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Dose–response relationships and statistical performance of a battery of bacterial gene profiling assays

  • Environmental Biotechnology
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Abstract

Because of increasing awareness and legislative demands, there is a demand for the development and use of biological assays for the assessment of the toxicity of chemicals, environmental samples. Recently, a growing number of bacterial reporter assays have been developed and implemented. Nevertheless, little data is published on the performance of these assays in terms of analytical parameters. We present results on a battery of 14 transgenic Escherichia coli strains carrying different promoter::reporter fusions. Growth characteristics and basal expression levels were modeled and fitted, data show that growth curves should be taken into account during test development. Our study shows that the induction profiles reflect the mode of action, e.g., paraquat clearly induces the soxRS operon. The sensitivity of the assay compares well to that of whole organism tests, e.g., fish and Daphnia for polar organics. Metal toxicity is detected less efficiently, e.g., cadmium is detected near the LC50 of carp, considered a relatively insensitive species towards cadmium. The assay variability ranges from 10 to 40% depending on the strain, comparable to that of other bioassays. The variability was shown to be determined by the intrinsic traits of the promoter–strain combination, not by operating conditions.

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References

  • Baranyi J, Mcclure PJ, Sutherland JP, Roberts TA (1993) Modelling bacterial-growth responses. J Ind Microbiol 12:190–194

    Article  Google Scholar 

  • Bechor O, Smulski DR, Van Dyk TK, LaRossa RA, Belkin S (2002) Recombinant microorganisms as environmental biosensors: pollutants detection by Escherichia coli bearing fabA':: lux fusions. J Biotechnol 94:125–132

    Article  CAS  PubMed  Google Scholar 

  • Bierkens J, Klein G, Corbisier P, Van den Heuvel R, Verschaeve L, Weltens R, Schoeters G (1998) Comparative sensitivity of 20 bioassays for soil quality. Chemosphere 37:2935–2947

    Article  CAS  PubMed  Google Scholar 

  • Bobeldijk I, Brandt A, Wullings B, Noij T (2001) High-performance liquid chromatography-ToxPrint: chromatographic analysis with a novel (geno)toxicity detection. J Chromatogr A918:277–291

    Article  Google Scholar 

  • Brams A, Buchet JP, Crutzenfayt MC, Demeester C, Lauwerys R, Leonard A (1987) A comparative-study, with 40 chemicals, of the efficiency of the Salmonella assay and the sos chromotest (Kit Procedure). Toxicol Lett 38:123–133

    Article  CAS  PubMed  Google Scholar 

  • Byford JR, Shaw LE, Drew MGB, Pope GS, Sauer MJ, Darbre PD (2002) Oestrogenic activity of parabens in MCF7 human breast cancer cells. J Steroid Biochem Mol Biol 80:49–60

    Article  CAS  PubMed  Google Scholar 

  • Carls N, Schiestl RH (1994) Evaluation of the yeast DEL assay with 10 compounds selected by the international program on chemical safety for the evaluation of short-term tests for carcinogens. Mutat Res 320:293–303

    Article  CAS  PubMed  Google Scholar 

  • Chilvers KF, Perry JD, James AL, Reed RH (2001) Synthesis and evaluation of novel fluorogenic substrates for the detection of bacterial B-galactosidase. J Appl Microbiol 91:1118–1130

    Article  CAS  PubMed  Google Scholar 

  • Choi K, Meier PG (2001) Toxicity evaluation of metal plating wastewater employing the Microtox((R)) assay: a comparison with cladocerans and fish. Environ Toxicol 16:136–141

    Article  CAS  PubMed  Google Scholar 

  • Del Nobile MA, Altieri C, Corbo MR, Sinigaglia M, La Notte E (2003) Development of a structured model for batch cultures of lactic acid bacteria. J Ind Microbiol Biotech 30:421–426

