Specific and selective probes for Staphylococcus aureus from phage-displayed random peptide libraries
2017
De Plano, Laura M. | Carnazza, Santina | Messina, Grazia M.L. | Rizzo, Maria Giovanna | Marletta, G. (Giovanni) | Guglielmino, Salvatore P.P.
Staphylococcus aureus is a major human pathogen causing health care-associated and community-associated infections. Early diagnosis is essential to prevent disease progression and to reduce complications that can be serious.In this study, we selected, from a 9-mer phage peptide library, a phage clone displaying peptide capable of specific binding to S. aureus cell surface, namely St.au9IVS5 (sequence peptide RVRSAPSSS).The ability of the isolated phage clone to interact specifically with S. aureus and the efficacy of its bacteria-binding properties were established by using enzyme linked immune-sorbent assay (ELISA). We also demonstrated by Western blot analysis that the most reactive and selective phage peptide binds a 78KDa protein on the bacterial cell surface. Furthermore, we observed selectivity of phage–bacteria-binding allowing to identify clinical isolates of S. aureus in comparison with a panel of other bacterial species.In order to explore the possibility of realizing a selective bacteria biosensor device, based on immobilization of affinity-selected phage, we have studied the physisorbed phage deposition onto a mica surface. Atomic Force Microscopy (AFM) was used to determine the organization of phage on mica surface and then the binding performance of mica-physisorbed phage to bacterial target was evaluated during the time by fluorescent microscopy. The system is able to bind specifically about 50% of S. aureus cells after 15′ and 90% after one hour. Due to specificity and rapidness, this biosensing strategy paves the way to the further development of new cheap biosensors to be used in developing countries, as lab-on-chip (LOC) to detect bacterial agents in clinical diagnostics applications.
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