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The VITOTOX test is a new bacterial genotoxicity test that was previously shown to be very rapid and sensitive. Initially only one Salmonella typhimurium strain (TA104 recN2-4) was used in the test. In this paper we introduce a second strain (TA104pr1) that can be used as an internal control to further enhance the reliability of the test. We demonstrate the usefulness of this pr1 strain in genotoxicity and toxicity testing. We also report on the results of a study where the VITOTOX test was performed on newly synthesized pharmaceutical compounds, or intermediate products in the synthesis of drug candidates. We demonstrate that the test gives identical results when performed independently in two different laboratories and that it correlates well with either the Ames test or SOS chromotest.

A new test to detect genotoxicity, that we refer to as the VITOTOX test, was developed. Four gene fusions that are based on the Escherichia coli recN promoter were constructed and evaluated for their SOS response-dependent induction. The wild-type recN promoter, a derivative mutated in the second LexA binding site, a derivative with a mutated -35 region, and a derivative from which both the second LexA binding site and the -35 region were mutated, were cloned upstream of the promoterless Vibrio fischeri luxCDABE operon of pMOL877, in such a way that lux became under transcriptional control of the recN promoter derivatives. The inducibility by the SOS response of the promoter constructs was tested in both E. coli and in the Ames test Salmonella typhimurium strains TA98, TA100 and TA104. In all strains, the highest sensitivity and induction was observed with the plasmids pMOL1067 and pMOL1068, that contain the lux operon under control of the recN promoter mutated in the second LexA binding site, or a recN promoter with a mutated -35 region, respectively. Therefore, strains containing pMOL1067 or pMOL1068 were further used for genotoxicity testing. With the VITOTOX test, genotoxicity was detected within 1-4 h. The VITOTOX test is very sensitive: for most products tested, the minimal detectable concentration (MDC) values were considerably lower (5 to > 100 times) than those described for the Ames test and the SOS chromotest. A good correlation was observed with the results from the Ames tests, but certain PAHs that are not mutagenic in the Ames test were genotoxic in the VITOTOX test. With the VITOTOX strains, the kinetics of SOS induction can be determined. This feature made it possible to distinguish between compounds in mixtures of genotoxic products so long as they had different induction kinetics.