Mammary pathogenic E. coli (MPEC) form intracellular bacterial communities (IBC) in murine mammary alveolar epithelium. Lactating C57BL/6 TLR2−/− (A, B) or C3H/HeJ (C, D) mice 24 h after challenge by approximately 1000 CFUs via the teat canal. Mammary gland cryosections stained with DAPI (blue) and phalloidin-TRITC (red) (A, B), thin sections (1 µm) stained with toluidine blue (C) and TEM (D). Scale bars 50 µm (A, B), 10 µm (C), and 1000 nm (D). IBC of GFP-producing bacteria are demonstrated using confocal microscopy (showing a single Z-stack; white arrows in A, B), toluidine blue staining (white arrows in C) and TEM (black arrow in D). Inflammation is characterized by massive neutrophil recruitment into the alveolar spaces (yellow arrows in A, C) interacting with free and phagocytosed bacteria (GFP in A, B and toluidine blue in C). All images are representative of the entire sample. The histological morphology and pathology results were very similar for each gland in a given mouse and between mice.