title: Method for rapid detection of viable Escherichia coli in water using real-time NASBA.
abstract: A rapid real-time NASBA method was developed for detection of Escherichia coli in water samples. In this method, a fragment of the clpB-mRNA is amplified and a specific molecular beacon probe is used to detect the amplified mRNA fragment during the NASBA reaction. The method was shown to be specific and sensitive (1 viable E. coli in 100ml) and can be performed within 3-4h. Different inactivation processes (starvation, heat, UV-irradiation and chlorine) were employed to study the relationship between culturability and the ability to detect E. coli using NASBA. Detection of clpB-mRNA correlated with culturability after starvation or chlorine treatment. After UV-irradiation or heat-inactivation, detection of the increase in production of clpB-mRNA in viable E. coli cells after heat-shock induction correlated with culturability. Application of the NASBA method on tap water, treated sewage and surface water samples showed that culture and NASBA yielded comparable results in these different matrices. This study demonstrates that the NASBA method has high potential as a rapid test for microbiological water quality monitoring.
authors.affiliation: KWR, Watercycle Research Institute, P.O. Box 1072, 3430 BB Nieuwegein, The Netherlands. leo.heijnen@kwrwater.nl
authors.forename: Leo
authors.initials: L
authors.lastname: Heijnen
authors.forename: Gertjan
authors.initials: G
authors.lastname: Medema
mesh_terms: D049071:Endopeptidase Clp; D004784:Environmental Monitoring / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification* / Q000502:physiology*; D029968:Escherichia coli Proteins / Q000235:genetics; D006360:Heat-Shock Proteins / Q000235:genetics; D021161:Self-Sustained Sequence Replication / Q000379:methods*; D014871:Water Microbiology
chemical_list: D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins; D049071:Endopeptidase Clp; C069775:ClpB protein, E coli