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Investigation of role of IS elements in conversion to hypermutability and emergence of drug resistance in Pseudomonas aeruginosa from CF patients in Cape Town, South Africa

Abstract number: P2024

Evans J.C., Motilal N., Segal H.

Objectives: Cystic fibrosis (CF) patients frequently succumb to infections with P. aeruginosa, in part due to its ability to adapt to the CF lung environment. Continual antibiotic therapy drives the emergence of multi-drug resistant (MDR) P. aeruginosa, further compromising the treatment of CF patients.

A plethora of insertion sequence (IS) elements have been described in P. aeruginosa. Some IS elements play a role in emergence of antibiotic resistance through insertional inactivation of genes. Additionally, IS6100 causes large chromosomal inversions (LCIs) that disrupt genes involved in CF lung pathogenesis. IS6100 is of particular concern as it is associated with conversion to hypermutability, resulting in increased rates of spontaneous mutations, often associated with MDR.

The aim of this study was to determine whether IS elements are associated with resistance, particularly to imipenem (IPM), and whether IS6100 plays a role in conversion to hypermutability in P. aeruginosa strains from CF patients in Cape Town.

Methods: CF and non-CF P. aeruginosa isolates (52) obtained from patients at two local hospitals during 2000–2006 were included in the study.

PCR amplification of oprD from all IPM-resistant strains (75) was carried out to detect insertions that could account for imipenem resistance.

Strains shown to contain IS6100 by PCR were included in Southern hybridisation experiments to determine IS6100 copy number.

Mutator assays were carried out and strains were considered hypermutable if their mutation rates were more than 20 times that of P. aeruginosa PAO1.

Results: PCR amplification of oprD in 74 isolates yielded a product of the expected size (1343bp), while the remaining isolate, strain 8, yielded a product of 2400bp. Sequence analysis revealed that oprD from strain 8 was interrupted by insertion of a novel IS element, designated ISPa26.

IS6100 was detected in 2 CF and 11 non-CF isolates. Interestingly, strain 6 harbours at least three copies of IS6100, which is associated with LCIs.

Of the CF P. aeruginosa isolates, 4 were hypermutable. However, isolates containing IS6100 had mutation rates comparable to P. aeruginosa PAO1.

Conclusion: Insertional inactivation of oprD by ISPa26 accounts for IPM resistance in strain 8.

As IS6100 was not associated with conversion to hypermutability, further investigation into the genetic environment of IS6100 is necessary as transmission of this element to CF isolates may negatively impact care and treatment of CF patients.

Session Details

Date: 19/04/2008
Time: 00:00-00:00
Session name: 18th European Congress of Clinical Microbiology and Infectious Diseases
Subject:
Location: Barcelona, Spain
Presentation type:
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