COMPARISON OF METHODS FOR THE DETECTION OF CULTURABLE AND VBNC ESCHERICHIA COLI 0157:H7 IN COMPLEX DRINKING WATER BIOFILMS

摘要

Vcrocytotoxigenic Escherichia coli 0157:H7 is responsible for outbreaks worldwide, causing haemorrhagic colitis characterised by abdominal pain and bloody diarrhoea, and can lead to haemolytic uraemic syndrome (HUS) which can, in turn, result in kidney failure. Outbreaks are often linked to inadequately cooked or prepared meat and other foodstuffs.1 E. coli 0157:H7 has also been responsible for a number of serious waterborne outbreaks2 including more than 2000 cases (with 7 deaths) in Walkerton, Canada in 2000.3 Previous work has shown E. coli 0157:H7 to survive in potable water for extended periods of time4 and to be incorporated into associated biofilms. It is known that bacteria can enter a viable but non-culturable (VBNC) state where they are sub-lethally stressed and their metabolic activity reduced, meaning they can be missed by standard detection methods such as culture-based analyses. In the current study, alternative methods have been compared with culturing. The approaches tested were the use of a combined method of direct viable count (DVC; a cell elongation based assay) with peptide nucleic acid (PNA) probes in a fluorescence in situ hybridation (FISH) assay and a real-time quantitative PCR (qPCR) method utilising propidium monoazide (PMA).