Publications by authors named "Brock Kingstad-Bakke"

18 Publications

  • Page 1 of 1

CCR2 Regulates Vaccine-Induced Mucosal T-Cell Memory to Influenza A Virus.

J Virol 2021 May 5. Epub 2021 May 5.

Department of Pathobiological Sciences, University of Wisconsin-Madison, Madison, 53706, WI, USA

Elicitation of lung tissue-resident memory CD8 T cells (Ts) is a goal of T-cell based vaccines against respiratory viral pathogens, such as influenza A virus (IAV). Chemokine receptor 2 (CCR2)-dependent monocyte trafficking plays an essential role in the establishment of CD8 Ts in lungs of IAV-infected mice. Here, we used a combination adjuvant-based subunit vaccine strategy that evokes multifaceted (T1/T17/T1/T17) IAV nucleoprotein-specific lung Ts, to determine whether CCR2 and monocyte infiltration are essential for vaccine-induced T development and protective immunity to IAV in lungs. Following intranasal vaccination, neutrophils, monocytes, conventional DCs and monocyte-derived dendritic cells internalized and processed vaccine antigen in lungs. We found that Basic Leucine Zipper ATF-Like Transcription Factor 3 (BATF-3)-dependent DCs were essential for eliciting T cell responses, but CCR2 deficiency enhanced the differentiation of CD127/KLRG-1, OX40CD62L and mucosally imprinted CD69CD103 effector and memory CD8 T cells in lungs and airways of vaccinated mice. Mechanistically, increased development of lung Ts, induced by CCR2 deficiency was linked to dampened expression of T-bet, but not altered TCF-1 levels or T cell receptor signaling in CD8 T cells. T1/T17 functional programming, parenchymal localization of CD8/CD4 effector and memory T cells, recall T cell responses and protective immunity to a lethal IAV infection were unaffected in CCR2-deficient mice. Taken together, we identified a negative regulatory role for CCR2 and monocyte trafficking in mucosal imprinting and differentiation of vaccine-induced Ts. Mechanistic insights from this study may aid the development of T-cell-based vaccines against respiratory viral pathogens including IAV and SARS-CoV-2.While antibody-based immunity to influenza A virus (IAV) is type and sub-type specific, lung and airway-resident memory T cells that recognize conserved epitopes in the internal viral proteins are known to provide heterosubtypic immunity. Hence, broadly protective IAV vaccines need to elicit robust T-cell memory in the respiratory tract. We have developed a combination adjuvant-based IAV nucleoprotein vaccine that elicits strong CD4 and CD8 T cell memory in lungs and protects against H1N1 and H5N1 strains of IAV. In this study, we examined the mechanisms that control vaccine-induced protective memory T cells in the respiratory tract. We found that trafficking of monocytes into lungs might limit the development of anti-viral lung-resident memory T cells, following intranasal vaccination. These findings suggested that strategies that limit monocyte infiltration can potentiate vaccine-induced frontline T-cell immunity to respiratory viruses, such as IAV and SARS-CoV-2.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JVI.00530-21DOI Listing
May 2021

CCR2 Regulates Vaccine-Induced Mucosal T-Cell Memory to Influenza A Virus.

bioRxiv 2021 Mar 25. Epub 2021 Mar 25.

Elicitation of lung tissue-resident memory CD8 T cells (T s) is a goal of T-cell based vaccines against respiratory viral pathogens such as influenza A virus (IAV). Chemokine receptor 2 (CCR2)-dependent monocyte trafficking plays an essential role in the establishment of CD8 T s in lungs of IAV-infected mice. Here, we used a combination adjuvant-based subunit vaccine strategy that evokes multifaceted (T 1/T 17/T 1/T 17) IAV nucleoprotein-specific lung T s, to determine whether CCR2 and monocyte infiltration are essential for vaccine-induced T development and protective immunity to IAV in lungs. Following intranasal vaccination, neutrophils, monocytes, conventional dendrtitic cells (DCs) and monocyte-derived DCs internalized and processed vaccine antigen in lungs. We also found that Basic Leucine Zipper ATF-Like Transcription Factor 3 (BATF-3)-dependent DCs were essential for eliciting T cell responses, but CCR2 deficiency enhanced the differentiation of CD127 /KLRG-1 , OX40 CD62L and mucosally imprinted CD69 CD103 effector and memory CD8 T cells in lungs and airways of vaccinated mice. Mechanistically, increased development of lung T s, induced by CCR2 deficiency was linked to dampened expression of T-bet, but not altered TCF-1 levels or T cell receptor signaling in CD8 T cells. T1/T17 functional programming, parenchymal localization of CD8/CD4 effector and memory T cells, recall T cell responses and protective immunity to a lethal IAV infection were unaffected in CCR2-deficient mice. Taken together, we identified a negative regulatory role for CCR2 and monocyte trafficking in mucosal imprinting and differentiation of vaccine-induced T s. Mechanistic insights from this study may aid the development of T-cell-based vaccines against respiratory viral pathogens including IAV and SARS-CoV-2.