    Article  CAS  Google Scholar 

  • Delihas N, Forst S (2001) MicF: an antisense RNA gene involved in response of Escherichia coli to global stress factors. J Mol Biol 313(1):1–12

    Article  CAS  PubMed  Google Scholar 

  • Diez A, Gustavsson N, Nystrom T (2000) The universal stress protein A of Escherichia coli is required for the resistance to DNA damaging agents and is regulated by a RecA/FtsK dependent regulatory pathway. Mol Microbiol 36(6):1494–1503

    Article  CAS  PubMed  Google Scholar 

  • Doherty FG (2001) A review of the Microtox (R) toxicity test system for assessing the toxicity of sediments and soils. Water Qual Res J Can 36:475–518

    Article  CAS  Google Scholar 

  • Eder E, Favre A, Stichtmann C, Deininger C (1989) Induction Of Sfia Sos function by peroxides using 3 different Escherichia coli strains. Toxicol Lett 48:225–234

    Article  CAS  PubMed  Google Scholar 

  • Elowitz MB, Levine AJ, Siggia ED, Swain PS (2002) Stochastic gene expression in a single cell. Science 297:1183–1186

    Article  CAS  PubMed  Google Scholar 

  • Fawcett WP, Wolf RE (1995) Genetic definition of the Escherichia coli zwf “soxbox”, the DNA binding site for SoxS-mediated induction of glucose 6-phosphate dehydrogenase in response to superoxide. J Bacteriol 177(7):1742–1750

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Gabrielson J, Kuhn I, Colque-Navarro P, Hart M, Iversen A, McKenzie D, Mollby R (2003) Microplate-based microbial assay for risk assessment and (eco)toxic fingerprinting of chemicals. Anal Chim Act 485:121–130

    Article  CAS  Google Scholar 

  • Galhardo RS, Almeida CEB, Leitao AC, Cabral-Neto JB (2000) Repair of DNA lesions induced by hydrogen peroxide in the presence of iron chelators in Escherichia coli: participation of endonuclease IV and Fpg. J Bacteriol 182(7):1964–1968

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Giesy J, Hilscherova K, Jones PD, Kannan K, Machala M (2002) Cell bioassays for detection of aryl hydrocarbon and estrogen receptor mediated activity in environmental samples. Marine Pollut Bull 45:3–16

    Article  CAS  Google Scholar 

  • Gompertz B (1825) On the nature of the function expressive of the law of human mortality. Phil Trans 27:510–519

    Google Scholar 

  • Huisman O, Dari R, Gottesman S (1984) Cell-division control in Escherichia coli: specific induction of the SOS function SfiA protein is sufficient to block septation. Proc Natl Acad Sci USA 81(14):4490–4494

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Isnard P, Flammarion P, Roman G, Babut M, Bastien P, Bintein S, Essermeant L, Ferard JF, Gallotti-Schmitt S, Saouter E, Saroli M, Thiebaud H, Tomassone R, Vindimian E (2001) Statistical analysis of regulatory ecotoxicity tests. Chemosphere 45:659–669

    Article  CAS  PubMed  Google Scholar 

  • Kaiser KLE, Palabrica VS (1991) Photobacterium phosphoreum toxicity data index. Water Pollut Res J Can 26:361–431

    Article  CAS  Google Scholar 

  • Kato T, Watanabe M, Ohta T (1994) Induction of the sos response and mutations by reactive oxygen-generating compounds in various Escherichia coli mutants defective in the mutm, muty or soxrs loci. Mutagenesis 9:245–251

    Article  CAS  PubMed  Google Scholar 

  • Kenyon CJ, Brent R, Ptashne M, Walker GC (1982) Regulation of damage-inducible genes in Escherichia coli. J Mol Biol 160(3):445–457

    Article  CAS  PubMed  Google Scholar 

  • Kim G, Oh TJ (2000) 2-mercaptoethylamine, radioprotector, inhibits the induction of the oxidative stress inducible (soi) gene by paraquat in Escherichia coli. Pharmacol Res 42(5):429–433

