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

<i>Ab Initio</i> Atomistic Thermodynamics Study of the (001) Surface of LiCoO₂ in a Water Environment and Implications for Reactivity under Ambient Conditions

We use GGA + <i>U</i> methodology to model the bulk and surface structure of varying stoichiometries of the (001) surface of LiCoO₂. The DFT energies obtained for these surface-slab models are used for two thermodynamic analyses to assess the relative stabilities of different surface configurations, including hydroxylation. In the first approach, surface free energies are calculated within a thermodynamic framework, and the second approach is a surface-solvent ion exchange model. We find that, for both models, the −CoO–H_1/2 surface is the most stable structure near the O-rich limit, which corresponds to ambient conditions. We find that surfaces terminated with Li are higher in energy, and we go on to show that H and Li behave differently on the (001) LiCoO₂ surface. The optimized geometries show that terminal Li and H occupy nonequivalent surface sites. In terms of electronic structure, Li and H terminations exhibit distinct bandgap characters, and there is also a distinctive distribution of charge at the surface. We go on to probe how the variable Li and H terminations affect reactivity, as probed through phosphate adsorption studies

Evidence for Considerable Metal Cation Concentrations from Lithium Intercalation Compounds in the Nano–Bio Interface Gap

An experimental investigation of how electrostatics and ion dissolution impact the interaction between nanosheets of lithium intercalation compounds and supported lipid bilayers has revealed evidence for considerable metal cation concentrations in the nanosheets–bilayer (the “nano–bio interface”) gap. Specifically, elevated concentrations of aqueous metal ions in the 1 mg/L concentration regime produce vibrational sum frequency generation signal intensity changes that are commensurate with the induction of compositional membrane asymmetry. This outcome is consistent with the notion that the induction of bilayer asymmetry by LiCoO₂ nanosheets occurs through a noncontact mechanism that primarily involves the interaction of negatively charged lipids with dissolved ions concentrated within the electrical double layers present in the nano–bio interface gap. Our findings provide a possible avenue for redesign strategies that mitigate noncontact interactions between nanomaterials and biological interfaces, enabling the design of new energy storage materials with reduced environmental impacts

Influence of Nanoparticle Morphology on Ion Release and Biological Impact of Nickel Manganese Cobalt Oxide (NMC) Complex Oxide Nanomaterials

Lithium intercalation compounds such as nickel manganese cobalt oxides (Li_<i>x</i>Ni_<i>y</i>Mn_<i>z</i>Co_{1–<i>y</i>–<i>z</i>}O₂, 0 < <i>x</i>, <i>y</i>, <i>z</i> < 1, or NMCs) are complex transition metal oxides of increasing interest in nanoscale form for applications in electrochemical energy storage and as tunable catalysts. These materials exhibit sheetlike structures that expose low-energy basal planes and higher-energy edge planes in relative amounts that vary with the nanoparticle morphology. Yet there is little understanding of how differences in nanoparticle morphology and exposed crystal planes affect the biological impact of this class of technologically relevant nanomaterials. We investigated how changing nanoparticle morphology from two-dimensional (001)-oriented nanosheets to three-dimensional nanoblocks affects the release of ions and the resulting biological impact using <i>Shewanella oneidensis</i> MR-1 as a model organism. NMC nanoparticles were synthesized in sheetlike morphology and then converted to block morphologies by heating, leading to two morphologies of identical chemical composition that were compared to a commercially available NMC. Ion dissolution studies reveal that NMC nanomaterials release transition metal ions incongruently (Ni > Co > Mn) in amounts that vary with nanoparticle morphology. However, when normalized by the specific surface areas, the rates of release of each transition metal from flakes, blocks, and commercial material were equivalent. Similarly, the impact on <i>S. oneidensis</i> MR-1 was different when using mass-based dosing but was nearly identical using surface-area-normalized exposure dosing. Our results show that even though nanosheets and nanoblocks expose different crystal faces with significantly different surface energies, the rate of ion release is independent of the distribution of crystal faces exposed and depends only on the total surface area exposed. These data suggest that the key protonation steps that control release of transition metals do not depend on the degree of coordination of the initially exposed surface, providing insights into the molecular-level factors that influence the environmental impact of complex metal oxide nanomaterials. Our results have significant implications for establishing methodologies to assess toxicity of reactive nanomaterials

Alteration of Membrane Compositional Asymmetry by LiCoO₂ Nanosheets

Given the projected massive presence of redox-active nanomaterials in the next generation of consumer electronics and electric vehicle batteries, they are likely to eventually come in contact with cell membranes, with biological consequences that are currently not known. Here, we present nonlinear optical studies showing that lithium nickel manganese cobalt oxide nanosheets carrying a negative ζ-potential have no discernible consequences for lipid alignment and interleaflet composition in supported lipid bilayers formed from zwitterionic and negatively charged lipids. In contrast, lithiated and delithiated LiCoO₂ nanosheets having positive and neutral ζ-potentials, respectively, alter the compositional asymmetry of the two membrane leaflets, and bilayer asymmetry remains disturbed even after rinsing. The insight that some cobalt oxide nanoformulations induce alterations to the compositional asymmetry in idealized model membranes may represent an important step toward assessing the biological consequences of their predicted widespread use

Growth-Based Bacterial Viability Assay for Interference-Free and High-Throughput Toxicity Screening of Nanomaterials

Current high-throughput approaches evaluating toxicity of chemical agents toward bacteria typically rely on optical assays, such as luminescence and absorbance, to probe the viability of the bacteria. However, when applied to toxicity induced by nanomaterials, scattering and absorbance from the nanomaterials act as interferences that complicate quantitative analysis. Herein, we describe a bacterial viability assay that is free of optical interference from nanomaterials and can be performed in a high-throughput format on 96-well plates. In this assay, bacteria were exposed to various materials and then diluted by a large factor into fresh growth medium. The large dilution ensured minimal optical interference from the nanomaterial when reading optical density, and the residue left from the exposure mixture after dilution was confirmed not to impact the bacterial growth profile. The fractions of viable cells after exposure were allowed to grow in fresh medium to generate measurable growth curves. Bacterial viability was then quantitatively correlated to the delay of bacterial growth compared to a reference regarded as 100% viable cells; data analysis was inspired by that in quantitative polymerase chain reactions, where the delay in the amplification curve is correlated to the starting amount of the template nucleic acid. Fast and robust data analysis was achieved by developing computer algorithms carried out using R. This method was tested on four bacterial strains, including both Gram-negative and Gram-positive bacteria, showing great potential for application to all culturable bacterial strains. With the increasing diversity of engineered nanomaterials being considered for large-scale use, this high-throughput screening method will facilitate rapid screening of nanomaterial toxicity and thus inform the risk assessment of nanoparticles in a timely fashion