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The capsular group B meningococcal (MenB) four component vaccine (4CMenB) has been licensed for the prevention of invasive disease caused by MenB. The vaccine causes fever in infants, particularly when given in combination (concomitant) with other routinely-administered vaccines (routine), such as the standard diphtheria, tetanus, pertussis (DTP)-containing vaccine. To assess the suitability of a mouse immunisation model to study this phenomenon, we monitored temperature in mice after a second dose of routine vaccines, with or without 4CMenB, and compared the results with those in humans. Using this mouse model, we explored the reactogenicity of 4CMenB components by measuring changes in temperature, cytokines, and gene expression induced by 4CMenB, one of its components, wild-type or attenuated endotoxin outer membrane vesicles (OMVs), or lipopolysaccharide (LPS). A significant rise (p < 0.01) in temperature was observed in mice immunised with 4CMenB, wild-type OMVs, and LPS. RNA-sequencing of mouse whole blood revealed a gene signature shared by the 4CMenB, OMV, and LPS groups consisting of bacterial pattern recognition receptors and neutrophil activation marker genes. Sequencing of neutrophils isolated after concomitant 4CMenB identified cells expressing the OMV-associated genes Plek and Lcp1. Immunisation with 4CMenB or OMVs led to increased IL-6 in serum and significant upregulation (p < 0.0001) of prostaglandin-synthesising enzymes on brain tissue. These data demonstrate the suitability of a mouse model for assessing vaccine reactogenicity and strongly indicate that the fever following vaccination with 4CMenB in human infants is induced by endotoxin contained in the OMV component of the vaccine.

Gram-negative bacteria are refractory to the action of many antibiotics due to their impermeable outer membrane. An important player of the immune system is the complement system, a protein network in serum that directly kills Gram-negative bacteria through pore-formation by the Membrane Attack Complexes (MAC). We here show that the MAC rapidly perforates the outer membrane but that inner membrane damage, which is essential for killing, is relatively slow. Importantly, we demonstrate that MAC-induced outer membrane damage sensitizes Gram-negative bacteria to otherwise ineffective, Gram-positive-specific, antimicrobials. Synergy between serum and nisin was observed for 22 out of 53 tested Gram-negative clinical isolates and for multi-drug resistant (MDR) blood isolates. The in vivo relevance of this process is further highlighted by the fact that blood sensitizes a MDR K. pneumoniae strain to vancomycin. Altogether, these data imply that antibiotics that are considered ineffective to treat infections with Gram-negatives may have different functional outcomes in patients, due to the presence of the complement system.