Original Paper

Analytical and Bioanalytical Chemistry

, Volume 400, Issue 9, pp 3013-3024

First online:

Microbial genotoxicity bioreporters based on sulA activation

  • Alva BiranAffiliated withDepartment of Plant and Environmental Sciences, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem
  • , Hadar Ben YoavAffiliated withDepartment of Physical Electronics, Faculty of Engineering, Tel Aviv University
  • , Sharon Yagur-KrollAffiliated withDepartment of Plant and Environmental Sciences, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem
  • , Rami PedahzurAffiliated withDepartment of Plant and Environmental Sciences, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem
  • , Sebastian BuchingerAffiliated withDivision of Qualitative Hydrology, German Federal Institute of Hydrology (BfG)
  • , Yosi Shacham-DiamandAffiliated withDepartment of Physical Electronics, Faculty of Engineering, Tel Aviv University
  • , Georg ReifferscheidAffiliated withDivision of Qualitative Hydrology, German Federal Institute of Hydrology (BfG)
  • , Shimshon BelkinAffiliated withDepartment of Plant and Environmental Sciences, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem Email author 

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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.

Keywords

Genotoxicity detection Whole-cell biosensors Environmental monitoring Electrochemistry Bioluminescence Escherichia coli