Epidemic Territorial Spread of IncP-2-Type VIM-2 Carbapenemase-Encoding Megaplasmids in Nosocomial Pseudomonas aeruginosa Populations

Abstract

In 2003-04 first five VIM-2 metallo-β-lactamase (MBL)-producing Pseudomonas aeruginosa (MPPA) isolates with an In4-like integron In461 (aadB–blaVIM-2–aadA6) on conjugative plasmids were identified in three hospitals in Poland. In 2005-15 MPPA much expanded in the country, and as many as 80 isolates in a collection of 454 MPPA (∼18%) had In461, being one of two most common MBL-encoding integrons. The organisms occurred in 49 hospitals in 33 cities of 11/16 main administrative regions. PFGE and MLST classified them into 55 pulsotypes and 35 STs, respectively, revealing their remarkable genetic diversity overall, with only a few small clonal clusters. The S1 nuclease/hybridization assays and mating of 63 representative isolates showed that ∼85% of these had large In461-carrying plasmids of ∼350-550 kb, usually self-transmitting with high efficiency (∼10−1-10−2 per donor cell). The plasmids from 19 isolates were sequenced, and subjected to structural and SNP-based phylogenetic analysis. These formed a subgroup within a family of IncP-2-type megaplasmids, observed worldwide in pseudomonads from various environments and conferring resistance/tolerance to multiple stress factors, including antibiotics. Their microdiversity in Poland arose mainly from acquisition of different accessory fragments, as well as new resistance genes and multiplication of these. Short-read sequence and/or PCR mapping confirmed the In461-carrying plasmids in the remaining isolates to be the IncP-2 types. The study demonstrated a large-scale epidemic spread of multi-drug resistance plasmids in P. aeruginosa populations, creating an epidemiological threat. It contributes to the knowledge on IncP-2 types, which are interesting research objects in resistance epidemiology, environmental microbiology and biotechnology.

Plasmid fitness costs are caused by specific genetic conflicts enabling resolution by compensatory mutation

“…For example, pQBR103 and pQBR57 are distant relatives of a large family of clinically important megaplasmids that appear to be disseminating multidrug resistance in clinical isolates of P . aeruginosa from opportunistic infections [ 101 , 102 ]. In those strains, these megaplasmids appear to be stable even without the selective pressure of antimicrobials, suggesting that those strains are either preadapted for megaplasmid carriage or have undergone prior compensatory evolution.…”

“…Advances in long-read sequencing technologies are beginning to expose the diversity, ubiquity, and complexity of megaplasmids in diverse bacterial genera, and, therefore, our findings are likely to apply beyond this specific case of mercury resistance in a soil Pseudomonad (e.g., [99,100]). For example, pQBR103 and pQBR57 are distant relatives of a large family of clinically important megaplasmids that appear to be disseminating multidrug resistance in clinical isolates of P. aeruginosa from opportunistic infections [101,102]. In those strains, these megaplasmids appear to be stable even without the selective pressure of antimicrobials, suggesting that those strains are either preadapted for megaplasmid carriage or have undergone prior compensatory evolution.…”

What makes a megaplasmid?

“…Though rare [ 13 , 82 ], some plasmids carry multiple conjugative and replication machineries (e.g. [ 79 ]), potentially extending the range of conditions in which they can transfer and hosts in which they can be maintained, with bigger plasmids more likely to carry multiple replication modules (electronic supplementary material, figure S4). Horizontal transmission can also be enhanced by plasmid-encoded pili and fimbriae that promote biofilm formation and cell-cell adhesion [ 83 , 84 ].…”

“…500 kb) known as IncP-2, which, in nosocomial isolates, carried antimicrobial resistance, and in soil isolates carried pathways for degradation of complex organic compounds such as camphor and octane [ 136 ]. Spurred on by the availability of long-read sequencing—often necessary to resolve their complex or repetitive structures [ 24 , 26 ]—recent years have seen an increase in the number of related Pseudomonas megaplasmid sequences [ 24 , 25 , 79 , 137 ], challenging the prevailing viewpoint that plasmids make only a minor contribution to antimicrobial resistance in Pseudomonas [ 138 ]. Found in clinical samples, soil isolates, and industrial processes, this family exemplifies the mosaic nature of megaplasmids and their ability to confer locally-adaptive traits.…”

Plasmid fitness costs are caused by specific genetic conflicts

“…94,95]. For example, megaplasmids pQBR103 and pQBR57 are distant relatives of a large family of clinically important plasmids that appear to be disseminating multidrug resistance in clinical isolates of Pseudomonas aeruginosa from opportunistic infections [96,97]. In those strains, megaplasmids appear to be stable even without the selective pressure of antimicrobials, suggesting that those strains are either pre-adapted for megaplasmid carriage, or have undergone compensatory evolution.…”