OvHV-2 Glycoprotein B Delivered by a Recombinant BoHV-4 Is Immunogenic and Induces Partial Protection against Sheep-Associated Malignant Catarrhal Fever in a Rabbit Model

An efficacious vaccine for sheep-associated malignant catarrhal fever (SA-MCF) is important for the livestock industry. Research towards SA-MCF vaccine development is hindered by the absence of culture systems to propagate the causative agent, ovine herpesvirus-2 (OvHV-2), which means its genome cannot be experimentally modified to generate an attenuated vaccine strain. Alternative approaches for vaccine development are needed to deliver OvHV-2 antigens. Bovine herpesvirus 4 (BoHV-4) has been evaluated as a vaccine vector for several viral antigens with promising results. In this study, we genetically engineered BoHV-4 to express OvHV-2 glycoprotein B (gB) and evaluated its efficacy as an SA-MCF vaccine using a rabbit model. The construction of a viable recombinant virus (BoHV-4-AΔTK-OvHV-2-gB) and confirmation of OvHV-2 gB expression were performed in vitro. The immunization of rabbits with BoHV-4-AΔTK-OvHV-2-gB elicited strong humoral responses to OvHV-2 gB, including neutralizing antibodies. Following intra-nasal challenge with a lethal dose of OvHV-2, 42.9% of the OvHV-2 gB vaccinated rabbits were protected against SA-MCF, while all rabbits in the mock-vaccinated group succumbed to SA-MCF. Overall, OvHV-2 gB delivered by the recombinant BoHV-4 was immunogenic and partly protective against SA-MCF in rabbits. These are promising results towards an SA-MCF vaccine; however, improvements are needed to increase protection rates.

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Replacement of Glycoprotein B in Alcelaphine Herpesvirus 1 by Its Ovine Herpesvirus 2 Homolog : Implications in Vaccine Development for Sheep-Associated Malignant Catarrhal Fever

mSphere ◽

10.1128/msphere.00108-16 ◽

2016 ◽

Vol 1 (4) ◽

Cited By ~ 3

Author(s):

Cristina W. Cunha ◽

Naomi S. Taus ◽

Benjamin G. Dewals ◽

Alain Vanderplasschen ◽

Donald P. Knowles ◽

...

Keyword(s):

Diagnostic Tool ◽

Vaccine Development ◽

Vaccine Candidate ◽

Chimeric Virus ◽

Glycoprotein B ◽

Malignant Catarrhal Fever ◽

Vaccine Target ◽

Herpesvirus 1

ABSTRACT Vaccine development is a top priority in malignant catarrhal fever (MCF) research. In the case of sheep-associated MCF (SA-MCF) caused by ovine herpesvirus 2 (OvHV-2), progress toward this objective has been hindered by the absence of methods to attenuate or modify the virus, since it cannot be propagated in vitro. As an alternative for vaccine development, in this study, we tested the hypothesis that one of the SA-MCF vaccine candidate targets, OvHV-2 glycoprotein B (gB), could be expressed by a nonpathogenic alcelaphine herpesvirus 1 (AlHV-1) and then evaluated the potential of the AlHV-1/OvHV-2 chimera to be used as a vaccine and a diagnostic tool. Vaccine development is a top priority in malignant catarrhal fever (MCF) research. In the case of sheep-associated MCF (SA-MCF) caused by ovine herpesvirus 2 (OvHV-2), progress toward this objective has been hindered by the absence of methods to attenuate or modify the virus, since it cannot be propagated in vitro. As an alternative for vaccine development, in this study, we tested the hypothesis that one of the SA-MCF vaccine candidate targets, OvHV-2 glycoprotein B (gB), could be expressed by a nonpathogenic alcelaphine herpesvirus 1 (AlHV-1) and then evaluated the potential of the AlHV-1/OvHV-2 chimera to be used as a vaccine and a diagnostic tool. The construction and characterization of an AlHV-1/OvHV-2 chimeric virus that is nonpathogenic and expresses an OvHV-2 vaccine target are significant steps toward the development of an SA-MCF vaccine and also provide a valuable means to study OvHV-2 biology.

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A Bacterially-Expressed Recombinant Envelope Protein from Usutu Virus Induces Neutralizing Antibodies in Rabbits

Vaccines ◽

10.3390/vaccines9020157 ◽

2021 ◽

Vol 9 (2) ◽

pp. 157

Author(s):

Kinga Böszörményi ◽

Janet Hirsch ◽

Gwendoline Kiemenyi Kayere ◽

Zahra Fagrouch ◽

Nicole Heijmans ◽

...

