Photodynamic therapy (PDT) is considered a very promising therapeutic modality for antimicrobial therapy. Although several studies have demonstrated that Gram-positive bacteria are very sensitive to PDT, Gram-negative bacteria are more resistant to photodynamic action. This difference is due to a different cell wall structure. Gram-negative bacteria have an outer cell membrane containing lipopolysaccharides (LPS) that hinder the binding of photosensitizer molecules, protecting the bacterial cells from chemical attacks. Combination of the lipopolysaccharides-binding activity of Concanavalin A (ConA) with the photodynamic properties of Rose Bengal (RB) holds the potential of an innovative protein platform for targeted photodynamic therapy against Gram-negative bacteria. A ConA-RB bioconjugate was synthesized and characterized. Approximately 2.4 RB molecules were conjugated per ConA monomer. The conjugation of RB to ConA determines a decrease of the singlet oxygen generation and an increase of superoxide and peroxide production. The photokilling efficacy of the ConA-RB bioconjugate was demonstrated in a planktonic culture of E. coli. Irradiation with white light from a LED lamp produced a dose-dependent photokilling of bacteria. ConA-RB conjugates exhibited a consistent improvement over RB (up to 117-fold). The improved uptake of the photosensitizer explains the enhanced PDT effect accompanying increased membrane damages induced by the ConA-RB conjugate. The approach can be readily generalized (i) using different photo/sonosensitizers, (ii) to target other pathogens characterized by cell membranes containing lipopolysaccharides (LPS).
Keywords: Antimicrobial photodynamic therapy; Bioconjugation; Carbohydrate-lectin recognition process; Concanavalin A; Gram-negative bacteria targeting; Rose Bengal.
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