Piscirickettsiosis is the most prevalent bacterial disease affecting seawater salmon in Chilean salmon industry. Antibiotic therapy is the first alternative to counteract infections caused by Piscirickettsia salmonis. The presence of bacterial biofilms on materials commonly used in salmon farming may be critical for understanding the bacterial persistence in the environment. In the present study, the CDC Biofilm Reactor® was used to investigate the effect of sub‐ and over‐MIC of florfenicol on both the pre‐formed biofilm and the biofilm formation by P. salmonis under the antibiotic stimuli on Nylon and high‐density polyethylene (HDPE) surfaces. This study demonstrated that FLO, at sub‐ and over‐MIC doses, decreases biofilm‐embedded live bacteria in the P. salmonis isolates evaluated. However, it was shown that in the P. salmonis Ps007 strain the presence of sub‐MIC of FLO reduced its biofilm formation on HDPE surfaces; however, biofilm persists on Nylon surfaces. These results demonstrated that P. salmonis isolates behave differently against FLO and also, depending on the surface materials. Therefore, it remains a challenge to find an effective strategy to control the biofilm formation of P. salmonis, and certainly other marine pathogens that affect the sustainability of the Chilean salmon industry.