Development of an enzyme assay for rapid assessment of Escherichia coli in seawaters

Abstract

Aims: An analytical protocol has been developed and applied for the detection of glucuronidase activity in marine waters as a rapid alternative approach to assess the microbiological quality of seawaters. Methods and Results: The fluorogenic substrate 4-methylumbelliferyl-b-D-glucuronide is cleaved to a fluorescent product, methylumbelliferone, by the enzyme b-glucuronidase, specific to Escherichia coli and closely related enterobacterial species (Shigella). The results suggest that this test is related to E. coli numbers, as estimated by immunofluorescence, more significantly than to faecal coliform numbers, obtained from culture media. Conclusions: The determination of the potential rate of glucuronidase activity may be used as a diagnostic tool for the indirect estimation of the presence of E. coli in seawaters. Significance and Impact of the Study: The method may be particularly useful in the early warning of seawater pollution, allowing the screening of coastal areas with different contamination levels in reduced time.

A biosensor platform for rapid detection of E. coli in drinking water

“…The high levels of activity observed in some cultures may indicate that their starved metabolic state lead to an increase in bacterial enzymatic activities hydrolyzing the fluorogenic substrate rapidly. Caruso et al (2002) reported that full development of enzymatic activities start at lag phase and is required for the enzyme expression.…”

“…Enzyme activities are subject to the physiological status of bacteria and that under nutritional and light stresses, a fraction of cells may gradually lose its culturability, although remaining metabolically active (Caruso et al 2002). Besides, E. coli β-Dglucuronidase (GUD) activities are very sensitive to temperature.…”

“…β-D-glucuronidase (GUD) activity is characteristic of most strains of E. coli and, hence, has been widely used to monitor the presence of this indicator organism in environmental samples, particularly water (Berg and Fiksdal 1988;Edberg et al 2000;Caruso et al 2002). Approximately 97% of E. coli strains have GUD activity and almost all other coliform bacteria lack this enzyme (Caruso et al 2002).…”

“…Approximately 97% of E. coli strains have GUD activity and almost all other coliform bacteria lack this enzyme (Caruso et al 2002). Studies have shown that E. coli can preserves an active metabolism without being able to grow on culture media (Roszak and Colwell 1987;George et al 2001).…”

“…Approximately 97% of E. coli strains have GUD activity and almost all other coliform bacteria lack this enzyme (Caruso et al 2002). The hydrolysis of MUG releases fluorescent 4-methylumbelliferyl (4MU) and the intensity of the measured fluorescent signal is proportionate to the amount of enzyme present, showing a correlation to the number of E. coli present in the sample (Fiksdal and Tryland 2008).…”