Research Article: Dynamic in vivo mutations within the ica operon during persistence of Staphylococcus aureus in the airways of cystic fibrosis patients

Date Published: November 30, 2016

Publisher: Public Library of Science

Author(s): Bianca Schwartbeck, Johannes Birtel, Janina Treffon, Lars Langhanki, Alexander Mellmann, Devika Kale, Janina Kahl, Nina Hirschhausen, Claudia Neumann, Jean C. Lee, Friedrich Götz, Holger Rohde, Hanae Henke, Peter Küster, Georg Peters, Barbara C. Kahl, Alice Prince.

http://doi.org/10.1371/journal.ppat.1006024

Abstract

Cystic fibrosis (CF) is associated with chronic bacterial airway infections leading to lung insufficiency and decreased life expectancy. Staphylococcus aureus is one of the most prevalent pathogens isolated from the airways of CF patients. Mucoid colony morphology has been described for Pseudomonas aeruginosa, the most common pathogen in CF, but not for S. aureus. From the airways of 8 of 313 CF patients (2.5%) mucoid S. aureus isolates (n = 115) were cultured with a mean persistence of 29 months (range 1 month, 126 months). In contrast to non-mucoid S. aureus, mucoid isolates were strong biofilm formers. The upstream region of the ica operon, which encodes the proteins responsible for the synthesis of the polysaccharide intercellular adhesin (PIA), of mucoid isolates was sequenced. Spa-types of mucoid and non-mucoid strains were identical, but differed between patients. Mucoid isolates carried a 5 bp deletion in the intergenic region between icaR and icaA. During long-term persistence, from two patients subsequent non-mucoid isolates (n = 12) with 5 bp deletions were cultured, which did not produce biofilm. Sequencing of the entire ica operon identified compensatory mutations in various ica-genes including icaA (n = 7), icaD (n = 3) and icaC (n = 2). Six sequential isolates of each of these two patients with non-mucoid and mucoid phenotypes were subjected to whole genome sequencing revealing a very close relationship of the individual patient’s isolates. Transformation of strains with vectors expressing the respective wild-type genes restored mucoidy. In contrast to the non-mucoid phenotype, mucoid strains were protected against neutrophilic killing and survived better under starvation conditions. In conclusion, the special conditions present in CF airways seem to facilitate ongoing mutations in the ica operon during S. aureus persistence.

Partial Text

Cystic fibrosis (CF) is one of the most common hereditary diseases in the Caucasian population caused by mutations of an important chloride channel (cystic fibrosis transmembrane regulator) and affects worldwide approximately 70,000 people [1]. The mutation leads to impaired mucociliary clearance by airway epithelial cells with ensuing recurrent suppurative bacterial infections [2]. Staphylococcus aureus is one of the first and today the most frequent isolated pathogen, which can be recovered from the airways of CF patients with increasing prevalence rates most likely due to early eradication strategies directed against Pseudomonas aeruginosa, which was the leading pathogen in CF for decades and which has been shown to be responsible for lung function decline [1,3]. Mucoid isolates of P. aeruginosa occur in late stages of CF after the patients experienced long-term persistence of non-mucoid P. aeruginosa phenotypes [4]. The recovery of mucoid isolates has been shown to play a greater role in lung disease progression than the recovery of non-mucoid P. aeruginosa isolates [4]. The underlying mechanism for mucoidy is caused by overproduction of alginate due to a mutation in the mucA gene [5,6].

Although S. aureus chronically colonizes and infects the airways of CF patients, there are no previous reports of mucoid phenotypes of this pathogen recovered from the airways of these patients. Therefore, after occasional culture of mucoid S. aureus phenotypes from the specimens of some CF patients, we determined the prevalence of mucoid S. aureus isolates in S. aureus-positive patients from two independent prospective longitudinal studies, the persistence of mucoid isolates in the airways of these CF patients and the underlying molecular mechanism for mucoidy.

 

Source:

http://doi.org/10.1371/journal.ppat.1006024