Keyword(s):

Envelope Protein ◽

Neutralizing Antibodies ◽

Size Exclusion ◽

Vitro Assay ◽

Usutu Virus ◽

Transmission Electron ◽

Exclusion Chromatography ◽

Efficacious Vaccine ◽

Human Infections

Background: Recently, an emerging flavivirus, Usutu virus (USUV), has caused an epidemic among birds in Europe, resulting in a massive die-off in Eurasian blackbirds. Currently found only in Europe and Africa, it can be envisioned that Usutu virus will follow the path of other flaviviruses, like West Nile virus and Zika virus, and will spread via its mosquito vectors and bird hosts to other parts of the world. Several cases of human infections by Usutu virus have already been published. Anticipating this spread, development of an efficacious vaccine would be highly desirable. Method: This study describes the production in E. coli, purification, and refolding of a partial USUV envelope protein. Prior to immunization, the protein was characterized using size exclusion chromatography, transmission electron microscopy and dynamic light scattering, showing the limited presence of virus-like structures, indicating that the protein solution is probably a mixture of mono and multimeric envelope proteins. Results: Immunizations of two rabbits with the refolded E-protein fraction, mixed with a strong adjuvant, resulted in the generation of neutralizing antibodies, as evidenced in an in vitro assay. Discussion: The way forward towards a subunit vaccine against Usutu virus infection is discussed.

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Effects of Size and Surface Properties of Nanodiamonds on the Immunogenicity of Plant-Based H5 Protein of A/H5N1 Virus in Mice

Nanomaterials ◽

10.3390/nano11061597 ◽

2021 ◽

Vol 11 (6) ◽

pp. 1597

Author(s):

Thuong Thi Ho ◽

Van Thi Pham ◽

Tra Thi Nguyen ◽

Vy Thai Trinh ◽

Tram Vi ◽

...

Keyword(s):

High Pressure ◽

Neutralizing Antibodies ◽

H5n1 Virus ◽

Vaccine Development ◽

Particle Characterization ◽

Innovative Strategy ◽

Specific Igg ◽

Positively Charged

Nanodiamond (ND) has recently emerged as a potential nanomaterial for nanovaccine development. Here, a plant-based haemagglutinin protein (H5.c2) of A/H5N1 virus was conjugated with detonation NDs (DND) of 3.7 nm in diameter (ND4), and high-pressure and high-temperature (HPHT) oxidative NDs of ~40–70 nm (ND40) and ~100–250 nm (ND100) in diameter. Our results revealed that the surface charge, but not the size of NDs, is crucial to the protein conjugation, as well as the in vitro and in vivo behaviors of H5.c2:ND conjugates. Positively charged ND4 does not effectively form stable conjugates with H5.c2, and has no impact on the immunogenicity of the protein both in vitro and in vivo. In contrast, the negatively oxidized NDs (ND40 and ND100) are excellent protein antigen carriers. When compared to free H5.c2, H5.c2:ND40, and H5.c2:ND100 conjugates are highly immunogenic with hemagglutination titers that are both 16 times higher than that of the free H5.c2 protein. Notably, H5.c2:ND40 and H5.c2:ND100 conjugates induce over 3-folds stronger production of both H5.c2-specific-IgG and neutralizing antibodies against A/H5N1 than free H5.c2 in mice. These findings support the innovative strategy of using negatively oxidized ND particles as novel antigen carriers for vaccine development, while also highlighting the importance of particle characterization before use.

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Virus-Neutralizing Monoclonal Antibody Expressed in Milk of Transgenic Mice Provides Full Protection against Virus-Induced Encephalitis

Journal of Virology ◽

10.1128/jvi.75.6.2803-2809.2001 ◽

2001 ◽

Vol 75 (6) ◽

pp. 2803-2809 ◽

Cited By ~ 21

Author(s):

Andreas F. Kolb ◽

Lecia Pewe ◽

John Webster ◽

Stanley Perlman ◽

C. Bruce A. Whitelaw ◽

...

