodies). We then summarize the currently available COVID-19 vaccines and therapeutics based on nanotechnology.

Advances in mRNA and other vaccines against MERS-CoV.

Middle East respiratory syndrome coronavirus (MERS-CoV) is a highly pathogenic human coronavirus (CoV). Belonging to the same beta-CoV genus as severe acute respiratory syndrome coronavirus-1 (SARS-CoV-1) and SARS-CoV-2, MERS-CoV has a significantly higher fatality rate with limited human-to-human transmissibility. MERS-CoV causes sporadic outbreaks, but no vaccines have yet been approved for use in humans, thus calling for continued efforts to develop effective vaccines against this important CoV. Similar to SARS-CoV-1 and SARS-CoV-2, MERS-CoV contains 4 structural proteins, among which the surface spike (S) protein has been used as a core component in the majority of currently developed MERS-CoV vaccines. Here, we illustrate the importance of the MERS-CoV S protein as a key vaccine target and provide an update on the currently developed MERS-CoV vaccines, including those based on DNAs, proteins, virus-like particles or nanoparticles, and viral vectors. Additionally, we describe approaches for designing MERS-CoV mRNA vaccines and explore the role and importance of naturally occurring pseudo-nucleosides in the design of effective MERS-CoV mRNA vaccines. This review also provides useful insights into designing and evaluating mRNA vaccines against other viral pathogens.