Overwhelming growth of bacterial biofilms on different metal-based pipeline materials are intractable and pose a serious threat to public health when tap water flows though these pipelines. Indeed, the underlying mechanism of biofilm growth on the surface of different pipeline materials deserves detailed exploration to provide subsequent implementation strategies for biofilm control. Thus, in this study, how bacteria response to their encounters was explored, when they inhabit different metal-based pipeline substrates. Results revealed that bacteria proliferated when they grew on stainless steel (SS) and titanium sheet (Ti), quickly developing into bacterial biofilms. In contrast, the abundance of bacteria on copper (Cu) and nickel foam (Ni) substates decreased sharply by 4–5 logs within 24 h. The morphological shrinkage and shortening of bacterial cells, as well as a sudden 64-fold increase of carbohydrate content in extracellular polymeric substances (EPS), were observed on Cu substrate. Furthermore, generation of reactive oxygen species and fluctuation of enzymatic activity demonstrated the destruction of redox equilibrium in bacteria. Bacteria cultured on Cu substrate showed the strongest response, followed by Ni, SS and Ti. The oxidative stress increased quickly during the growth of bacterial biofilm, and almost all tested metal transporter-related genes were upregulated by 2–11 folds on Cu, which were higher than on other substrates (1–2 folds for SS and Ti, 2–9 folds for Ni). Finally, these behaviors were compared under the biofilm regulatory molecular network. This work may facilitate better understanding different response mechanisms during bacterial biofilm colonization on metal-based pipelines and provide implications for subsequent biofilm control. Display omitted •Bacteria activity drops sharply on Cu and Ni, as compared on SS and Ti substrates.•Bacteria can adapt to SS and Ti easily to form biofilms.•Only survived bacteria can develop biofilms onto Cu and Ni substrates.•Bacteria grown onto Cu substrate have the most drastic defense activity.•All behaviors of biofilm bacteria are performed under the molecular regulatory network.