Metabolic regulation of denitrification in the opportunistic pathogen Pseudomonas aeruginosa

Pseudomonas aeruginosa is a World Health Organization Priority 1 human pathogen and is one of the leading bacterial species implicated in nosocomial infections, posing a significant risk to immunocompromised patients due to its high level of intrinsic antimicrobial resistance and diverse metabolic physiology. Denitrification allows P. aeruginosa to utilise N-oxides as alternative electron acceptors to enable maintenance of proton motive force and thus the generation of ATP under nutrient- and oxygen-limited scenarios, and may also act as a defence mechanism against nitrosative attack by host immune cells. Traditionally, denitrification is thought of as an anaerobic process. However, recent work in the Welch laboratory has shown that when grown aerobically with acetate as a sole carbon source, genes involved in the denitrification pathway are highly up-regulated, despite there being no change in the transcript level of known regulators of denitrification. This is intriguing since fatty acids, such as acetate, are a preferred energy source for P. aeruginosa during infection, and furthermore, the known regulators of denitrification are inactivated in the presence of oxygen. In this work, I investigate the regulation of denitrification under aerobic conditions. The data revealed that the oft-reported master regulator of (anaerobic) denitrification, Anr, is of limited utility under aerobic conditions. I then examined how a loss of denitrification capabilities affects the redox state of the cell. I hypothesised that under conditions that promote rapid growth, I would observe an increase in NADH accumulation, and that this might be decreased through the use of aerobic denitrification. I found that this redox-balancing activity was independent of Anr, but was dependent on its subordinate, Dnr, and the NarG nitrate reductase. I also investigated the impact of denitrification on P. aeruginosa survival in a polymicrobial community using an in vitro continuous-flow system developed in the Welch laboratory with subsequent analysis using a Lotka-Volterra competition model. This analysis revealed that denitrification mutants are considerably less “tolerant” towards competing species and begin to rapidly dominate the culture medium, highlighting a potentially significant role for denitrification in maintaining stable population dynamics during an infection scenario.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2022

Author dc:creator

• Trigg, Stephen

Advisor dc:contributor.advisor

• Welch, Martin

• Pseudomonas aeruginosa
• denitrification
• metabolism
• microbiology
• bacteria

Licence

• https://www.rioxx.net/licenses/all-rights-reserved/

Language dc:language

eng

Author Identifier

0000-0002-8031-1516

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/346800