title: Rapid amperometric detection of Escherichia coli in wastewater by measuring β-D glucuronidase activity with disposable carbon sensors.
abstract: An assay on the indirect amperometric quantification of the β-D-Glucuronidase (GLUase) activity was developed for the rapid and specific detection of Escherichia coli (E. coli) in complex environmental samples. The p-aminophenyl β-D-glucopyranoside (PAPG) was selected as an electrochemical substrate for GLUase measurement and the p-aminophenol (PAP) released during the enzymatic hydrolysis was monitored by cyclic voltammetry with disposable carbon screen-printed sensors. The intensity of the measured anodic peak current was proportional to the amount of GLUase, and therefore to the number of E. coli in the tested sample. Once the substrate concentration and pH values optimized, a GLUase detection limit of 10 ng mL(-1) was achieved. Using a procedure involving a filtration step of the bacteria followed by their incubation with the substrate solution containing both the nonionic detergent Triton X-100 as permeabilization agent and the culture media Luria broth to monitor the growth, filtered bacterial cells ranging from 5 × 10(4) to 10(8) UFC/membrane were detected within 3 h. The amperometric assay was applied to the determination of fecal contamination in raw and treated wastewater samples and it was successfully compared with conventional bacterial plating methods and uidA gene quantitative PCR. Owing to its ability to perform measurements in turbid media, the GLUase amperometric method is a reliable tool for the rapid and decentralized quantification of viable but also nonculturable E. coli in complex environmental samples.
authors.affiliation: Université Bourgogne Franche-Comté, UMR1347 Agroécologie, 17 Rue Sully, 21000 Dijon, France. Electronic address: Murielle.dequaire@u-bourgogne.fr.
authors.forename: Murielle
authors.initials: M
authors.lastname: Rochelet
authors.affiliation: Université Bourgogne Franche-Comté, UMR1347 Agroécologie, 17 Rue Sully, 21000 Dijon, France.
authors.forename: Sébastien
authors.initials: S
authors.lastname: Solanas
authors.affiliation: Université Bourgogne Franche-Comté, UMR1347 Agroécologie, 17 Rue Sully, 21000 Dijon, France.
authors.forename: Laetitia
authors.initials: L
authors.lastname: Betelli
authors.affiliation: Université Bourgogne Franche-Comté, UMR1347 Agroécologie, 17 Rue Sully, 21000 Dijon, France.
authors.forename: Benoît
authors.initials: B
authors.lastname: Chantemesse
authors.affiliation: Université Bourgogne Franche-Comté, UMR1347 Agroécologie, 17 Rue Sully, 21000 Dijon, France.
authors.forename: Fabienne
authors.initials: F
authors.lastname: Vienney
authors.affiliation: INRA, UMR1347 Agroécologie, 17 Rue Sully, 21000 Dijon, France.
authors.forename: Alain
authors.initials: A
authors.lastname: Hartmann
mesh_terms: D000627:Aminophenols / Q000032:analysis* / Q000378:metabolism; D002244:Carbon / Q000737:chemistry*; D055664:Electrochemical Techniques; D004566:Electrodes; D004926:Escherichia coli / Q000201:enzymology / Q000302:isolation & purification*; D005243:Feces / Q000737:chemistry; D005966:Glucuronidase / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D062065:Wastewater / Q000382:microbiology*
chemical_list: D000627:Aminophenols; D002244:Carbon; D005966:Glucuronidase; D062065:Wastewater; C026729:4-aminophenol
keywords: Electrochemistry; Escherichia coli; Sensors; Wastewater analysis; p-aminophenol; β-D glucuronidase