Use of epifluorescence microscopy to assess the effectiveness of phage P100 in controlling Listeria monocytogenes biofilms on stainless steel surfaces

Abstract

One source of foodborne listeriosis is related to a virulent strain established in the food-processing environment. We used direct epifluorescence microscopy (DEM) to evaluate the effectiveness of phage P100 in controlling Listeria monocytogenes biofilms on stainless steel surfaces, in wet conditions at room temperature. Biofilms that were allowed to develop for 72 h were subsequently treated with different concentrations of phage (5, 6, 7, and 8 log PFU/ml). L. monocytogenes was monitored up to 48 h, using both DEM and cultivation method. When L. monocytogenes was monitored using a cultivation method, the first significant reduction was observed after phage treatment with 7 and 8 log PFU/ml at 8 h. Subsequently, these treatments achieved undetectable levels of pathogen at 48 h, with a mean reduction of 5.29 log CFU/cm2. Conversely, when samples were evaluated using DEM in treatments with 6, 7, and 8 log PFU/ml, although disaggregation of biofilms could be observed after 8 h, viable cells were still present up to 48 h (maximal reduction 1.5 log units). The phage titer remained stable or increased up to 2.59 log units during the study period. In conclusion, phage P100 may provide an adjuvant measure to control L. monocytogenes biofilm on stainless steel surfaces. However, phage treatment must be used in combination with other hygienization measures to increase efficacy. In this study, DEM was a good tool to quickly and accurately assess the real effect of phage P100 on L. monocytogenes biofilms.