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Pathogenicity of Drinking Water Isolates of Heterotrophic Bacteria with Putative Virulence Factors

Petterson, Lynne
EPA Office of Research and Development
Start date
End date
Develop data for use in assessing the efficacy of the in vitro virulence tests in predicting pathogenicity of heterotrophic bacteria and in assessing risk of infection by opportunistic pathogens in potable water and to attempt to identify possible opportunistic pathogens among the heterotrophic bacteria that occur in drinking water, while determining the influence of type of source water or treatment technology on the occurrence of opportunistic pathogens.
Although the heterotrophic plate count (HPC) bacteria normally found in potable water are not a threat to the healthy population, some of them may be opportunistic pathogens that could cause adverse health effects in individuals with impaired immune systems. Earlier studies of these organisms have suggested that a small percentage of them express in vitro characteristics normally associated with virulence and may, therefore, be opportunistic pathogens. Heterotrophic bacteria isolated by Dr. Pierre Payment from the distribution system in Montreal express in vitro virulence characteristics with greater frequency than those typically found in chlorinated drinking water. These organisms are being tested in immunocompromised mice to determine whether they truly have pathogenic potential. The primary purpose of these studies is to ascertain the accuracy of the in vitro tests developed in our laboratory in predicting pathogenicity as established in animal models. If these tests are highly accurate in predicting pathogenicity, the need for future animal testing would be greatly reduced. A second purpose is to possibly identify currently unrecognized opportunistic pathogens among Dr. Payments isolates. Mice subjected to immunosuppression by carrageenan and cyclophosphamide are challenged with low numbers of heterotrophic bacteria and observed for signs of infection or for death If any mice become infected, the organism causing the infection will be isolated from their livers or spleens; and the identity of the infecting organism as the challenge strain will be verified by cultural or (if necessary) molecular methods. The data will be used to determine how well the various combinations of in vitro virulence tests correlate with pathogenicity as shown in an animal model. If any currently unknown opportunistic pathogens are discovered they will be studied in greater detail in a separate task. It seems possible that some of the other slow-growing bacteria could be capable of causing illness in highly susceptible individuals. Heterotrophic bacteria occur in even greater numbers and in more varieties in untreated ground waters and in potable waters treated by ozonation or chloramination; thus, it is essential to screen all of these types of potable waters for possible opportunistic pathogens. Representative samples of potable water from communities using various types of treatment and communities using untreated groundwater will be analyzed for opportunistic pathogens by concentrating water samples and injecting the concentrates into immunocompromised mice. The samples will be from points in the distribution systems in which the HPC bacteria are most numerous rather than sites containing typical numbers. Point-of-use (POU) filters will be installed at sites containing typical numbers and types of HPC bacteria and concentrates of the bacteria that amplify and are eluted from the filters will also be tested in the compromised mice. The number of samples per community and the number of communities to be sampled per region will be determined from the results of Task 0378. If any mice become infected, the organism causing the infection will be isolated from the mice and the presence of that strain in the water sample will be verified by cultural and, if necessary, by molecular DNA fingerprinting methods. The data will be used to identify possible risks that these various types of water present to immunocompromised people who are exposed to them.
Project number
Bacterial Pathogens
Sanitation and Quality Standards