Keyword(s):

Transgenic Mice ◽

Neutralizing Antibodies ◽

Defense Mechanism ◽

Viral Infections ◽

Hepatitis Virus ◽

Recombinant Antibody ◽

Lethal Dose ◽

Neutralizing Antibody ◽

Murine Hepatitis Virus

ABSTRACT Neutralizing antibodies represent a major host defense mechanism against viral infections. In mammals, passive immunity is provided by neutralizing antibodies passed to the offspring via the placenta or the milk as immunoglobulin G and secreted immunoglobulin A. With the long-term goal of producing virus-resistant livestock, we have generated mice carrying transgenes that encode the light and heavy chains of an antibody that is able to neutralize the neurotropic JHM strain of murine hepatitis virus (MHV-JHM). MHV-JHM causes acute encephalitis and acute and chronic demyelination in susceptible strains of mice and rats. Transgene expression was targeted to the lactating mammary gland by using the ovine β-lactoglobulin promoter. Milk from these transgenic mice contained up to 0.7 mg of recombinant antibody/ml. In vitro analysis of milk derived from different transgenic lines revealed a linear correlation between antibody expression and virus-neutralizing activity, indicating that the recombinant antibody is the major determinant of MHV-JHM neutralization in murine milk. Offspring of transgenic and control mice were challenged with a lethal dose of MHV-JHM. Litters suckling nontransgenic dams succumbed to fatal encephalitis, whereas litters suckling transgenic dams were fully protected against challenge, irrespective of whether they were transgenic. This demonstrates that a single neutralizing antibody expressed in the milk of transgenic mice is sufficient to completely protect suckling offspring against MHV-JHM-induced encephalitis.

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Application of Median Lethal Concentration (LC50) of Pathogenic Microorganisms and Their Antigens in Vaccine Development

10.21203/rs.2.23298/v2 ◽

2020 ◽

Author(s):

Saganuwan Alhaji Saganuwan

Keyword(s):

Staphylococcus Aureus ◽

Disease Virus ◽

Vaccine Development ◽

Hepatitis A Virus ◽

Lethal Dose ◽

Vaccine Dose ◽

Foot And Mouth Disease ◽

Bacterial Colony ◽

Mouth Disease Virus

Abstract Objective: Lack of ideal mathematical models to qualify and quantify both pathogenicity, and virulence is a dreadful setback in development of new antimicrobials and vaccines against resistance pathogenic microorganisms.Hence, the modified arithmetical formula of Reed and Muenchhas been integrated with other formulas and used to determine bacterial colony forming unit/ viral concentration, virulence and immunogenicity from LC50s established in the laboratories. Results: Microorganisms’antigens tested are Staphylococcus aureus, Streptococcuspneumonia, Pseudomonas aeruginosa in mice and rat, Edwardsiellaictaluri, Aeromonashydrophila, Aeromonasveronii in fish, New Castle Disease virus in chicken, Sheep Pox Virus, Foot-and-Mouth Disease Virus and Hepatitis A virus in vitro, respectively. The LC50s for the pathogens using different routes of administrations are 1.93 x 103(sheep poxvirus) and 1.75 x 1010 for Staphylococcus aureus (ATCC29213) in rat respectively. Titre index (TI) equals N log10 LC50 and provides protection against lethal dose in graded fashion which translates to protection index. N is the number of vaccine dose that could neutralize the LC50. Hence, parasite inoculum of 103 to 1011could be used as basis for determination of median lethal dose(LD50), LC50 and median bacterial concentrations (BC50)determination, pathogenic dose for immune stimulation should be sought at concentrations less than LC10.

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Determination of Median Lethal Concentrations (Lc50) of Pathogenic Antigens and Their Applications in Vaccine Development

10.21203/rs.2.23298/v1 ◽

2020 ◽

Author(s):

Saganuwan Alhaji Saganuwan

Keyword(s):

Staphylococcus Aureus ◽

Disease Virus ◽

Vaccine Development ◽

Hepatitis A Virus ◽

Lethal Dose ◽

Vaccine Dose ◽

Foot And Mouth Disease ◽

Mouth Disease Virus

Abstract Objective: Lack of ideal mathematical models to qualify and quantify both pathogenicity, and virulence is a dreadful setback in development of new antimicrobials and vaccines against resistance pathogenic microorganisms. Hence, the modified arithmetical formula of Reed and Muench has been integrated with other formulas and used for determination of antigen concentration and parasites inoculums that would kill 50% of test animals (LC50).Results: Microorganisms’ antigens tested are Staphylococcus aureus, Streptococcus pneumonia, Pseudomonas aeruginosa in mice and rat, Edwardsiella ictaluri, Aeromonas hydrophila, Aeromonas veronii in fish, New Castle Disease virus in chicken, Sheep Pox Virus, Foot-and-Mouth Disease Virus and Hepatitis A virus in vitro, respectively. The LC50s for the pathogens using different routes of administrations are 1.93 x 103 (sheep poxvirus) and 1.75 x 1010 for Staphylococcus aureus (ATCC29213) in rat respectively. N is the number of vaccine dose that could neutralize the LC50.Titre index (TI) equals N log10 LC50 and provides protection against lethal dose in graded fashion which translates to protection index. Hence, parasite inoculum of 103 to 1011 could be used as basis for median lethal dose (LD50), LC50 and median bacterial concentrations (BC50) determination, pathogenic dose for immune stimulation should be sought at concentrations less than LC10.

