Distinct Antibacterial Activities of Nanosized Cationic Liposomes against Gram-Negative Bacteria Correlate with Their Heterogeneous Fusion Interactions

Nanosized liposomes are promising vehicles for delivering antibiotics to bacterial pathogens. Their fusion with bacterial membranes is hypothesized to enhance the potency of some antibiotics, but the mechanisms by which liposomes interact with bacteria are poorly understood. Liposome fusion with bacteria is typically detected using ensemble-averaged approaches based on the presumption that bacteria of the same species interact with liposomes homogeneously. Here, we challenge this presumption with observations of heterogeneous fusion interactions between cationic liposomes and the Gram-negative bacterium Escherichia coli. Importantly, the different interactions observed lead to different antibacterial outcomes. In one subpopulation, liposomes fused with both the outer and inner membranes of E. coli. In the second subpopulation, they fused only with the outer membrane. Liposomes caused permeabilization of the bacterial cell envelope and cell death only when they fuse with both the outer and inner membranes of E. coli. Compared to fusion, electrostatic binding of liposomes with the bacterial membrane has a significantly less effect in inhibiting bacterial growth. We further found that the fusion interactions of liposomes with bacteria vary across bacterial species. For the entire population of Vibrio cholerae studied, liposomes fused with only the outer membrane and thus had a negligible inhibitory effect. Our results demonstrate the significant level of heterogeneity in the interaction between nanomaterials and bacteria and highlight the consequent need to assess such interactions by mechanistic analysis at the level of the single cell.