Staphylococcus aureus biofilm infection impairs efferocytosis in wound-macrophages via induction of a MARCOhiMERTKlo subset.

Chronic, non-healing wounds are sustained by Staphylococcus aureus biofilms, yet how biofilm reprograms the immune cells tasked with resolving injury has remained unclear. We show that biofilm-derived soluble cues, not bacterial contact, generate a previously undescribed biofilm-associated macrophage (BAM) subset marked by high Macrophage receptor with collagenous structure (MARCO) and suppressed MERTK, which recognizes apoptotic cells but fails to engulf them. Using isogenic S. aureus USA300 variants of graded biofilm capacity, only high biofilm-conditioned medium elicited this state in human macrophages; MARCO neutralization restored corpse clearance. Mechanistically, biofilm factors drove phospho-C/EBPβ into the nucleus to repress MERTK, severing sensing from engulfment. Single-cell and Xenium spatial transcriptomics, with cytometry by time of flight (CyTOF) and PhenoCycler, localized BAMs to biofilm-proximal niches as a hybrid CD64+CD163+ state outside M1/M2 categories, populating 60%-80% of infected chronic wounds. In vivo, myeloid-restricted MARCO overexpression by tissue nanotransfection recapitulated impaired efferocytosis and persistent inflammation, nominating MARCOhiMERTKlo macrophages as a druggable checkpoint in biofilm-associated disease.