K-123. Distinct Transcriptional Responses of the Escherichia coli O157:H7 Spinach Outbreak Strain and the Sakai Strain Exposed to Bovine Mammary Epithelial Cells (MAC-T)

G. S. Abu-Ali, L. M. Ouellette, T. S. Whittam;
Michigan State Univ., East Lansing, MI.

E. coli O157:H7, responsible for many outbreaks and sporadic cases of foodborne illness, caused a multistate outbreak in 2006 that was linked to consumption of contaminated spinach. In all, 204 cases were reported from 26 states and 1 case in Canada. The high hospitalization rate observed in the spinach outbreak suggests that this particular O157:H7 strain is hyper-virulent. To test this hypothesis, we compared the transcriptome of the Spinach strain to the Sakai strain from the 1996 Japan outbreak. Exponentially growing strains were exposed to MAC-T monolayers for 30 minutes. Both strains have been shown to intimately adhere to MAC-T cells. RNA was harvested from pathogenic O157:H7 cells preceding intimate attachment to MAC-T cells, from 5 independent cultures of both strains. cDNA dye-swap hybridizations were performed on oligo-arrays that probe for 6088 ORFs from O157:H7 Sakai, EDL933, pO157 plasmid and K12 MG1655. Analysis of variance (R/MAANOVA, Fs test) identified 915 genes significantly differentially expressed (p<0.05). Of those, 206 genes had a fold change difference between 1.5 and 8.3. From the locus of enterocyte effacement (LEE), 37 genes were upregulated in the Spinach strain of which eae, tir, espA, espB and sepZ were confirmed by qRT-PCR. In addition, adhesins aidA-I (ECs1396) and ECs0350, encoding an HmwA-like protein, were upregulated in the Spinach strain. qRT-PCR also confirmed an increased expression of Shiga toxin 2 genes, stx2a and stx2b, in the Spinach strain. Upregulation of a LysR-like regulator (ECs0309) in the Spinach strain, associated with expression of virulence in a number of species, was consistent with an overall increase in transcription of virulence genes in this strain. In Sakai, 14 genes involved in flagellation and chemotaxis were found to be upregulated, pointing to a temporally discrepant regulation of the LEE and the flagella in the two strains. The results suggest that genetic traits have increased expression of specific virulence factors in the Spinach strain which may account for variation in virulence among O157:H7 clades and help elucidate why certain O157 strains cause more serious disease.