TheAchromobacterType 3 secretion system drives pyroptosis and immunopathology via independent activation of NLRC4 and NLRP3 inflammasomes

Abstract

AbstractAchromobacterspecies are newly recognized opportunistic, pro-inflammatory Gram-negative pathogens in immunocompromised individuals, but how they interact with the innate immune system to drive inflammation is poorly understood. We createdsctV(Type 3 Secretion System baseplate) mutants in threeAchromobacterclinical isolates from two species and showed that all three required the T3SS to induce cell death in human macrophages. Mutating other critical T3SS components also abolished cell death, which was restored by genetic complementation. Cell death ofAchromobacter-infected macrophages was contact-dependent, enhanced by bacterial internalisation, and caused by inflammasome-dependent pyroptosis (typified by Gasdermin-D cleavage and IL-1β secretion). Macrophages deficient in the inflammasome sensors NLRC4 or NLRP3 underwent pyroptosis upon bacterial internalization but those deficient in both NLRC4 and NLRP3 did not, suggesting either sensor can mediate pyroptosis induction in a T3SS-dependent manner. Detailed analysis of the intracellular trafficking of one isolate indicated that the intracellular bacteria reside in an acidic LAMP-1/dextran-positive membrane compartment. Using an intranasal mouse infection model, we observed thatAchromobacterdamages lung structure and causes severe illness, contingent on a functional T3SS. Together, we demonstrate thatAchromobacterspecies can survive phagocytosis by macrophages and promote macrophage cell death and inflammation by redundant mechanisms of pyroptosis induction.