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Chimeric virus-like particles of universal antigen epitopes of coronavirus and phage Qβ coat protein trigger the production of neutralizing antibodies

Current Topics in Medicinal Chemistry ◽

10.2174/1568026621666210618145411 ◽

2021 ◽

Vol 21 ◽

Author(s):

Yukun Guo ◽

Ruizhen Guo ◽

Yingxian Ma ◽

Wenru Chang ◽

Shengli Ming ◽

...

Keyword(s):

Coat Protein ◽

Neutralizing Antibodies ◽

Vaccine Development ◽

Genetic Recombination ◽

Fluorescent Label ◽

Salting Out ◽

Virus Like Particles ◽

Chimeric Vlps ◽

Universal Epitope

Background: Virus-like particles (VLPs) are non-genetic multimeric nanoparticles synthesized through in vitro or in vivo self-assembly of one or more viral structural proteins. Immunogenicity and safety of VLPs make them ideal candidates for vaccine development and efficient nanocarriers for foreign antigens or adjuvants to activate the immune system. Aims: The present study aimed to design and synthesize a chimeric VLP vaccine of the phage Qbeta (Qβ) coat protein presenting the universal epitope of the coronavirus. Methods: The RNA phage Qβ coat protein was designed and synthesized, denoted as Qbeta. The CoV epitope, a universal epitope of coronavirus, was inserted into the C-terminal of Qbeta using genetic recombination, which was designated as Qbeta-CoV. The N-terminal of Qbeta-CoV was successively inserted into the TEV restriction site using mCherry red fluorescent label and modified affinity-purified histidine label 6xHE, which was denoted as HE-Qbeta-CoV. Isopropyl β-D-1-thiogalactopyranoside (IPTG) assessment revealed the expression of Qbeta, Qbeta-CoV, and HE-Qbeta-CoV in the BL21 (DE3) cells. The fusion protein was purified by salting out using ammonium sulfate and affinity chromatography. The morphology of particles was observed using electron microscopy. The female BALB/C mice were immunized intraperitoneally with the Qbeta-CoV and HE-Qbeta-CoV chimeric VLPs vaccines. Their sera were collected for the detection of antibody level and antibody titer using ELISA. The serum is used for the neutralization test of the three viruses of MHV, PEDV, and PDCoV. Results: The results revealed that the fusion proteins Qbeta, Qbeta-CoV, and HE-Qbeta-CoV could all obtain successful expression. Particles with high purity were obtained after purification; the chimeric particles of Qbeta-CoV and HE-Qbeta-CoV were found to be similar to Qbeta particles in morphology and formed chimeric VLPs. In addition, two chimeric VLP vaccines induced specific antibody responses in mice, and the antibodies showed certain neutralizing activity. Conclusion: The successful construction of the chimeric VLPs of the phage Qβ coat protein presenting the universal epitope of coronavirus provides a vaccine form with potential clinical applications for the treatment of coronavirus disease.

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Protection from cytomegalovirus viremia following glycoprotein B vaccination is not dependent on neutralizing antibodies

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1800224115 ◽

2018 ◽

Vol 115 (24) ◽

pp. 6273-6278 ◽

Cited By ~ 53

Author(s):

Ilona Baraniak ◽

Barbara Kropff ◽

Lyn Ambrose ◽

Megan McIntosh ◽

Gary R. McLean ◽

...

Keyword(s):

Antibody Response ◽

Neutralizing Antibodies ◽

De Novo ◽

Natural Infection ◽

Neutralizing Antibody ◽

Glycoprotein B ◽

Solid Organ ◽

Viral Glycoprotein ◽

Viral Spread

Human cytomegalovirus (HCMV) is an important pathogen in transplant patients and in congenital infection. Previously, we demonstrated that vaccination with a recombinant viral glycoprotein B (gB)/MF59 adjuvant formulation before solid organ transplant reduced viral load parameters post transplant. Reduced posttransplant viremia was directly correlated with antibody titers against gB consistent with a humoral response against gB being important. Here we show that sera from the vaccinated seronegative patients displayed little evidence of a neutralizing antibody response against cell-free HCMV in vitro. Additionally, sera from seronegative vaccine recipients had minimal effect on the replication of a strain of HCMV engineered to be cell-associated in a viral spread assay. Furthermore, although natural infection can induce antibody-dependent cellular cytotoxicity (ADCC) responses, serological analysis of seronegative vaccinees again presented no evidence of a substantial ADCC-promoting antibody response being generated de novo. Finally, analyses for responses against major antigenic domains of gB following vaccination were variable, and their pattern was distinct compared with natural infection. Taken together, these data argue that the protective effect elicited by the gB vaccine is via a mechanism of action in seronegative vaccinees that cannot be explained by neutralization or the induction of ADCC. More generally, these data, which are derived from a human challenge model that demonstrated that the gB vaccine is protective, highlight the need for more sophisticated analyses of new HCMV vaccines over and above the quantification of an ability to induce potent neutralizing antibody responses in vitro.

