Immune responsiveness influences Staphylococcus aureus persistence via citrate metabolic flux during prosthetic joint infection 2257191

Abstract

Abstract Introduction Prosthetic joint infection (PJI) is a complication following arthroplasty that occurs in 1-3% of patients, with ∼50% of cases caused by Staphylococcus aureus (S. aureus) that forms biofilm. Biofilms are difficult to eradicate due to their recalcitrance to antibiotics and immune-mediated clearance, resulting in chronic symptoms that are debilitating for PJI patients. Bacterial-derived metabolites play a central role in biofilm infection, where several tricarboxylic acid cycle (TCA) intermediates are key for biofilm development. However, little information is available regarding how leukocytes influence S. aureus TCA cycle activity and resultant adaptations by the bacteria, which can inform future therapeutic targets to promote biofilm clearance. Methods S. aureus strains lacking the TCA cycle enzymes, citrate synthase (ΔgltA), aconitase (ΔacnA), isocitrate dehydrogenase (Δicd), and a double mutant lacking citrate synthesis and transport (ΔgltA/ΔcitM) were assessed for intracellular survival in macrophages (Mϕs) and a mouse model of PJI. Results Both ΔacnA and Δicd had increased survival within Mϕs at 4 h; however, this was transient, with intracellular burdens reaching that of WT S. aureus by 24 h. The early survival advantage of α”acnA and α”icd strains was partially driven by mitochondrial reactive oxygen species (mtROS), since the mtROS inhibitor MitoTEMPO negated this phenotype. In a mouse model of PJI, bacterial burden was significantly decreased with α”gltA compared to WT at day 7 post-infection. This difference was transient, as no changes in biofilm burden were observed at day 14. However, the ΔgltA/ΔcitM strain displayed reduced bacterial burden at day 14. Conclusion Citrate synthesis is important for early biofilm infection; however, S. aureus employs compensatory mechanisms such as citrate import to overcome TCA cycle loss. Future studies will explore how leukocytes reprogram S. aureus TCA cycle activity and how S. aureus adapts to immune pressure. Funding Source UNMC Fellowship; UNMC Neuroimmunology of Disease Training Program T32 Topic Categories Microbial, Parasitic, and Fungal Immunology (MPF)