Effect of Lactobacillus plantarum supernatants on bacterial strains isolated from infected chronic wounds

RAMOS AN; SESTO CABRAL ME; CABRERA CA; VALDEZ JC

Rosario

Congreso; Reunión Internacional de Ciencia Farmacéuticas; 2012

RICIFA

Introduction: Chronic wounds are those detained in the inflammatory phase of healing mainly by the presence of biofilm-forming bacteria. Some authors claim that the opportunistic bacterium Pseudomonas aeruginosa is present in all wounds and would be the cause of chronicity and predisposing factor for other infections (1). Previously we demonstrated that the culture supernatant of Lactobacillus plantarum has anti-pathogenic effect on P. aeruginosa inhibiting in vitro viability, biofilm, quorum sensing, adhesion and virulence factors (2). In addition, our medical team demonstrated in clinical investigations that L. plantarum supernatants have interesting pro-healing properties in mice and humans (3). Therefore we proposed the use of these supernatants in the treatment of chronic wounds infected with P. aeruginosa. The aim of this work was to study the effect of L. plantarum supernatants on the principal bacterial strains isolated from chronic wounds in order to extrapolate the treatment to any infected chronic wound. We evaluated in vitro effect of L. plantarum on the viability, adhesion and biofilm (inhibition and disruption) of the different strains tested. Materials and methods: 1) Bacterial strains: Staphylococcus aureus, Staphylococcus epidermidis, Serratia marcescens, Streptococcus pyogenes, Enterococcus faecalis. 2) Viability: halos of inhibition and minimum inhibitory concentration 50% (MIC50) in microculture kinetics. 3) Inhibition and disruption of biofilm: 96-wells microplates as substrate and quantification (crystal violet technique). 4) Formed biomass per unit area: Pearls of polypropylene as substrate. In all cases MRS (culture medium for L. plantarum) was used as negative control and Gentamicin solution 8 mg/mL was used as positive control. Results: L. plantarum supernatant is able to inhibit growth in all strains tested. In all cases the MIC50 is only 5% except for S. marcescens where it reaches a value of 15%. Besides the supernatant significantly inhibited adhesion and biofilm formation of all tested strains both microplates and polypropylene beads with inhibition percentages of 50 ± 8 for P. aeruginosa, 45 ± 4 for S. aureus, 33 ± 5 for S. epidermidis, 25 ± 3 for S. pyogenes, 15 ± 4 for S. marcescens and 10 ± 2 for E. faecalis. Conclusions: The presence of organic acids such as lactic and acetic acid, bacteriocins (plantaricinas) and other organic molecules explain supernatants great bacteriostatic and bactericidal capacity. However the supernatant only inhibited biofilm of aerobic and aero-tolerant bacteria, while microaerophilic bacteria were more resistant. The components responsible for the inhibition of P. aeruginosa biofilm (DNAase, ions, surfactants, autoinducers type 2) do not act in the same way on the biofilm of the other strains, possibly because their respective matrix have different chemical compositions. We propose the use of L. plantarum supernatants in the infected chronic wounds treatment in a strain-dependent manner. References: 1) Bjarnsholt T, Kirketerp-MK, et al. Why chronic wounds will not heal: a novel hypothesis. Wound Rep Reg (2008)16, 2?10. 2) Ramos AN, Sesto Cabral ME, et al. Antipathogenic properties of Lactobacillus plantarum on Pseudomonas aeruginosa: the potential use of its supernatants in the treatment of infected chronic wounds. Wound Rep Reg (2012) 20, 552-562. 3) Valdez JC, Peral M, et al. Interference of Lactobacillus plantarum on Pseudomonas aeruginosa in vitro and in infected burns. The potential use of probiotic in wound treatment?. Clinical Microbiology and Infection (2005) 11, 472-479.