Heavy metals such as zinc (Zn) and copper (Cu) may be associated with antibiotic resistance dissemination. Our aim was to investigate whether sub-lethal dosage of Zn and Cu may enhance plasmid transfer and subsequently resistance genes dissemination. Plasmid conjugation frequencies (PCF) were performed with Escherichia coli strains bearing IncL-bla _OXA-48 , IncA/C-bla _CMY-2 , IncI1-bla _CTX-M-1 , IncF-bla _CTX-M-1 , and IncX3-bla _NDM-5 as donors. Mating-out assays were performed with sub-dosages of zinc oxide (ZnO) and Cu sulfate (CuSO ₄ ). Quantification of the SOS response-associated gene expression levels and of the production of reactive oxygen species were determined. Increased PCF was observed for IncL, IncA/C, and IncX3 when treated with ZnO. PCF was only increased for IncL when treated with CuSO ₄ . The ROS production presented an overall positive correlation with PCF after treatment with ZnO for IncL, IncA/C, and IncX3. For CuSO ₄ treatment, the same was observed only for IncL. No increase was observed for expression of SOS response-associated genes under CuSO ₄ treatment, and under ZnO treatment, we observed an increase in SOS response-associated genes only for IncX3. Our data showed that sub-dosages of ZnO and CuSO ₄ could significantly enhance PCF in E. coli, with a more marked effect observed with IncL, IncA/C, and IncX3 scaffolds. Our study suggested that use of certain heavy metals is not the panacea for avoiding use of antibiotics in order to prevent the dissemination of antibiotic resistance.