Cystic Fibrosis Transmembrane Conductance Regulator and Pseudomonas

Cystic fibrosis (CF) lung disease is characterized by chronic airway infection and a heightened immune response leading to airway damage and bronchiectasis (1). Pseudomonas aeruginosa is the species of bacteria most commonly associated with CF, with almost 80% of adults chronically infected (2). Chronic P. aeruginosa infection is associated with more rapid decline in lung function and decreased survival (3, 4). Inhaled antipseudomonal antibiotics are effective in eradicating early P. aeruginosa infection and appear to delay the onset of chronic P. aeruginosa infection (5). Current treatment guidelines focus on early detection and attempted eradication of P. aeruginosa prior to establishment of chronic infection (6). Once chronic infection develops, eradication is rarely achieved, and the goals of inhaled antibiotic therapy change to bacterial load suppression rather than eradication. Despite chronic infection, resistance to eradication by antibiotics, and high airway bacterial loads, invasive P. aeruginosa infection is rare in CF (7).

The primary defect in CF is absence or diminished production of the CF transmembrane conductance regulator (CFTR) protein due to mutations in the CFTR gene (1). CFTR protein functions as a cell membrane channel responsible for chloride and other ion transport across epithelial cell linings in the lungs, gastrointestinal tract, and pancreas. Lack of CFTR in the lung is thought to lead to a dehydrated airway surface liquid layer, mucus abnormalities, and altered immune response. Efforts to treat P. aeruginosa infection have been hampered by a lack of understanding of how defects in CFTR protein lead to chronic P. aeruginosa infection. Several mechanisms have been proposed. Arguably, the most accepted hypothesis is that dehydration of the airway surface liquid due to sodium hyperabsorption leads to impaired mucociliary clearance, bacterial stasis, and poor clearance of bacteria; the propensity for P. aeruginosa infection, however, is not well explained by this mechanism (8). Increased adherence of P. aeruginosa to CFTR-deficient airway epithelial cells has been proposed, but results of experimental studies are mixed, and most P. aeruginosa found in vivo is not attached to the epithelium (9). CFTR dysfunction has also been implicated in the exaggerated proinflammatory, neutrophil-dominated, immune response seen in patients with CF, but why this immune response is ineffective at eradication P. aeruginosa is incompletely understood (10). Biofilm formation by bacterial species such as P. aeruginosa has been proposed as a mechanism of chronic infection and poor susceptibility to antibiotics. However, P. aeruginosa isolates from patients with CF with chronic infection are variable in their ability to form biofilms. Recently aggregates of P. aeruginosa similar in characteristics to biofilms but nonadherent have been proposed as a mechanism of chronic infection in CF (11).

Clinical observational data clearly supports a link between CFTR dysfunction and chronic P. aeruginosa infection. Chronic P. aeruginosa is more common in those with absent CFTR function (two severe CFTR mutations) compared with those with residual CFTR function (one or two mild CFTR mutations). Even in early disease, risk of P. aeruginosa acquisition is increased in those with absent CFTR function (12). In addition, individuals with non-CF bronchiectasis develop chronic P. aeruginosa infection at lower rates compared with those with CF-related bronchiectasis (13). CFTR dysfunction thus appears to favor chronic P. aeruginosa infection that is difficult to eradicate. Understanding the link between CFTR and P. aeruginosa may lead to novel treatment approaches.

In this issue of the Journal, Staudinger and colleagues (pp. 812–824) address several key questions regarding the link between CFTR and chronic P. aeruginosa infection (14). They first investigated how airway conditions associated with CFTR dysfunction, increased mucus density, and altered biochemical properties of sputum might lead to the phenotypic eradication-resistant, antibiotic-tolerant, and noninvasive chronic P. aeruginosa infection in CF. They specifically focused on the growth of bacteria in aggregates rather than biofilms, and explored airway and bacterial features that would promote aggregate formation. Using models of low- and high-density agar gels, plus the addition of soluble components of CF sputum, they found that both high-density gels and soluble sputum components promoted P. aeruginosa growth in aggregates. Hypothesizing that neutrophil elastase (NE) within the sputum may drive aggregate formation, they performed a series of experiments to disrupt NE activity within sputum, restore activity using purified NE, and then disrupt NE again with elastase inhibitors. Aggregate formation occurred in the presence of NE but was prevented when NE activity was diminished. Using genetically immotile P. aeruginosa and motility studies of bacteria exposed to NE, they showed that decreased bacterial motility may be the common mechanism by which increased density mucus and NE lead to aggregate formation.

Staudinger and colleagues also showed that P. aeruginosa grown in aggregates had increased resistance to killing by NE and antibiotic tolerance compared with dispersed bacteria. Tobramycin appeared to provide a competitive advantage to nonmotile P. aeruginosa. These studies suggest that aggregate formation may increase antibiotic tolerance even in genetically susceptible bacteria and that chronic use of antibiotics may push infections toward this more tolerant phenotype. Importantly, genetic mutations preventing biofilm formation, found in many late CF P. aeruginosa isolates, did not prevent aggregates from developing. To explore whether aggregate formation could reduce invasiveness, they studied the effect of aggregates on epithelial cells and in an in vivo wound model and found attenuated immune response and decreased mortality in the presence of aggregates. Thus, conditions in the CF airway due to CFTR dysfunction and resultant treatment, including dense mucus, high levels of NE, and exposure to chronic antibiotics, appear to favor nonmotile P. aeruginosa, promoting aggregate formation. These aggregates may at least partially explain the eradication-resistant, antibiotic-tolerant, and noninvasive infection seen in CF.

Despite advances in early eradication approaches and chronic suppressive therapy with inhaled antibiotics, P. aeruginosa infection remains a significant problem in CF and contributes to much of the morbidity and mortality. The work by Staudinger and colleagues provides important insight into how the basic defect in CF, CFTR dysfunction, may foster an environment ideal for chronic P. aeruginosa infection. Yet, we are still left with the question of why the CF airway is susceptible to P. aeruginosa in particular. One limitation of the study is that it focuses exclusively on P. aeruginosa, whereas many other bacteria, including Staphylococcus aureus, other gram-negative bacteria, nontuberculous mycobacterium, and, more recently, obligate anaerobes, are known or suspected to contribute to CF lung disease (15, 16). Infection is frequently polymicrobial; thus, how polymicrobial infections impact aggregate formation is an area that needs further study. Another avenue of research is whether restoration of CFTR function can reverse the pressure on P. aeruginosa to form aggregates once chronic infection has developed. As demonstrated by Staudinger and colleagues, the mechanisms linking CFTR dysfunction and chronic P. aeruginosa infection are complex and multifaceted. Perhaps our best hope for true eradication of P. aeruginosa is through improved CFTR function, a possibility now with the development of CFTR modulators.