Importance: While antibody-based immunity to influenza A virus (IAV) is type and sub-type specific, lung and airway-resident memory T cells that recognize conserved epitopes in the internal viral proteins are known to provide heterosubtypic immunity. Hence, broadly protective IAV vaccines need to elicit robust T-cell memory in the respiratory tract. We have developed a combination adjuvant-based IAV nucleoprotein vaccine that elicits strong CD4 and CD8 T cell memory in lungs and protects against H1N1 and H5N1 strains of IAV. In this study, we examined the mechanisms that control vaccine-induced protective memory T cells in the respiratory tract. We found that trafficking of monocytes into lungs might limit the development of anti-viral lung-resident memory T cells, following intranasal vaccination. These findings suggested that strategies that limit monocyte infiltration can potentiate vaccine-induced frontline T-cell immunity to respiratory viruses such as IAV and SARS-CoV-2.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/2021.03.24.436901DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8010722PMC
March 2021

Combination Adjuvants Affect the Magnitude of Effector-Like Memory CD8 T Cells and Protection against Listeriosis.

Infect Immun 2021 Mar 29. Epub 2021 Mar 29.

Department of Pathobiological Sciences, University of Wisconsin-Madison, Madison, 53706, WI, USA

Development of T-cell-based subunit protein vaccines against diseases, such as tuberculosis and malaria, remains a challenge for immunologists. Here, we have identified a nano-emulsion adjuvant Adjuplex (ADJ), which enhanced dendritic cell (DC) cross-presentation and elicited effective memory T cell-based immunity to We further evaluated whether cross-presentation induced by ADJ, can be combined with the immunomodulatory effects of TLR agonists (CpG or glucopyranosyl lipid adjuvant [GLA]) to evoke systemic CD8 T cell-based immunity to Mechanistically, vaccination with ADJ, alone or in combination with CpG or GLA augmented activation and antigen uptake by CD103 migratory and CD8α resident DCs and up-regulated CD69 expression on B and T lymphocytes in vaccine-draining lymph nodes. By engaging basic leucine zipper ATF-like transcription factor 3-dependent cross-presenting DCs, ADJ potently elicited effector CD8 T cells that differentiated into granzyme B-expressing CD27 effector-like memory CD8 T cells, which provided effective immunity to LM in spleen and liver. CpG or GLA alone did not elicit effector-like memory CD8 T cells and induced moderate protection in spleen, but not in the liver. Surprisingly, combining CpG or GLA with ADJ reduced the number of ADJ-induced memory CD8 T cells and compromised protective immunity to LM, especially in the liver. Taken together, data presented in this manuscript provides a glimpse of protective CD8 T cell memory differentiation induced by a nano-emulsion adjuvant and demonstrates the unexpected negative effects of TLR signaling on the magnitude of CD8 T cell memory and protective immunity to LM, a model intracellular pathogen.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/IAI.00768-20DOI Listing
March 2021

Polymeric Pathogen-Like Particles-Based Combination Adjuvants Elicit Potent Mucosal T Cell Immunity to Influenza A Virus.

Front Immunol 2020 4;11:559382. Epub 2021 Mar 4.

Department of Pathobiological Sciences, University of Wisconsin-Madison, Madison, WI, United States.

