Linking heavy metal(loid) stress to biofilm formation and conjugation: A critical review of experimental evidence

The environmental dissemination of antibiotic resistance genes (ARGs) and heavy metal resistance genes (HMRGs) is increasingly driven by complex interactions between metal contaminants and bacterial communities. Conjugative plasmids and biofilm formation are central to these processes, serving as key vectors and hotspots for horizontal gene transfer (HGT). This review systematically evaluates the influence of heavy metal(loid)s on bacterial biofilm development and plasmid-mediated ARG transfer by analyzing 63 peer-reviewed studies published between 2005 and 2026. Emphasis is placed on metal-specific and concentration-dependent effects, experimental conditions, and variability among bacterial strains. Essential metals, including copper (Cu) and zinc (Zn), frequently promoted conjugation and biofilm development at sub-inhibitory levels, whereas higher concentrations often induced toxicity and inhibited biofilm formation. Conjugative transfer was also enhanced by non-essential metal(loid)s, such as cadmium (Cd), mercury (Hg), arsenic (As), and silver (Ag) in certain studies, with Hg showing increases of up to approximately 4.5-fold. Modulatory factors, including reactive oxygen species (ROS) and changes in membrane permeability, were identified as critical mechanisms that may influence HGT. This review highlights the need for mechanistic and environmentally relevant investigations that integrate metal–biofilm–conjugation dynamics to gain deeper insight into the propagation of ARGs under realistic exposure conditions.