Copper nanoparticles and copper ions promote horizontal transfer of plasmid-mediated multi-antibiotic resistance genes across bacterial genera

Abstract

The spread of antibiotic resistance has become a major concern for public health. As emerging contaminants, various metallic nanoparticles (NPs) and ionic heavy metals have been ubiquitously detected in various environments. Although previous studies have indicated NPs and ionic heavy metals could exhibit co-selection effects for antibiotic resistance, little is known about whether and how they could promote antibiotic resistance spread via horizontal gene transfer across bacterial genera. This study, we report both CuO NPs and copper ions (Cu²⁺) could stimulate the conjugative transfer of multiple-drug resistance genes. When exposing bacteria to CuO NPs or Cu²⁺ at environmental-relevant and sub-inhibitory concentrations (e.g., 1-100 μmol/L), conjugation frequencies of plasmid-encoded antibiotic resistance genes across genera (i.e., from Escherichia coli to Pseudomonas putida) were significantly enhanced (p < 0.05). The over-production of reactive oxygen species played a crucial role in promoting conjugative transfer. Genome-wide RNA and protein sequencing suggested expressional levels of genes and proteins related to oxidative stress, cell membrane permeability, and pilus generation were significantly up-regulated under CuO NPs and Cu²⁺ exposure (p < 0.05). This study provides insights in the contributions of NPs and heavy metals on the spread of antibiotic resistance.

Keywords: Antibiotic resistance genes; Conjugative transfer; Copper ions; Copper oxide nanoparticles; Genome-wide RNA sequencing; Horizontal gene transfer.