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

Abstract

The 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 bacterial pathogenesis and host response remains unexplored. Here, we simultaneously investigate proteomic changes in A. baumannii and macrophages following infection in the absence or presence of the polymyxins. We discover that macrophages and polymyxins exhibit complementary effects to disarm several stress tolerance and survival strategies in A. baumannii, including oxidative stress resistance, copper tolerance, bacterial iron acquisition and stringent response regulation systems. Using the spoT mutant strains, we demonstrate that bacterial cells with defects in stringent response exhibit enhanced susceptibility to polymyxin killing and reduced survival in infected mice, compared to the wild-type strain. Together, our findings highlight that better understanding of host-pathogen-antibiotic interplay is critical for optimization of antibiotic use in patients and the discovery of new antimicrobial strategy to tackle multidrug-resistant bacterial infections. Author summary Bacterial response towards antibiotics is sensitive to the surrounding environment; however, how the host immune microenvironment affects the response of opportunistic pathogen Acinetobacter baumannii towards polymyxins remains largely unexplored. In this study, we established a host-pathogen-antibiotic tripartite in vitro model to examine the complex tripartite molecular interplay from both bacteria and macrophages perspectives, mimicking the physiological infection-treatment condition. This is the first comprehensive proteomics dataset for A. baumannii interacting with host cells, with an extraordinary coverage (i.e., 50%) of A. baumannii proteome even in infection conditions. For the first time, we report that macrophages and polymyxins utilize complementary mechanisms to disarm several A. baumannii stress tolerance strategies to persist in the macrophages. Our host-pathogen-antibiotic mechanistic study and the discovery of potential druggable targets (e.g., bacterial stringent stress response regulator SpoT) represent a significant advance, paving way to antibiotic optimization and drug discovery to tackle multidrug-resistant bacterial infections.