UC Berkeley

Diarrheagenic pathotypes of Escherichia coli are responsible for a substantial number of childhood deaths each year, and Helicobacter pylori causes gastric diseases in both children and adults worldwide. Free-living protozoa have been implicated in the survival and transport of pathogens in the environment, but the relationship between non-Shiga toxin-producing E. coli or H. pylori and free-living ciliated protozoa has not been characterized. Likewise, there is a paucity of research regarding the ability of these pathogenic bacteria to survive phagocytosis by amoebae.

A literature review revealed two articles demonstrating survival of E. coli O157:H7 or an environmental isolate of E. coli in Acanthamoeba and one article reporting survival of H. pylori in Acanthamoeba. There have been several reports of E. coli O157:H7 or genetically modified laboratory strains surviving digestion by the ciliate, Tetrahymena, and no investigations on the resistance of H. pylori to digestion by this protozoan.

Therefore, for the present study, six diarrheagenic serotypes of E. coli and an isolate of H. pylori were evaluated for their susceptibility to digestion by Tetrahymena. Escherichia coli and H. pylori were fed separately to Tetrahymena strain MB125, and the ciliate's egested products were examined for viable pathogens. Viability was assessed by the BacLight^TM LIVE/DEAD^TM assay, by a cell elongation method, and by colony counts. Three H. pylori strains were used: one expressing the green fluorescent protein, an ATCC strain originating from a clinical isolate, and a fresh clinical isolate of H. pylori, and each was fed separately to Acanthamoeba polyphaga. Viability of H. pylori in Acanthamoeba was assessed with: the BacLight^TM LIVE/DEAD^TM assay, fluorescent in situ hybridization, and quantification of amplified gene products unique to H. pylori over time, determined by a real time polymerase chain reaction test.

All six pathogenic E. coli serotypes survived digestion by Tetrahymena and emerged as viable cells in fecal pellets, whereas H. pylori was digested. Growth of E. coli on agar plates indicated that the bacteria were able to replicate after passage through the ciliate. Transmission electron micrographs of E. coli cells as intact rods versus degraded H. pylori cells corroborated these results. Scanning electron microscopy revealed a net-like matrix around intact E. coli cells in fecal pellets. Helicobacter pylori did not survive phagocytosis by Acanthamoeba.

These results suggest a possible role for Tetrahymena and its egested fecal pellets in the survival and dissemination of diarrheagenic E. coliin the environment. This bacterial-protozoan association may increase opportunities for transmission of these bacterial pathogens to mammalian hosts including humans.