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Improved vaccines and antiviral agents that provide better, broader protection against seasonal and emerging influenza viruses are needed. The viral surface glycoprotein hemagglutinin (HA) is a primary target for the development of universal influenza vaccines and therapeutic antibodies. The other major surface antigen, neuraminidase (NA), has been less well studied as a potential target and fewer broadly reactive anti-NA antibodies have been identified. In this study, we isolate three human monoclonal antibodies that recognize NA from A/H1N1 subtypes, and find that one of them, clone DA03E17, binds to the NA of A/H3N2, A/H5N1, A/H7N9, B/Ancestral-lineage, B/Yamagata-lineage, and B/Victoria-lineage viruses. DA03E17 inhibits the neuraminidase activity by direct binding to the enzyme active site, and provides in vitro and in vivo protection against infection with several types of influenza virus. This clone could, therefore, be useful as a broadly protective therapeutic agent. Moreover, the neutralizing epitope of DA03E17 could be useful in the development of an NA-based universal influenza vaccine., (© 2022. The Author(s).)

Vaccination is the first line of protection against influenza virus infection in humans. Although inactivated and live-attenuated vaccines are available, each vaccine has drawbacks in terms of immunogenicity and safety. To overcome these issues, our group has developed a replication-incompetent PB2-knockout (PB2-KO) influenza virus that replicates only in PB2-expressing cells. Here we generated PB2-KO viruses possessing the hemagglutinin (HA) and neuraminidase (NA) segments from H1N1pdm09 or type B viruses and tested their vaccine potential. The two PB2-KO viruses propagated efficiently in PB2-expressing cells, and expressed chimeric HA as expected. Virus-specific IgG and IgA antibodies were detected in mice immunized with the viruses, and the immunized mice showed milder clinical signs and/or lower virus replication levels in the respiratory tract upon virus challenge. Our results indicate that these PB2-KO viruses have potential as vaccine candidates., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Background: Secondary bacterial infections after influenza can be a serious problem, especially in young children and the elderly, yet the efficacy of current vaccines is limited. Earlier work demonstrated that a replication-incompetent PB2-knockout (PB2-KO) influenza virus possessing a foreign gene in the coding region of its PB2 segment can serve as a platform for a bivalent vaccine., Methods: In the current study, we generated the PB2-KO virus expressing pneumococcal surface protein A (PspA), PB2-KO-PspA virus, the replication of which is restricted to PB2-expressing cells. We then examined the protective efficacy of intranasal immunization with this virus as a bivalent vaccine in a mouse model., Results: High levels of influenza virus-specific and PspA-specific antibodies were induced in the serum and airways of immunized mice. The intranasally immunized mice were protected from lethal doses of influenza virus or Streptococcus pneumoniae. These mice were also completely protected from secondary pneumococcal pneumonia after influenza virus infection., Conclusions: These findings indicate that our recombinant influenza virus serves as a novel and powerful bivalent vaccine against primary and secondary pneumococcal pneumonia as well as influenza., (© The Author 2015. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Because vaccination is an effective means to protect humans from influenza viruses, extensive efforts have been made to develop not only new vaccines, but also for new adjuvants to enhance the efficacy of existing inactivated vaccines. Here, we examined the adjuvanticity of synthetic hemozoin, a synthetic version of the malarial by-product hemozoin, on the vaccine efficacy of inactivated whole influenza viruses in a mouse model. We found that mice immunized twice with hemozoin-adjuvanted inactivated A/California/04/2009 (H1N1pdm09) or A/Vietnam/1203/2004 (H5N1) virus elicited higher virus-specific antibody responses than did mice immunized with non-adjuvanted counterparts. Furthermore, mice immunized with hemozoin-adjuvanted inactivated viruses were better protected from lethal challenge with influenza viruses than were mice immunized with non-adjuvanted inactivated vaccines. Our results show that hemozoin improves the immunogenicity of inactivated influenza viruses, and is thus a promising adjuvant for inactivated whole virion influenza vaccines., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Influenza virus and human parainfluenza virus (HPIV) are major etiologic agents of acute respiratory illness in young children. Inactivated and live attenuated influenza vaccines are approved in several countries, yet no vaccine is licensed for HPIV. We previously showed that a replication-incompetent PB2-knockout (PB2-KO) virus that possesses a reporter gene in the coding region of the PB2 segment can serve as a platform for a bivalent vaccine. To develop a bivalent vaccine against influenza and parainfluenza virus, here, we generated a PB2-KO virus possessing the hemagglutinin-neuraminidase (HN) glycoprotein of HPIV type 3 (HPIV3), a major surface antigen of HPIV, in its PB2 segment. We confirmed that this virus replicated only in PB2-expressing cells and expressed HN. We then examined the efficacy of this virus as a bivalent vaccine in a hamster model. High levels of virus-specific IgG antibodies in sera and IgA, IgG, and IgM antibodies in bronchoalveolar lavage fluids against both influenza virus and HPIV3 were detected from hamsters immunized with this virus. The neutralizing capability of these serum antibodies was also confirmed. Moreover, the immunized hamsters were completely protected from virus challenge with influenza virus or HPIV3. These results indicate that PB2-KO virus expressing the HN of HPIV3 has the potential to be a novel bivalent vaccine against influenza and human parainfluenza viruses., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Vaccination is an effective means to protect against influenza virus. Although inactivated and live-attenuated vaccines are currently available, each vaccine has disadvantages (e.g., immunogenicity and safety issues). To overcome these problems, we previously developed a replication-incompetent PB2-knockout (PB2-KO) influenza virus that replicates only in PB2 protein-expressing cells. Here, we generated two PB2-KO viruses whose PB2-coding regions were replaced with the HA genes of either A/California/04/2009 (H1N1pdm09) or A/Vietnam/1203/2004 (H5N1). The resultant viruses comparably, or in some cases more efficiently, induced virus-specific antibodies in the serum, nasal wash, and bronchoalveolar lavage fluid of mice relative to a conventional formalin-inactivated vaccine. Furthermore, mice immunized with these PB2-KO viruses were protected from lethal challenges with not only the backbone virus strain but also strains from which their foreign HAs originated, indicating that PB2-KO viruses with antigenically different HAs could serve as bivalent influenza vaccines.

