Influence of the Spatial Distribution of Cationic Functional Groups at Nanoparticle Surfaces on Bacterial Viability and Membrane Interactions.

While positively charged nanomaterials induce cytotoxicity in many organisms, much less is known about how the spatial distribution and presentation of molecular surface charge impact nanoparticle-biological interactions. We systematically functionalized diamond nanoparticle surfaces with five different cationic surface molecules having different molecular structures and conformations, including four small ligands and one polymer, and we then probed the molecular-level interaction between these nanoparticles and bacterial cells. Shewanella oneidensis MR-1 was used as a model bacterial cell system to investigate how the molecular length and conformation of cationic surface charges influence their interactions with the Gram-negative bacterial membranes. Nuclear magnetic resonance (NMR) and X-ray photoelectron spectroscopy (XPS) demonstrate the covalent modification of the nanoparticle surface with the desired cationic organic monolayers. Surprisingly, bacterial growth-based viability (GBV) and membrane damage assays both show only minimal biological impact by the NPs functionalized with short cationic ligands within the concentration range tested, yet NPs covalently linked to a cationic polymer induce strong cytotoxicity, including reduced cellular viability and significant membrane damage at the same concentration of cationic groups. Transmission electron microscopy (TEM) images of these NP-exposed bacterial cells show that NPs functionalized with cationic polymers induce significant membrane distortion and the production of outer membrane vesicle-like features, while NPs bearing short cationic ligands only exhibit weak membrane association. Our results demonstrate that the spatial distribution of molecular charge plays a key role in controlling the interaction of cationic nanoparticles with bacterial cell membranes and the subsequent biological impact. Nanoparticles functionalized with ligands having different lengths and conformations can have large differences in interactions even while having nearly identical zeta potentials. While the zeta potential is a convenient and commonly used measure of nanoparticle charge, it does not capture essential differences in molecular-level nanoparticle properties that control their biological impact.

Chronic exposure to complex metal oxide nanoparticles elicits rapid resistance in Shewanella oneidensis MR-1.

Engineered nanoparticles are incorporated into numerous emerging technologies because of their unique physical and chemical properties. Many of these properties facilitate novel interactions, including both intentional and accidental effects on biological systems. Silver-containing particles are widely used as antimicrobial agents and recent evidence indicates that bacteria rapidly become resistant to these nanoparticles. Much less studied is the chronic exposure of bacteria to particles that were not designed to interact with microorganisms. For example, previous work has demonstrated that the lithium intercalated battery cathode nanosheet, nickel manganese cobalt oxide (NMC), is cytotoxic and causes a significant delay in growth of Shewanella oneidensis MR-1 upon acute exposure. Here, we report that S. oneidensis MR-1 rapidly adapts to chronic NMC exposure and is subsequently able to survive in much higher concentrations of these particles, providing the first evidence of permanent bacterial resistance following exposure to nanoparticles that were not intended as antibacterial agents. We also found that when NMC-adapted bacteria were subjected to only the metal ions released from this material, their specific growth rates were higher than when exposed to the nanoparticle. As such, we provide here the first demonstration of bacterial resistance to complex metal oxide nanoparticles with an adaptation mechanism that cannot be fully explained by multi-metal adaptation. Importantly, this adaptation persists even after the organism has been grown in pristine media for multiple generations, indicating that S. oneidensis MR-1 has developed permanent resistance to NMC.