Opposing effects of acellular and whole cell pertussis vaccines on Bordetella pertussis biofilm formation, Siglec-F+ neutrophil recruitment and bacterial clearance in mouse nasal tissues.

Despite global vaccination, pertussis caused by Bordetella pertussis (Bp) is resurging. Pertussis resurgence is correlated with the switch from whole cell vaccines (wPV) that elicit TH1/TH17 polarized immune responses to acellular pertussis vaccines (aPV) that elicit primarily TH2 polarized immune responses. One explanation for the increased incidence in aPV-immunized individuals is the lack of bacterial clearance from the nose. To understand the host and bacterial mechanisms that contribute to Bp persistence, we evaluated bacterial localization and the immune response in the nasal associated tissues (NT) of naïve and immunized mice following Bp challenge. Bp resided in the NT of unimmunized and aPV-immunized mice as biofilms. In contrast, Bp biofilms were not observed in wPV-immunized mice. Following infection, Siglec-F+ neutrophils, critical for eliminating Bp from the nose, were recruited to the nose at higher levels in wPV immunized mice compared to aPV immunized mice. Consistent with this observation, the neutrophil chemokine CXCL1 was only detected in the NT of wPV immunized mice. Importantly, the bacteria and immune cells were primarily localized within the NT and were not recovered by nasal lavage (NL). Together, our data suggest that the TH2 polarized immune response generated by aPV vaccination facilitates persistence in the NT by impeding the infiltration of immune effectors and the eradication of biofilms In contrast, the TH1/TH17 immune phenotype generated by wPV, recruits Siglec-F+ neutrophils that rapidly eliminate the bacterial burden and prevent biofilm establishment. Thus, our work shows that aPV and wPV have opposing effects on Bp biofilm formation in the respiratory tract and provides a mechanistic explanation for the inability of aPV vaccination to control bacterial numbers in the nose and prevent transmission.

Architecture and matrix assembly determinants of Bordetella pertussis biofilms on primary human airway epithelium.

Traditionally, whooping cough or pertussis caused by the obligate human pathogen Bordetella pertussis (Bp) is described as an acute disease with severe symptoms. However, many individuals who contract pertussis are either asymptomatic or show very mild symptoms and yet can serve as carriers and sources of bacterial transmission. Biofilms are an important survival mechanism for bacteria in human infections and disease. However, bacterial determinants that drive biofilm formation in humans are ill-defined. In the current study, we show that Bp infection of well-differentiated primary human bronchial epithelial cells leads to formation of bacterial aggregates, clusters, and highly structured biofilms which are colocalized with cilia. These findings mimic observations from pathological analyses of tissues from pertussis patients. Distinct arrangements (mono-, bi-, and tri-partite) of the polysaccharide Bps, extracellular DNA, and bacterial cells were visualized, suggesting complex heterogeneity in bacteria-matrix interactions. Analyses of mutant biofilms revealed positive roles in matrix production, cell cluster formation, and biofilm maturity for three critical Bp virulence factors: Bps, filamentous hemagglutinin, and adenylate cyclase toxin. Adherence assays identified Bps as a new Bp adhesin for primary human airway cells. Taken together, our results demonstrate the multi-factorial nature of the biofilm extracellular matrix and biofilm development process under conditions mimicking the human respiratory tract and highlight the importance of model systems resembling the natural host environment to investigate pathogenesis and potential therapeutic strategies.

Bps polysaccharide of Bordetella pertussis resists antimicrobial peptides by functioning as a dual surface shield and decoy and converts Escherichia coli into a respiratory pathogen.

Infections and disease caused by the obligate human pathogen Bordetella pertussis (Bp) are increasing, despite widespread vaccinations. The current acellular pertussis vaccines remain ineffective against nasopharyngeal colonization, carriage, and transmission. In this work, we tested the hypothesis that Bordetella polysaccharide (Bps), a member of the poly-ß-1,6-N-acetyl-D-glucosamine (PNAG/PGA) family of polysaccharides promotes respiratory tract colonization of Bp by resisting killing by antimicrobial peptides (AMPs). Genetic deletion of the bpsA-D locus, as well as treatment with the specific glycoside hydrolase Dispersin B, increased susceptibility to AMP-mediated killing. Bps was found to be both cell surface-associated and released during laboratory growth and mouse infections. Addition of bacterial supernatants containing Bps and purified Bps increased B. pertussis resistance to AMPs. By utilizing ELISA, immunoblot and flow cytometry assays, we show that Bps functions as a dual surface shield and decoy. Co-inoculation of C57BL/6J mice with a Bps-proficient strain enhanced respiratory tract survival of the Bps-deficient strain. In combination, the presented results highlight the critical role of Bps as a central driver of B. pertussis pathogenesis. Heterologous production of Bps in a non-pathogenic E. coli K12 strain increased AMP resistance in vitro, and augmented bacterial survival and pathology in the mouse respiratory tract. These studies can serve as a foundation for other PNAG/PGA polysaccharides and for the development of an effective Bp vaccine that includes Bps.