Immunogenicity and effectiveness of a bivalent influenza A/H1N2 vaccine strain against seasonal human influenza A viruses in mice

Abstract

Background: Recent studies and reports have documented the ability of the co-circulating seasonal influenza A/H1N1 (ancestor: 2009 pandemic H1N1) and A/H3N2 to exchange their genetic segments, generating a novel H1N2 strain in different geographical localities around the world with an ability to infect human. This raises concerns and triggers alarms to develop a multivalent vaccine that can protect against the documented H1- and H3-type human influenza A viruses (IAVs).

Results: Here, we generated a PR8-based vaccine strain that carries the HA gene segment from the contemporary H1N1 virus while the NA gene segment was derived from a currently circulating influenza A/H3N2 strain. A recombinant PR8-based H1N2 vaccine strain (rgH1N2), engineered by reassortment between influenza A/H1N1 and A/H3N2 to mimic the documented human influenza A/H1N2, was used for immunization to provoke immunogenicity and cross-antigenicity against the H1- and H3-type human IAVs and was evaluated for its immunogenicity and effectiveness in mice. Following challenge infection of rgH1N2-vaccinated mice with contemporary influenza A/H1N1 and A/H3N2, results revealed that rgH1N2-vaccinated mice showed less viral shedding, more survival, and less body weight loss compared to control unvaccinated groups and vaccinated mice with rgH1N1 and rgH3N2.

Conclusions: This study highlights the applicability of the PR8-based H1N2 vaccine strain to protect against seasonal IAVs and emphasizes the role of both surface proteins, HA and NA, to stimulate protective and neutralizing antibodies against circulating influenza A/H1N1 and A/H3N2 strains.

Keywords: Immunogenicity; Influenza; Influenza A/H1N1; Influenza A/H1N2; Influenza A/H3N2; Vaccine effectiveness.

Fig. 1

Genetic illustration of rescued viruses that were tested as candidate vaccine seed strains and their parent strains

Fig. 2

The experimental timeline includes initial immunostimulation, booster vaccination, and challenge infection of immunized mice for up to 7 weeks post-vaccination (WPV). Eight-week-old black/6 mice were immunized at zero time of the experiment, followed by a booster vaccination at week 2 post-initial immunization to stimulate neutralizing antibodies against candidate vaccine strain (CVS) in mice. Challenge infection of immunized mice with the parent strains (PS) was done at week 5 post-vaccination and followed up for 2 weeks post-infection

Fig. 3

Phylogenetic analysis of the full HA (a) and NA (b) genetic segments from the studied candidate influenza vaccine strain A/Egypt/NRC098/2019 (H1N1). The tree was rooted with A/California/07/2009. Our candidate vaccine strain A/Egypt/NRC098/2019 (H1N1) is indicated in blue color. Vaccine strains A/Brisbane/02/2018 (egg-based, 2019–2020 NH influenza season), A/Guangdong-Maonan/SWL1536/2019 (egg-based, 2020–2021 influenza season), A/Hawaii/70/2019 (cell-based, 2020–2021 influenza season), A/Victoria/2570/2019 (egg-based, 2021–2022 influenza season), and A/Wisconsin/588/2019 (cell-based, 2021–2022 influenza season) are indicated with black circles

Fig. 4

Phylogenetic analysis of the full HA (a) and NA (b) sequences from the studied candidate influenza vaccine strain A/Egypt/NRC107/2019 (H3N2). The tree is rooted with A/Perth/16/2009 (H3N2). Our studied candidate vaccine strain is indicated in blue color. Vaccine strains A/Kansas/14/2017 (egg-based, 2019–2020 season), A/Hong Kong/2671/2019 (egg-based, 2020–2021), A/Hong Kong/45/2019 (cell-based, 2020–2021 season), and A/Cambodia/e0826360/2020 (2021–2022 season) are indicated with black circles

Fig. 5

Graphical representation of the parental and rescued viruses that were tested as candidate vaccine seed strains

Fig. 6

Immunogenicity of inactivated candidate vaccine strains as measured by plaque reduction neutralization assay (PRNT) for the collected sera from vaccinated and control mice groups (n = 16 per group) at different time points post-vaccination. The sera plaque neutralization assay was done against influenza A/H1N1 (a) and A/H3N2 (b). Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test. The significant differences are indicated (*P < 0.05 and non-significant = ns)

Fig. 7

Survival rate and body weight losses of vaccinated mice (n = 8 per each virus challenge) against circulating IAVs. Vaccinated mice that were infected with influenza A/H3N2 followed by monitoring the survival rate percent (a) and body weight loss percent (b). The second group of vaccinated mice was challenged with influenza A/H1N1 and protectiveness was investigated by following up on the survival rate percent (c) and body weight loss percent (d)

Fig. 8

Viral shedding or viral loads in lungs, nasal terminates, and nasal washes of the vaccinated and control mice (n = 8). The viral loads in the lungs and nasal terminates were detected in challenged mice with influenza A/H3N2 (a) and A/H1N1 (b) at day 3 post-infection. The viral loads in nasal washes were detected on days 3, 5, and 7 post-infection with H3N2 (c) or H1N1 (d) using semi-quantitative rtRT-PCR