Antibodies recognizing protective pertussis toxin epitopes are preferentially elicited by natural infection versus acellular immunization

Abstract

Despite more than 50 years of vaccination, disease caused by the bacterium Bordetella pertussis persists, with rates increasing in industrialized countries over the past decade. This rise may be attributed to several factors, including increased surveillance, emergence of vaccine escape variants, waning immunity in adults, and the introduction of acellular subunit vaccines, which include chemically detoxified pertussis toxin (PTd). Two potently protective epitopes on pertussis toxin (PTx) are recognized by the monoclonal antibodies 1B7 and 11E6, which inhibit catalytic and cell-binding activities, respectively. In order to determine whether the PTx exposure route affects antibody responses to these epitopes, we analyzed sera from 30 adults with confirmed pertussis exposure and from 30 recently vaccinated adults for specific anti-PTx antibody responses and in vitro CHO cell neutralization titers. While overall titers against PTx and the genetically detoxified variant, PTg, containing the R9K and E129G substitutions, were similar in the two groups, titers against specific epitopes depended on the exposure route. Natural infection resulted in significantly higher titers of anti-PTx-subunit 1, 1B7-like, and 11E6-like antibodies, while acellular vaccination resulted in significantly higher titers of antibodies recognizing PTd. We also observed a correlation between in vitro protection and the presence of 1B7-like and 11E6-like antibodies. Notably, chemical detoxification, as opposed to genetic inactivation, alters the PTx tertiary and quaternary structure, thereby affecting conformational epitopes and recognition of PTx by 1B7 and 11E6. The lower levels of serum antibodies recognizing clinically relevant epitopes after vaccination with PTd support inclusion of PTg in future vaccines.

Fig. 1.

ELISA curves shows relative binding affinities for pertussis toxin (PTx [diaf]]), chemically detoxified PTx (PTd [■]), and genetically detoxified PTx (PTg [trif]), containing the R9K and E129G amino acid substitutions. (A) Binding of the human immune serum preparation P-IGIV. (B) Binding of the 1B7 monoclonal antibody and an IgG2a isotype control. (C) Binding of the 11E6 monoclonal antibody.

Fig. 2.

Comparison of vaccinated and exposed serum responses to PT variants, including enzymatically active PTx, genetically inactivated PTg, and chemically detoxified PTd, as measured by indirect ELISA. Significance is indicated as follows: *, P < 0.1 (by two-tailed t test for paired samples, comparing exposed and vaccinated populations for each PT variant). ■, exposed samples; □, vaccinated samples.

Fig. 3.

Summary of CHO cell neutralization assay. (A) Correlation between PTx antibody titers and protective CHO cell neutralization titers. (B) CHO cell protection assay, which was not statistically significant as determined by two-tailed t test for paired samples. ■, exposed samples; □, vaccinated samples.

Fig. 4.

Correlation between subsets of PTx-specific antibodies. (A) PTx-S1-220 versus PTx; (B) 1B7-like antibodies versus PTx; (C) 11E6-like antibodies versus PTx; (D) 1B7-like antibodies versus 11E6-like antibodies.

Fig. 5.

Summary of vaccinated and exposed serum PTx-specific responses. Assays included the indirect ELISAs to measure overall responses to PTx-S1-220 and competition assays to monitor the presence of 1B7- and 11E6-like antibodies. Significance is indicated as follows: *, P < 0.1; ***, P < 0.001 (by two-tailed t test for paired samples, comparing exposed and vaccinated populations within each assay). ■, exposed samples; □, vaccinated samples.

Fig. 6.

Correlation between IC₅₀ of protective antibodies and protective dilution in CHO assay. (A) 1B7-like antibodies; (B) 11E6-like antibodies.