Limited information exists on how bacterial resistance to antibiotics is acquired and altered in response to short-term metal stress, and what the prevailing pathways are. Here the precursor mechanisms of development of bacterial antibiotic resistance mediated by oxidative stress induce under sub-lethal Cu2+ exposure were explored. The results showed that the overall level of antibiotic resistance in wild-type Escherichia coli and antibiotic-resistant E. coli was enhanced under 4 and 20 mg/L Cu2+ exposure, as demonstrated by the 2- to 8-fold increase in minimum inhibitory concentration (MIC). The MIC correlated with the increase of the cellular ROS generation and the enhancement of the antioxidant enzyme activity (p < 0.05), suggesting that changes in antibiotic resistance under sub-lethal Cu2+ exposure could be associated with oxidative stress. Likewise, enhanced cell membrane permeability and an increase in the number of bacteria entering the viable but non culturable (VBNC) state contributed to bacterial resistance to antibiotics. Moreover, the variance partitioning analysis demonstrated that the alterations of the antibiotic resistance phenotype of wild-type E. coli was mainly caused by oxidative stress-mediated increase in cell membrane permeability and entry into the VBNC state. The development of antibiotic resistance in resistant E. coli was primarily attributed to changes in the abundance and horizontal transfer ability of its antibiotic resistance genes, both of which contributed up to 20 %. Taken together the results allowed to propose a conseptual scheme on developing bacterial antibiotic resistance mediated by oxidative stress under sub-lethal Cu2+ exposure. This result provided a strong basis for reduction of early bacterial resistance.