Neutralization of pertussis toxin by a single antibody prevents clinical pertussis in neonatal baboons.

Pertussis continues to cause considerable infant mortality world-wide, which could be addressed in part by passive immunization strategies. Antibody hu1B7 is a candidate therapeutic that potently neutralizes pertussis toxin in vitro, prevents leukocytosis in mice and treats established disease in weanling baboons as part of an antibody cocktail. Here, we evaluated the potential for hu1B7 and an extended half-life hu1B7 variant to prevent death, leukocytosis and other clinical symptoms in a newborn baboon model that mimics many aspects of human disease. We administered a single antibody dose to newborn baboons five weeks prior to experimental infection. While all animals were heavily colonized with Bordetella pertussis, prophylaxed animals showed significantly greater survival (P < 0.005), delayed and suppressed leukocytosis (P < 0.01) and enhanced clinical outcomes, including coughing (P < 0.01), as compared to controls. Together, this work demonstrates that a single neutralizing anti-PTx antibody is sufficient to prevent clinical pertussis symptoms.

Isolated CyaA-RTX subdomain from Bordetella pertussis: Structural and functional implications for its interaction with target erythrocyte membranes.

The 126-kDa Bordetella pertussis CyaA-hemolysin (CyaA-Hly) was previously expressed in Escherichia coli as a soluble precursor that can be acylated to retain hemolytic activity. Here, we investigated structural and functional characteristics of a ∼100-kDa isolated RTX (Repeat-in-ToXin) subdomain (CyaA-RTX) of CyaA-Hly. Initially, we succeeded in producing a large amount with high purity of the His-tagged CyaA-RTX fragment and in establishing the interaction of acylated CyaA-Hly with sheep red blood cell (sRBC) membranes by immuno-localization. Following pre-incubation of sRBCs with non-acylated CyaA-Hly or with the CyaA-RTX fragment that itself produces no hemolytic activity, there was a dramatic decrease in CyaA-Hly-induced hemolysis. When CyaA-RTX was pre-incubated with anti-CyaA-RTX antisera, the capability of CyaA-RTX to neutralize the hemolytic activity of CyaA-Hly was greatly decreased. A homology-based model of the 100-kDa CyaA-RTX subdomain revealed a loop structure in Linker II sharing sequence similarity to human WW domains. Sequence alignment of Linker II with the human WW-domain family revealed highly conserved aromatic residues important for protein-protein interactions. Altogether, our present study demonstrates that the recombinant CyaA-RTX subdomain retains its functionality with respect to binding to target erythrocyte membranes and the WW-homologous region in Linker II conceivably serves as a functional segment required for receptor-binding activity.

Pertussis whole cell vaccine: relation between intracerebral protection in mice and antibody response to pertussis toxin, filamentous hemagglutinin and adenylate cyclase.

N:NIH mice were vaccinated according to the WHO recommendations for the potency test with the Second International Standard for Pertussis Vaccine (ISPV). Blood for serological investigation was taken from the animals on day 14 post immunization before intracerebral challenge with Bordetella pertussis 18323 was done. The relationship between anti-pertussis toxin, anti-filamentous hemagglutinin and anti-adenylate cyclase antibody levels as measured by ELISA and protection from intracerebral challenge was studied. The proportion of surviving mice increased in correlation with increasing anti-PT titres; a protective level of 4 ELISA units/ml was found. Such relationship between protection against intracerebral challenge and antibody titres was not found for anti-FHA nor for anti-AC antibodies, thus suggesting that these antibodies do not play an important role in protection in this model. The excellent correlation between anti-PT antibody titres and protection suggests that the measure of anti-PT response could be a useful tool for estimating the potency of whole-cell vaccines. The development of an alternative method for testing the potency of pertussis whole-cell vaccines based on the anti-PT response should be considered.