Evaluating the growth potential of pathogenic bacteria in water

Abstract

The degree to which a water sample can potentially support the growth of human pathogens was evaluated. For this purpose, a pathogen growth potential (PGP) bioassay was developed based on the principles of conventional assimilable organic carbon (AOC) determination, but using pure cultures of selected pathogenic bacteria (Escherichia coli O157, Vibrio cholerae, or Pseudomonas aeruginosa) as the inoculum. We evaluated 19 water samples collected after different treatment steps from two drinking water production plants and a wastewater treatment plant and from ozone-treated river water. Each pathogen was batch grown to stationary phase in sterile water samples, and the concentration of cells produced was measured using flow cytometry. In addition, the fraction of AOC consumed by each pathogen was estimated. Pathogen growth did not correlate with dissolved organic carbon (DOC) concentration and correlated only weakly with the concentration of AOC. Furthermore, the three pathogens never grew to the same final concentration in any water sample, and the relative ratio of the cultures to each other was unique in each sample. These results suggest that the extent of pathogen growth is affected not only by the concentration but also by the composition of AOC. Through this bioassay, PGP can be included as a parameter in water treatment system design, control, and operation. Additionally, a multilevel concept that integrates the results from the bioassay into the bigger framework of pathogen growth in water is discussed. The proposed approach provides a first step for including pathogen growth into microbial risk assessment.

FIG. 1.

Schematic overview of the developed assay investigating the growth potential of pathogenic bacteria in different water samples. Abbreviations: FCM, flow cytometric measurement.

FIG. 2.

Growth of the three pathogens, E. coli O157 (white squares), V. cholerae (black circles), and P. aeruginosa (gray triangles), in water samples taken after the individual treatment steps from drinking water treatment plant A. The growth potential, i.e., net grown cells (A), as well as the fraction of estimated consumed assimilable organic carbon (AOC) for each pathogen (B) are shown. The samples or treatment steps are shown in order from left to right as follows: RW, untreated (raw) water; 1. Oz, the first ozonation; RSF, rapid sand filtration; 2. Oz, the second ozonation; GAC + SSF, granular activated carbon filtration and slow sand filtration. The error bars indicate the standard deviations for three replicate samples.

FIG. 3.

Growth of the three pathogens, E. coli O157 (white squares), V. cholerae (black circles), and P. aeruginosa (gray triangles), in samples taken after the individual treatment steps from drinking water treatment plant B. The growth potential, i.e., net grown cells (A), as well as the fraction of estimated consumed assimilable organic carbon (AOC) for each pathogen (B) are shown. The samples or treatment steps are shown in order from left to right as follows: RW, untreated (raw) water; 1.Oz, the first ozonation; RSF, rapid sand filtration; 2.Oz, the second ozonation; GAC, granular activated carbon filtration; Cl, chlorination. The error bars indicate the standard deviations for three replicate samples.

FIG. 4.

Growth of the three pathogens, E. coli O157 (white squares), V. cholerae (black circles), and P. aeruginosa (gray triangles), in samples taken after the individual treatment steps from a wastewater water treatment plant. The growth potential, i.e., net grown cells (A), as well as the fraction of estimated consumed assimilable organic carbon (AOC) for each pathogen (B) are shown. The samples or treatment steps are shown in order from left to right as follows: RW, untreated (raw) water; Bio, biological treatment; Oz, ozonation; SSF, slow sand filtration. The error bars indicate the standard deviations for three replicate samples.

FIG. 5.

Growth of the three pathogens, E. coli O157 (white squares), V. cholerae (black circles), and P. aeruginosa (gray triangles), in river water exposed to ozone. The growth potential, i.e., net grown cells (A), as well as the fraction of estimated consumed assimilable organic carbon (AOC) for each pathogen (B) are shown. RW, untreated (raw) river water; ct 1 to ct 10, degree of oxidation (mg·min/liter). Error bars indicate the standard deviations for three replicate samples.

FIG. 6.

Schematic overview of the three different levels that are important in assessing the risk of pathogen growth in water. For explanation, see the discussion of Fig. 6 in the Discussion section. PGP, pathogen growth potential.