Probing Nano-Specific Interactions Between Bacteria and Antimicrobial Nanoparticles Using Microbial Community Changes and Gene Expression

Antimicrobial engineered nanomaterials (ENM) are increasingly incorporated into products despite limited understanding of the interactions between ENMs and bacteria that lead to toxic impacts. The hazard posed by increasing environmental release of antimicrobial ENMs is also poorly characterized. The overall objective of this thesis is to inform questions about the types of interactions that lead to an ENM inducing bacterial toxicity. Many antimicrobial ENMs are soluble, and the ion plays an important role in their toxicity. Some believe that, beyond release of ions, ENM toxicity is expected to derive from a nanoparticle (NP)-specific effect. This research compares bacterial responses to ENMs, their metal salts, and/or their transformed species within different experimental settings to improve our understanding of the interactions that enable ENM bacterial toxicity. The first objective is to characterize the potential hazard posed by pristine and transformed antimicrobial ENMs on microbial communities within a complex environmental system. One pair of ENMs (Ag0 and Ag2S) led to differential short-term impacts on surficial sediment microbial communities, while the other did not (CuO and CuS), showing that ENM transformation does not universally lead to distinct impacts. The metal ion (Cu2+) had a more profound microbial community impact than did any of the four ENMs. By 300 days the microbial community structure and composition re-converged, suggesting minimal long-term impacts of high pulse inputs of antimicrobial ENMs on microbial communities within complex environments. The second objective is to identify NP-specific effects of a common antimicrobial ENM on a model bacterium. Analysis of transcriptional responses identified NP-specific induction of a membrane stress responsive gene, providing evidence of a NP-specific effect. Otherwise, our results suggest that CuO NP toxicity triggers the same stress responses as does Cu2+, but at more moderate levels. Two ion treatments with the same total Cu input – one with pulse addition and one with gradual addition that was meant to better represent the slow dissolution of the CuO NP – led to temporally distinct responses. This calls for the use of more representative ion controls for comparison against soluble NP impacts in future nanotoxicity studies. The third objective is to investigate the potential use of CuO ENMs to reduce virulence and growth of an emerging bacterial pathogen. CuO NP exposure led to reduction in relative expression of three Staphylococcus aureus virulence factor genes, especially in methicillinresistant S. aureus (MRSA) clinical isolates. Growth was inhibited at high CuO NP concentrations for all four isolates, too. Comparison across all genes assayed showed isolatespecific transcriptional responses, but with NP- and ion-induced responses showing clear differences for each isolate, too. Altogether, this research contributes novel knowledge that will guide efforts to characterize potential hazard from release of ENMs into the environment and to apply ENMs for effective antibacterial treatment.