Kurt E. Schaecher
#Alternative corresponding author
Bethesda, Maryland, USA
Email:patrick.t.mcgann4.ctr@mail.mil
Walter Reed Army Institute of Research, 2S35
mcr1in the USA
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Running Title:Colistin resistance in the USA
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1 1 2 1 3# Clifford , Mary Hinkle , Timothy Whitman , Emil Lesho , and Kurt E. Schaecher
1#* 1* 1 1 1 Patrick McGann , Erik Snesrud , Rosslyn Maybank , Brendan Corey , Ana C. Ong , Robert
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# Corresponding author
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Escherichia coliHarboringmcr1andblaCTX-Mon a Novel IncF Plasmid: First report of
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AAC Accepted Manuscript Posted Online 26 May 2016 Antimicrob. Agents Chemother. doi:10.1128/AAC.01103-16 Copyright © 2016 McGann et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license.
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Department of Pathology,
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1 Multidrug-resistant Organism Repository and Surveillance Network, Walter Reed Army
Walter Reed National Military Medical Center
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*PMG and ES contributed equally to this manuscript
Patrick Mc Gann, PhD
Email:kurt.e.schaecher.ml@mail.mil
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Institute of Research, Silver Spring, Maryland, USA.
2 Department of Infectious Diseases, Walter Reed National Military Medical Center, MD, USA .
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Silver Spring, Maryland 20910, USA
Phone: (301) 319 9912
3 Department of Pathology, Walter Reed National Military Medical Center, MD, USA.
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The recent discovery of a plasmid-borne colistin resistance gene,mcr1, heralds the emergence
of truly pan-drug resistant bacteria (1). The gene has been found primarily inEscherichia coli,
but has also been identified in other members of theEnterobacteriaceaefrom human, animal,
food and environmental samples on every continent (2-5). In response to this threat, starting in
May 2016, all extended-spectrum β-lactamase (ESBL)-producingE.coliclinical isolates
submitted to the clinical microbiology laboratory at the Walter Reed National Military Medical
Center (WRNMMC) were tested for resistance to colistin by E-test. Herein we report the
presence ofmcr1in anE. colicultured from a patient with a urinary tract infection (UTI) in the
United States.
E. coliMRSN 388634 was cultured from the urine of a 49 year-old female who presented to a
th clinic in Pennsylvania on April 26 2016 with symptoms indicative of a UTI. The isolate was
forwarded to WRNMMC, where susceptibility testing indicated an ESBL phenotype (Table 1).
The isolate was included in the first 6 ESBL-producingE.coliselected for colistin susceptibility
testing, and it was the only isolate to have a MIC of colistin of 4µg/ml (all others had MICs ≤
0.25 µ/ml). Colistin MIC was confirmed by microbroth dilution andmcr1detected by real-time
PCR (6). Whole genome sequencing (WGS) of MRSN 388634 was performed on a PacBio RS II
and Miseq benchtop sequencer.
E. coliMRSN 388634 belonged to ST457, a rareE.coliST first identified in 2008 from a urine
culture in the UK (7). It was subsequently identified from a bloodstream culture in Italy, where it
was found to harbor the carbapenemase geneblaKPC-3andblaCTX-M-55(8). MRSN 388634 carried
15 antibiotic resistance genes, but no carbapenemases, that were harbored on two plasmids
(Table 2).
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The first plasmid, pMR0516mcr, was 225,707 bp in size and belonged to incompatibility group
F18:A-:B1 (9). BLAST analysis indicated that pMR0516mcr represented a novel IncF plasmid.
Notably, it shares 89 Kb of homologous sequence with pHNSHP45-2, amcr1carrying IncHI2
plasmid described by Liu and colleagues (1). This shared sequence containsmcr1in association
with ISApl1(1), but in pMR0516mcr it is in a different location and orientation (Figure 1).
pMR0516mcr also carried 7 additional antibiotic resistance genes, including the ESBL gene
blaCTX-M-55(Table 2). The second plasmid, pMR0416ctx, was ~47 Kb in size and was assigned to
IncN (Table 2). It carried 7 antibiotic resistance genes includingblaCTX-M-14. A complete
description of both plasmids is under preparation.
