Abstract
A faster and simpler method to monitor the photoinactivation process of Escherichia coli involving the use of recombinant bioluminescent bacteria is described here. Escherichia coli cells were transformed with luxCDABE genes from the marine bioluminescent bacterium Vibrio fischeri and the recombinant bioluminescent indicator strain was used to assess, in real time, the effect of three cationic meso-substituted porphyrin derivatives on their metabolic activity, under artificial (40 W m(-2)) and solar irradiation (approximately 620 W m(-2)). The photoinactivation of bioluminescent E. coli is effective (>4 log bioluminescence decrease) with the three porphyrins used, the tricationic porphyrin Tri-Py+-Me-PF being the most efficient compound. The photoinactivation process is efficient both with solar and artificial light, for the three porphyrins tested. The results show that bioluminescence analysis is an efficient and sensitive approach being, in addition, more affordable, faster, cheaper and much less laborious than conventional methods. This approach can be used as a screening method for bacterial photoinactivation studies in vitro and also for the monitoring of the efficiency of novel photosensitizer molecules. As far as we know, this is the first study involving the use of bioluminescent bacteria to monitor the antibacterial activity of porphyrins under environmental conditions.
Highlights
The growing reduction of water resources due to environmental pollution has become a major public health concern
We proposed to develop a rapid method to assess the antibacterial eVect of meso-substituted porphyrins based on the metabolic activity of recombinant bioluminescent Escherichia coli (E. coli) under artiWcial and solar irradiation
For the lower concentrations used (0.5 and 1.0 M), the tricationic porphyrins (Tri-Py+Me-PF and Tri-Py+-Me-CO2Me) are more eYcient than the tetracationic one (Tetra-Py+-Me) (P < 0.05, analysis of variance (ANOVA)). They caused more than 4 log decrease in bioluminescence after 270 min of irradiation, while at these concentrations and after the same period, the tetracationic porphyrin (Tetra-Py+-Me) shows only a 0.50 and a 0.65 log decrease in bioluminescence (Fig. 5a)
Summary
The growing reduction of water resources due to environmental pollution has become a major public health concern. To overcome the high costs and the diYculty in implementing these techniques, alternative physico-chemical methods have been studied, namely the photodynamic antimicrobial therapies [1,2,3] This somewhat new technique uses a light source (sunlight or artiWcial light), an oxidizing agent (molecular oxygen dissolved in water) and an intermediary agent [named photosensitizer (PS)], able to absorb and transfer the energy of the light source to molecular oxygen leading to the formation of highly cytotoxic reactive oxygen species (1O2, OH, O2¡, H2O2) [4]. The potential use of this approach for water disinfection has already been studied, showing that photoinactivation (PI) of bacteria in drinking [3] and residual waters [1, 6] is possible under artiWcial or solar irradiation.
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