Eliciting durable and protective T cell-mediated immunity in the respiratory mucosa remains a significant challenge. Polylactic-co-glycolic acid (PLGA)-based cationic pathogen-like particles (PLPs) loaded with TLR agonists mimic biophysical properties of microbes and hence, simulate pathogen-pattern recognition receptor interactions to safely and effectively stimulate innate immune responses. We generated micro particle PLPs loaded with TLR4 (glucopyranosyl lipid adjuvant, GLA) or TLR9 (CpG) agonists, and formulated them with and without a mucosal delivery enhancing carbomer-based nanoemulsion adjuvant (ADJ). These adjuvants delivered intranasally to mice elicited high numbers of influenza nucleoprotein (NP)-specific CD8+ and CD4+ effector and tissue-resident memory T cells (Ts) in lungs and airways. PLPs delivering TLR4 versus TLR9 agonists drove phenotypically and functionally distinct populations of effector and memory T cells. While PLPs loaded with CpG or GLA provided immunity, combining the adjuvanticity of PLP-GLA and ADJ markedly enhanced the development of airway and lung Ts and CD4 and CD8 T cell-dependent immunity to influenza virus. Further, balanced CD8 (Tc1/Tc17) and CD4 (Th1/Th17) recall responses were linked to effective influenza virus control. These studies provide mechanistic insights into vaccine-induced pulmonary T cell immunity and pave the way for the development of a universal influenza and SARS-CoV-2 vaccines.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fimmu.2020.559382DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7986715PMC
April 2021

Structural basis of Blastomyces Endoglucanase-2 adjuvancy in anti-fungal and -viral immunity.

PLoS Pathog 2021 Mar 18;17(3):e1009324. Epub 2021 Mar 18.

Department of Pediatrics, University of Wisconsin School of Medicine and Public Health, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.

The development of safe subunit vaccines requires adjuvants that augment immunogenicity of non-replicating protein-based antigens. Current vaccines against infectious diseases preferentially induce protective antibodies driven by adjuvants such as alum. However, the contribution of antibody to host defense is limited for certain classes of infectious diseases such as fungi, whereas animal studies and clinical observations implicate cellular immunity as an essential component of the resolution of fungal pathogens. Here, we decipher the structural bases of a newly identified glycoprotein ligand of Dectin-2 with potent adjuvancy, Blastomyces endoglucanase-2 (Bl-Eng2). We also pinpoint the developmental steps of antigen-specific CD4+ and CD8+ T responses augmented by Bl-Eng2 including expansion, differentiation and tissue residency. Dectin-2 ligation led to successful systemic and mucosal vaccination against invasive fungal infection and Influenza A infection, respectively. O-linked glycans on Bl-Eng2 applied at the skin and respiratory mucosa greatly augment vaccine subunit- induced protective immunity against lethal influenza and fungal pulmonary challenge.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.ppat.1009324DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8009368PMC
March 2021

Carbomer-based adjuvant elicits CD8 T-cell immunity by inducing a distinct metabolic state in cross-presenting dendritic cells.

PLoS Pathog 2021 01 14;17(1):e1009168. Epub 2021 Jan 14.

Department of Pathobiological Sciences, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.

There is a critical need for adjuvants that can safely elicit potent and durable T cell-based immunity to intracellular pathogens. Here, we report that parenteral vaccination with a carbomer-based adjuvant, Adjuplex (ADJ), stimulated robust CD8 T-cell responses to subunit antigens and afforded effective immunity against respiratory challenge with a virus and a systemic intracellular bacterial infection. Studies to understand the metabolic and molecular basis for ADJ's effect on antigen cross-presentation by dendritic cells (DCs) revealed several unique and distinctive mechanisms. ADJ-stimulated DCs produced IL-1β and IL-18, suggestive of inflammasome activation, but in vivo activation of CD8 T cells was unaffected in caspase 1-deficient mice. Cross-presentation induced by TLR agonists requires a critical switch to anabolic metabolism, but ADJ enhanced cross presentation without this metabolic switch in DCs. Instead, ADJ induced in DCs, an unique metabolic state, typified by dampened oxidative phosphorylation and basal levels of glycolysis. In the absence of increased glycolytic flux, ADJ modulated multiple steps in the cytosolic pathway of cross-presentation by enabling accumulation of degraded antigen, reducing endosomal acidity and promoting antigen localization to early endosomes. Further, by increasing ROS production and lipid peroxidation, ADJ promoted antigen escape from endosomes to the cytosol for degradation by proteasomes into peptides for MHC I loading by TAP-dependent pathways. Furthermore, we found that induction of lipid bodies (LBs) and alterations in LB composition mediated by ADJ were also critical for DC cross-presentation. Collectively, our model challenges the prevailing metabolic paradigm by suggesting that DCs can perform effective DC cross-presentation, independent of glycolysis to induce robust T cell-dependent protective immunity to intracellular pathogens. These findings have strong implications in the rational development of safe and effective immune adjuvants to potentiate robust T-cell based immunity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.ppat.1009168DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7840022PMC
January 2021

Programming Multifaceted Pulmonary T Cell Immunity by Combination Adjuvants.

