blaIMP-27 on transferable plasmids in Proteus mirabilis and Providencia rettgeri

Abstract

Objectives: A carbapenem-resistant Providencia rettgeri (PR1) isolate was recovered from a wound infection in Missouri, USA. This isolate possessed an EDTA-inhibitable carbapenemase that was unidentified using the Xpert CARBA-R assay. Our objective was to elucidate the molecular determinant of carbapenem resistance in this isolate. We then sought to test the transmissibility of bla_IMP-27 loci in clinical P. rettgeri and Proteus mirabilis isolates.

Methods: In October 2016 the novel ambler Class B carbapenemase bla_IMP-27, was reported in two different Proteus mirabilis (PM185 and PM187) isolates. Broth mating assays for transfer of carbapenemase activity were performed for the three clinical isolates with recipient sodium azide-resistant Escherichia coli J53. Antibiotic susceptibility testing and phenotypic carbapenemase activity testing were performed on the clinical isolates, J53 and transconjugants using the Kirby-Bauer disc diffusion method according to CLSI guidelines. Plasmid DNA from PM187, PR1 and their transconjugants were used as input for Nextera Illumina sequencing libraries and sequenced on a NextSeq platform.

Results: PR1 was resistant to both imipenem and meropenem. PM187 and PR1 could transfer resistance to E. coli through plasmid conjugation (pPM187 and pPR1). pPM187 had a virB/virD4 type IV secretion system whereas pPR1 had a traB/traD type IV secretion system.

Conclusion: Two of three bla_IMP-27-bearing clinical isolates tested could conjugate resistance into E. coli. The resulting transconjugants became positive for phenotypic carbapenemase production but did not pass clinical resistance breakpoints. bla_IMP-27 can be transmitted on different plasmid replicon types that rely on distinct classes of type IV secretion system for horizontal transfer.

Keywords: Carbapenem resistance; Horizontal gene transfer; Proteus mirabilis; Providencia rettgeri; bla(IMP).

Figure 1

(A) Annotated plasmid diagram from DNAPlotter of pPM187 (36,843 bp) displaying bla_IMP-27 co-localized with a Class II integron gene cassette and type IV secretion system. The inner most ring shows GC plot, the second ring shows GC skew, the third ring represents open reading frames in the forward direction, and the fourth ring (adjacent to the nucleotide position counter) indicates open reading frames in the reverse direction. (B) Annotated plasmid diagram from DNAPlotter of pPR1 (107,365 bp) displaying bla_IMP-27 co-localized with a Class II integron gene cassette, a tra operon, and additional resistance genes.