Correlative proteomics identify the key roles of stress tolerance strategies in Acinetobacter baumannii in response to polymyxin and macrophages

Opportunistic pathogen Acinetobacter baumannii possesses stress tolerance strategies against host innate immunity and antibiotic killing. However, how the host-pathogen-antibiotic interaction affects the overall molecular regulation of pro- and anti-pathogenic machineries remains unexplored. Here, we simultaneously investigate proteomic changes in A. baumannii and macrophages following infection in the absence or presence of the last-line polymyxins. We discover that macrophages and polymyxins exhibit complementary effects to disarm several A. baumannii stress tolerance and survival strategies, including oxidative stress resistance, copper tolerance, bacterial iron acquisition and stringent response regulation systems. Using bacterial mutants with impaired stringent response associated (p)ppGpp synthetase/hydrolase spoT we demonstrate that spot mutants exhibit significantly enhanced susceptibility to polymyxin killing and reduced in vivo survivability compared to the wild-type strain. Together, our findings highlight that improved understanding of host-pathogen-antibiotic interplay is critical for optimisation of antibiotic use in patients and discovery of new antibiotics to tackle multidrug-resistant bacterial infections.