Harnessing the Klebsiella

: Macrophages arms race

Abstract

Klebsiella pneumoniae is an emerging pathogen for which there are limited therapeutic options. K. pneumoniae survives in macrophages within specialised Klebsiella containing vacuoles (KCV) by subverting phagosome maturation. Here, we have investigated the metabolic adaptation of K. pneumoniae to its intracellular lifestyle.  Isogenic mutants were constructed for key enzymes from each metabolic pathway and assessed for intracellular survival. We found that the glyoxylate shunt was dispensable for K. pneumoniae survival within the KCV, but disruption of glycolysis, gluconeogenesis and pentose phosphate pathway impaired survival within macrophages. From the tricarboxylic acid cycle, fumarate reductase and isocitrate and malate dehydrogenases play a role in intracellular survival whereas neither aconitase nor citrate synthase do. Klebsiella-controlled blockage of phagolysosome maturation was compromised for most that showed an intracellular survival phenotype, but 6-phosphofructokinase and 6-phosphogluconate dehydrogenase did not affect this pathway, possibly due to a nutritional deficit. Disruption of these enzymes also resulted in recruitment of Rab14 to the KCV, unlike the mutants that were unable to prevent phagolysosomal fusion. The ability, or lack thereof, to recruit Rab14 to the KCV was reflected in Kp52145 and the isogenic mutants ability to activate AKT in macrophages. Klebsiella mutants of fructose bisphosphate aldolase, malate and 6-phosphogluconate dehydrogenases induced host inflammatory response upon infection above that of wild type Kp52145. The remaining mutants with intracellular survival deficits did not elicit immune responses beyond that seen with Kp52145 infection. Interestingly, alterations in the metabolic profile of Kp52145 did not affect the induction of IL-10 in macrophages. We determined macrophage metabolism upon Klebsiella infection using a pharmacologic approach. Inhibition of glycolysis negatively impacted Klebsiella intracellular survival, whereas fatty acid oxidation or oxidative phosphorylation inhibition did not.  We have also demonstrated that macrophage glycolysis is required by the host to illicit an immune response to infection. Mechanistically, we determined that the STAT6 and IL-10 are exploited to aid intracellular survival in macrophages.  In conclusion, we have uncovered the metabolic profile of intracellular K. pneumoniae and how the macrophage is rewired upon infection. 

Thesis embargoed until 31 July 2023

Date of Award	Jul 2022
Original language	English
Awarding Institution	Queen's University Belfast
Sponsors	Northern Ireland Department for the Economy
Supervisor	Jose Bengoechea (Supervisor) & Paul Moynagh (Supervisor)