ICEs Are the Main Reservoirs of the Ciprofloxacin-Modifying crpP Gene in Pseudomonas aeruginosa

Abstract

The ciprofloxacin-modifying crpP gene was recently identified in a plasmid isolated from a Pseudomonas aeruginosa clinical isolate. Homologues of this gene were also identified in Escherichia coli, Klebsiella pneumoniae and Acinetobacter baumannii. We set out to explore the mobile elements involved in the acquisition and spread of this gene in publicly available and complete genomes of Pseudomonas spp. All Pseudomonas complete genomes were downloaded from NCBI's Refseq library and were inspected for the presence of the crpP gene. The mobile elements carrying this gene were further characterized. The crpP gene was identified only in P. aeruginosa, in more than half of the complete chromosomes (61.9%, n = 133/215) belonging to 52 sequence types, of which the high-risk clone ST111 was the most frequent. We identified 136 crpP-harboring integrative and conjugative elements (ICEs), with 93.4% belonging to the mating-pair formation G (MPF_G) family. The ICEs were integrated at the end of a tRNA^Lys gene and were all flanked by highly conserved 45-bp direct repeats. The crpP-carrying ICEs contain 26 core genes (2.2% of all 1193 genes found in all the ICEs together), which are present in 99% or more of the crpP-harboring ICEs. The most frequently encoded traits on these ICEs include replication, transcription, intracellular trafficking and cell motility. Our work suggests that ICEs are the main vectors promoting the dissemination of the ciprofloxacin-modifying crpP gene in P. aeruginosa.

Keywords: Pseudomonas aeruginosa; ciprofloxacin resistance; crpP; integrative and conjugative elements; pangenome.

Figure 1

Approximately-maximum-likelihood phylogenetic tree base on the core genome alignment of 215 complete P. aeruginosa genomes. Branches and labels from crpP-positive hits are colored blue and the branches have twice the standard width. Branch lengths are ignored. Bootstrap values between 0.9 and 1 are represented by orange circles.

Figure 2

Distribution of the GC content of the chromosomes and integrative and conjugative elements (ICEs).

Figure 3

Pie chart of the breakdown of genes and the number of P. aeruginosa genomes in which they are present. Core genes are present in 99% or more of the genomes, soft core genes in 95% or more of the genomes and less than 99%, shell genes in between 15% and 95% and cloud genes in less than 15% of the genomes. This figure was created using the contributed Python script roary_plots.py in https://github.com/sanger-pathogens/Roary/blob/master/contrib/roary_plots/roary_plots.py.

Figure 4

Approximately-maximum-likelihood phylogenetic tree base on the alignment of the 26 core genes identified in the crpP-harboring ICEs. Genome accession numbers containing more than one crpP-harboring ICE are highlighted in green. Branch lengths are ignored. Bootstrap values between 0.9 and 1 are represented by orange circles.

Figure 5

Functional annotation of proteins encoded by the crpP-harboring ICEs.