6 Bonnin RA, Nordmann P, Potron A et al.

Carbapenem-hydrolyzing
GES-type extended-spectrum b-lactamase in Acinetobacter baumannii.
Antimicrob Agents Chemother 2011; 55: 34954.
J Antimicrob Chemother 2012
doi:10.1093/jac/dks041
Advance Access publication 16 February 2012
Emergence of NDM-1-producing
Acinetobacter baumannii in Belgium
Pierre Bogaerts
1
*, Roberta Rezende de Castro
1
,
Sandrine Roisin
2
, Ariane Deplano
2
, Te-Din Huang
1
,
Marie Hallin
2
, Olivier Denis
2
and Youri Glupczynski
1
1
National Reference Laboratory of Antimicrobial Resistance in
Gram-negative Bacteria, Cliniques universitaires CHU UCL de
Mont-Godinne, Yvoir, Belgium;
2
Associated National Reference
Laboratory of Antimicrobial Resistance in Gram-negative Bacteria,
Ho pital Universitaire Erasme, Universite Libre de Bruxelles (ULB),
Brussels, Belgium
*Corresponding author. Laboratory of Bacteriology, Cliniques
Universitaires, UCL de Mont-Godinne, Av. Dr Therasse 1, 5530 Yvoir,
Belgium. Tel: +32-(0)81-42-32-41; Fax: +32-(0)81-42-32-04;
E-mail: pierre.bogaerts@uclouvain.be
Keywords: metallo-b-lactamases, carbapenemases, resistance,
New Delhi
Sir,
New Delhi metallo-b-lactamase (NDM) is a class B b-lactamase
that confers resistance to virtually all b-lactams, including
carbapenems. Initially reported in Sweden in Enterobacteriaceae
isolates from a patient transferred from India, NDM-1 is currently
spreading worldwide, including Belgium, especially in Enterobac-
teriaceae.
1
Recently the presence of bla
NDM-1
was also reported
in Acinetobacter baumannii isolates from India, China and
Germany, suggesting a broad host range distribution of this
gene.
24
Here, we report an NDM-1-producing A. baumannii
detected in Belgium that was isolated from a patient following
medical repatriation from Algeria.
In early 2011, a young male patient was admitted to the
emergency ward of an Algerian hospital for severe cranial
and thoracic trauma following a road trafc accident. He pre-
sented with a Glasgow Coma Score (GCS) of 07/15 and was
rapidly intubated, ventilated and transferred to the intensive
care unit (ICU).
While hospitalized in Algeria, he experienced three successive,
distinct episodes of bloodstream infection, due to Acinetobacter
spp., Candida albicans and Klebsiella pneumoniae, but no clinical
information was available concerning the possible sources of
these infections. In August, the patient was transferred, in a
vegetative state (GCS of 02/15), to the ICU of a Belgian hospital.
He was immediately placed in a single room with contact isola-
tion precautions.
Screening for asymptomatic intestinal carriage of
carbapenemase producers was carried out upon admission, by
rectal swabbing cultured by direct plating on ChromID
TM
ESBL
Agar (bioMerieux, Marcy lE

toile, France). The recovered isolate

was identied as A. baumannii (Ab 11314) by MicroFlex LT (Bruker
Daltonik, Bremen, Germany). By disc diffusion, Ab 11314 was
found to be multiresistant and synergy was observed in a double
disc test using imipenem and EDTA (420 mg), suggesting involve-
ment of a metallo-b-lactamase in resistance to b-lactams. Use
of the CLSI microdilution method conrmed that the isolate was
resistant to b-lactams including ceftazidime (MIC .64 mg/L),
cefepime (MIC .64 mg/L), imipenem (MIC 4 mg/L; Etest MIC
.32 mg/L), meropenem (MIC 4 mg/L; Etest MIC .32 mg/L), ami-
kacin (MIC .32 mg/L) and ciprooxacin (MIC .32 mg/L), and sus-
ceptibletotigecycline (MIC0.125 mg/L), colistin(MIC0.5 mg/L) and
minocycline (MIC ,2 mg/L).
