Intraclonal Genome Stability of the Metallo-β-lactamase SPM-1-producing Pseudomonas aeruginosa ST277, an Endemic Clone Disseminated in Brazilian Hospitals

Abstract

Carbapenems represent the mainstay therapy for the treatment of serious P. aeruginosa infections. However, the emergence of carbapenem resistance has jeopardized the clinical use of this important class of compounds. The production of SPM-1 metallo-β-lactamase has been the most common mechanism of carbapenem resistance identified in P. aeruginosa isolated from Brazilian medical centers. Interestingly, a single SPM-1-producing P. aeruginosa clone belonging to the ST277 has been widely spread within the Brazilian territory. In the current study, we performed a next-generation sequencing of six SPM-1-producing P. aeruginosa ST277 isolates. The core genome contains 5899 coding genes relative to the reference strain P. aeruginosa PAO1. A total of 26 genomic islands were detected in these isolates. We identified remarkable elements inside these genomic islands, such as copies of the blaSPM−1 gene conferring resistance to carbapenems and a type I-C CRISPR-Cas system, which is involved in protection of the chromosome against foreign DNA. In addition, we identified single nucleotide polymorphisms causing amino acid changes in antimicrobial resistance and virulence-related genes. Together, these factors could contribute to the marked resistance and persistence of the SPM-1-producing P. aeruginosa ST277 clone. A comparison of the SPM-1-producing P. aeruginosa ST277 genomes showed that their core genome has a high level nucleotide similarity and synteny conservation. The variability observed was mainly due to acquisition of genomic islands carrying several antibiotic resistance genes.

Comparative genome analysis of a multidrug-resistant Pseudomonas aeruginosa sequence type 277 clone that harbours two copies of the blaSPM-1 gene and multiple single nucleotide polymorphisms in other resistance-associated genes

“…Another player in shaping the bacterial genome is the CRISPR-Cas system. The type I-C found in CCBH4851 genome was previously described in other ST-277 bacteria as well as in members of ST-235 [25,40]. A recent study suggests the CRISPR-Cas systems' protective effect is more evident at the population level than at an evolutionary scale [41].…”

“…The P. aeruginosa CCBH4851 genome contained 19 integrated genomic islands with a total of 705 CDSs (Fig 2, yellow highlighted regions; S3 Table). Eighteen islands were homologous to P. aeruginosa genome islands (PAGIs) previously found in other clones belonging to ST-277 [25,26]. An additional region found by IslandViewer was less than 5,000 bp, but had a G+C content of 55.13% and encoded 5 proteins including an integrase and an HTH domain-containing protein.…”

“…Previous studies demonstrate ST277 strains have a highly conserved DNA sequence and share several virulence and antimicrobial resistance-related features. However, to date, the presence of bla SPM-1 gene conferring resistance to carbapenem is a singular trait of Brazilian clones [4,25]. The purpose of this study was to reassemble and to re-annotate CCBH4851 genome in order to perform a thorough characterization and a genomic comparison with other P. aeruginosa strains.…”

“…In addition, the second CRISPR array was located adjacent to an intact CRISPR-associated gene cluster, cas2 (PA4851 25885), cas1 (PA4851 25890), cas4 (PA4851 25895), cas7 (PA4851 25900), cas8c (PA4851 25905), cas5 (PA4851 25910), cas3 (PA4851 25915). These cas genes were related to the type I-C CRISPR-Cas system and were located inside the PAGI-34, as previously described in other ST-277 clones [25].…”

Gene regulatory network inference and analysis of multidrug-resistant Pseudomonas aeruginosa

“…Most of SPM-producing isolates belong to clone ST277, as indicated through multilocus sequence typing (MLST). This clone has been associated with hospital outbreaks in several Brazilian states, and have already been found in hospital sewage and rivers (8)(9)(10) . Over the past years, researchers have applied mathematical methods in order to generate computational models used to study several organisms' behavior, contributing to the development of new products, improvement and acceleration of existing health policies, and research of novel ways of overcoming scientific challenges.…”