Pseudomonas aeruginosa Alginate Benefits Staphylococcus aureus?

In this issue of Journal of Bacteriology, Price et al. show that the Pseudomonas aeruginosa-produced exopolysaccharide alginate protects Staphylococcus aureus by dampening the expression of P. aeruginosa virulence products that usually inhibit S. aureus respiration and cell membrane integrity when the two organisms compete in other environments (C. E. Price, D. G. Brown, D. H. Limoli, V. V. Phelan, and G.

ABSTRACT

In this issue of Journal of Bacteriology, Price et al. show that the Pseudomonas aeruginosa-produced exopolysaccharide alginate protects Staphylococcus aureus by dampening the expression of P. aeruginosa virulence products that usually inhibit S. aureus respiration and cell membrane integrity when the two organisms compete in other environments (C. E. Price, D. G. Brown, D. H. Limoli, V. V. Phelan, and G. A. O’Toole, J Bacteriol 202:e00559-19, 2020, https://doi.org/10.1128/jb.00559-19). This is the first report that exogenously added alginate affects P. aeruginosa competition and provides a partial explanation for S. aureus and P. aeruginosa coinfections in cystic fibrosis.

TEXT

In this issue of the Journal of Bacteriology, Price et al. show that the Pseudomonas aeruginosa-produced exopolysaccharide alginate protects Staphylococcus aureus by dampening the expression of P. aeruginosa virulence genes whose products usually inhibit S. aureus when the two organisms compete in other environments. The authors show that exogenous alginate, regardless of the source, protects S. aureus from P. aeruginosa in both planktonic and biofilm coculture models. Furthermore, the authors demonstrate that coculture of mucoid P. aeruginosa with nonmucoid P. aeruginosa strains can mitigate S. aureus killing by the nonmucoid P. aeruginosa strain. Interestingly, the authors also show that some clinical mucoid P. aeruginosa isolates retain their ability to kill S. aureus, indicating that there are strain-specific variations for these observations. The authors thus provide at least a partial explanation for the ∼30% of cystic fibrosis (CF) patients who are coinfected with both organisms and demonstrate that alginate protects not only P. aeruginosa but also S. aureus. Their results also indicate that exogenous alginate affects nonmucoid and mucoid P. aeruginosa gene expression (1).

The lungs of CF children are readily colonized by S. aureus during the early years of life, with colonization by P. aeruginosa during the mid- to late teenage years. S. aureus colonization is associated with a higher probability of secondary P. aeruginosa infection. P. aeruginosa colonization will eventually outcompete S. aureus and other microbes present in the CF lung to become the predominant pathogen (2, 3). Coinfection with P. aeruginosa and S. aureus ultimately results in a poor clinical outcome for the patient, including a reduced median forced expiratory volume in 1 s (FEV1) and increased rates of pulmonary exacerbation (4). During the process of coinfection and treatment, P. aeruginosa will eventually be pushed into a mucoid phenotype through the acquisition of mucA (algN) mutations (5, 6). Several studies have documented that the mucA mutation exists within several mucoid clinical CF isolates (7, 8). The result of this mutation is the release of an extracytoplasmic sigma factor (ECF) called AlgU (9), AlgT (10), or σ²² (11) and is the Escherichia coli RpoE orthologue (12). AlgU/T is required for the transcription of the P. aeruginosa alginate biosynthetic pathway (9, 13–15). Alginate is a secreted anionic exopolysaccharide composed of various proportions of 1,4-linked β-d-mannuronic acid (M) and its C-5 epimer α-l-guluronic acid (G), which is naturally produced by some bacteria and brown seaweeds (16). Significant effort to understand alginate biosynthesis has been due to its role in bacterial pathogenesis as well as the possibility to produce tailor-made alginates exhibiting material properties suitable for various medical and industrial applications. In the context of pathogenesis, alginate provides mucoid P. aeruginosa strains several advantages over nonmucoid strains (17, 18). These advantages include resistance to phagocytosis (19), resistance to killing by polymorphonuclear cells (20), scavenging of free radicals released by macrophages (21), and resistance to antibiotics by reducing diffusion to the organism (22). Additionally, mucoid strains grow slowly, are generally nonmotile, and express low levels of protease exotoxins and siderophores (23–28). Ultimately, P. aeruginosa alginate production creates a survival advantage as it mediates the formation of persistent biofilms during chronic infections (29, 30). Until very recently, the only documented advantages that alginate provided were to P. aeruginosa.

A few studies have described the in vitro and in vivo interactions between S. aureus and P. aeruginosa using CF lung infection models and sputum samples (31–33). These studies led to the conclusion that the microorganisms are antagonistic in vitro (34–36); however, in vivo models showed contradictory results (35, 37) despite clinical evidence that there may be synergy between them. For the in vitro work, Filkins et al. utilized a P. aeruginosa-S. aureus coinfection model on a human CF bronchial epithelial (CFBE) cell line to show that P. aeruginosa drove the S. aureus expression profile from that of aerobic respiration to that of fermentation. The fermentative respiration was dependent on P. aeruginosa production of both 2-heptyl-4-hydroxyquinoline N-oxide (HQNO) and siderophores. The authors observed that initially, S. aureus and P. aeruginosa coexisted; however, extended coculture reduced S. aureus viability. Additionally, the authors showed that S. aureus small-colony-variant (SCV) genetic mutant strains, which have defects in their electron transport chain, experience reduced killing by P. aeruginosa compared to their wild-type parent strains, indicating that P. aeruginosa HQNO and siderophores act specifically to inhibit the S. aureus aerobic electron transport chain (32). Nguyen and Oglesby-Sherrouse extended this further by observing that the iron-regulated antimicrobial activity of P. aeruginosa against S. aureus was due to the cumulative effects of multiple alkyl quinolone (AQ) metabolites, both the production and activity of which are modulated by environmental iron levels (28). Yang et al. utilized coculture biofilms of S. aureus with P. aeruginosa mutants in a flow chamber system and showed that wild-type P. aeruginosa PAO1 facilitates S. aureus microcolony formation (27). In contrast, P. aeruginosa mucA (presumably mucoid) and rpoN mutants did not facilitate S. aureus microcolony formation and tended to outcompete S. aureus in coculture biofilms. Interestingly, the data from Yang et al. are in agreement with those from Filkins et al. when a mucoid P. aeruginosa strain, FRD1, was used on a CFBE cell coculture, and both outcompeted S. aureus. Some additional understanding of mucoid P. aeruginosa interactions with S. aureus was provided by Baldan et al., who showed that early CF P. aeruginosa isolates were competitive with S. aureus, whereas late CF isolates were not as competitive with S. aureus. The authors surmised that the secondary “pathoadaptive” mutations acquired by P. aeruginosa made the organism less competitive (31). Interestingly, the authors did not mention that S. aureus found in the same lung environment and exposed to the same innate and adaptive immune responses and antibiotics likely underwent pathoadaptive mutations as well. For instance, were S. aureus SCVs tested? Alginate production is the best-studied pathoadaptive virulence factor made by P. aeruginosa and has been shown to be one result of the onslaught of antibiotics and oxidative stress that is endured by P. aeruginosa in the CF lung (18, 38).

Altogether, it appears that mucoid strains can both antagonize and synergize with S. aureus. Whether antagonism or synergism is most important remains an open question and will probably be determined by a multitude of factors, including the patient’s immune status, the efficacy of antibiotic treatment, or even other organisms that are found in the CF lung. The current work has brought to light that alginate helps S. aureus under certain conditions by dampening P. aeruginosa gene expression, with the surprise being that alginate can do this exogenously.

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