Karla Bosse-Plois
PhD Student
Hospital and Environmental Hygiene
Institute of Hygiene
Robert-Koch-Str. 41
D-48149 Münster, Germany
Tel.: +49 251 83-59965
Education
- Since 2024:
PhD studies in the Hospital and Environmental Hygiene Group, Institute of Hygiene, University Hospital Münster, Germany - 2020-2023:
Master of Science (M.Sc.) in Microbiology, Rheinische Friedrich-Wilhelms-Universität Bonn, Germany. - 2016-2020
Bachelor of Science (B.Sc.) in Biology, University of Cologne, Germany.
Work experience
- 2022-2023
Scientific assistant in the care of laboratory animals, Bonn University Hospital, Institute for Medical Microbiology, Immunology and Parasitology - 2023
Scientific assistant in accounting, Bonn University Hospital, Institute for Pathology
Supervisors
- Prof. Dr. Alexander Mellmann, Hospital and Environmental Hygiene Group, Institute for Hygiene, University Hospital Münster, Germany
- Prof. Dr. Stefan Ludwig, Molecular Infection Biology Group, Institute of Virology, University of Münster, Germany
- Prof. Dr. Petra Dersch, Molecular Infection Biology Group, Institute of Infectiology, University of Münster, Germany
Research interests
- Biology of host-pathogen interactions
- Genome plasticity
- Impact of environmental pressures on EHEC
PhD project description
Escherichia coli (E. coli) is one of the most frequent commensal colonizers of the intestine of mammals and warm-blooded animals. In the digestive tract, commensal strains inhabit the mucus layer that covers the epithelial cells of the large intestine, as part of the healthy microbiome. However, pathogenic E. coli cause several million infections and thousands of deaths every year. In 2011, the largest outbreak of enterohemorrhagic E. coli (EHEC) O104:H4 yet recorded caused almost 4000 cases of acute gastroenteritis with an unusually high progression to haemolytic uremic syndrome (HUS) in adults, leading to 54 deaths. Interestingly, the outbreak strain was identified to be more closely related to enteroaggregative E. coli (EAEC) strain 55989 than to other EHEC strains and to carry virulence factors specific to both pathogroups. More precisely, EHEC O104:H4 produces Stx2a, the cardinal factor of EHEC responsible for cellular cytotoxicity. Furthermore, the outbreak strain holds an EAEC-specific pAA plasmid, which encodes genes responsible for the characteristic “stacked-brick” adherence to cells, while lacking the typical EHEC mode of attachment. To summarise, virulence factors from both EAEC and EHEC together with novel tools are estimated to be responsible for the outstanding virulence of EHEC O104:H4. Alternative sigma factor RpoS is transcribed upon specific environmental stimuli in E. coli. While the housekeeping sigma factor regulates the expression of essential genes and pathways, RpoS is a global regulator of stress specific adaptions of the organism. Acting as an activator and repressor, RpoS serves as a tool to react to nutrient limitations or other stressors. Strikingly, a severe heterogeneity of rpoS gene sequence and function was surveyed even on strain level in natural and laboratory populations of E. coli. Intriguingly, a laboratory-acquired single nucleotide polymorphism (SNP) in the start codon of rpoS of a clinical isolate from EHEC O104:H4 was identified in our lab (paper under revision). The bacteria were characterised by an enhanced expression of virulence factors, even though RpoS levels were reduced 5-fold. A second, upstream-located ATG was detected, accompanied by a shortened Shine-Dalgarno sequence that might act as a less efficient ribosome-binding site. It remains uncertain, which start codon drives rpoS expression and the effect of polymorphism on the function, virulence, and fitness of EHEC O104:H4. Thus, the impact of the annotated ATG, the mutated ATG>ATA and the upstream ATG on RpoS are to be characterised. Therefore, yfp-fusion constructs using recombineering will be established in collaboration with Prof. Ulrich Dobrindt (ICB). YFP amounts will be determined by Western blot, kinetic measurements in a multiplate reader, fluorescence microscopy and eventually single cell flow cytometry. Additionally, the N-terminal sequence of RpoS in E. coli EHEC O104:H4 wild type and ATG>ATA mutant will be directly identified by 2D gel electrophoresis and subsequent mass spectrometry in collaboration with Prof. Simone König (IZKF Core Unit Proteomics). Next, the function of wild type and ATG>ATA RpoS will be analysed in in vitro transcription assays. Therefore, the recombinant RpoS variants will be purified, mixed with E. coli RNA polymerase core enzyme and in vitro transcription of circular or linearized templates assessed. The template will be a high copy number plasmid carrying the rpoS-dependent proV operon, fused to yfp. Samples will then be subjected to northern blot analysis. Lastly, the impact of additional RpoS polymorphism of two more patient isolates collected during the 2011 outbreak will be defined by re-sequencing to confirm the nucleotide sequence and screen for additional mutations. Their polymorphisms include a premature stop codon and the mutation of the C-terminal region. Corresponding rpoS alleles will be expressed heterologously in E. coli O104:H4 ΔrpoS and the effect on protein amounts, virulence gene expression and metabolism monitored by western blot analysis, cell culture experiments and Biolog microarrays. Additionally, whole genome sequence data from available EHEC patient isolates will be screened for rpoS polymorphism and their global impact on gene expression compared by RNA sequencing. For Illumina sequencing, cDNA libraries will be generated and sequenced reads will be mapped in relation to EHEC O104:H4 and E. coli K12
Publications
Hansen P, Haubenthal T, Reiter C, Kniewel J, Bosse-Plois K, Niemann HH, von Bargen K, Haas A. 2023. Differential Effects of Rhodococcus equi Virulence-Associated Proteins on Macrophages and Artificial Lipid Membranes. Microbiol Spectr 11:e03417-22. https://doi.org/10.1128/spectrum.03417-22