SOS-LUX- and LAC-FLUORO-TEST for the quantification of genotoxic and/or cytotoxic effects of heavy metal salts

The constantly increasing industrial, agricultural and domestic activities with their growing risk of contaminating fresh water and ground water by discharged compounds lead to an increasing concern focused on the quality of water. Because of their simplicity and sensitivity, bacterial tests play an important role in the detection and screening of genotoxins or cytotoxins in water.One of those bacterial tests is the SOS-LUX- and LAC-FLUORO-TEST, which is a combination of two bioassays, that simultaneously measures the genotoxicity (SOS-LUX-TEST) and the cytotoxicity (LAC-FLUORO-TEST) of substances and mixtures of substances.The SOS-LUX-TEST is based on genetically modified Salmonella typhimurium TA1535 bacteria, which have been transformed with the plasmid pPLS-1 carrying the promoterless lux operon of Photobacterium leiognathi as reporter element under the control of a DNA damage-dependent SOS promoter from ColD as sensing element. This system reacts in a dose-dependent manner to agents which induce DNA damages inside these bacterial cells with the production of bioluminescence that can easily be measured.The analogous LAC-FLUORO-TEST has been developed for the detection of cellular responses to cytotoxins. It is based on the constitutive expression of green fluorescent protein (GFP) mediated by the bacterial protein expression vector pGFPuv (Clontech, Palo Alto, USA). In response to cytotoxic agents, this system reacts with a dose-dependent reduction of GFP-fluorescence.A panel of recombinant S. typhimurium strains carrying either the SOS-LUX plasmid or the fluorescence-mediating lac-GFPuv plasmid was used to determine in parallel on one microplate the genotoxic and the cytotoxic potential of heavy metal salts like K₂Cr₂O₇, CrCl₃, ZnSO₄, CuSO₄, NiSO₄, KH₂AsO₄ and As₂O₃ at the same time. Light and fluorescence emission of untreated and chemical-treated cells were measured in a microplate luminometer-fluorometer-photometer combination and the luminescence induction as well as the fluorescence reduction were used to determine the genotoxic and/or cytotoxic potential of the heavy metal salts.