, 2007), were upregulated in our microarray BEZ235 in vitro study, including stx1A, stx1B, and stx2A. Shiga toxins have been shown to play a role in E. coli O157:H7 survival within grazing protozoa, and it has been postulated that the maintenance of shiga toxin genes is important for the protozoal-bacterial interaction and not the mammalian host interaction (Steinberg
& Levin, 2007). The role of the LEE-encoded Type III secretion system (T3SS) in the development of E. coli O157:H7 attaching and effacing lesions and translocation of effectors is well documented (Knutton et al., 1998; Roe et al., 2003; Tobe et al., 2006). Our microarray analysis indicated that genes that comprise part of the LEE-encoded T3SS (espAD, escF) on the transcript expADB-escF-Z5102-Z5104 were upregulated. This result was further confirmed by qRT-PCR analysis estimating the upregulation of espA to be 5.1-fold. A second T3SS described in E. coli O157:H7 (Ideses et al., 2005) that shares homology with
the Salmonella pathogenicity island 1 (eivCAEGF) was also upregulated. Two LEE-encoded and seven non-LEE-encoded T3SS translocated factor transcripts were upregulated. In summary, the analysis of transcript changes in E. coli O157:H7 during its interaction with A. castellanii show that E. coli upregulates genes involved in the response to various stressful environments including iron deprivation and oxidative stress. These results are purely based on transcript levels and check details need to be confirmed at the protein level. The phenotype changes required for protozoa survival may also include changes in the surface architecture and the translocation of effector molecules by T3SS. Some virulence genes were also upregulated, which suggests that protozoa may serve as the environment that selects for and helps to maintain virulence genes that result in colonization and disease outbreaks in mammalian populations. We would like to thank Dr Greg Phillips for providing A. castellanii.
This study was funded in part by a contract to F.C.M. from the United States Department of Agriculture (Specific Cooperative Agreement 58-3625-2-127). Table S1. Differentially expressed genes of Escherichia coli O157:H7 within Acesulfame Potassium Acanthamoeba. Please note: Wiley-Blackwell is not responsible for the content or functionality of any supporting materials supplied by the authors. Any queries (other than missing material) should be directed to the corresponding author for the article. “
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