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Master
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Exploration of P. aeruginosa metal homeostasis systems during phagocytosis AND Zucchero project: Investigating the emergence of phage-resistant P. aeruginosa mutants following phage therapy

DirectorsPerron, Karl
Number of pages71
Defense date2023-08-26
Abstract

Pseudomonas aeruginosa is a Gram-negative bacterium that carries a large repertoire of genes involved in pathogenicity as well as in adaptations to environmental changes. This versatility allows the pathogen to survive in several environments like metal-excess and antibiotic-supplemented ones. Besides, both are linked transcriptionnally with different metal-induced regulators having effects on the expression of antibiotic-resistance genes. The bacterium also possesses high intrinsic resistance to antibiotics because of a low outer-membrane permeability and multiple efflux systems that it can regulate. It is an opportunistic pathogen that thrives in hospital-related environments and induces various nosocomial infections. These characteristics place the bacteria in the ranks of the ESKAPE pathogens, for which urgent new therapies' discovery is needed.

Boosts of zinc and copper take place in the phagosome and have multiple antibacterial effects. The phagocytosis of P. aeruginosa was therefore studied to observe and explore the different gene regulations as well as their importance in this singular environment. The activation of the different zinc and copper-related systems as well as their effect on the antibiotic-resistance genes like oprD and mexPQ-opmE was indeed observed. Investigation about both genes' role in the phagosome was also conducted, however, no link with intraphagosomal survival was highlighted. The Mex systems was also investigated for its plausible homologous function with E. coli's copper efflux systems CusABC. Despite previous studies showing a role in copper resistance, none was found in this work. This was backed-up by in silico phylogenetic assays in the litterature. Survival of different metal-homeostasis-related mutants was also examined but non showed significantly different survival rates than a WT PAO1 one.

A special case in phage therapy was also assessed in this thesis. The emergence of phage-resistant bacteria (Pae_0271) after he in vivo use of a bacteriophage cocktail, "Pyophage", was reported and strains were collected at distinct stages of the treatment to find the cause of this resistance. Moreover, different attempts at isolating a novel phage capable of lysing these phage-resistant bacteria were conducted without success. A large deletion (148.1 kb) in the retrieved strain Pae_0271 was found to be the cause of the phage resistance through the loss of the galU gene. Its phenotype could also be explained by the loss of the hmgA gene, which led to the production of pyomelanin, a brown/reddish pigment. Other genes linked with antibiotic resistance were also comprised in this deletion, but their loss did not provide any restoration of the sensitivity.

eng
Keywords
  • Pseudomonas aeruginosa
  • Macrophages
  • Antibiotics
  • Metals
  • Phage therapy
  • Resistance
Citation (ISO format)
BAUDIN, Raphaël Alain. Exploration of <i>P. aeruginosa</i> metal homeostasis systems during phagocytosis AND Zucchero project: Investigating the emergence of phage-resistant <i>P. aeruginosa</i> mutants following phage therapy. 2023.
Main files (1)
Master thesis
accessLevelRestricted
Identifiers
  • PID : unige:171175
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Creation09/01/2023 3:11:17 PM
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