The transcriptome of Escherichia coli O157:H7 reveals a role for oxidative stress resistance in its survival from predation by Tetrahymena

Authors: George, Andree; Rehfuss, Marc; Parker, Craig; Brandl, Maria

Submitted to: FEMS Microbiology Ecology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/26/2020
Publication Date: 2/3/2020
Citation: George, A.S., Rehfuss, M.Y., Parker, C., Brandl, M. 2020. The transcriptome of Escherichia coli O157:H7 reveals a role for oxidative stress resistance in its survival from predation by Tetrahymena. FEMS Microbiology Ecology. 96(3). https://doi.org/10.1093/femsec/fiaa014.
DOI: https://doi.org/10.1093/femsec/fiaa014

Interpretive Summary: Outbreaks of Shiga toxin-producing E. coli associated with consumption of fresh produce remain a serious food safety concern. The survival of these pathogens in water is an important aspect of produce contamination in the agricultural environment. E. coli O157:H7 (EcO157) has the ability to survive digestion by the free-living ciliate Tetrahymena spp. We investigated the transcriptome of EcO157 EDL933 during interaction with Tetrahymena by microarray analysis and observed that genes involved in mitigating reactive oxygen stress, including aphC and aphF, katG, and emrR were significantly upregulated in the vacuoles of the protist. We hypothesized that the oxidative stress response in EcO157 partly mediates its resistance to the Tetrahymena digestive process. A double mutant of EcO157 with deletions in katG and ahpCF was more inhibited than the wild type strain by peroxides. The double mutant also had reduced viability after ingestion and excretion as fecal pellets by Tetrahymena. Our results support the hypothesis that resistance to ROS plays a role in the interaction of EcO157 with Tetrahymena in aqueous habitats. As the occurrence of produce-associated outbreaks of foodborne illness continues to impact public health and the produce industry, our findings provide information relevant to the survival of this important foodborne pathogen in agricultural environments related to produce production.

Technical Abstract: E. coli O157:H7 (EcO157) has the ability to survive digestion by the free-living ciliate Tetrahymena spp. We investigated the transcriptome of EcO157 EDL933 during interaction with Tetrahymena by microarray analysis and observed that genes involved in mitigating reactive oxygen stress, including aphC and aphF, katG, and emrR were significantly upregulated in the vacuoles of the protist. We hypothesized that the oxidative stress response in EcO157 partly mediates its resistance to the Tetrahymena digestive process. The presence of reactive oxygen species (ROS) in the vacuoles was confirmed by epifluorescence microscopy using CELLROX orange reagent. Deletion mutants in oxidative stress genes katG and ahpCF of EDL933 were constructed and showed a significant increase in inhibition compared with the wild type strain (P<0.05) upon exposure to hydrogen peroxide (H2O2) and the organic peroxide, cumene hydroperoxide, respectively, in a disk diffusion assay. A double mutant in KatG and AhpCF was inhibited by both peroxides. Bacterial cells and protist fecal pellets from co-cultures of Tetrahymena with the EDL933 GFP-labeled KatG-AhpCF-minus mutant and wild type strain were labeled with propidium iodide for evaluation of EDL933 cell viability by epifluorescence microscopy. Distribution of the fraction of dead EDL933 cells in fecal pellets of Tetrahymena indicated a decrease in cell viability of the mutant cells compared with that of the wild type strain. Overall, these data support our hypothesis that resistance to ROS plays a role in the interaction of EcO157 with Tetrahymena and its survival in aqueous habitats.