    Article  CAS  PubMed  Google Scholar 

  • Kitagawa M, Wada C, Yoshioka S, Yura T (1991) Expression of clpb, an anolog of the ATP-dependent protease regulatory subunit in Escherichia coli, is controlled by a heat-shock sigma-factor (SIGMA-32). J Bacteriol 173:4247–4253

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Klamer HJC, Villerius LA, Roelsma J, DeMaagd PGJ, Opperhuizen A (1997) Genotoxicity testing using the Mutatox(TM) assay: evaluation of benzo[a]pyrene as a positive control. Environ Toxicol Chem 16:857–861

    Article  CAS  Google Scholar 

  • Kvint K, Nachin L, Diez A, Nystrom T (2003) The bacterial universal stress protein: function and regulation. Curr Opin Microbiol 6:140–145

    Article  CAS  PubMed  Google Scholar 

  • Landini P, Hajec LI, Volkert MR (1994) Structure and transcriptional regulation of the Escherichia coli adaptive response gene aidB. J Bacteriol 176(21):6583–6589

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lanzer M, Bujard H (1988) Promoters largely determine the efficiency of repressor action. Proc Natl Acad Sci 85:8973–8977

    Article  Google Scholar 

  • Lee S, Sowa ME, Choi JM, Tsai FTF (2004) The ClpB/Hsp104 molecular chaperone—a protein disaggragating machine. J Struct Biol 146:99–105

    Article  CAS  PubMed  Google Scholar 

  • Liebert CA, Hall RM, Summers AO (1999) Transposon Tn21, flagship of the floating genome. Microbiol Mol Biol Rev 63:507–522

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Livrelli V, Lee IW, Summers AO (1993) In vivo DNA-protein interactions at the divergent mercury resistance (mer) promoters. 1. metalloregulatory protein merR mutants. J Biol Chem 268(4):2623–2631

    Article  CAS  PubMed  Google Scholar 

  • MerschSundermann V, Klopman G, Rosenkranz HS (1996) Chemical structure and genotoxicity: studies of the SOS chromotest. Mutat Res Rev Genet Toxicol 340:81–91

    Article  Google Scholar 

  • Oda Y, Nakamura S, Oki I, Kato T, Shinagawa H (1985) Evaluation of the new system (Umu-test) for the detection of environmental mutagens and carcinogens. Mutat Res 147:219–229

    Article  CAS  PubMed  Google Scholar 

  • Orser CS, Foong FCF, capaldi SR, Nalezny W, Mackay W, Benjamin M, Farr SB (1995) Use of prokaryotic stress promotors as indicators of the mechanisms of chemical toxicity. In Vitro Toxicol 8:71–85

    CAS  Google Scholar 

  • Peeters ETHM, Dewitte A, Koelmans AA, van der Velden JA, den Besten PJ (2001) Evaluation of bioassays versus contaminant concentrations in explaining the macroinvertebrate community structure in the Rhine-Meuse delta, the Netherlands. Environ Toxicol Chem 20:2883–2891

    Article  CAS  PubMed  Google Scholar 

  • Quillardet P, Huisman O, Dari R, Hofnung M (1982) SOS chromotest, a direct assay of induction of an sos function in Escherichia coli k-12 to measure genotoxicity. Proc Natl Acad Sci USA 79:5971–5975

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Quillardet P, Debellecombe C, Hofnung M (1985) The SOS chromotest, a colorimetric bacterial assay for genotoxins—validation-study with 83 compounds. Mutat Res 147:79–95

    Article  CAS  PubMed  Google Scholar 

  • Rila JP, Eisentraeger A (2003) Application of bioassays for risk characterisation and remediation control of soils polluted with nitroaromatics and PAHs. Water Air Soil Pollut 148:223–242

