The effect of different proteases on staphylococcal biofilm
Abstract number: P1803
Cafiso V., Bertuccio T., Stefani S., Papa R., Scoarughi G.L., Cellini A., Artini M., Selan L.
Objectives: The metalloprotease serratiopeptidase (SPEP) has been proved to be effective in the treatment of biofilm of different Gram-positive bacterial species belonging to the Listeria and Staphylococcus genera. Furthermore, SPEP has been used as an anti-inflammatory agent for over 30 years. The analysis of surface protein profiles of treated and untreated bacteria revealed that SPEP modulates specifically the protein pattern acting on different adhesins and autolysins. In order to highlight the mechanism of action of SPEP, different proteases, including metalloproteases and serin-proteases, were tested and compared for their capability to impair biofilm formation and to modulate protein expression of different staphylococcal strains.
Methods: Six Staphylococcus strains were studied, 3 S. epidermidis and 3 S. aureus. Biofilm formation and surface protein pattern was evaluated in the presence of the following proteases: SPEP, carboxypeptidase-A, proteinase-K, trypsin, chymotrypsin. Biofilm growth was assessed by the Christensen method. Proteins were analyzed by SDS-PAGE and zymography to evaluate modifications of the expression of autolytic enzymes. The presence of ica locus and of genes involved in adhesion and autolytic pattern was revealed by PCR. agr-typing was carried out by RT-PCR.
Results: The effect of tested proteases was not related to the protease category but was strain-dependent. SPEP, reducing the biofilm growth in 4 of 6 strains tested, is the more effective; in particular the effect seemed to be proportional to biofilm production. Moreover SPEP treatment showed a limited modification of surface protein patterns as demonstrated by SDS-PAGE and zymogram analysis, while treatment with serin-proteases induced a complete digestion of most of them. A correlation between the action of SPEP and agr-type of different bacterial strains is ongoing.
Conclusion: The degradation of staphylococcal surface proteins does not necessarily result in biofilm inhibition. Moreover, the action of proteases is not related to the presence of icaADCB locus. In the strain more sensitive to SPEP action, S. aureus 6538P, we investigated the agrD expression, which markedly increases following SPEP treatment.
|Session name:||19th European Congress of Clinical Microbiology and Infectious Diseases|
|Location:||Helsinki, Finland, 16 - 19 May 2009|
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