title: Protocol for the recovery and detection of Escherichia coli in environmental water samples.
abstract: To achieve active management of bathing areas and to reduce risk associated with the presence of fecal pollution, tests capable of rapid on-site assessment of microbiological water quality are required. A protocol for the recovery and detection of fecal pollution indicator bacteria, E. coli, using β-glucuronidase (GUS) activity was developed. The developed protocol involves two main steps: sample preparation and GUS activity measurement. In the sample preparation step, syringe filters were used with a dual purpose, for the recovery and pre-concentration of E. coli from the water matrix and as μL reactors for bacteria lysis and GUS extraction. Subsequently, GUS activity was measured using a continuous fluorometric method developed previously. The optimum GUS recovery conditions for the sample preparation step were found to be 100 μL PELB (supplemented with 1 mg mL-1lysozyme and 20 mM DTT) at 37 °C for 30 min. The protocol was evaluated on environmental samples (fresh and seawater) against an establish GUS assay method (Coliplage®). GUS activities corresponding to samples containing as low as 26 MPN E. coli 100 mL-1were detected for the seawater sample and as low as 110 MPN E. coli 100 mL-1for the freshwater samples. By comparison with the Coliplage®method, this protocol offered an improvement in the measured GUS activities of 3.1 fold for freshwater samples and 4.1 fold for seawater samples. Furthermore, the protocol developed here, has a time-to-result of 75 min, and successfully addresses the requirement for tests capable of rapid assessment of microbiological water quality.
authors.affiliation: DCU Water Institute, National Centre for Sensor Research (NCSR), School of Chemical Sciences, Dublin City University, Dublin, Ireland.
authors.forename: Ciprian
authors.initials: C
authors.lastname: Briciu-Burghina
authors.affiliation: DCU Water Institute, National Centre for Sensor Research (NCSR), School of Chemical Sciences, Dublin City University, Dublin, Ireland.
authors.forename: Brendan
authors.initials: B
authors.lastname: Heery
authors.affiliation: DCU Water Institute, National Centre for Sensor Research (NCSR), School of Chemical Sciences, Dublin City University, Dublin, Ireland. Electronic address: fiona.regan@dcu.ie.
authors.forename: Fiona
authors.initials: F
authors.lastname: Regan
mesh_terms: D004926:Escherichia coli / Q000302:isolation & purification*; D005243:Feces; D005618:Fresh Water / Q000382:microbiology; D005966:Glucuronidase; D012623:Seawater / Q000382:microbiology; D014871:Water Microbiology; D014876:Water Pollution; D060753:Water Quality
chemical_list: D005966:Glucuronidase
keywords: E. coli; Fecal pollution; Fluorescence; Rapid test; β-glucuronidase (GUS)