title: Microbial genotoxicity bioreporters based on sulA activation.
abstract: A bacterial genotoxicity reporter strain was constructed in which the tightly controlled strong promoter of the Escherichia coli SOS response gene sulA was fused to the alkaline phosphatase-coding phoA reporter gene. The bioreporter responded in a dose-dependent manner to three model DNA-damaging agents-hydrogen peroxide, nalidixic acid (NA), and mitomycin C (MMC)-detected 30-60 min after exposure. Detection thresholds were 0.15 μM for MMC, 7.5 μM for nalidixic acid, and approximately 50 μM for hydrogen peroxide. A similar response to NA was observed when the bioreporter was integrated into a specially designed, portable electrochemical detection platform. Reporter sensitivity was further enhanced by single and double knockout mutations that enhanced cell membrane permeability (rfaE) and inhibited DNA damage repair mechanisms (umuD, uvrA). The rfaE mutants displayed a five- and tenfold increase in sensitivity to MMC and NA, respectively, while the uvrA mutation was advantageous in the detection of hydrogen peroxide. A similar sensitivity was displayed by the double rfaE/uvrA mutant when challenged with the pre-genotoxic agents 2-amino-3-methylimidazo[4,5-f]quinoline and 2-aminoanthracene following metabolic activation with an S9 mammalian liver fraction.
authors.affiliation: Department of Plant and Environmental Sciences, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Israel.
authors.forename: Alva
authors.initials: A
authors.lastname: Biran
authors.forename: Hadar
authors.initials: H
authors.lastname: Ben Yoav
authors.forename: Sharon
authors.initials: S
authors.lastname: Yagur-Kroll
authors.forename: Rami
authors.initials: R
authors.lastname: Pedahzur
authors.forename: Sebastian
authors.initials: S
authors.lastname: Buchinger
authors.forename: Yosi
authors.initials: Y
authors.lastname: Shacham-Diamand
authors.forename: Georg
authors.initials: G
authors.lastname: Reifferscheid
authors.forename: Shimshon
authors.initials: S
authors.lastname: Belkin
mesh_terms: D000469:Alkaline Phosphatase / Q000235:genetics / Q000378:metabolism; D000818:Animals; D015374:Biosensing Techniques / Q000379:methods*; D055664:Electrochemical Techniques / Q000379:methods; D004926:Escherichia coli / Q000187:drug effects* / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism; D017930:Genes, Reporter; D006861:Hydrogen Peroxide / Q000633:toxicity; D016685:Mitomycin / Q000633:toxicity; D009152:Mutagenicity Tests / Q000379:methods*; D009153:Mutagens / Q000633:toxicity*; D009268:Nalidixic Acid / Q000633:toxicity; D011401:Promoter Regions, Genetic; D013014:SOS Response, Genetics
chemical_list: D029968:Escherichia coli Proteins; D009153:Mutagens; C053139:sulA protein, E coli; D009268:Nalidixic Acid; D016685:Mitomycin; D006861:Hydrogen Peroxide; D000469:Alkaline Phosphatase