Assessment of a new technique combining a viability test, whole-cell hybridization and laser-scanning cytometry for the direct counting of viable Enterobacteriaceae cells in drinking water.

A new direct approach, called direct viable count (DVC)-FISH-ScanRDI, combining viability measurement, specific detection and sensitive enumeration of highly diluted Enterobacteriaceae cells, was assessed during the summer in water samples from a North American drinking water treatment plant and its distribution system. Major results of this field investigation show a higher sensitivity of the DVC-FISH-ScanRDI approach in enumerating viable Enterobacteriaceae cells in distributed drinking water, relative to a culture-based method, and the increased concentration of viable but non-culturable (VBNC) Enterobacteriaceae cells in distributed water for temperatures above 18 degrees C.

Biofilm responses to ageing and to a high phosphate load in a bench-scale drinking water system.

The effects of ageing and of phosphate load on drinking water biofilms developed on a polycarbonate substratum in the pseudo-equilibrium state have been evaluated. Phosphate was added in an amount higher than the stochiometric nutrient requirements of bacteria, at concentrations commonly applied in a drinking water distribution system for corrosion control. Multiple parameters were monitored: heterotrophic plate counts (HPCs), total direct counts (TDCs) and potential exoproteolytic activity (PEPA) in order to characterise changes in bacterial biofilms. The total carbohydrate, amino acid and phosphate contents of biofilms were analysed to characterise and monitor the biochemical composition of the biofilm.The three enumeration methods showed that a pseudo-equilibrium state was reached after 7 weeks of colonisation after which, the bacterial growth rate in the biofilm was 0.1 log per week on average. Bulk phosphate addition doubled the phosphate in the biofilm, but did not affect the other biological, physiological or chemical parameters measured. Polysaccharides increased in the biofilm with ageing and the dynamics of individual carbohydrate synthesis also varied with the age of the biofilm. Once pseudo-equilibrium, it was found that the total proteins were globally constant, whereas the spectra of some individual amino acids of the proteins had significantly changed.

Rapid and sensitive enumeration of viable diluted cells of members of the family enterobacteriaceae in freshwater and drinking water.

Water quality assessment involves the specific, sensitive, and rapid detection of bacterial indicators and pathogens in water samples, including viable but nonculturable (VBNC) cells. This work evaluates the specificity and sensitivity of a new method which combines a fluorescent in situ hybridization (FISH) approach with a physiological assay (direct viable count [DVC]) for the direct enumeration, at the single-cell level, of highly diluted viable cells of members of the family Enterobacteriaceae in freshwater and drinking water after membrane filtration. The approach (DVC-FISH) uses a new direct detection device, the laser scanning cytometer (Scan RDI). Combining the DVC-FISH method on a membrane with Scan RDI detection makes it possible to detect as few as one targeted cell in approximately 10(8) nontargeted cells spread over the membrane. The ability of this new approach to detect and enumerate VBNC enterobacterial cells in freshwater and drinking water distribution systems was investigated and is discussed.