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Rescue of pyrimidine-defective Pseudomonas aeruginosa through metabolic complementation.

ABSTRACT: Chronic infections harbor multiple pathogens where dynamic interactions between members of the polymicrobial community play a major role in determining the infection outcome. For example, in a nutrient-rich polymicrobial infection, bacteria have the potential to undergo evolutionary changes that impair their ability to synthesize essential metabolites. This adaptation may facilitate metabolic interdependencies between neighboring pathogens and lead to difficult-to-treat chronic infections. Our research group previously demonstrated that Pseudomonas aeruginosa (PA) and Staphylococcus aureus (SA), typically considered classical competitors, can adopt a cooperative lifestyle through bi-directional purine exchange medicated by exogenous DNA (eDNA) release. To further validate our initial findings, in this study, we investigated the potential exchange of pyrimidine between PA and other pathogens, which is another constituent of DNA. In our findings, we observed that a pyrimidine-deficient transposon mutant strain of PA showed improved growth when co-cultured with wild-type PA, SA, Acinetobacter baumannii (AB), and Enterococcus faecalis (EF). Additionally, improved fitness of pyrimidine-deficient PA was further observed in chemical complementation with eDNA and uridine-5'-monophosphate. Interestingly, the rescue of PA growth through eDNA complementation is not as effective as in intact cells, such as SA, AB, EF, and wild-type PA, implying that eDNA is a lesser contributor to this metabolic complementation. Also, the exchange mechanism between pathogens involves more active mechanisms beyond simple eDNA or metabolite release. Our data further highlights the importance of cell-to-cell contact for effective and increased metabolic complementation.

Importance

This research holds crucial implications for combating chronic infections, where multiple pathogens coexist and interact within the same environment. By uncovering the dynamic exchange of pyrimidines between Pseudomonas aeruginosa (PA) and Staphylococcus aureus (SA), our study reveals a previously unrecognized aspect of interspecies cooperation. The observed enhanced growth of a pyrimidine-deficient PA strain when co-cultured with SA suggests potential avenues for understanding and disrupting bacterial metabolic interdependencies in chronic infection settings. Furthermore, our findings highlight the mechanisms involved in metabolic exchange, emphasizing the importance of cell-to-cell contact. This research explored essential metabolic interactions to address the challenges posed by difficult-to-treat chronic infections.

SUBMITTER: Mahmud HA

PROVIDER: S-EPMC11302043 | biostudies-literature | 2024 Jul

REPOSITORIES: biostudies-literature

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Rescue of pyrimidine-defective <i>Pseudomonas aeruginosa</i> through metabolic complementation.

Mahmud Hafij Al HA Garcia Randy R Garcia Alexsis A Baishya Jiwasmika J Wakeman Catherine A CA

Microbiology spectrum 20240711

Chronic infections harbor multiple pathogens where dynamic interactions between members of the polymicrobial community play a major role in determining the infection outcome. For example, in a nutrient-rich polymicrobial infection, bacteria have the potential to undergo evolutionary changes that impair their ability to synthesize essential metabolites. This adaptation may facilitate metabolic interdependencies between neighboring pathogens and lead to difficult-to-treat chronic infections. Our r ...[more]

PMID: 38990029

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Project description:Mucoidy due to alginate overproduction by the Gram-negative bacterium Pseudomonas aeruginosa facilitates chronic lung infections in patients with cystic fibrosis (CF). We previously reported that disruption in de novo synthesis of pyrimidines resulted in conversion to a nonmucoid small-colony variant (SCV) in the mucoid P. aeruginosa strain (PAO581), which has a truncated anti-sigma factor, MucA25, that cannot sequester sigma factor AlgU (AlgT). Here, we showed that supplementation with the nitrogenous bases uracil or cytosine in growth medium complemented the SCV to normal growth, and nonmucoidy to mucoidy, in these mucA25 mutants. This conversion was associated with an increase in intracellular levels of UMP and UTP suggesting that nucleotide restoration occurred via a salvage pathway. In addition, supplemented pyrimidines caused an increase in activity of the alginate biosynthesis promoter (P algD ), but had no effect on P algU , which controls transcription of algU Cytosolic levels of AlgU were not influenced by uracil supplementation, yet levels of RpoN, a sigma factor that regulates nitrogen metabolism, increased with disruption of pyrimidine synthesis and decreased after supplementation of uracil. This suggested that an elevated level of RpoN in SCV may block alginate biosynthesis. To support this, we observed that overexpressing rpoN resulted in a phenotypic switch to nonmucoidy in PAO581 and in mucoid clinical isolates. Furthermore, transcription of an RpoN-regulated promoter increased in the mutants and decreased after uracil supplementation. These results suggest that the balance of RpoN and AlgU levels may regulate growth from SCV to mucoidy through sigma factor competition for P algDIMPORTANCE Chronic lung infections with P. aeruginosa are the main cause of morbidity and mortality in patients with cystic fibrosis. This bacterium overproduces a capsular polysaccharide called alginate (also known as mucoidy), which aids in bacterial persistence in the lungs and in resistance to therapeutic regimens and host immune responses. The current study explores a previously unknown link between pyrimidine biosynthesis and mucoidy at the level of transcriptional regulation. Identifying/characterizing this link could provide novel targets for the control of bacterial growth and mucoidy. Inhibiting mucoidy may improve antimicrobial efficacy and facilitate host defenses to clear the noncapsulated P. aeruginosa bacteria, leading to improved prognosis for patients with cystic fibrosis.