Cell Rep Med 2020 Sep;1(6):100095

Department of Pathobiological Sciences, University of Wisconsin-Madison, Madison, WI, USA.

Induction of protective mucosal T cell memory remains a formidable challenge to vaccinologists. Using a combination adjuvant strategy that elicits potent CD8 and CD4 T cell responses, we define the tenets of vaccine-induced pulmonary T cell immunity. An acrylic-acid-based adjuvant (ADJ), in combination with Toll-like receptor (TLR) agonists glucopyranosyl lipid adjuvant (GLA) or CpG, promotes mucosal imprinting but engages distinct transcription programs to drive different degrees of terminal differentiation and disparate polarization of T1/T1/T17/T17 effector/memory T cells. Combination of ADJ with GLA, but not CpG, dampens T cell receptor (TCR) signaling, mitigates terminal differentiation of effectors, and enhances the development of CD4 and CD8 T cells that protect against H1N1 and H5N1 influenza viruses. Mechanistically, vaccine-elicited CD4 T cells play a vital role in optimal programming of CD8 T and viral control. Taken together, these findings provide further insights into vaccine-induced multifaceted mucosal T cell immunity with implications in the development of vaccines against respiratorypathogens, including influenza virus and SARS-CoV-2.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.xcrm.2020.100095DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7508055PMC
September 2020

A Novel Mucosal Adjuvant System for Immunization against Avian Coronavirus Causing Infectious Bronchitis.

J Virol 2020 09 15;94(19). Epub 2020 Sep 15.

Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin, Madison, Wisconsin, USA

Infectious bronchitis (IB) caused by infectious bronchitis virus (IBV) is currently a major threat to chicken health, with multiple outbreaks being reported in the United States over the past decade. Modified live virus (MLV) vaccines used in the field can persist and provide the genetic material needed for recombination and emergence of novel IBV serotypes. Inactivated and subunit vaccines overcome some of the limitations of MLV with no risk of virulence reversion and emergence of new virulent serotypes. However, these vaccines are weakly immunogenic and poorly protective. There is an urgent need to develop more effective vaccines that can elicit a robust, long-lasting immune response. In this study, we evaluate a novel adjuvant system developed from Quil-A and chitosan (QAC) for the intranasal delivery of nucleic acid immunogens to improve protective efficacy. The QAC adjuvant system forms nanocarriers (<100 nm) that efficiently encapsulate nucleic acid cargo, exhibit sustained release of payload, and can stably transfect cells. Encapsulation of plasmid DNA vaccine expressing IBV nucleocapsid (N) protein by the QAC adjuvant system (pQAC-N) enhanced immunogenicity, as evidenced by robust induction of adaptive humoral and cellular immune responses postvaccination and postchallenge. Birds immunized with pQAC-N showed reduced clinical severity and viral shedding postchallenge on par with protection observed with current commercial vaccines without the associated safety concerns. Presented results indicate that the QAC adjuvant system can offer a safer alternative to the use of live vaccines against avian and other emerging coronaviruses. According to 2017 U.S. agriculture statistics, the combined value of production and sales from broilers, eggs, turkeys, and chicks was $42.8 billion. Of this number, broiler sales comprised 67% of the industry value, with the production of >50 billion pounds of chicken meat. The economic success of the poultry industry in the United States hinges on the extensive use of vaccines to control infectious bronchitis virus (IBV) and other poultry pathogens. The majority of vaccines currently licensed for poultry health include both modified live vaccine and inactivated pathogens. Despite their proven efficacy, modified live vaccine constructs take time to produce and could revert to virulence, which limits their safety. The significance of our research stems from the development of a safer and potent alternative mucosal vaccine to replace live vaccines against IBV and other emerging coronaviruses.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JVI.01016-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7495365PMC
September 2020

Effective mosaic-based nanovaccines against avian influenza in poultry.

Vaccine 2019 08 9;37(35):5051-5058. Epub 2019 Jul 9.

Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin, Madison, WI, USA; Pan Genome Systems, Madison, WI, USA. Electronic address:

Avian influenza virus (AIV) is an extraordinarily diverse pathogen that causes significant morbidity in domesticated poultry populations and threatens human life with looming pandemic potential. Controlling avian influenza in susceptible populations requires highly effective, economical and broadly reactive vaccines. Several AIV vaccines have proven insufficient despite their wide use, and better technologies are needed to improve their immunogenicity and broaden effectiveness. Previously, we developed a "mosaic" H5 subtype hemagglutinin (HA) AIV vaccine and demonstrated its broad protection against diverse highly pathogenic H5N1 and seasonal H1N1 virus strains in mouse and non-human primate models. There is a significant interest in developing effective and safe vaccines against AIV that cannot contribute to the emergence of new strains of the virus once circulating in poultry. Here, we report on the development of an H5 mosaic (H5M) vaccine antigen formulated with polyanhydride nanoparticles (PAN) that provide sustained release of encapsulated antigens. H5M vaccine constructs were immunogenic whether delivered by the modified virus Ankara (MVA) strain or encapsulated within PAN. Both humoral and cellular immune responses were generated in both specific-pathogen free (SPF) and commercial chicks. Importantly, chicks vaccinated by H5M constructs were protected in terms of viral shedding from divergent challenge with a low pathogenicity avian influenza (LPAI) strain at 8 weeks post-vaccination. In addition, protective levels of humoral immunity were generated against highly pathogenic avian influenza (HPAI) of the similar H5N1 and genetically dissimilar H5N2 viruses. Overall, the developed platform technologies (MVA vector and PAN encapsulation) were safe and provided high levels of sustained protection against AIV in chickens. Such approaches could be used to design more efficacious vaccines against other important poultry infections.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.vaccine.2019.06.077DOI Listing
August 2019

Virally-vectored vaccine candidates against white-nose syndrome induce anti-fungal immune response in little brown bats (Myotis lucifugus).

Sci Rep 2019 05 1;9(1):6788. Epub 2019 May 1.

Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin - Madison, Madison, Wisconsin, USA.

White-nose syndrome (WNS) caused by the fungus, Pseudogymnoascus destructans (Pd) has killed millions of North American hibernating bats. Currently, methods to prevent the disease are limited. We conducted two trials to assess potential WNS vaccine candidates in wild-caught Myotis lucifugus. In a pilot study, we immunized bats with one of four vaccine treatments or phosphate-buffered saline (PBS) as a control and challenged them with Pd upon transfer into hibernation chambers. Bats in one vaccine-treated group, that received raccoon poxviruses (RCN) expressing Pd calnexin (CAL) and serine protease (SP), developed WNS at a lower rate (1/10) than other treatments combined (14/23), although samples sizes were small. The results of a second similar trial provided additional support for this observation. Bats vaccinated orally or by injection with RCN-CAL and RCN-SP survived Pd challenge at a significantly higher rate (P = 0.01) than controls. Using RT-PCR and flow cytometry, combined with fluorescent in situ hybridization, we determined that expression of IFN-γ transcripts and the number of CD4 + T-helper cells transcribing this gene were elevated (P < 0.10) in stimulated lymphocytes from surviving vaccinees (n = 15) compared to controls (n = 3). We conclude that vaccination with virally-vectored Pd antigens induced antifungal immunity that could potentially protect bats against WNS.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-019-43210-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6494898PMC
May 2019

Zika virus like particles elicit protective antibodies in mice.

PLoS Negl Trop Dis 2018 02 5;12(2):e0006210. Epub 2018 Feb 5.

Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin, Madison, WI, United States of America.

Mosquito-borne Zika virus (ZIKV) typically causes a mild and self-limiting illness known as Zika fever. Since its recent emergence in 2014 in the American continent, ZIKV infection during pregnancy has been closely associated with a wide range of congenital abnormalities. To date, no vaccines or antivirals are publicly available. We developed Zika virus-like particles (VLPs) and evaluated their immunogenicity and protective efficacy in mouse models. ZIKV VLPs (ZIKVLPs) formulated with alum were injected into 6-8-week-old interferon deficient AG129 mice as well as wild type BALB/c mice. Control mice received PBS/alum. Animals were challenged with 200 PFU (>1000 AG129 LD50s) of ZIKV strain H/PF/2013. All vaccinated mice survived with no morbidity or weight loss while control animals either died at 9 days post challenge (AG129) or had increased viremia (BALB/c). Neutralizing antibodies were observed in all ZIKVLP vaccinated mice. The role of neutralizing antibodies in protecting mice was demonstrated by passive transfer. Our findings demonstrate the protective efficacy of the ZIKVLP vaccine and highlight the important role that neutralizing antibodies play in protection against ZIKV infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.pntd.0006210DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5814096PMC
February 2018

A modified vaccinia Ankara vaccine vector expressing a mosaic H5 hemagglutinin reduces viral shedding in rhesus macaques.

