Friday, April 19, 2024
4/19/2024
PROJECT SUMMARY/ABSTRACT Staphylococcus aureus causes community-acquired and healthcare-associated pneumonias that drive a decline in lung function and an increased risk of mortality, especially in patients with preexisting lung disease. These infections are difficult to eradicate because S. aureus can transition to adaptive phenotypes like persister cells and biofilm, which protect the bacteria against host phagocytes and antimicrobial factors, as well as antibiotics. A key component of the host immune response to pathogens is the production of regulatory metabolites like itaconate, which is synthesized in immune cells by the enzyme Immune-Responsive Gene 1 (Irg1). Itaconate balances pro- and anti-inflammatory signaling in host immune cells and exerts metabolic stress on bacteria. We recently demonstrated that itaconate inhibits S. aureus glycolysis and restructures staphylococcal metabolism to promote biofilm formation. Itaconate also limited the use of key energy-producing pathways, suggesting that it may promote the formation of antibiotic-tolerant persister cells. The role of itaconate in driving S. aureus lung infections cannot be completely understood without also investigating its impact on the host response to S. aureus. Preliminary data demonstrate that Irg1 is highly expressed in neutrophils and is associated with increased pro-inflammatory cytokine production during S. aureus lung infection, which differs from the anti- inflammatory effects of itaconate that have been defined in other cell types and infection models. This proposal aims to further investigate itaconate as a central mediator of the host-pathogen dynamic during S. aureus lung infection and a potential driver of S. aureus adaptation to the lung. Aim 1 will define the role of itaconate in regulating the host immune response to S. aureus by using metabolomics, phagocytosis assays, oxidative burst assays, and scRNA-sequencing to 1) determine if neutrophils are a major source of itaconate during S. aureus infection, 2) define the impact of itaconate on neutrophil effector function, and 3) identify the pathways that are differentially regulated by itaconate in neutrophils and other immune cell populations during in vivo infection. Aim 2 will define the role of itaconate in driving S. aureus adaptation to the lung by using bacterial qRT-PCR, ATP quantification, antibiotic tolerance assays, and flow cytometry to 1) determine if itaconate drives transcriptional changes that promote persister cell formation in vivo, 2) establish that itaconate exposure drives reduced energy metabolism and increased antibiotic tolerance ex vivo, and 3) quantify S. aureus division and growth in response to itaconate in vivo. Together, these studies will define the role of host immunometabolism in driving inflammation and bacterial persistence during S. aureus lung infection.
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