P-019. Pseudomonas aeruginosa Secretes a Substance that Prevents and Disrupts Listeria Biofilms

L. E. Davey, H. Schraft;
Lakehead Univ., Thunder Bay, ON, CANADA.

Listeria monocytogenes (LM), a food-borne pathogen associated with severe disease, forms resistant biofilms which enable it to persist in food processing environments. The use of microbially secreted substances has been suggested as a novel method of biofilm control. Conflicting reports regarding the influence of Pseudomonads on the growth of Listeria biofilms led us to investigate the effect of substances secreted by Pseudomonas aeruginosa PAO1 on L. monocytogenes EGDe biofilms. To investigate the effect of PAO1 secreted substances on LM biofilms, filter sterilized supernatants of 48h PAO1 cultures grown in tryptic soy broth (TSB) at 37°C were mixed 1:1 with 2X concentrated TSB to produce Pseudomonas conditioned media (PCM). LM biofilms were grown on stainless steel 316 coupons (SS) at 22°C and 37°C, and in polystyrene microtiter plates (PS) at 22°C. LM biofilms were exposed to the PCM at 0, 3, and 24h after biofilm initiation and incubated for an additional 24h to determine the effect of PAO1 at different stages of biofilm development. Attached cells were quantified on SS by drop-plating and on PS by quantitative crystal-violet staining. At 22°C, PCM reduced attachment of LM to SS by up to 1.7 Log CFU cm-2 compared to controls, and decreases > 1 Log CFU cm-2 were observed regardless of biofilm development stage at the time of exposure to the PCM. Treatment on PS eliminated virtually all LM biofilm (> tenfold reduction in the uptake of crystal violet vs. controls). LM biofilms on PS responded similarly to exposure to PCM at all stages of development. These findings were temperature dependent as treatment with PCM at 37°C did not significantly change LM biofilms on SS. Thus, P. aeruginosa secretes a substance that prevents the initiation and growth of LM biofilms, and that disperses mature biofilms. The role of temperature in the effect of PCM suggests that this reduction in biofilm formation may involve the flagella of LM. Identification of the substance(s) produced by P. aeruginosa that are able to disperse the LM biofilms should provide additional insight into their mode of action, and may represent a novel approach to biofilm control.