A non-chemical solution is needed to control biofilms in water and wastewater treatment systems. High-frequency alternating electric field application offers an alternative approach that does not involve undesired electrode surface reactions. However, the effect of high-frequency alternating electric fields on bacterial cells in the attached-growth mode remains unexplored. This study investigated the impact of such fields on two stages of the biofilm development process: the initial adhesion phase (stage 1) and the early development phase (stage 2). Experiments were conducted using Escherichia coli and Pseudomonas aeruginosa in a three-channel flow cell exposed to alternating electric fields (3.05 V/cm, 20 MHz). The primary outcome of this study demonstrated that alternating electric fields decreased adhered cell numbers at both stages due to their inhibitory effect on growth. The alternating electric fields also triggered cell detachment after the initial attachment stage but not in mature biofilms. Interestingly, despite a reduction in cell counts, the amount of total biofilm biomass remained unaffected, which was likely due to increased cell size via cell elongation compensating for the decrease in numbers. No synergistic effects with respect to hydrodynamic forces were observed. These findings highlight the potential applicability of alternating electric fields to biofilm control and provide implications for water and wastewater engineering applications.