Cultural, morphological and biochemical properties of Salmonella typhimurium indicator bacteria used in the Ames test and their applicability for the assessment of the test system quality

Abstract

Introduction. The Ames test has been widely used to study the mutagenicity of chemicals. In view of the implementation of a quality management system in the test facilities, the cultural, morphological and biochemical properties of strains used in the Salmonella/microsome test system were studied. An assessment of their applicability as additional characteristics of quality of the tester strain was made. Materials and methods. The study of cultural and morphological characteristics of S. typhimurium strains was carried out using the commercial media. The biochemical properties of S. typhimurium cultures were evaluated using a Vitek-2 system. Results. Studies have shown that the procedure of marker verification for confirming the reference properties of tester strains, in addition to evaluation of the spontaneous mutation level and phenotypic characteristics (-his phenotype, the presence or absence of the R-factor plasmids, rfa- and ∆uvrB mutations), can be extended by periodically assessing their biochemical properties. Most of the biochemical characteristics of bacteria of the Salmonella typhimurium serovar are inherent for tester strains. Compared to the wild-type bacteria atypical features of tester strains are a negative reaction in the hydrogen sulfide production test, positive activity of tyrosine arylamidase, the ability to utilize 5-keto-D-gluconate, unstable activity of alpha-galactosidase. Differential selective media used to maintain wild-type Salmonella cultures are not universal for the growth of strains of the Salmonella/microsome test system. When choosing media for cytotoxicity, survival, or sterility assessment, it is necessary to take into consideration the growth characteristics of the Salmonella tester strains on different media: the ability to grow on selective media decreases in the series of Endo-agar > Salmonella Shigella -agar > Bismuth-sulfite agar > Ploskirev’s agar. Limitations. The research is limited to the study of cultural, morphological and biochemical characteristics of S. typhimurium, but not Escherichia coli. Conclusion. The described cultural and morphological properties of tested strains on different media can be used to assess cytotoxicity/survival of treated cultures performing studies according to OECD 471. The use of additional biochemical markers of authenticity and morphological properties of tester strains will help to ensure the quality of studies using the Ames test in the processes of conservation, reproduction, and routine testing.