    Article  CAS  Google Scholar 

  • Safe SH, Pallaroni L, Yoon K, Gaido K, Ross S, McDonnell D (2002) Problems for risk assessment of endocrine-active estrogenic compounds. Environ Health Perspect 110:925–929

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sakagami Y, Yamazaki H, Ogasawara N, Yokoyama H, Ose Y, Sato T (1988) The evaluation of genotoxic activities of disinfectants and their metabolites by Umu test. Mutat Res 209:155–160

    Article  CAS  PubMed  Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (1989) Molecular Cloning, A Laboratory manual. Cold Spring Harbor Laboratory

  • Smith BT, Walker GC (1998) Mutagenesis and more: umuDC and the Escherichia coli SOS response. Genet 148(4):1599–1610

    Article  CAS  Google Scholar 

  • Swain PS, Elowitz MB, Siggia ED (2002) Intrinsic and extrinsic contributions to stochasticity in gene expression. Proc Natl Acad Sci 99:12795–12800

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Tartaglia LA, Storz G, Ames BN (1989) Identification and molecular analysis of oxyr-regulated promoters important for the bacterial adaptation to oxidative stress. J Molec Biol 210(4):709–719

    Article  CAS  PubMed  Google Scholar 

  • Trim AH, Marcus JM (1990) Integration of long-term fish kill data with ambient water-quality monitoring data and application to water-quality management. Environ Man 14:389–396

    Google Scholar 

  • vanderLelie D, Regniers L, Borremans B, Provoost A, Verschaeve L (1997) The VITOTOX(R) test, an SOS bioluminescence Salmonella typhimurium test to measure genotoxicity kinetics. Mutat Res Genet Toxicol Environ Mutagen 389:279–290

    Article  CAS  Google Scholar 

  • Weibull W (1939) A statistical distribution function of wide applicability. J Appl Mech 18:293

    Article  Google Scholar 

  • White PA, Rasmussen JB, Blaise C (1996) A semi-automated, microplate version of the SOS Chromotest for the analysis of complex environmental extracts. Mutat Res 360:51–74

    Article  CAS  PubMed  Google Scholar 

  • Witkin EM (1991) RecA protein in the SOS response – milestones and mysteries. Biochimie 73(2–3):133–141

    Article  CAS  PubMed  Google Scholar 

  • Yim HH, Brems RL, Villarejo M (1994) Molecular characterization of the promoter of OsmY, an RPOS dependent gene. J Bacteriol 176(1):100–107

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zwietering MH, Jongenburger I, Rombouts FM, Vantriet K (1990) Modeling of the bacterial growth curve. Appl Environ Microbiol 56:1875–1881

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Authors and Affiliations

  1. Department of Biology, Ecophysiology, Biochemistry and Toxicology Group, University of Antwerp, Groenenborgerlaan 171/U7, B-2020, Antwerp, Belgium

    F. Dardenne, I. Nobels, W. De Coen & R. Blust

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  1. F. Dardenne
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  2. I. Nobels
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  3. W. De Coen
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  4. R. Blust
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Correspondence to F. Dardenne.

Electronic Supplementary Material

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S 1

Model compounds and their primary lesions and expected primary targets according to their mode of action (DOC 28 kb).

S 2

Tukey’s honest significant difference test on all intra-assay coefficients of variation. Figures listed in bold designate significant differences (p < 0.05) (DOC 37 kb).

S 3

Growth rates of undosed controls during the exposure phase of the assays. The boxes denote the interquartile range, the hinges the 25 and 75% quartile, and the cross hairs the median value (DOC 31 kb).

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Dardenne, F., Nobels, I., De Coen, W. et al. Dose–response relationships and statistical performance of a battery of bacterial gene profiling assays. Appl Microbiol Biotechnol 75, 223–234 (2007). https://doi.org/10.1007/s00253-006-0808-5

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  • DOI: https://doi.org/10.1007/s00253-006-0808-5

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