H5N1 highly pathogenic avian influenza viruses evolved into several clades, leading to appreciably distinct antigenicities of their hemagglutinins. As such, candidate H5N1 pre-pandemic vaccines for human use should be sought. Here, to evaluate fundamental immunogenic variations between H5N1 vaccines, we prepared four inactivated H5N1 test vaccines from different phylogenetic clades (clade 1, 2.1, 2.2, and 2.3.4) in accordance with the WHO recommendation, and tested their cross-clade immunity in a mouse model by vaccination followed by challenge with heterologous virulent viruses. All H5N1 vaccines tested provided full or partial cross-clade protective immunity, except one clade 2.2-based vaccine, which did not protect mice from clade 2.3.4 virus challenge. Among the test vaccines, a clade 2.1-based vaccine possessed the broadest-spectrum cross-immunity. These results suggest that currently stockpiled pre-pandemic vaccines, especially clade 2.1-based vaccines, will likely be useful as backup vaccines in a pandemic situation, even one involving antigenic-drifted viruses.

Seed viruses used to produce inactivated H5N1 influenza vaccines are recombinant viruses with modified avirulent-type hemagglutinin (HA) and intact neuraminidase (NA) genes, both derived from an H5N1 isolate, and all remaining genes from the PR8 strain, which grows well in eggs. However, some reassortants grow suboptimally in eggs, imposing obstacles to timely, cost-efficient vaccine production. Here, we demonstrate that our PR8 strain supports better in ovo growth than the PR8 strain used for the WHO-recommended seed virus, NIBRG-14. Moreover, inclusion of an alternative NA protein further enhanced viral growth in eggs. These findings suggest that our H5N1 vaccine candidates would increase the availability of H5N1 vaccine doses at the onset of a new pandemic.

Background: Secondary bacterial infections after influenza can be a serious problem, especially in young children and the elderly, yet the efficacy of current vaccines is limited. Earlier work demonstrated that a replication-incompetent PB2-knockout (PB2-KO) influenza virus possessing a foreign gene in the coding region of its PB2 segment can serve as a platform for a bivalent vaccine.Methods: In the current study, we generated the PB2-KO virus expressing pneumococcal surface protein A (PspA), PB2-KO-PspA virus, the replication of which is restricted to PB2-expressing cells. We then examined the protective efficacy of intranasal immunization with this virus as a bivalent vaccine in a mouse model.Results: High levels of influenza virus-specific and PspA-specific antibodies were induced in the serum and airways of immunized mice. The intranasally immunized mice were protected from lethal doses of influenza virus or Streptococcus pneumoniae. These mice were also completely protected from secondary pneumococcal pneumonia after influenza virus infection.Conclusions: These findings indicate that our recombinant influenza virus serves as a novel and powerful bivalent vaccine against primary and secondary pneumococcal pneumonia as well as influenza. [ABSTRACT FROM AUTHOR]