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Population structure of Pseudomonas aeruginosa from five Mediterranean countries: evidence for frequent recombination and epidemic occurrence of CC235.

ABSTRACT: Several studies in recent years have provided evidence that Pseudomonas aeruginosa has a non-clonal population structure punctuated by highly successful epidemic clones or clonal complexes. The role of recombination in the diversification of P. aeruginosa clones has been suggested, but not yet demonstrated using multi-locus sequence typing (MLST). Isolates of P. aeruginosa from five Mediterranean countries (n = 141) were subjected to pulsed-field gel electrophoresis (PFGE), serotyping and PCR targeting the virulence genes exoS and exoU. The occurrence of multi-resistance (≥ 3 antipseudomonal drugs) was analyzed with disk diffusion according to EUCAST. MLST was performed on a subset of strains (n = 110) most of them had a distinct PFGE variant. MLST data were analyzed with Bionumerics 6.0, using minimal spanning tree (MST) as well as eBURST. Measurement of clonality was assessed by the standardized index of association (I(A) (S)). Evidence of recombination was estimated by ClonalFrame as well as SplitsTree4.0. The MST analysis connected 70 sequence types, among which ST235 was by far the most common. ST235 was very frequently associated with the O11 serotype, and frequently displayed multi-resistance and the virulence genotype exoS⁻/exoU⁺. ClonalFrame linked several groups previously identified by eBURST and MST, and provided insight to the evolutionary events occurring in the population; the recombination/mutation ratio was found to be 8.4. A Neighbor-Net analysis based on the concatenated sequences revealed a complex network, providing evidence of frequent recombination. The index of association when all the strains were considered indicated a freely recombining population. P. aeruginosa isolates from the Mediterranean countries display an epidemic population structure, particularly dominated by ST235-O11, which has earlier also been coupled to the spread of ß-lactamases in many countries.

SUBMITTER: Maatallah M

PROVIDER: S-EPMC3184967 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:From May 1997 to December 2001, a serotype O:6 multidrug-resistant strain of Pseudomonas aeruginosa colonized or infected 201 patients in the University Hospital of Besançon (France). The susceptibility profile of this epidemic clone to fluoroquinolones and aminoglycosides was relatively stable during the outbreak but showed important isolate-to-isolate variations (up to 64-fold) in the MICs of beta-lactams. Analysis of 18 genotypically related isolates selected on a quarterly basis demonstrated alterations in the two DNA topoisomerases II and IV (Thr83-->Ile in GyrA and Ser87-->Leu in ParC) and production of an ANT(2")-I enzyme. Although constitutively overproduced in these bacteria, the MexXY efflux system did not appear to contribute significantly to aminoglycoside resistance. beta-Lactam resistance was associated with derepression of intrinsic AmpC beta-lactamase (with isolate-to-isolate variations of up to 58-fold) and sporadic deficiency in a 46-kDa protein identified as the carbapenem-selective porin OprD. Of the 18 isolates, 14 were also found to overproduce the efflux system MexAB-OprM as a result of alteration of the repressor protein MexR (His107-->Pro). However, complementation experiments with the cloned mexR gene demonstrated that MexAB-OprM contributed only marginally to beta-lactam and fluoroquinolone resistance. Of the four isolates exhibiting wild-type MexAB-OprM expression despite the MexR alteration, two appeared to harbor secondary mutations in the mexA-mexR intergenic region and one harbored secondary mutations in the putative ribosome binding site located upstream of the mexAB oprM operon. In conclusion, this study shows that many mechanisms were involved in the multiresistance phenotype of this highly epidemic strain of P. aeruginosa. Our results also demonstrate that the clone sporadically underwent substantial genetic and phenotypic variations during the course of the outbreak, perhaps in relation to local or individual selective drug pressures.

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Population structure of Pseudomonas aeruginosa from five Mediterranean countries: evidence for frequent recombination and epidemic occurrence of CC235.

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