Direct sputum analysis of Pseudomonas aeruginosa macrorestriction fragment genotypes in patients with cystic fibrosis

Breitenstein, Stefanie; Walter, S.; Bosshammer, J; Römling, Ute; Tümmler, Burkhard

doi:10.1007/s004300050050

Cited by 22 publications

(17 citation statements)

References 44 publications

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“…The presented results are consistent with those generated from previous investigations of P. aeruginosa in chronically infected CF patients [17,22,23]. These studies have all reported that, over time, most adult CF patients tend to harbour a single clone or subclone of P. aeruginosa during periods of clinical stability, and that the same bacterial clone retrieved during periods of clinical stability is also retrieved during pulmonary exacerbations [17].…”

Section: Discussionsupporting

confidence: 91%

Sputum versus bronchoscopy for diagnosis of Pseudomonas aeruginosa biofilms in cystic fibrosis

Aaron

Kottachchi²,

Ferris³

et al. 2004

European Respiratory Journal

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The present authors hypothesised that bronchoscopy with protected specimen brush may sample biofilm-forming bacteria adherent to the airway wall, whereas traditional sputum collection may not.Pseudomonas aeruginosa obtained from sputum, bronchoalveolar lavage and protected brush, taken from the right upper lung bronchus of 12 adult patients with cystic fibrosis, were compared. Retrieved bacteria were genotyped, and grown in planktonic cultures and as biofilms, and susceptibilities to individual antibiotics and to antibiotic combinations were determined.Bacterial cultures obtained using bronchoscopy did not yield any new strains of bacteria that were not also found in sputum. A total of 10 patients (83%) had a single strain of P. aeruginosa found using sputum, bronchoalveolar lavage and protected brush techniques, and two patients (17%) had two strains recovered in sputum, but only one strain was recovered using bronchoscopic techniques. Susceptibility to single antibiotics and to antibiotic combinations were not different between planktonically or biofilmgrown bacteria derived from sputum, as compared to those obtained by bronchoalveolar lavage and protected brush.In conclusion, sputum collection provides as much information as bronchoscopy for characterising the genotype and antibiotic susceptibility of chronic Pseudomonas aeruginosa infection in patients with stable cystic fibrosis. Eur Respir J 2004; 24: 631-637.

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Section: Discussionsupporting

confidence: 91%

Sputum versus bronchoscopy for diagnosis of Pseudomonas aeruginosa biofilms in cystic fibrosis

Aaron

Kottachchi²,

Ferris³

et al. 2004

European Respiratory Journal

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“…By sequencing several strains of the same clone, we now demonstrate that interclonal recombination contributes to intraclonal variation and the generation of mosaic genes. The colonized CF lung is characterized by high numbers of P. aeruginosa and only a few other bacterial competitors (Breitenstein et al, 1997). Such a scenario may facilitate the exchange of homologous genes between two or more P. aeruginosa clones.…”

Section: Discussionmentioning

confidence: 99%

Sequence diversity of the mucABD locus in Pseudomonas aeruginosa isolates from patients with cystic fibrosis

et al. 2006

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The mucA gene of the muc operon, which is instrumental in the control of the biosynthesis of the exopolysaccharide alginate, is a hotspot of mutation in Pseudomonas aeruginosa, a micro-organism that chronically colonizes the airways of individuals with cystic fibrosis (CF). The mucA, mucB and mucD genes were sequenced in nine environmental isolates from aquatic habitats, and in 37 P. aeruginosa strains isolated from 10 patients with CF, at onset or at a late stage of chronic airway colonization, in order to elucidate whether there was any association between mutation and background genotype. The 61 identified single nucleotide polymorphisms (SNPs) segregated into 18 mucABD genotypes. Acquired and de novo stop mucA mutations were present in 14 isolates (38 %) of five mucABD genotypes. ΔG430 was the most frequent and recurrent mucA mutation detected in four genotypes. The classification of strains by mucABD genotype was generally concordant with that by genome-wide SpeI fragment pattern or multilocus SNP genotypes. The exceptions point to intragenic mosaicism and interclonal recombination as major forces for intraclonal evolution at the mucABD locus.

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“…Furthermore, a wide phenotypic variation has been observed in P. aeruginosa clinical isolates (Govan & Deretic, 1996;Häußler et al, 2003), despite the fact that chronically infected CF patients harbour only one or a few genotypes (Breitenstein et al, 1997) and that there seems to be no genetic or functional differences between environmental and clinical strains (Alonso et al, 1999). These observations suggest that in P. aeruginosa, phenotypic diversification would constitute a relevant survival strategy associated with pathogenesis.…”

Section: Discussionmentioning

confidence: 99%

Emergence of phenotypic variants upon mismatch repair disruption in Pseudomonas aeruginosa

et al. 2004

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MutS is part of the bacterial mismatch repair system that corrects point mutations and small insertions/deletions that fail to be proof-read by DNA polymerase activity. In this work it is shown that the disruption of the P. aeruginosa mutS gene generates the emergence of diverse colony morphologies in contrast with its parental wild-type strain that displayed monomorphic colonies. Interestingly, two of the mutS morphotypes emerged at a high frequency and in a reproducible way and were selected for subsequent characterization. One of them displayed a nearly wild-type morphology while the other notably showed, compared with the wild-type strain, increased production of pyocyanin and pyoverdin, lower excretion of LasB protease and novel motility characteristics, mainly related to swarming. Furthermore, it was reproducibly observed that, after prolonged incubation in liquid culture, the pigmented variant consistently emerged from the mutS wild-type-like variant displaying a reproducible event. It is also shown that these P. aeruginosa mutS morphotypes not only displayed an increase in the frequency of antibiotic-resistant mutants, as described for clinical P. aeruginosa mutator isolates, but also generated mutants whose antibiotic-resistant levels were higher than those measured from spontaneous resistant mutants derived from wild-type cells. It was also found that both morphotypes showed a decreased cytotoxic capacity compared to the wild-type strain, leading to the emergence of invasive variants. By using mutated versions of a tetracycline resistance gene, the mutS mutant showed a 70-fold increase in the reversion frequency of a +1 frameshift mutation with respect to its parental wild-type strain, allowing the suggestion that the phenotypical diversity generated in the mutS population could be produced in part by frameshift mutations. Finally, since morphotypical diversification has also been described in clinical isolates, the possibility that this mutS diversification was related to the high frequency hypermutability observed in P. aeruginosa CF isolates is discussed.

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Direct sputum analysis of Pseudomonas aeruginosa macrorestriction fragment genotypes in patients with cystic fibrosis

Cited by 22 publications

References 44 publications

Sputum versus bronchoscopy for diagnosis of Pseudomonas aeruginosa biofilms in cystic fibrosis

Sputum versus bronchoscopy for diagnosis of Pseudomonas aeruginosa biofilms in cystic fibrosis

Sequence diversity of the mucABD locus in Pseudomonas aeruginosa isolates from patients with cystic fibrosis

Emergence of phenotypic variants upon mismatch repair disruption in Pseudomonas aeruginosa

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