Photoinactivation of planktonic and biofilm bacterial cultures employing delta-aminolevulinic acid.

CASAS, A; VALLECORSA, P; DI VENOSA, G; ORLANDO, GABRIEL; SAENZ D; BUZZOLA F; CALVO, G; CERVINI BOHM G

Villa Carlos Paz, Cordoba

Congreso; XIII Encuentro Latinoamericano de Fotoquímica y Fotobiología; XIII ELAFOT.; 2017

Grupo Argentino de Fotobiología

BacterialPhoto-Inactivation (BPI) is an antimicrobial treatment that uses aphotosensitizing molecule (FS), which is then irradiated with visible lightgenerating reactive oxygen species and the consequent damage to themicroorganisms. On the other hand, antibiotic-resistant infections represent animportant clinical problem, most due to the formation of biofilms.Consequently, the need arises for new alternative therapies to the use ofantibiotics. In this work we evaluated δ-aminolevulinic acid (ALA) action as aprecursor of photosensitive porphyrins [1-2]] in planktonic cultures andbiofilms of Staphylococcus aureusATCC 25923 [4], Staphylococcus epidermidis,Escherichia coli [3] and Pseudomonas aeruginosa ATCC 27853. We workedwith 24-h biofilms grown over polystyrene plates and planktonic cultures withan OD600 0.7. After incubation in the dark at different doses of ALA, we irradiated thebacteria with a non-coherent white light source. The determination of theeffectiveness of the treatments was performed counting colony forming units(CFU/ml). In addition, the amount and type of porphyrins synthesized in thedifferent treatments were determined fluorimetrically and by HPLC. The BPI withconcentrations between 1 and 2 mM of ALA, reduces 6 logs the viability of theplanktonic and biofilms cultures of the two Gram positive strains analyzed,relative to the untreated controls. In the case of the Gram negative strains,only P. aeruginosa showed sensitivityto ALA-BPI in planktonic culture, reducing its viability by 5 logs.Porphyrinsfrom Gram positive bacteria peaks at concentrations of 1 mM or 2 mM for planktonic or biofilmcultures respectively, concentrations leading to the maximal phototoxicity. Inaddition, in the case of S. aureus, achange in the pattern of porphyrins synthesized can be observed upon increasingALAconcentrations, which is in line with a decrease of the photoinactivation rate.Inconclusion, the results obtained here indicate that BPI employing ALA is an effectivetreatment, alternative to antibiotics, for Gram positive bacteria in cultureand biofilms. In addition, the sensitivity of this treatment is directlyrelated to the pattern and amount of porphyrins synthesized. However,optimization of the BPI treatment in Gram negative bacteria is necessary.[1]Fotinos N, Convert M, Piffaretti JC, Gurny R, Lange N. Antimicrob AgentsChemother. 2008 Apr;52(4):1366-73. doi: 10.1128/AAC.01372-07.[2]Harris F, Pierpoint L. Med Res Rev. 2012, 32(6):1292-1327. doi:10.1002/med.20251.[3] SzocsK, Gabor F, Csik G and Fidy J. B, J. Photochem. Photobiol., B, 1999, 50, 8?17.[4] ZhangQZ, Zhao KQ, Wu Y, Li XH, Yang C, Guo LM, Liu CH, Qu D, Zheng CQ. PLoS One.2017, 12(3):e0174627