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Schlagwörter:
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Zusammenfassung:
Introduction: Bartonella bacilliformis is the causative agent of Carrion's disease, a vector-borne biphasic illness restricted to the South American Andes. The pathogen infects human erythrocytes causing severe hemolytic anemia (Oroya fever) with high mortality rates in untreated patients. In a second chronic phase, the infection of endothelial cells results in the formation of blood-filled nodular lesions at skin sites (verruga peruana). Underlying molecular mechanisms of host infection are still unclear. Trimeric autotransporter adhesins (TAAs) play an essential role in bacterial pathogenicity and are encoded in various Bartonella spp. We are interested in Bartonella bacilliformis adhesins A (BbadA) that has been identified as a TAA.
Objectives: The objective of this work is the identification and characterization of immunodominant proteins of B. bacilliformis and the genetic and functional characterization of BbadA and Flagellin. Furthermore, potential target proteins will be analyzed for diagnostic and therapeutic use.
Material & Methods: Molecular genetic strategies using bacterial mutants (transposon mutagenesis library), recombinant protein expression strategies (heterologous expression library) and a reverse vaccinology approach are used to identify immunodominant proteins and pathogenicity factors of B. bacilliformis. Deletion mutants of bbadA and flagellin were generated using homologous recombination techniques. Furthermore, infection experiments with erythrocytes and endothelial cells will be performed to characterize the role of BbadA in the infection process.
Results & Conclusion: A genomic B. bacilliformis expression library was established in E. coli and screened by using anti B. bacilliformis sera. Western blot analysis with reactive mutants revealed immunodominance of recombinant BbadA and Flagellin. The genomic deletion of bbadA and flagellin was confirmed by PCR and sequencing. Electron microscopy showed the presence of BbadA on the surface of B. bacilliformis but not in ΔbbadA. Furthermore, the deletion of bbadA leads to a reduction in cell adhesion to endothelial cells.
