The mouse paramyxovirus Sendai, which is capable of limited replication in human bronchial epithelial cells without causing disease, is well suited for the development of vector-based intranasal ...vaccines against respiratory infections, including SARS-CoV-2. Using the Moscow strain of the Sendai virus, we developed a vaccine construct, Sen-Sdelta(M), which expresses the full-length spike (S) protein of the SARS-CoV-2 delta variant. A single intranasal delivery of Sen-Sdelta(M) to Syrian hamsters and BALB/c mice induced high titers of virus-neutralizing antibodies specific to the SARS-CoV-2 delta variant. A significant T-cell response, as determined by IFN-γ ELISpot and ICS methods, was also demonstrated in the mouse model. Mice and hamsters vaccinated with Sen-Sdelta(M) were well protected against SARS-CoV-2 challenge. The viral load in the lungs and nasal turbinates, measured by RT-qPCR and TCID50 assay, decreased dramatically in vaccinated groups. The most prominent effect was revealed in a highly sensitive hamster model, where no tissue samples contained detectable levels of infectious SARS-CoV-2. These results indicate that Sen-Sdelta(M) is a promising candidate as a single-dose intranasal vaccine against SARS-CoV-2, including variants of concern.
Introduction. Intranasal vaccination using live vector vaccines based on non-pathogenic or slightly pathogenic viruses is the one of the most convenient, safe and effective ways to prevent ...respiratory infections, including COVID-19. Sendai virus is the best suited for this purpose, since it is respiratory virus and is capable of limited replication in human bronchial epithelial cells without causing disease.
The aim of the work is to design and study the vaccine properties of recombinant Sendai virus, Moscow strain, expressing secreted receptor-binding domain of SARS-CoV-2 Delta strain S protein (RBDdelta) during a single intranasal immunization.
Materials and methods. Recombinant Sendai virus carrying insertion of RBDdelta transgene between P and M genes was constructed using reverse genetics and synthetic biology methods. Expression of RBDdelta was analyzed by Western blot. Vaccine properties were studied in two models: Syrian hamsters and BALB/c mice. Immunogenicity was evaluated by ELISA and virus-neutralization assays. Protectiveness was assessed by quantitation of SARS-CoV-2 RNA in RT-PCR and histological analysis of the lungs.
Results. Based on Sendai virus Moscow strain, a recombinant Sen-RBDdelta(M) was constructed that expressed a secreted RBDdelta immunologically identical to natural SARS-CoV-2 protein. A single intranasal administration of Sen-RBDdelta(M) to hamsters and mice significantly, by 15 and 107 times, respectively, reduced replicative activity of SARS-CoV-2 in lungs of animals, preventing the development of pneumonia. An effective induction of virus-neutralizing antibodies has also been demonstrated in mice.
Conclusion. Sen-RBDdelta(M) is a promising vaccine construct against SARS-CoV-2 infection and has a protective properties even after a single intranasal introduction.
Although the primary structures of class 1 polypeptide
release factors (RF1 and RF2 in prokaryotes, eRF1 in eukaryotes)
are known, the molecular basis by which they function in
translational ...termination remains obscure. Because all
class 1 RFs promote a stop-codon-dependent and ribosome-dependent
hydrolysis of peptidyl-tRNAs, one may anticipate that this
common function relies on a common structural motif(s).
We have compared amino acid sequences of the available
class 1 RFs and found a novel, common, unique, and strictly
conserved GGQ motif that should be in a loop (coil) conformation
as deduced by programs predicting protein secondary structure.
Site-directed mutagenesis of the human eRF1 as a representative
of class 1 RFs shows that substitution of both glycyl residues
in this motif, G183 and G184, causes complete inactivation
of the protein as a release factor toward all three stop
codons, whereas two adjacent amino acid residues, G181
and R182, are functionally nonessential. Inactive human
eRF1 mutants compete in release assays with wild-type eRF1
and strongly inhibit their release activity. Mutations
of the glycyl residues in this motif do not affect another
function, the ability of eRF1 together with the ribosome
to induce GTPase activity of human eRF3, a class 2 RF.
We assume that the novel highly conserved GGQ motif is
implicated directly or indirectly in the activity of class
1 RFs in translation termination.
Most of the live vaccine doses of vaccinia virus donated to the Intensified Smallpox Eradication Programme after 1971 were prepared using the L-IVP strain. A mixture of three clones of the L-IVP ...strain was sequenced using MySEQ. Consensus sequence similarity with the vaccinia virus Lister strain is 99.5%.
Accurate measurement of tumor size and margins is crucial for successful oncotherapy. In the last decade, non-invasive imaging modalities, including optical imaging using non-radioactive substrates, ...deep-tissue imaging with radioactive substrates, and magnetic resonance imaging have been developed. Reporter genes play the most important role among visualization tools; their expression in tumors and metastases makes it possible to track changes in the tumor growth and gauge therapy effectiveness. Oncolytic viruses are often chosen as a vector for delivering reporter genes into tumor cells, since oncolytic viruses are tumor-specific, meaning that they infect and lyse tumor cells without damaging normal cells. The choice of reporter transgenes for genetic modification of oncolytic viruses depends on the study objectives and imaging methods used. Optical imaging techniques are suitable for in vitro studies and small animal models, while deep-tissue imaging techniques are used to evaluate virotherapy in large animals and humans. For optical imaging, transgenes of fluorescent proteins, luciferases, and tyrosinases are used; for deep-tissue imaging, the most promising transgene is the sodium/iodide symporter (NIS), which ensures an accumulation of radioactive isotopes in virus-infected tumor cells. Currently, NIS is the only reporter transgene that has been shown to be effective in monitoring tumor virotherapy not only in preclinical but also in clinical studies.
The use of a scanning flow cytometer (SFC) to study the evolution of monomers, dimers and higher multimers of latex particles at the initial stage of the immunoagglutination is described. The SFC can ...measure the light-scattering pattern (indicatrix) of an individual particle over an angular range of 10–60°. A comparison of the experimentally measured and theoretically calculated indicatrices allows one to discriminate different types of latex particles (i.e. monomers, dimers, etc.) and, therefore, to study the evolution of immunoagglutination process. Validity of the approach was verified by simultaneous measurements of light-scattering patterns and fluorescence from individual polymer particles. Immunoagglutination was initiated by mixing bovine serum albumin (BSA)-covered latex particles (of 1.8 μm in diameter) with anti-BSA IgG. The analysis of experimental data was performed on the basis of a mathematical model of diffusion-limited immunoagglutination aggregation with a steric factor. The steric factor was determined by the size and the number of binding sites on the surface of a latex particle. The obtained data are in good agreement with the proposed mathematical modeling.
The use of a scanning flow cytometer (SFC) to study the evolution of monomers, dimers and higher multimers of latex particles at the initial stage of the immunoagglutination is described. The SFC can ...measure the light-scattering pattern (indicatrix) of an individual particle over an angular range of 10-60 deg. A comparison of the experimentally measured and theoretically calculated indicatrices allows one to discriminate different types of latex particles (i.e. monomers, dimers, etc.) and, therefore, to study the evolution of immunoagglutination process. Validity of the approach was verified by simultaneous measurements of light-scattering patterns and fluorescence from individual polymer particles. Immunoagglutination was initiated by mixing bovine serum albumin (BSA)-covered latex particles (of 1.8 um in diameter) with anti-BSA IgG. The analysis of experimental data was performed on the basis of a mathematical model of diffusion-limited immunoagglutination aggregation with a steric factor. The steric factor was determined by the size and the number of binding sites on the surface of a latex particle. The obtained data are in good agreement with the proposed mathematical modeling.
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