Alternative method in Galleria mellonella larvae to study biofilm infection and treatment.

In vivo studies are crucial decision-maker step in order to translate in vitro data to an applied therapy. Considering this we describe a simple method that analyzes and quantifies biofilm formation inside the Galleria mellonella larvae. Toothbrush bristles were employed as an abiotic surface to mimic a medical device. A standardized inoculum of Staphylococcus aureus was systemically injected in the larvae together with the insertion of a bristle in the last proleg pair. After incubation adhered cells were detached from bristles and quantified by colony-forming units (CFU) counting using staphylococci-selective medium. About 3 × 10 CFU of S. aureus were recovered from bristles and scanning electron microscopy (SEM) images confirmed biofilm formation. Control group did not show adherent bacteria, as demonstrated by absence of CFU counting and SEM images, indicating that the insertion procedure is free of bacterial contamination. We present a feasible method to evaluate bacterial biofilm formation in vivo that in the near future can be used to evaluate antibiofilm compounds.