Dataset Information

Use of subtractive hybridization to identify a diagnostic probe for a cystic fibrosis epidemic strain of Pseudomonas aeruginosa.

ABSTRACT: A multiresistant strain of Pseudomonas aeruginosa is widespread among cystic fibrosis (CF) patients attending clinics in Liverpool, United Kingdom. Suppression subtractive hybridization was used to identify sequences present in the Liverpool CF epidemic strain but absent from strain PAO1. Using dot blot and PCR amplification assays, the prevalence of such sequences among a panel of CF isolates was determined. Several sequences were found only in the Liverpool epidemic strain. Some sequences were present in the Liverpool epidemic strain and in a minority of other isolates, including sequences with homology to genes implicated in O6 serotype and siderophore production. The Liverpool epidemic strain and 81% of nonepidemic isolates contained a sequence identified as part of the PAGI-1 genomic island. Other strains implicated in epidemic spread, which were from Manchester, United Kingdom, and Melbourne, Australia, were also screened. None of the sequences identified was present in the Manchester strain. However, one of two Melbourne strains contained some of the sequences found in the Liverpool epidemic strain. All isolates implicated in epidemic spread and 76% of sporadic isolates contained the exoS gene. A sequence present in all isolates of the Liverpool epidemic strain was used to develop a diagnostic PCR test for identification of the strain from colonies or directly from sputum samples.

SUBMITTER: Parsons YN

PROVIDER: S-EPMC154653 | biostudies-literature | 2002 Dec

REPOSITORIES: biostudies-literature

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Use of subtractive hybridization to identify a diagnostic probe for a cystic fibrosis epidemic strain of Pseudomonas aeruginosa.

Parsons Yasmin N YN Panagea Stavroula S Smart Catherine H M CH Walshaw Martin J MJ Hart C Anthony CA Winstanley Craig C

Journal of clinical microbiology 20021201 12

A multiresistant strain of Pseudomonas aeruginosa is widespread among cystic fibrosis (CF) patients attending clinics in Liverpool, United Kingdom. Suppression subtractive hybridization was used to identify sequences present in the Liverpool CF epidemic strain but absent from strain PAO1. Using dot blot and PCR amplification assays, the prevalence of such sequences among a panel of CF isolates was determined. Several sequences were found only in the Liverpool epidemic strain. Some sequences were ...[more]

PMID: 12454160

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Project description:Epidemic strains of Pseudomonas aeruginosa have been found worldwide among the cystic fibrosis (CF) patient population. Using pulse-field gel electrophoresis, the Prairie Epidemic Strain (PES) has recently been found in one-third of patients attending the Calgary Adult CF Clinic in Canada. Using multi-locus sequence typing, PES isolates from unrelated patients were found to consistently have ST192. Though most patients acquired PES prior to enrolling in the clinic, some patients were observed to experience strain replacement upon transitioning to the clinic whereby local non-epidemic P. aeruginosa isolates were displaced by PES. Here we genotypically and phenotypically compared PES to other P. aeruginosa epidemic strains (OES) found around the world as well as local non-epidemic CF P. aeruginosa isolates in order to characterize PES. Since some epidemic strains are associated with worse clinical outcomes, we assessed the pathogenic potential of PES to determine if these isolates are virulent, shared properties with OES, and if its phenotypic properties may offer a competitive advantage in displacing local non-epidemic isolates during strain replacement. As such, we conducted a comparative analysis using fourteen phenotypic traits, including virulence factor production, biofilm formation, planktonic growth, mucoidy, and antibiotic susceptibility to characterize PES, OES, and local non-epidemic isolates. We observed that PES and OES could be differentiated from local non-epidemic isolates based on biofilm growth with PES isolates being more mucoid. Pairwise comparisons indicated that PES produced significantly higher levels of proteases and formed better biofilms than OES but were more susceptible to antibiotic treatment. Amongst five patients experiencing strain replacement, we found that super-infecting PES produced lower levels of proteases and elastases but were more resistant to antibiotics compared to the displaced non-epidemic isolates. This comparative analysis is the first to be completed on a large scale between groups of epidemic and non-epidemic CF P. aeruginosa isolates.

Project description:Pseudomonas aeruginosa is an opportunistic pathogen which is evolving resistance to many currently used antibiotics. While much research has been devoted to the roles of pathogenic P. aeruginosa in cystic fibrosis (CF) patients, less is known of its ecological properties. P. aeruginosa dominates the lungs during chronic infection in CF patients, yet its abundance in some environments is less than that of other diverse groups of pseudomonads. Here, we sought to determine if clinical isolates of P. aeruginosa are vulnerable to environmental pseudomonads that dominate soil and water habitats in one-to-one competitions which may provide a source of inhibitory factors. We isolated a total of 330 pseudomonads from diverse habitats of soil and freshwater ecosystems and competed these strains against one another to determine their capacity for antagonistic activity. Over 900 individual inhibitory events were observed. Extending the analysis to P. aeruginosa isolates revealed that clinical isolates, including ones with increased alginate production, were susceptible to competition by multiple environmental strains. We performed transposon mutagenesis on one isolate and identified an ∼14.8-kb locus involved in antagonistic activity. Only two other environmental isolates were observed to carry the locus, suggesting the presence of additional unique compounds or interactions among other isolates involved in outcompeting P. aeruginosa This collection of strains represents a source of compounds that are active against multiple pathogenic strains. With the evolution of resistance of P. aeruginosa to currently used antibiotics, these environmental strains provide opportunities for novel compound discovery against drug-resistant clinical strains.ImportanceWe demonstrate that clinical CF-derived isolates of P. aeruginosa are susceptible to competition in the presence of environmental pseudomonads. We observed that many diverse environmental strains exhibited varied antagonistic profiles against a panel of clinical P. aeruginosa isolates, suggesting the presence of distinct mechanisms of inhibition among these ecological strains. Understanding the properties of these antagonistic events offers the potential for discoveries of antimicrobial compounds or metabolic pathways important to the development of novel treatments for P. aeruginosa infections.

Dataset Information

Use of subtractive hybridization to identify a diagnostic probe for a cystic fibrosis epidemic strain of Pseudomonas aeruginosa.

Publications

Use of subtractive hybridization to identify a diagnostic probe for a cystic fibrosis epidemic strain of Pseudomonas aeruginosa.

Similar Datasets

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