Original antigenic sin priming of influenza virus hemagglutinin stalk antibodies

Abstract

Immunity to influenza viruses can be long-lived, but reinfections with antigenically distinct viral strains and subtypes are common. Reinfections can boost antibody responses against viral strains first encountered in childhood through a process termed "original antigenic sin." It is unknown how initial childhood exposures affect the induction of antibodies against the hemagglutinin (HA) stalk domain of influenza viruses. This is an important consideration since broadly reactive HA stalk antibodies can protect against infection, and universal vaccine platforms are being developed to induce these antibodies. Here we show that experimentally infected ferrets and naturally infected humans establish strong "immunological imprints" against HA stalk antigens first encountered during primary influenza virus infections. We found that HA stalk antibodies are surprisingly boosted upon subsequent infections with antigenically distinct influenza A virus subtypes. Paradoxically, these heterosubtypic-boosted HA stalk antibodies do not bind efficiently to the boosting influenza virus strain. Our results demonstrate that an individual's HA stalk antibody response is dependent on the specific subtype of influenza virus that they first encounter early in life. We propose that humans are susceptible to heterosubtypic influenza virus infections later in life since these viruses boost HA stalk antibodies that do not bind efficiently to the boosting antigen.

Keywords: hemagglutinin; influenza virus; original antigenic sin.

Fig. 1.

Heterosubtypic boosting of HA stalk antibodies in ferrets. Ferrets were infected with H1N1 or H3N2 and then reinfected with H1N1, H3N2, or influenza B virus (IBV) 79 to 84 d later. Sera were collected at different times after the primary (1°) and secondary (2°) infections, and serological assays were completed. (A–C) Animals were infected with H1N1 and then reinfected with H3N2 (blue), infected with H3N2, and then reinfected with H1N1 (red) or infected with H1N1 and reinfected with IBV (gray). (D–F) Animals were infected with H1N1 and then reinfected with H3N2 (blue), infected with H3N2 and then reinfected with H1N1 (red), or infected with H1N1 and reinfected with IBV (gray). (A) Total H1 antibodies were quantified by ELISA, (B) H1 head-reactive antibodies were quantified by HAI assays, (C) H1 stalk-reactive antibodies were quantified by ELISA, (D) total H3 antibodies were quantified by ELISA, (E) H3 head-reactive antibodies were quantified by HAI assays, and (F) H3 stalk-reactive antibodies were quantified by ELISA. Data are presented as endpoint titrations or µg/mL equivalents based on a standard monoclonal antibody. Three to six ferrets were included for each experimental group. Data are shown as mean ± SEM.

Fig. 2.

Heterosubtypic-boosted HA stalk antibodies from ferrets do not bind efficiently to the boosting antigen. Serum samples were collected from ferrets before and 7, 14, and 28 d after secondary (2°) heterosubtypic influenza virus infection. Samples were incubated with 293F cells expressing H1 HA (white), H3 HA (gray), or no HA (black), and (A) H1 stalk and (B) H3 stalk antibody levels remaining in the unabsorbed fraction were quantified by ELISA. Serum from three to six ferrets was included for each experimental group. Data are shown as mean ± SEM. Titers were compared using a one-way ANOVA with Tukey’s posttest at each time point on log-transformed data (*P < 0.05).

Fig. 3.

H3N2 influenza infection boosts H1 stalk antibodies in children with evidence of prior H1N1 exposure. Serum samples were collected from 18 children before and after PCR-confirmed H3N2 infection in 2017. Samples were stratified into different experimental groups based on prior H1 and H3 exposure history, which we inferred based on H1 and H3 antibody reactivity prior to H3N2 infection in 2017 (SI Appendix, Fig. S3). We quantified (A–C) H3 stalk and (D–F) H1 stalk antibodies in serum from children with no evidence of prior exposure (n = 7; A and D), H3N2 prior exposure (n = 6; B and E), and H1N1 prior exposure (n = 5; C and F). Each circle represents a sample from one individual. Data are shown as mean ± SEM. Pre- and postinfection antibody titers were compared using a paired t test on log-transformed data (*P < 0.05).

Fig. 4.

Heterosubtypic-boosted HA stalk antibodies from children do not bind efficiently to the boosting antigen. Serum samples were collected from 18 children before and after PCR-confirmed H3N2 infection in 2017. Samples were stratified into different experimental groups based on prior H1 and H3 exposure history, which we inferred based on H1 and H3 antibody reactivity prior to H3N2 infection in 2017 (SI Appendix, Fig. S3). We collected samples from individuals with (A) no evidence of prior exposure (n = 7), (B) evidence of prior H3N2 exposure (n = 6), and (C) evidence of prior H1N1 exposure (n = 5). Samples were incubated with 293F cells expressing H1 HA (white), H3 HA (gray), or no HA (black), and H1 stalk antibody levels remaining in the unabsorbed fraction were quantified by ELISA. Each circle represents a sample from one individual. Data are shown as mean ± SEM. Antibody titers were compared using a one-way ANOVA with Tukey’s posttest on log-transformed data (*P < 0.05).