A Newly Identified Spike Protein Targeted Linear B-Cell Epitope Based Dissolvable Microneedle Array Successfully Eliciting Neutralizing Activities against SARS-CoV-2 Wild-Type Strain in Mice.

Vaccination is a cost-effective medical intervention. Inactivated whole virusor large protein fragments-based severe acute respiratory syndrome coronavirus (SARS-CoV-2) vaccines have high unnecessary antigenic load to induce allergenicity and/orreactogenicity, which can be avoided by peptide vaccines of short peptide fragments that may induce highly targeted immune response. However, epitope identification and peptide delivery remain the major obstacles in developing peptide vaccines. Here, a multi-source data integrated linear B-cell epitope screening strategy is presented and a linear B-cell epitope enriched hotspot region is identified in Spike protein, from which a monomeric peptide vaccine (Epitope25) is developed and applied to subcutaneously immunize wildtype BALB/c mice. Indirect ELISA assay reveals specific and dose-dependent binding between Epitope25 and serum IgG antibodies from immunized mice. The neutralizing activity of sera from vaccinated mice is validated by pseudo and live SARS-CoV-2 wild-type strain neutralization assays. Then a dissolvable microneedle array (DMNA) is developed to pain-freely deliver Epitope25. Compared with intramuscular injection, DMNA and subcutaneous injection elicit neutralizing activities against SARS-CoV-2 wild-type strain as demonstrated by live SARS-CoV-2 virus neutralization assay. No obvious damages are found in major organs of immunized mice. This study may lay the foundation for developing linear B-cell epitope-based vaccines against SARS-CoV-2.

Evolutionary plasticity of zoonotic porcine Deltacoronavirus (PDCoV): genetic characteristics and geographic distribution.

The emergence and rapid spread of the acute respiratory syndrome coronavirus-2 have confirmed that animal coronaviruses represent a potential zoonotic source. Porcine deltacoronavirus is a worldwide evolving enteropathogen of swine, detected first in Hong Kong, China, before its global identification. Following the recent detection of PDCoV in humans, we attempted in this report to re-examine the status of PDCoV phylogenetic classification and evolutionary characteristics. A dataset of 166 complete PDCoV genomes was analyzed using the Maximum Likelihood method in IQ-TREE with the best-fitting model GTR + F + I + G4, revealing two major genogroups (GI and GII), with further seven and two sub-genogroups, (GI a-g) and (GII a-b), respectively. PDCoV strains collected in China exhibited the broadest genetic diversity, distributed in all subgenotypes. Thirty-one potential natural recombination events were identified, 19 of which occurred between China strains, and seven involved at least one China strain as a parental sequence. Importantly, we identified a human Haiti PDCoV strain as recombinant, alarming a possible future spillover that could become a critical threat to human health. The similarity and recombination analysis showed that PDCoV spike ORF is highly variable compared to ORFs encoding other structural proteins. Prediction of linear B cell epitopes of the spike glycoprotein and the 3D structural mapping of amino acid variations of two representative strains of GI and GII showed that the receptor-binding domain (RBD) of spike glycoprotein underwent a significant antigenic drift, suggesting its contribution in the genetic diversity and the wider spread of PDCoV.

Predicting Antigenic Peptides from Rocio Virus NS1 Protein for Immunodiagnostic Testing Using Immunoinformatics and Molecular Dynamics Simulation.

The mosquito-borne disease caused by the Rocio virus is a neglected threat, and new immune inputs for serological testing are urgently required for diagnosis in low-resource settings and epidemiological surveillance. We used in silico approaches to identify a specific antigenic peptide (p_ROCV2) in the NS1 protein of the Rocio virus that was theoretically predicted to be stable and exposed on its surface, where it demonstrated key properties allowing it to interact with antibodies. These findings related to the molecular dynamics of this peptide provide important insights for advancing diagnostic platforms and investigating therapeutic alternatives.

Rabbit Monoclonal Antibody Specifically Recognizing a Linear Epitope in the RBD of SARS-CoV-2 Spike Protein.

To date, SARS-CoV-2 pandemic has caused more than 188 million infections and 4.06 million deaths worldwide. The receptor-binding domain (RBD) of the SARS-CoV-2 spike protein has been regarded as an important target for vaccine and therapeutics development because it plays a key role in binding the human cell receptor ACE2 that is required for viral entry. However, it is not easy to detect RBD in Western blot using polyclonal antibody, suggesting that RBD may form a complicated conformation under native condition and bear rare linear epitope. So far, no linear epitope on RBD is reported. Thus, a monoclonal antibody (mAb) that recognizes linear epitope on RBD will become valuable. In the present study, an RBD-specific rabbit antibody named 9E1 was isolated from peripheral blood mononuclear cells (PBMC) of immunized rabbit by RBD-specific single B cell sorting and mapped to a highly conserved linear epitope within twelve amino acids 480CNGVEGFNCYFP491 on RBD. 9E1 works well in Western blot on S protein and immunohistochemistry on the SARS-CoV-2 infected tissue sections. The results demonstrated that 9E1 can be used as a useful tool for pathological and functional studies of SARS-CoV-2.