Inactivated Whole Virus Particle Influenza Vaccine Induces Anti-Neuraminidase Antibodies That May Contribute to Cross-Protection against Heterologous Virus Infection

Abstract

Despite the use of vaccines, seasonal influenza remains a risk to public health. We previously proposed the inactivated whole virus particle vaccine (WPV) as an alternative to the widely used split vaccine (SV) for the control of seasonal and pandemic influenza based on the superior priming potency of WPV to that of SV. In this study, we further examined and compared the immunological potency of monovalent WPV and SV of A/California/7/2009 (X-179A) (H1N1) pdm09 (CA/09) to generate immune responses against heterologous viruses, A/Singapore/GP1908/2015 (IVR-180) (H1N1) pdm09 (SG/15), and A/duck/Hokkaido/Vac-3/2007 (H5N1) (DH/07) in mice. Following challenge with a lethal dose of heterologous SG/15, lower virus titer in the lungs and milder weight loss were observed in WPV-vaccinated mice than in SV-vaccinated ones. To investigate the factors responsible for the differences in the protective effect against SG/15, the sera of vaccinated mice were analyzed by hemagglutination-inhibition (HI) and neuraminidase-inhibition (NI) assays to evaluate the antibodies induced against viral hemagglutinin (HA) and neuraminidase (NA), respectively. While the two vaccines induced similar levels of HI antibodies against SG/15 after the second vaccination, only WPV-vaccinated mice induced significantly higher titers of NI antibodies against the strain. Furthermore, given the significant elevation of NI antibody titers against DH/07, an H5N1 avian influenza virus, WPV was also demonstrated to induce NA-inhibiting antibodies that recognize NA of divergent strains. This could be explained by the higher conservation of epitopes of NA among strains than for HA. Taking these findings together, NA-specific antibodies induced by WPV may have contributed to better protection from infection with heterologous influenza virus SG/15, compared with SV. The present results indicate that WPV is an effective vaccine for inducing antibodies against both HA and NA of heterologous viruses and may be a useful vaccine to conquer vaccine strain mismatch.

Immunogenicity and protective efficacy of a co-formulated two-in-one inactivated whole virus particle COVID-19/influenza vaccine

“…In our study, we showed that a single inoculation of unadjuvanted COVID-19 WPVs significantly prevented body weight loss, gene induction of pro-inflammatory molecules, and reduced virus titers in the lungs following SARS-CoV-2 infection, suggesting that our vaccine is sufficiently immunogenic and protective in mice, comparable to other studies using approved inactivated COVID-19 vaccines. The immunogenicity and protectivity of Flu WPV were comparable to those of the seasonal influenza vaccines used in our previous studies 32 , 42 , 43 , thereby further validating our vaccine design, product quality, and reliability. Furthermore, given comparable antibody titers in mice vaccinated with co-formulated WPV and individual WPVs against both viruses, co-formulation of either monovalent or quadrivalent Flu WPV with Co WPV does not seem to interfere with each other in inducing antibodies against SARS-CoV-2 and seasonal influenza viruses.…”

“…Following inactivation, the virus was concentrated by centrifugation at 53,900 g , 4 °C for 2 h then purified by sucrose density ultracentrifugation at 107,000 g , 4 °C for 1.5 h. The total protein concentration was measured using Nanodrop One spectrophotometer (Thermo Fisher Scientific). Flu WPV was prepared from A/California (H1N1) as previously described 43 . qFlu WPV containing the 2018–2019 virus strains [A/Singapore (H1N1), A/Singapore (H3N2), B/Phuket (Yamagata), B/Maryland (Victoria)] was kindly provided by KM biologics Co. Ltd (Kumamoto, Japan).…”

“…The neutralization assay was performed to measure neutralizing antibody titers in the serum of vaccinated mice as previously described for influenza viruses 42 , 43 . To measure neutralizing antibody titers against SARS-CoV-2, monolayers of Vero TMPRSS2 cells were prepared in 6-well plates by seeding 1.425 × 10 6 cells/well in 3 mL DMEM and incubated overnight at 37 °C in 5% CO 2 .…”

“…At 3 and 5 dpi, some mice were humanely euthanized, and lung tissues (right lobes) were harvested for measurement of virus titers and gene expression of proinflammatory cytokines and chemokines. Influenza viral loads in the lungs were determined as described previously 32 , 43 . To measure the SARS-CoV-2 viral load, monolayers of Vero TMPRSS2 cells were prepared by seeding 1.425 × 10 6 cells in 3 mL DMEM and incubated overnight at 37 °C in 5% CO 2 .…”

A single immunization with H5N1 virus-like particle vaccine protects chickens against divergent H5N1 influenza viruses and vaccine efficacy is determined by adjuvant and dosage

“…In this study, the H5N1 VLP vaccine required a large dose of antigen to achieve full protection against antigenically divergent H5N1 virus challenge, which needed further modification. Studies have shown that neuraminidase (NA) [ 41 ], nucleoprotein (NP) [ 20 ], influenza conserved epitopes such as HA-stalk, M2e [ 42 ], Toll-like receptor ligands [ 19 , 34 ], and immunological proteins adjuvants such as LTB [ 19 ] contribute to cross-protection against heterologous virus challenge. These immunological components can be incorporated into H5N1 VLP to enhance the cross-protection capacity of H5N1 VLP vaccines.…”