Analysis of b-lactamase-coding genes by an MDR CT102
array (Check-Points, Wageningen, The Netherlands) and by
PCR sequencing targeting metallo-b-lactamases and OXA-
carbapenemases revealed the presence of bla
NDM-1
. Plasmid
extraction was performed with a Qiagen plasmid kit, and this
revealed a single 174 kb untypeable plasmid.
5
This plasmid was ef-
ciently electroporated and transferred by mating experiments to
the A. baumannii recipient strain N9040364, and it conferred
resistance to aminoglycosides (gentamicin, tobramycin and
amikacin) only (data not shown). The bla
NDM-1
gene was not asso-
ciated with this plasmid suggesting that it was located in the
chromosome. PCR mapping and sequencing revealed that bla
NDM-1
was located between two direct repeats of the ISAba125 element
in a transposon similar to the one in A. baumannii Ab 161/07
reported from Germany (accession no. HQ857107).
2
Nevertheless,
a PCR experiment performed with Ab 161/07 as a positive control
revealed that in Ab 11314, in contrast, the mfs gene (a
shikimate-transporter-coding gene belonging to the major facilita-
tor superfamily) was not disrupted, indicating that the ISAba125
transposon was inserted elsewhere in the chromosome. By multi-
locus sequence typing (MLST) (http:// pubmlst.org/abaumannii/), A.
baumannii Ab 11314 was shown to belong to sequence type (ST)
92 (allelic prole: 1-3-3-2-2-7-3), the founding genotype of
clonal complex (CC) 92, which includes European clone 2 (EU2)
and worldwide lineage 2 (WW2). CC92 is one of the most prevalent
CCs worldwide, mostly represented by OXA-23-producing strains.
Sequencing analysis of the intrinsic bla
OXA-51
-like and bla
ADC
genes showed that this strain harboured bla
OXA-64
and bla
ADC-26
,
neither of which was downstream of ISAba1 or ISAba125. These
two intrinsic genes were identical to those recovered in the
NDM-1-expressing A. baumannii Ab 161/07, but nevertheless pre-
sented a very different MLST prole (1-15-2-28-1-new allele-32),
not related to any CC described up to now.
This clinical isolate is the rst characterized NDM-1-producing
A. baumannii from Belgium. As yet, there have been only a few
reports of NDM-producing A. baumannii, and the only case
reported in Europe was from Germany in a patient transferred
from Serbia in 2007.
2
In the present case, the NDM-1-producing
A. baumannii isolate originated from North Africa (Algeria) where
NDM-1 has not yet been described in A. baumannii, although
another single case of A. baumannii producing an NDM-1
variant (NDM-2) was recently reported from Egypt.
6
This brief
report further highlights the wide distribution of NDM-producing
organisms around the world, their constant evolving spread
Research letters
1552

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and underlines the importance of systematic screening upon ad-
mission and early application of strict barrier precautions for any
transferred patient, whatever the country of origin.
Acknowledgements
We are very grateful to Yvonne Pfeifer for providing us with the German
A. baumannii Ab 161/07 NDM-1-producing isolate.
Funding
This work was supported by EU grant FP7-HEALTH-2009-SINGLE-STAGE
TEMPOtest-QC, project 241742.
Transparency declarations
None to declare.
References
1 Nordmann P, Poirel L, Toleman M et al. Does broad-spectrum b-lactam
resistance due to NDM-1 herald the end of the antibiotic era for
the treatment of infections caused by Gram-negative bacteria? J
Antimicrob Chemother 2011; 66: 68992.
2 Pfeifer Y, Wilharm G, Zander E et al. Molecular characterization of
bla
NDM-1
in an Acinetobacter baumannii strain isolated in Germany in
2007. J Antimicob Chemother 2011; 66: 19982001.
3 Chen Y, Zhou ZH, Jiang Y et al. Emergence of NDM-1-producing
Acinetobacter baumannii in China. J Antimicrob Chemother 2011; 66:
12559.
4 Karthikeyan K, Thirunarayan MA, Krishnan P. Coexistence of bla
OXA-23
with bla
NDM-1
and armA in clinical isolates of Acinetobacter baumannii
from India. J Antimicrob Chemother 2010; 65: 22534.
5 Bertini A, Poirel L, Mugnier PD et al. Characterization and PCR-based
replicon typing of resistance plasmids in Acinetobacter baumannii.
