Phage Mediated Biofilm Disruption

The ability of phages SJM3 and LASTA to lyse host cells in preformed biofilm on porous glass beads was investigated as previously described [22 (link),38 (link)] with minor modifications. Glass beads (diameter 4 mm, pore size 60 μm, and surface area of ~60 cm²; VitraPor; ROBU, Hattert, Germany) were statically incubated with 100-times-diluted overnight culture of Ni9 for 24 h at 37 °C. Further on, beads were washed three times with saline solution and separated in single wells of a 24-well plate (Tissue Culture Plate; Sarstedt). Each was treated with 10⁹ PFU/mL phage suspension (LASTA, SJM3 or cocktail) at predicted MOI 100 in LB medium for 24 h. After 24 h, treated beads and beads from the untreated control were washed with saline, vortexed in microtubes and, subsequently, sonicated in an ultrasound water bath and vortexed once again in order to detach biofilm-embedded bacteria [39 (link)]. CFUs of biofilm before treatment, biofilm after phage treatments and biofilm with fresh medium were counted on LA plates.