A recombinant Escherichia coli sensor strain for the detection of tetracyclines.

Abstract

A bioluminescent Escherichia coli K-12 strain for the specific detection of the tetracycline group of antibiotics is described. A sensor plasmid, containing five genes from bacterial luciferase operon of Photorhabdus luminescens inserted under the control of tetracycline-responsive elements of the transposon Tn10, was constructed. Usage of the full-length luciferase operon in the sensor resulted in tetracycline-dependent light production without additions, i.e., self-luminescent phenotype, since all the substrates were intrinsically produced by the recombinant organism. The time needed for optimal induction of light emission was 90 min. Maximal induction of approximately 100-fold over uninduced levels by using 20 ng of tetracycline, and picomole sensitivities for the seven different tetracyclines tested, were obtained without added Mg2+ ions. The higher the pH and the magnesium ion concentration in the assay medium the higher was the amount of membrane-impermeable tetracycline-Mg2+ chelate complex. In consequence, by adjusting the pH and the Mg2+ ion concentration, the sensitivity of the assay can be modified for different analytical purposes. Different non-tetracycline antibiotics did not cause induction of light emission.