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Recombinant Bovine Herpesvirus 4 (BoHV-4) Expressing Glycoprotein D of BoHV-1 Is Immunogenic and Elicits Serum-Neutralizing Antibodies against BoHV-1 in a Rabbit Model

Clinical and Vaccine Immunology ◽

10.1128/cvi.00200-06 ◽

2006 ◽

Vol 13 (11) ◽

pp. 1246-1254 ◽

Cited By ~ 13

Author(s):

Gaetano Donofrio ◽

Sandro Cavirani ◽

Alain Vanderplasschen ◽

Laurent Gillet ◽

Cesidio Filippo Flammini

Keyword(s):

Neutralizing Antibodies ◽

Animal Species ◽

Hepatitis Virus ◽

Genetic Material ◽

Rabbit Model ◽

Bovine Herpesvirus ◽

Woodchuck Hepatitis Virus ◽

Glycoprotein D ◽

Bovine Herpesvirus 4 ◽

Herpesvirus 4

ABSTRACT Several biological characteristics of bovine herpesvirus 4 (BoHV-4) make it a good candidate as a gene delivery vector for vaccination purposes. These characteristics include little or no pathogenicity, unlikely oncogenicity, the capability to accommodate large amounts of foreign genetic material, the ability to infect several cell types coming from different animal species, and the ability to maintain transgene expression in both undifferentiated and differentiated cells. Starting from BoHV-4 cloned as a bacterial artificial chromosome (BAC), we used MuA transposase-mediated in vitro transposition to generate recombinant BoHV-4 expressing the immunodominant glycoprotein D (gD) of BoHV-1, one of the most important pathogens of cattle. Although a cis-acting element from woodchuck hepatitis virus (the woodchuck hepatitis virus posttranscriptional regulatory element [WPRE]) in the 3′ end of the gD expression cassette was required for maximal gD expression from plasmids in transient transfection assays, this element was not necessary for efficient expression of gD from recombinant BoHV-4 genomes. BoHV-4 recombinants containing gD expression cassettes with or without the WPRE expressed gD at similarly high levels. Several cell lines originating from different animal species expressed gD when infected with BoHV-4 recombinants. When rabbits were immunized with one of the recombinants, high levels of serum neutralizing antibodies against BoHV-1 were generated. This work is one of the first demonstrations of the use BoHV-4 as a vector for vaccine purposes and may provide the basis for BoHV-1 vaccination of cattle with recombinant BoHV-4.

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Cellular Targeting of Engineered Heterologous Antigens Is a Determinant Factor for Bovine Herpesvirus 4-Based Vaccine Vector Development

Clinical and Vaccine Immunology ◽

10.1128/cvi.00224-09 ◽

2009 ◽

Vol 16 (11) ◽

pp. 1675-1686 ◽

Cited By ~ 20

Author(s):

Gaetano Donofrio ◽

Valentina Franceschi ◽

Antonio Capocefalo ◽

Simone Taddei ◽

Chiara Sartori ◽

...

Keyword(s):

Recombinant Virus ◽

Neutralizing Antibodies ◽

Vaccine Development ◽

Bovine Herpesvirus ◽

Artificial Chromosome ◽

Diarrhea Virus ◽

Chimeric Peptide ◽

Bovine Herpesvirus 4 ◽

Herpesvirus 4 ◽

Protein Cell

ABSTRACT In a previous study, an apathogenic strain of bovine herpesvirus 4 (BoHV-4) cloned as a bacterial artificial chromosome and expressing a chimeric peptide (gE2/gD) as a secreted form was described. Recombinant virus-inoculated animals produced antibodies against bovine viral diarrhea virus (BVDV) gE2 and BoHV-1 gD. However, neutralizing antibodies were produced only against BVDV, not against BoHV-1. In the present work a recombinant BoHV-4 expressing a membrane-linked form of gE2/gD chimeric peptide was constructed, and inoculated rabbits produced serum-neutralizing antibodies against both BVDV and BoHV-1. Protein cell sorting and targeting are a very important issue when immunodominant antigens are engineered for recombinant virus vaccine development.

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