A novel cystic fibrosis-mimetic Pseudomonas auxotrophic vaccine is protective in vivo and is associated with IL-17/IgA mucosal Immunity.

Pseudomonas aeruginosa (P.a) is a Gram-negative opportunistic pathogen that poses a major global health threat, particularly in immunocompromised individuals, patients with cystic fibrosis, and those with burn injuries or ventilator-associated pneumonia. Despite intense efforts, no vaccine is available for human use. In this context, live attenuated vaccines represent a promising but underexplored approach, offering the potential to elicit robust, long-lasting, and multifaceted immune responses including inducing trained immunity. Here, we sub-cultured ΔLasB-PAO1 (a P.a strain we have previously shown to have reduced virulence) in artificial sputum medium, a culture medium mimicking cystic fibrosis (CF) sputum in which bacteria often show auxotrophy. We showed that such strain ("V" for vaccine) was auxotrophic, less virulent, and had characteristics of "CF-like strains." Crucially, "V" induced both local (IgA) and systemic humoral responses as well as memory Th17 immune responses, and could, when administered in the lung, but not intra-muscularly, protect mice against a lethal PAO1-WT infection. Overall, the present study demonstrates that our vaccine formulation, in addition to providing an advantageous auxotrophic phenotype adapted to the CF setting, was efficient, when given mucosally and was associated with a pathway involving Th17 responses and secretory IgA, a critical barrier that neutralizes pathogens before tissue invasion.