The differential importance of mutations within AmpD in cephalosporin resistance of Enterobacter aerogenes and Enterobacter cloacae.
Mechanisms leading to carbapenem and cephalosporin resistance were sought in Enterobacter aerogenes isolates that were highly resistant to carbapenems but had no known carbapenemase. Results were compared with recent work examining carbapenem-resistant Enterobacter cloacae. Eighteen carbapenem-resistant E. aerogenes were screened for known β-lactamase and carbapenemase genes, and novel carbapenemases were sought in whole-genome sequencing (WGS) data of the three most resistant isolates. For all isolates, ampC, ampR, ampD and the porin genes omp35 and omp36 were investigated by Sanger sequencing or from available WGS data. Expression of ampC and porin genes was measured in comparison with cephalosporin- and carbapenem-susceptible control strains by reverse transcriptase PCR, with porin translation also detected by SDS-PAGE. Loss of Omp35, primarily due to decreased transcription (up to 250×), was observed in ertapenem-resistant isolates (MICs ≥ 2 mg/L), whereas meropenem resistance (MICs ≥ 4 mg/L) was observed in those isolates also showing decreased or no production of Omp36. Loss of Omp36 was due to combinations of premature translation termination or reduced transcription. In contrast to E. cloacae, cephalosporin resistance in E. aerogenes was not associated with lesions in AmpD. High-level cefepime resistance (MIC = 32 mg/L) was caused by a novel modification in the H-10 helix of AmpC in one isolate. The differential importance of AmpD lesions in cephalosporin resistance in E. cloacae and E. aerogenes underlines the differences between these contrasting members of the Enterobacter genus. Porin loss resulted in high-level carbapenem resistance with gradual loss of Omp36, which led to high-level meropenem resistance.
Antimicrobial Resistance and Healthcare Associated Infections (AMRHAI) Reference Unit, National Infection Service, Public Health England, 61 Colindale Avenue, London NW9 5EQ, UK. Electronic address: b.baboueeflury@bluewin.ch.
Baharak
B
Babouee Flury
Antimicrobial Resistance and Healthcare Associated Infections (AMRHAI) Reference Unit, National Infection Service, Public Health England, 61 Colindale Avenue, London NW9 5EQ, UK.
Matthew J
MJ
Ellington
Antimicrobial Resistance and Healthcare Associated Infections (AMRHAI) Reference Unit, National Infection Service, Public Health England, 61 Colindale Avenue, London NW9 5EQ, UK.
Katie L
KL
Hopkins
Antimicrobial Resistance and Healthcare Associated Infections (AMRHAI) Reference Unit, National Infection Service, Public Health England, 61 Colindale Avenue, London NW9 5EQ, UK.
Jane F
JF
Turton
Antimicrobial Resistance and Healthcare Associated Infections (AMRHAI) Reference Unit, National Infection Service, Public Health England, 61 Colindale Avenue, London NW9 5EQ, UK.
Michel
M
Doumith
Antimicrobial Resistance and Healthcare Associated Infections (AMRHAI) Reference Unit, National Infection Service, Public Health England, 61 Colindale Avenue, London NW9 5EQ, UK.
Neil
N
Woodford
D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics*; D018550:Cephalosporin Resistance; D002511:Cephalosporins / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D021902:Enterobacter aerogenes / Q000187:drug effects* / Q000235:genetics; D016972:Enterobacter cloacae / Q000187:drug effects* / Q000235:genetics; D020869:Gene Expression Profiling; D006801:Humans; D008826:Microbial Sensitivity Tests; D009154:Mutation; D009238:N-Acetylmuramoyl-L-alanine Amidase / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D002511:Cephalosporins; C062460:AmpD protein, Bacteria; D009238:N-Acetylmuramoyl-L-alanine Amidase
Carbapenem resistance; Enterobacter aerogenes; Outer membrane protein; ampC; ampD