Antimicrob Agents Chemother 2010; 54: 416877.
6 Kaase M, Nordmann P, Wichelhaus TA et al. NDM-2 carbapenemase in
Acinetobacter baumannii from Egypt. J Antimicrob Chemother 2011; 66:
12602.
J Antimicrob Chemother 2012
doi:10.1093/jac/dks095
Advance Access publication 21 March 2012
Emergence of NDM-1-producing
Enterobacteriaceae in China
Pak-Leung Ho
1
*, Zhen Li
1
, Eileen L. Lai
1
, Susan S. Chiu
2
and Vincent C. C. Cheng
1
1
Department of Microbiology and Carol Yu Centre for Infection,
The University of Hong Kong, Queen Mary Hospital, Hong Kong;
2
Department of Paediatrics and Adolescent Medicine,
The University of Hong Kong, Queen Mary Hospital, Hong Kong
*Corresponding author. Division of Infectious Diseases, Department of
Microbiology, The University of Hong Kong, Queen Mary Hospital,
Pokfulam Road, Pokfulam, Hong Kong SAR, China. Tel: +852-2855 4897;
Fax: +852-2855 1241; E-mail: plho@hkucc.hku.hk
Keywords: carbapenem resistance, carbapenemases,
cefotaximases, Escherichia coli
Sir,
The worldwide dissemination of bacteria producing New Delhi
metallo-b-lactamase (NDM) is a major public health issue.
1
Since NDM producers are primarily found among individuals
with a history of hospitalization or travel to the Indian subcontin-
ent, many hospitals have implemented microbiological screening
of patients with such an epidemiological history.
1,2
In mid-2011, the stool sample of a 1-year-old infant was
found to have carbapenem-resistant Enterobacteriaceae (CRE)
upon admission screening. The infant was admitted because of
cough and intermittent fever in the preceding 2 weeks. The
family had travelled to and stayed in Hunan Province, China in
the preceding month. Following onset of the symptoms, the
infant had been admitted to a hospital in Hunan for 3 days.
The patient was given a diagnosis of broncholitis and had been
treated with a course of intravenous cefoperazone. At presenta-
tion to our hospital, the patient had a fever of 388C, but the chest
examination was unremarkable. The patient was treated conser-
vatively and the fever resolved without further antibiotic treat-
ment. The patient was discharged 2 days later. In accordance
with the screening policy, stool samples were obtained for sur-
veillance culture.
In brief, a red-bean-size faecal pellet was suspended in saline.
A 10 mL aliquot of the suspension was then removed and plated
directly onto a ChromID ESBL plate (bioMerieux, Marcy lEtoile,
France). In addition, a broth enrichment step was performed
by inoculating another 10 mL aliquot of the faecal suspension
into nutrient broth with 1 mg/L meropenem, incubated at 378C
overnight and then subcultured onto a MacConkey agar plate
with 1 mg/L meropenem. All plates were incubated at 378C in
air for 20 h. All distinct colony types recovered from either the
chromogenic or the MacConkey media were investigated for evi-
dence of carbapenemase activity, using the combined disc
method and boronic acid or EDTA as inhibitor.
3
All isolates
were identied using VITEK 2 (bioMerieux), and antimicrobial sus-
ceptibility was determined using the CLSI disc diffusion method.
4
After the patients stool samples were found to carry CRE,
stool samples from the infants parents and other family
members were also cultured using the same methodology. A
total of four CRE isolates were recovered from the faecal
samples of the child and her mother (Table 1). Cultures of the
faecal samples from the other household members (father,
grandfather, grandmother and aunt) were negative. All four iso-
lates (two Escherichia coli, one Klebsiella pneumoniae and one
Enterobacter aerogenes) exhibited synergy with EDTA in the com-
bined disc testing. No synergy with boronic acid was observed.
PCR and sequencing, using previously described methods,
2
con-
rmed the presence of NDM-1 in the four CRE isolates. The pres-
ence of additional b-lactamase genes, including other
metallo-b-lactamase (IMP, VIM, GIM, SPM and SIM), CTX-M and
OXA-48-like genes, was also tested by PCR and sequencing.
5
This allowed identication of additional extended-spectrum
b-lactamase (ESBL) genes in the K. pneumoniae isolate
Research letters
1553
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