PLoS One 2017 3;12(8):e0181738. Epub 2017 Aug 3.

Department of Pathobiological Sciences, University of Wisconsin-Madison, School of Veterinary Medicine, Madison, Wisconsin, United States of America.

The rapid antigenic evolution of influenza viruses requires frequent vaccine reformulations. Due to the economic burden of continuous vaccine reformulation and the threat of new pandemics, there is intense interest in developing vaccines capable of eliciting broadly cross-reactive immunity to influenza viruses. We recently constructed a "mosaic" hemagglutinin (HA) based on subtype 5 HA (H5) and designed to stimulate cellular and humoral immunity to multiple influenza virus subtypes. Modified vaccinia Ankara (MVA) expressing this H5 mosaic (MVA-H5M) protected mice against multiple homosubtypic H5N1 strains and a heterosubtypic H1N1 virus. To assess its potential as a human vaccine we evaluated the ability of MVA-H5M to provide heterosubtypic immunity to influenza viruses in a non-human primate model. Rhesus macaques received an initial dose of either MVA-H5M or plasmid DNA encoding H5M, followed by a boost of MVA-H5M, and then were challenged, together with naïve controls, with the heterosubtypic virus A/California/04/2009 (H1N1pdm). Macaques receiving either vaccine regimen cleared H1N1pdm challenge faster than naïve controls. Vaccination with H5M elicited antibodies that bound H1N1pdm HA, but did not neutralize the H1N1pdm challenge virus. Plasma from vaccinated macaques activated NK cells in the presence of H1N1pdm HA, suggesting that vaccination elicited cross-reactive antibodies capable of mediating antibody-dependent cell-mediated cytotoxicity (ADCC). Although HA-specific T cell responses to the MVA-H5M vaccine were weak, responses after challenge were stronger in vaccinated macaques than in control animals. Together these data suggest that mosaic HA antigens may provide a means for inducing broadly cross-reactive immunity to influenza viruses.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0181738PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5542451PMC
August 2017

Infectivity of attenuated poxvirus vaccine vectors and immunogenicity of a raccoonpox vectored rabies vaccine in the Brazilian Free-tailed bat (Tadarida brasiliensis).

Vaccine 2016 10 17;34(44):5352-5358. Epub 2016 Sep 17.

U.S. Geological Survey, National Wildlife Health Center, 6006 Schroeder Rd., Madison, WI 53711, USA. Electronic address:

Bats (Order Chiroptera) are an abundant group of mammals with tremendous ecological value as insectivores and plant dispersers, but their role as reservoirs of zoonotic diseases has received more attention in the last decade. With the goal of managing disease in free-ranging bats, we tested modified vaccinia Ankara (MVA) and raccoon poxvirus (RCN) as potential vaccine vectors in the Brazilian Free-tailed bat (Tadarida brasiliensis), using biophotonic in vivo imaging and immunogenicity studies. Animals were administered recombinant poxviral vectors expressing the luciferase gene (MVA-luc, RCN-luc) through oronasal (ON) or intramuscular (IM) routes and subsequently monitored for bioluminescent signal indicative of viral infection. No clinical illness was noted after exposure to any of the vectors, and limited luciferase expression was observed. Higher and longer levels of expression were observed with the RCN-luc construct. When given IM, luciferase expression was limited to the site of injection, while ON exposure led to initial expression in the oral cavity, often followed by secondary replication at another location, likely the gastric mucosa or gastric associated lymphatic tissue. Viral DNA was detected in oral swabs up to 7 and 9 days post infection (dpi) for MVA and RCN, respectively. While no live virus was detected in oral swabs from MVA-infected bats, titers up to 3.88 x 10 PFU/ml were recovered from oral swabs of RCN-infected bats. Viral DNA was also detected in fecal samples from two bats inoculated IM with RCN, but no live virus was recovered. Finally, we examined the immunogenicity of a RCN based rabies vaccine (RCN-G) following ON administration. Significant rabies neutralizing antibody titers were detected in the serum of immunized bats using the rapid fluorescence focus inhibition test (RFFIT). These studies highlight the safety and immunogenicity of attenuated poxviruses and their potential use as vaccine vectors in bats.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.vaccine.2016.08.088DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5543807PMC
October 2016

Mosaic H5 Hemagglutinin Provides Broad Humoral and Cellular Immune Responses against Influenza Viruses.