To the best of our knowledge, this is the first report ofmcr1in the USA. The epidemiology of
MRSN 388634 is noteworthy; the isolate was submitted from a clinic in Pennsylvania, and the
patient reported no travel history within the prior 5 months. To date a further 20 ESBL-
producingE. colifrom patients at the WRNMMC have tested negative formcr1, and are colistin
sensitive. However, as testing has only been ongoing for 3 weeks, it remains unclear what the
true prevalence ofmcr1is in the population. The association betweenmcr1and IncF plasmids
is concerning as these plasmids are vehicles for the dissemination of antibiotic resistance and
virulence genes among theEnterobacteriaceae(9). Continued surveillance to determine the true
frequency for this gene in the USA is critical.
Nucleotide Accession Numbers.TheShort Read Archive (SRA) file for MRSN 388623 has
been deposited at Genbank under Accession number SRP075674.
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Acknowledgements.
This study was funded by the U.S. Army Medical Command, the Global Emerging Infections
Surveillance and Response System,and the Defence Medical Research and Development
Program. All authors declare no conflict of interest.
The identification of specific products or scientific instrumentation does not constitute
endorsement or implied endorsement on the part of the author, DoD, or any component agency.
While we generally excise references to products, companies, manufacturers, organizations, etc.
in government produced works, the abstracts produced and other similarly situated research
presents a special circumstance when such product inclusions become an integral part of the
scientific endeavour.
Material has been reviewed by the Walter Reed Army Institute of Research. There is no
objection to its presentation. The views expressed in this article are those of the author and do
not reflect the official policy of the Department of Army/Navy/Air Force, Department of
Defense, or the U.S. Government.
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could contribute to the global spread of themcr1gene. Euro surveillance21.
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Table 1. Antibiotic resistance profile of MRSN 388634
Antibiotic
Amikacin
Amoxacillin/clavulanate
Ampicillin
Aztreonam
Cefazolin
Cefepime
Ceftazidime
Ceftriaxone
Ciprofloxacin
Colistin
Ertapenem
Gentamicin
Imipenem
Levofloxacin
Meropenem
Nitrofurantoin
piperacillin-tazobactam
Tetracycline
Tobramycin
1 MIC (µg/ml)
≤8
16/8
>16
>16
>16
>16
>16
>32
>2
4
≤ 0.25
>8
≤ 0.25
>4
≤ 0.25
≤ 16
4/4
>8
>8
Trimethoprim/sulfamethoxazole >2/38
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1 MIC’s were determined using the BD Phoenix (BD Diagnostics Systems, MD, USA) with
panels NMIC/ID 133 except for colistin, which was performed using E-test and manual
microbroth dilution, and both gave MICs of colistin = 4µg/ml.
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Table 2 Characteristics of plasmids inE.coliMRSN 388634
Name
pMR0516mcr
pMR0416ctx
Size (Kb)
225.7
47
1 Inc
F18:A-:B+
N
2 Copy#
2
1
Antibiotic resistance genes
strA,strB,blaCTX-M-55,blaTEM-1B,mcr1,sul2, tet(A), dfrA14
aac(3)IVa,aph(4)Ia,blaCTX-M-14,fosA3,mph(A),floR,sul2
1 Plasmid Incompatibility (Inc) group, as determined by Plasmid Finder version 1.2 (10).
2 Average copy number per cell, normalized to the chromosomal read coverage.
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Figure 1.Comparison of the homologous region containingmcr1shared by pMR0516mcr and
pHNSHP45-2. Open arrows represent coding sequences (Green arrows,mcr1; white arrows,
ISapl1; purple arrows, metabolic function; blue arrows, plasmid replication and maintenance;
grey arrows, hypothetical and unclassified) and indicate direction of transcription. Arrow size is
proportional to the gene length. The grey and blue areas between pMR0516mcr and pHNSHP45-
2 indicate nucleotide identity >99.9% by BLASTN.
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