Koordinierung von Virulenzfaktoren und Lebensstilwechsel bei Pseudomonas aeruginosa anhand von Einzelzellanalysen

Pseudomonas aeruginosa, a versatile Gram-negative opportunistic pathogen, relies on multiple virulence mechanisms, including a Type III Secretion System (T3SS) and several Type VI Secretion Systems (T6SS), to establish infections. The bacterial universal second messenger cyclic di-guanylate (c-di-GMP) orchestrates the lifestyle transitions of Pseudomonas aeruginosa between motile and biofilm-associated states, and influences the expression of virulence traits. However, while previous research suggests that c-di-GMP downregulates the T3SS and upregulates the H1-T6SS, recent evidence challenges this simplicity. Besides, a comprehensive virulence picture of P. aeruginosa, especially the crosstalk existed among the virulence systems, has still not emerged. In this thesis, I combine the population-level and single-cell analysis to dissect the role of c-di-GMP in regulating secretion systems’ functions, as well as the virulence functional diversification of P. aeruginosa populations. The results show that on the single-cell level, as on the population level, high c-di-GMP levels lead to increased formation and activity of the H1-T6SS, while negatively influencing formation and activity of the T3SS. Moreover, this thesis shows a comparison of the c-di-GMP – T3SS relationship between Pseudomonas aeruginosa and Yersinia enterocolitica, which presents possible different virulence strategies in various pathogen species. The lifestyle transition, modulated by c-di-GMP, helps the pathogen to deal with diverse environments. Different virulence features, as well as their combinations, might reflect the survival strategy of the pathogens from another perspective. Therefore, the association among the presence of various virulence systems is also analyzed in this thesis. On the single-cell level, I demonstrate a previously uncharacterized division of labor within P. aeruginosa populations by presenting a cooperative relationship among T3SS and flagellum, as well as an antagonistic relationship between presence of the H1-T6SS and the T3SS, as well as the flagellum. This division of labor suggests a strategy for optimizing survival and pathogenicity under varying environmental conditions. An optogenetic switch capable of translocating cytosolic proteins to the cell membrane, thereby sequestering their functions, was developed and evaluated. This model offers a powerful tool for manipulating virulence systems such as T3SS and T6SS in P. aeruginosa, holding significant potential for advancing our understanding of P. aeruginosa as a pathogen and facilitating downstream applied bioscience studies. Collectively, this thesis provides a comprehensive and in-depth exploration of P. aeruginosa virulence, offering valuable insights that could inform future strategies to combat P. aeruginosa infections.