title: Enhanced Klebsiella pneumoniae Carbapenemase Expression from a Novel Tn4401Deletion.
abstract: TheKlebsiella pneumoniaecarbapenemase gene (blaKPC) is typically located within mobile transposon Tn4401Enhanced KPC expression has been associated with deletions in the putative promoter region upstream ofblaKPCIllumina sequences fromblaKPC-positive clinical isolates from a single institution were mapped to a Tn4401b reference sequence, which carries no deletions. The novel isoform Tn4401h (188-bp deletion [betweenistBandblaKPC]) was present in 14% (39/281) of clinical isolates. MICs showed thatEscherichia colistrains containing plasmids with Tn4401a and Tn4401h were more resistant to meropenem (≥16 and ≥16, respectively), ertapenem (≥8 and 4, respectively), and cefepime (≥64 and 4, respectively) thanE. colistrains with Tn4401b (0.5, ≤0.5, and ≤1, respectively). Quantitative real-time PCR (qRT-PCR) demonstrated that Tn4401a had a 16-fold increase and Tn4401h a 4-fold increase inblaKPCmRNA levels compared to the reference Tn4401b. AlacZreporter plasmid was used to test the activity of the promoter regions from the different variants, and the results showed that the Tn4401a and Tn4401h promoter sequences generated higher β-galactosidase activity than the corresponding Tn4401b sequence. Further dissection of the promoter region demonstrated that putative promoter P1 was not functional. The activity of the isolated P2 promoter was greatly enhanced by inclusion of the P1-P2 intervening sequence. These studies indicated that gene expression could be an important consideration in understanding resistance phenotypes predicted by genetic signatures in the context of sequencing-based rapid diagnostics.
authors.affiliation: Division of Infectious Diseases and International Health, Department of Medicine, University of Virginia Health System, Charlottesville, Virginia, USA.
authors.forename: Anita
authors.initials: A
authors.lastname: Cheruvanky
authors.affiliation: Modernizing Medical Microbiology Consortium, Nuffield Department of Clinical Medicine, John Radcliffe Hospital, Oxford University, Oxford, United Kingdom.
authors.forename: Nicole
authors.identifier: 0000-0002-4508-7969
authors.initials: N
authors.lastname: Stoesser
authors.affiliation: Modernizing Medical Microbiology Consortium, Nuffield Department of Clinical Medicine, John Radcliffe Hospital, Oxford University, Oxford, United Kingdom.
authors.forename: Anna E
authors.initials: AE
authors.lastname: Sheppard
authors.affiliation: Modernizing Medical Microbiology Consortium, Nuffield Department of Clinical Medicine, John Radcliffe Hospital, Oxford University, Oxford, United Kingdom.
authors.forename: Derrick W
authors.initials: DW
authors.lastname: Crook
authors.affiliation: Division of Infectious Diseases and International Health, Department of Medicine, University of Virginia Health System, Charlottesville, Virginia, USA.
authors.forename: Paul S
authors.initials: PS
authors.lastname: Hoffman
authors.affiliation: Division of Infectious Diseases and International Health, Department of Medicine, University of Virginia Health System, Charlottesville, Virginia, USA.|Department of Biologic Sciences, Shenandoah University, Winchester, Virginia, USA.
authors.forename: Erin
authors.initials: E
authors.lastname: Weddle
authors.affiliation: Clinical Microbiology, Department of Pathology, University of Virginia Health System, Charlottesville, Virginia, USA.
authors.forename: Joanne
authors.initials: J
authors.lastname: Carroll
authors.affiliation: Division of Infectious Diseases and International Health, Department of Medicine, University of Virginia Health System, Charlottesville, Virginia, USA.
authors.forename: Costi D
authors.initials: CD
authors.lastname: Sifri
authors.affiliation: Division of Infectious Diseases and International Health, Department of Medicine, University of Virginia Health System, Charlottesville, Virginia, USA.
authors.forename: Weidong
authors.initials: W
authors.lastname: Chai
authors.affiliation: Division of Infectious Diseases and International Health, Department of Medicine, University of Virginia Health System, Charlottesville, Virginia, USA.
authors.forename: Katie
authors.initials: K
authors.lastname: Barry
authors.affiliation: Division of Infectious Diseases and International Health, Department of Medicine, University of Virginia Health System, Charlottesville, Virginia, USA.
authors.forename: Girija
authors.initials: G
authors.lastname: Ramakrishnan
authors.affiliation: Division of Infectious Diseases and International Health, Department of Medicine, University of Virginia Health System, Charlottesville, Virginia, USA ajm5b@virginia.edu.|Clinical Microbiology, Department of Pathology, University of Virginia Health System, Charlottesville, Virginia, USA.
authors.forename: Amy J
authors.identifier: 0000-0002-3533-6351
authors.initials: AJ
authors.lastname: Mathers
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics*; D015780:Carbapenems / Q000494:pharmacology*; D000077723:Cefepime; D002511:Cephalosporins / Q000494:pharmacology; D004251:DNA Transposable Elements / Q000235:genetics*; D000077727:Ertapenem; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics; D006801:Humans; D007710:Klebsiella Infections / Q000382:microbiology; D007711:Klebsiella pneumoniae / Q000187:drug effects* / Q000235:genetics* / Q000302:isolation & purification; D000077731:Meropenem; D008826:Microbial Sensitivity Tests; D011401:Promoter Regions, Genetic / Q000235:genetics; D017384:Sequence Deletion / Q000235:genetics; D013845:Thienamycins / Q000494:pharmacology; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism; D001618:beta-Lactamases / Q000096:biosynthesis / Q000235:genetics*; D047090:beta-Lactams / Q000494:pharmacology
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D015780:Carbapenems; D002511:Cephalosporins; D004251:DNA Transposable Elements; D013845:Thienamycins; D047090:beta-Lactams; D000077723:Cefepime; D001616:beta-Galactosidase; D001618:beta-Lactamases; C063912:carbapenemase; D000077731:Meropenem; D000077727:Ertapenem
keywords: KPC; Tn4401; carbapenem-resistant Enterobacteriaceae; gene expression; meropenem; promoters