Multimodal Imaging Reveals Spatial Host-Pathogen Microenvironments in Escherichia coli Meningoencephalitis

Experimental bacterial meningitis is often analyzed as microbial burden and inflammation, but the tissue-level organization of host-pathogen interplay remains poorly resolved. Here, we use multimodal imaging to map Escherichia coli meningoencephalitis as a spatially organized process across infected rat brain tissue and cerebrospinal fluid (CSF). A low-dose intracerebral inoculum expands within 24 h into an anatomically structured infection involving ventricular, meningeal, and perivascular interfaces. Scanning electron microscopy reveals biofilm-like multicellular E. coli aggregates embedded in extracellular material, consistent with neutrophil extracellular traps. MALDI mass spectrometry imaging detects aerobactin and salmochelin-derived metabolites, whereas intact enterobactin is not detected despite favorable analytical sensitivity. These bacterial iron-acquisition signals partially overlap with calprotectin proteoforms, defining ventricular and periventricular metal-conflict microenvironments. Spatial peptidomics identifies infection-enriched antimicrobial territories dominated by rat neutrophil peptides RatNP-2, RatNP-3, and RatNP-4. Endogenous proenkephalin-derived neuropeptides are reduced in basal ganglia regions. Finally, cerebrospinal fluid captures both bacterial siderophores and host antimicrobial peptides with lipocalin-2 protein, linking tissue-resolved host-pathogen chemistry to a proximal diagnostic fluid. Together, these data show that experimental E. coli meningoencephalitis is a spatially organized host-pathogen process. Bacterial communities, nutritional immunity, antimicrobial peptides, and neuropeptide networks occupy distinct but connected CNS niches, components of which are recoverable in CSF.