Evolution of Pseudomonas aeruginosa in a single cystic fibrosis patient.

PAUL H. ROY; MARÍA FLORENCIA RAPISARDI; MAXIME DERASPE; VERONICA ALVAREZ; NAEEM IQBAL; KEN DEWAR; JESSICA WASSERSCHEID; CECILIA QUIROGA; DANIELA CENTRÓN

Washington

Conferencia; American Society for Microbiology's Conference on Pseudomonas; 2015

American Society for Microbiology'

Cystic fibrosis (CF) is the most common autosomal recessive hereditary disease in the Caucasian population and the leading cause of chronic lung disease in childhood. P. aeruginosa is frequently found infecting these patients, and is associated with low life expectancy and high deterioration of lung function. It has been shown that stages towards P. aeruginosa chronicity include both adaptation of a lineage and coexistence of different lineages. The genetic mechanisms that are involved in the adaptation of a lineage include mutations in specific genes and genome reduction. All these changes are necessary to develop persistence of P. aeruginosa in the lung of CF patients. From a phenotypic perspective, this chronicity can be seen as a switch of colonies to mucoidy, progression from the planktonic to biofilm growth, loss of motility, auxotrophy, hypermutability, significant contribution of some virulence factors such as elastase, LPS, alginate rhamnolipids, and rise of antimicrobial resistance. From a genomic perspective, deletions originate through both illegitimate and homologous recombination, with reductions reaching 8% of the entire genome. In Argentina, the life expectancy is 20 years, half of that of other countries such as Denmark (45 years), USA (38 years) or Europe (35 years). In this study we examined whether presence of epidemic and virulent clones of P. aeruginosa could be responsible for this low life expectancy. From a previous epidemiological study from our laboratory, we isolated several strains from a single patient during the course of this disease (Pae969 (10/2005), Pae975 (10/2006), Pae977(11/2006) and Pae981 (05/2007) for Illumina sequencing and analysis. The patient died after the last isolate Pae981. The last of these, Pae981, was sequenced using PacBio SMRT technology. The four strains were in the PAO1-LESB58 clade. Surprisingly, Pae975 and Pae981 proved to be almost isogenic. Comparative analysis of genomes showed the presence of 66 plasticity regions with greatest variability in RGP23. There were about 30 SNP?s (possibly more due to differences in the Ray and Spades assemblies of Pae975), the most important of which was a mutation in mucB resulting in mucoidy in Pae981. This strain also underwent a genome reduction in the region corresponding to PA1944-PA1982. Notably, Pae975 and Pae981 harbor large novel plasmids (332 kb in Pae981). These plasmids were highly similar except for an inserted 90 kb region including a class 1 integron in Pae981. This integron contained a blaVIM-2 gene cassette that confers resistance to carbapenems. The absence of this integron in Pae975 shows that P. aeruginosa adapted to the host by acquiring antimicrobial resistance genes while losing other genomic traits.