Determination of low-density Escherichia coli and Helicobacter pylori suspensions in water

Determination of low-density of bacteria, especially those of pathogenic strains in water, has proven difficult and challenging. Here, we developed and evaluated a lanthanum-based concentration method coupled with quantitative real-time PCR to concentrate and detect selected bacteria (Escherichia coli and Helicobacter pylori) in water. To improve qPCR efficiency, the flocs with enmeshed bacteria after chemical flocculation need to be dissolved before PCR detection. Ethylenediaminetetraacetic acid (EDTA) salt successfully dissolved the flocs from a lanthanum-based flocculation process, but not those from the traditional processes using chemicals such as FeCl3 or Al2(SO4)3. Lanthanum-based concentration coupled with real-time PCR successfully determined E. coli at a concentration of 15 cells/mL in raw and finished water and H. pylori at a concentration of about 1 cell/mL in the finished water prior to disinfection. The H. pylori detection sensitivity could be easily increased to 60 cells/L by reducing the final volume of the DNA samples from 3 mL to 60 μL. With the elimination of membrane-clogging problem that is often encountered in direct membrane filtration, the lanthanum-based chemical flocculation coupled with qPCR is a promising method for determination of low-density of microbial suspensions in water.

Schematic of lanthanum-based bacterial concentration and PCR detectionFigure optionsDownload high-quality image (69 K)Download as PowerPoint slideHighlights
► Lanthanum-based concentration coupled with qPCR successfully determined low-density bacteria.
► EDTA salt successfully dissolved the flocs resulting in good recovery of bacteria in water.
► EDTA did not dissolve the flocs when FeCl3 or Al2(SO4)3 was used as a coagulating agent.
► The new method could determine Helicobacter pylori at a concentration of 60 cells/L.