Investigations of the impact of different determinants on the virulence and fitness of EHEC O104:H4

• Applicant: Dr. Petya Berger
• Subject Area: Medical Microbiology and Mycology, Hygiene, Molecular Infection Biology
• Term: from 2015 to 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 276606594

Enterohemorrhagic Escherichia coli (EHEC) can cause severe foodborne illness in humans, which is typically characterized by bloody diarrhea and progresses to hemolytic uremic syndrome (HUS) in circa 10% of the cases. EHEC O104:H4 was identified as the causative agent of the largest German outbreak (May-July 2011) during which nearly 4000 people were infected, and of them 22% developed HUS. Besides having a chromosomally integrated Shiga toxin 2 (Stx2) encoding phage, the highly virulent EHEC O104:H4 expresses pAA plasmid-encoded aggregative adherence fimbriae I (AAF/I; a characteristic virulence feature of enteroaggregative E. coli), which are mediating its tight adherence to cultured human epithelial cells. In addition, EHEC O104:H4 displays an extended spectrum beta-lactamase (ESBL) phenotype mediated by the conjugative pESBL plasmid.Our research exploits state of the art transcriptomic approaches to gain further insight into the virulence and fitness determinants of EHEC O104:H4. In the first funding period, we analyzed the primary transcriptomes of the pAA and pESBL plasmids and gained further insights into their gene expression using differential RNA seq, which is a powerful method for mapping of transcription start sites and non-coding RNAs. In addition, using comparative RNA-seq we identified features which are shared between EHEC O104:H4 and other less pathogenic and commensal strains, but are differentially expressed under identical conditions. We hypothesized that such cases of differential gene expression may significantly contribute to EHEC O104:H4 virulence and fitness. Our transcriptome analysis revealed that under conditions simulating the gut central metabolic genes are downregulated in EHEC O104:H4 in comparison to the control strains included in our analysis. Further investigations revealed that lysogenizing E. coli K-12 MG1655 with the Stx2 phage of EHEC O104:H4 resulted in analogous changes in the transcriptome and phenotype. Therefore, in the second funding period we will analyze the impact of Stx2 phage carriage on EHEC O104:H4 host gene expression. Moreover, we will investigate the impact of Stx2 phage-dependent transcriptome on EHEC O104:H4 virulence and fitness and identify the phage-encoded factors mediating it. Last but not least, we will evaluate the biological function of several genes, which were found upregulated in EHEC O104:H4 in our analysis and were previously linked to E. coli virulence and fitness. We anticipate that our results will significantly contribute to our general understanding of EHEC pathogenicity and in particular be beneficial for risk assessment of emerging Stx hybrid strains. Moreover, the identification of common but differentially regulated E. coli determinants associated with EHEC virulence will be of relevance to public health by facilitating phenotypic characterization and surveillance of pathogenic strains and eventually therapy and/or prevention of the disease.

• DFG Programme: Research Grants
• International Connection: USA
• Cooperation Partner: Professor Gerald Koudelka, Ph.D.