Abstract
BackgroundBurkholderia cenocepacia, an opportunistic pathogen that causes lung infections in cystic fibrosis (CF) patients, is associated with rapid and usually fatal lung deterioration due to necrotizing pneumonia and sepsis, a condition known as cepacia syndrome. The key bacterial determinants associated with this poor clinical outcome in CF patients are not clear. In this study, the cytotoxicity and procoagulant activity of B. cenocepacia from the ET-12 lineage, that has been linked to the cepacia syndrome, and four clinical isolates recovered from CF patients with mild clinical courses were analysed in both in vitro and in vivo assays.MethodsB. cenocepacia-infected BEAS-2B epithelial respiratory cells were used to investigate the bacterial cytotoxicity assessed by the flow cytometric detection of cell staining with propidium iodide. Bacteria-induced procoagulant activity in cell cultures was assessed by a colorimetric assay and by the flow cytometric detection of tissue factor (TF)-bearing microparticles in cell culture supernatants. Bronchoalveolar lavage fluids (BALF) from intratracheally infected mice were assessed for bacterial proinflammatory and procoagulant activities as well as for bacterial cytotoxicity, by the detection of released lactate dehydrogenase.ResultsET-12 was significantly more cytotoxic to cell cultures but clinical isolates Cl-2, Cl-3 and Cl-4 exhibited also a cytotoxic profile. ET-12 and CI-2 were similarly able to generate a TF-dependent procoagulant environment in cell culture supernatant and to enhance the release of TF-bearing microparticles from infected cells. In the in vivo assay, all bacterial isolates disseminated from the mice lungs, but Cl-2 and Cl-4 exhibited the highest rates of recovery from mice livers. Interestingly, Cl-2 and Cl-4, together with ET-12, exhibited the highest cytotoxicity. All bacteria were similarly capable of generating a procoagulant and inflammatory environment in animal lungs.ConclusionB. cenocepacia were shown to exhibit cytotoxic and procoagulant activities potentially implicated in bacterial dissemination into the circulation and acute pulmonary decline detected in susceptible CF patients. Improved understanding of the mechanisms accounting for B. cenocepacia-induced clinical decline has the potential to indicate novel therapeutic strategies to be included in the care B. cenocepacia-infected patients.
Highlights
Burkholderia cenocepacia, an opportunistic pathogen that causes lung infections in cystic fibrosis (CF) patients, is associated with rapid and usually fatal lung deterioration due to necrotizing pneumonia and sepsis, a condition known as cepacia syndrome
B. cenocepacia did not modify the expression of tissue factor (TF) by infected cells but enhanced the release of TF into cell supernatants No significant difference between control and infected cultures in their percentage of TF-expressing cells could be detected (Fig. 2A), as well as their expression of TF mRNA
B. cenocepacia enhanced the release of TF-bearing microparticles Fig. 3A shows that the number of microparticles binding annexin V, a protein known for its interaction with negatively charged phosphatidylserine residues, was significantly higher in supernatants from electrophoretic type 12 (ET-12)- and Cl-2infected cells than in supernatants from control cultures
Summary
Burkholderia cenocepacia, an opportunistic pathogen that causes lung infections in cystic fibrosis (CF) patients, is associated with rapid and usually fatal lung deterioration due to necrotizing pneumonia and sepsis, a condition known as cepacia syndrome. Pulmonary colonization/ infection by these bacteria may persist for months or even years but a minority of patients exhibits a rapid clinical deterioration associated with severe respiratory inflammation, epithelial necrosis and invasive disease, a condition known as cepacia syndrome [1]. B. cenocepacia ability to induce a marked release of proinflammatory mediators [2,3,4] is likely to contribute to lung damage and respiratory failure but whether bacterial isolates recovered from patients with poor clinical prognosis exhibit differential virulence profile has been so far poorly investigated. Because the lungs of CF patients is characterized by a florid inflammatory response, we wonder whether alveolar clotting processes may be involved in the pathogenesis of pulmonary decline observed in a proportion of B. cenocepacia-infected CF patients
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