J Virol 2016 08 11;90(15):6771-6783. Epub 2016 Jul 11.

Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin-Madison, Madison, Wisconsin, USA

Unlabelled: The most effective way to prevent influenza virus infection is via vaccination. However, the constant mutation of influenza viruses due to antigenic drift and shift compromises vaccine efficacy. This represents a major challenge to the development of a cross-protective vaccine that can protect against circulating viral antigenic diversity. Using the modified vaccinia Ankara (MVA) virus, we had previously generated a recombinant vaccine against highly pathogenic avian influenza virus (H5N1) based on an in silico mosaic approach. This MVA-H5M construct protected mice against multiple clades of H5N1 and H1N1 viruses. We have now further characterized the immune responses using immunodepletion of T cells and passive serum transfer, and these studies indicate that antibodies are the main contributors in homosubtypic protection (H5N1 clades). Compared to a MVA construct expressing hemagglutinin (HA) from influenza virus A/VN/1203/04 (MVA-HA), the MVA-H5M vaccine markedly increased and broadened B cell and T cell responses against H5N1 virus. The MVA-H5M also provided effective protection with no morbidity against H5N1 challenge, whereas MVA-HA-vaccinated mice showed clinical signs and experienced significant weight loss. In addition, MVA-H5M induced CD8(+) T cell responses that play a major role in heterosubtypic protection (H1N1). Finally, expression of the H5M gene as either a DNA vaccine or a subunit protein protected mice against H5N1 challenge, indicating the effectiveness of the mosaic sequence without viral vectors for the development of a universal influenza vaccine.

Importance: Influenza viruses infect up to one billion people around the globe each year and are responsible for 300,000 to 500,000 deaths annually. Vaccines are still the main intervention to prevent infection, but they fail to provide effective protection against heterologous strains of viruses. We developed broadly reactive H5N1 vaccine based on an in silico mosaic approach and previously demonstrated that modified vaccinia Ankara expressing an H5 mosaic hemagglutinin prevented infection with multiple clades of H5N1 and limited severe disease after H1N1 infection. Further characterization revealed that antibody responses and T cells are main contributors to protection against H5N1 and H1N1 viruses, respectively. The vaccine also broadens both T cell and B cell responses compared to native H5 vaccine from influenza virus A/Vietnam/1203/04. Finally, delivering the H5 mosaic as a DNA vaccine or as a purified protein demonstrated effective protection similar to the viral vector approach.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JVI.00730-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4944288PMC
August 2016

Mucosal administration of raccoonpox virus expressing highly pathogenic avian H5N1 influenza neuraminidase is highly protective against H5N1 and seasonal influenza virus challenge.

Vaccine 2015 Sep 10;33(39):5155-62. Epub 2015 Aug 10.

Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin, Madison, WI 53706, USA. Electronic address:

We previously generated recombinant poxviruses expressing influenza antigens and studied their efficacy as potential highly pathogenic avian influenza (HPAI) vaccines in mice. While both modified vaccinia Ankara (MVA) and raccoon poxvirus (RCN) expressing hemagglutinin (HA) provided strong protection when administered by parenteral routes, only RCN-neuraminidase (NA) showed promise as a mucosal vaccine. In the present study we evaluated the efficacy of RCN-NA constructs by both intradermal (ID) and intranasal (IN) routes. Surprisingly, while RCN-NA completely protected mice when administered by the IN route, it failed to protect mice when administered by the ID route. After challenge, significantly less virus induced pathology was observed in the lungs of mice vaccinated with RCN-NA by the IN route as compared to the ID route. Furthermore, IN administration of RCN-NA elicited neutralizing antibodies detected in bronchoalveolar lavage (BAL) samples. We also determined the role of cellular immune responses in protection elicited by RCN-NA by depleting CD4 and CD8 T cells prior to challenge. Finally, we demonstrated for the first time that antibodies against NA can block viral entry in addition to viral spread in vitro. These studies demonstrate the importance of mucosal administration of RCN viral vectors for eliciting protective immune responses against the NA antigen.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.vaccine.2015.08.005DOI Listing
September 2015

Broad protection against avian influenza virus by using a modified vaccinia Ankara virus expressing a mosaic hemagglutinin gene.

J Virol 2014 Nov 10;88(22):13300-9. Epub 2014 Sep 10.

Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin-Madison, Madison, Wisconsin, USA

Unlabelled: A critical failure in our preparedness for an influenza pandemic is the lack of a universal vaccine. Influenza virus strains diverge by 1 to 2% per year, and commercially available vaccines often do not elicit protection from one year to the next, necessitating frequent formulation changes. This represents a major challenge to the development of a cross-protective vaccine that can protect against circulating viral antigenic diversity. We have constructed a recombinant modified vaccinia virus Ankara (MVA) that expresses an H5N1 mosaic hemagglutinin (H5M) (MVA-H5M). This mosaic was generated in silico using 2,145 field-sourced H5N1 isolates. A single dose of MVA-H5M provided 100% protection in mice against clade 0, 1, and 2 avian influenza viruses and also protected against seasonal H1N1 virus (A/Puerto Rico/8/34). It also provided short-term (10 days) and long-term (6 months) protection postvaccination. Both neutralizing antibodies and antigen-specific CD4(+) and CD8(+) T cells were still detected at 5 months postvaccination, suggesting that MVA-H5M provides long-lasting immunity.

Importance: Influenza viruses infect a billion people and cause up to 500,000 deaths every year. A major problem in combating influenza is the lack of broadly effective vaccines. One solution from the field of human immunodeficiency virus vaccinology involves a novel in silico mosaic approach that has been shown to provide broad and robust protection against highly variable viruses. Unlike a consensus algorithm which picks the most frequent residue at each position, the mosaic method chooses the most frequent T-cell epitopes and combines them to form a synthetic antigen. These studies demonstrated that a mosaic influenza virus H5 hemagglutinin expressed by a viral vector can elicit full protection against diverse H5N1 challenges as well as induce broader immunity than a wild-type hemagglutinin.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JVI.01532-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4249068PMC
November 2014

A Recombinant Raccoon Poxvirus Vaccine Expressing both Yersinia pestis F1 and Truncated V Antigens Protects Animals against Lethal Plague.

Vaccines (Basel) 2014 Oct 27;2(4):772-84. Epub 2014 Oct 27.

Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin, Madison, WI 53706, USA.

In previous studies, we demonstrated in mice and prairie dogs that simultaneous administration of two recombinant raccoon poxviruses (rRCN) expressing Yersinia pestis antigens (F1 and V307-a truncated version of the V protein) provided superior protection against plague challenge compared to individual single antigen constructs. To reduce costs of vaccine production and facilitate implementation of a sylvatic plague vaccine (SPV) control program for prairie dogs, a dual antigen construct is more desirable. Here we report the construction and characterization of a novel RCN-vectored vaccine that simultaneously expresses both F1 and V307 antigens. This dual antigen vaccine provided similar levels of protection against plague in both mice and prairie dogs as compared to simultaneous administration of the two single antigen constructs and was also shown to protect mice against an F1 negative strain of Y. pestis. The equivalent safety, immunogenicity and efficacy profile of the dual RCN-F1/V307 construct warrants further evaluation in field efficacy studies in sylvatic plague endemic areas.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/vaccines2040772DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4494250PMC
October 2014

Effects of route and coadministration of recombinant raccoon poxviruses on immune responses and protection against highly pathogenic avian influenza in mice.

Vaccine 2012 Oct 22;30(45):6402-8. Epub 2012 Aug 22.

Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin, Madison, WI 53706, USA.

We previously demonstrated that recombinant raccoonpox (RCN) virus could serve as a vector for an influenza vaccine. RCN constructs expressing the hemagglutinin (HA) from H5N1 viruses were immunogenic in chickens. In the current study, we generated several recombinant RCN constructs expressing influenza (H5N1) antigens and a molecular adjuvant (Heat-Labile enterotoxin B from E. coli: RCN-LTB), demonstrated their expression in vitro, and evaluated their ability to protect mice against H5N1 virus challenge. RCN-HA provided strong protection when administered intradermally (ID), but not intranasally (IN). Conversely, the RCN-neuraminidase (NA) construct was highly efficacious by the IN route and elicited high titers of neutralizing antibodies in mice. Vaccination by combined ID (RCN-HA) and IN (RCN-NA) routes offered mice the best protection against an IN challenge with heterologous H5N1 virus. However, protection was reduced when the different RCN constructs were pre-mixed, perhaps due to reduced expression of antigen.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.vaccine.2012.08.018DOI Listing
October 2012