Journal of Medical Microbiology (2012), 61, 353–360 DOI 10.1099/jmm.0.

036939-0

In vitro time-kill studies of antimicrobial agents

against blood isolates of imipenem-resistant
Acinetobacter baumannii, including colistin- or
tigecycline-resistant isolates
Kyong Ran Peck,13 Min Ja Kim,23 Ji Young Choi,33 Hong Sun Kim,3
Cheol-In Kang,1 Yong Kyun Cho,4 Dae Won Park,5 Hee Joo Lee,6
Mi Suk Lee7 and Kwan Soo Ko3,8
Correspondence 1
Division of Infectious Diseases, Samsung Medical Center, Sungkyunkwan University School of
Kwan Soo Ko Medicine, Seoul, Republic of Korea
ksko@skku.edu 2
Division of Infectious Diseases, Korea University Anam Hospital, Korea University,
College of Medicine, Seoul, Republic of Korea
3
Department of Molecular Cell Biology, Samsung Biomedical Research Institute,
Sungkyunkwan University School of Medicine, Suwon, Republic of Korea
4
Gachon University, Gil Hospital, Inchon, Republic of Korea
5
Division of Infectious Diseases, Korea University Ansan Hospital, Korea University,
College of Medicine, Ansan, Republic of Korea
6
Departments of Laboratory Medicine, School of Medicine, Kyung Hee University, Seoul,
Republic of Korea
7
Department of Internal Medicine, School of Medicine, Kyung Hee University, Seoul,
Republic of Korea
8
Asia Pacific Foundation for Infectious Diseases (APFID), Seoul, Republic of Korea

The emergence of colistin or tigecycline resistance as well as imipenem resistance in

Received 2 August 2011
Accepted 17 October 2011
Acinetobacter baumannii poses a great therapeutic challenge. The bactericidal and synergistic
effects of several combinations of antimicrobial agents against imipenem-, colistin- or tigecycline-
resistant A. baumannii isolates were investigated by in vitro time-kill experiments. Six imipenem-
resistant A. baumannii blood isolates were examined in this study, including colistin- and
tigecycline-susceptible, colistin-resistant but tigecycline-susceptible, and colistin-susceptible but
tigecycline-resistant isolates. Time-kill studies were performed using five antimicrobial agents
singly or in combinations (imipenem plus colistin, imipenem plus ampicillin-sulbactam, colistin plus
rifampicin, colistin plus tigecycline, and tigecycline plus rifampicin) at concentrations of 0.5¾ and
1¾ their MICs. Only imipenem was consistently effective as a single agent against all six A.
baumannii isolates. Although the effectiveness of combinations of 0.5¾ MIC antimicrobial agents
was inconsistent, combination regimens using 1¾ MIC of the antimicrobial agents displayed
excellent bactericidal activities against all six A. baumannii isolates. Among the combinations of
0.5¾ MIC antimicrobial agents, the combination of colistin and tigecycline showed synergistic or
bactericidal effects against four of the isolates. This in vitro time-kill analysis suggests that
antimicrobial combinations are effective for killing imipenem-resistant A. baumannii isolates, even
if they are simultaneously resistant to either colistin or tigecycline. However, the finding that the
combinations of 0.5¾ MIC antimicrobial agents were effective on only some isolates may warrant
further investigation of the doses of combination agents needed to kill resistant A. baumannii.

3These authors contributed equally to this work.

Abbreviation: MDR, multidrug resistant.

036939 G 2012 SGM Printed in Great Britain 353

K. R. Peck and others

INTRODUCTION baumannii (Perez et al., 2007; Peleg et al., 2008). However,

Acinetobacter baumannii has emerged as an important
nosocomial pathogen, especially in intensive care units
(Dijkshoorn et al., 2007). A. baumannii infections may be
difficult to treat due to the pathogen’s multidrug resistance.
Although carbapenems, including imipenem and merope-
nem, have been commonly used as the mainstay of
treatment for severe A. baumannii infections, carbapenem-
resistant isolates have emerged and disseminated world-
wide in recent years (Perez et al., 2007). With the exception
of polymyxins (such as polymyxin B and colistin) and
tigecycline, few alternative therapeutic options are available
(Munoz-Price & Weinstein, 2008). However, polymyxin-
resistant isolates of A. baumannii have also developed (Li
et al., 2006a; Park et al., 2009a), along with tigecycline-
resistant isolates (Capone et al., 2008; Park et al., 2009b).
Even pandrug-resistant (PDR) A. baumannii isolates,
displaying resistance to all antimicrobial agents, including
both polymyxins and tigecycline, have recently emerged
(Doi et al., 2009; Park et al., 2009c).
A. baumannii isolates cause bloodstream infection, noso-
comial-acquired pneumonia or ventilator-associated pneu-
monia in critically ill patients. Especially those with
inappropriate treatment are associated with higher mor-
tality (Falagas et al., 2006). However, the development of
new antimicrobial agents to combat A. baumannii infec-
tions has been slow. Thus, the use of combinations of two
or more agents has drawn attention as an option for
treating multidrug-resistant (MDR) A. baumannii infec-
tions (Munoz-Price & Weinstein, 2008; Peleg et al., 2008),
although the effectiveness of such combinations remains
controversial (Moland et al., 2008; Scheetz et al., 2007). In
addition to increasing eradication efficacy, combination
therapy may also help to prevent the emergence of resistant
populations (Pachón-Ibáñez et al., 2006). So far, several
combinations, such as imipenem and ampicillin-sulbac-
tam, rifampicin and polymyxin B, imipenem and poly-
myxin B, and colistin and rifampicin, have been reported
to be effective in vitro against carbapenem-resistant A.
studies on the effects of these combinations against
colistin- or tigecycline-resistant A. baumannii isolates are
very limited (Moland et al., 2008).
In this study, we investigated the synergistic and bacte-
ricidal effects of combinations of antimicrobial agents
against carbapenem-resistant A. baumannii blood isolates
that were also resistant to either colistin or tigecyc-
line by in vitro time-kill analysis using a microdilution
method.
METHODS
Bacterial isolates. Six representative imipenem-resistant A.
baumannii blood isolates that were isolated from intensive care
unit patients at four university hospitals in South Korea were
included in the present study (Table 1). All were also resistant to
meropenem. Two isolates (KRU-A-3 and KHU-4) were resistant to
carbapenems, but susceptible to polymyxins and tigecycline (COL-
S/TIG-S), two isolates (KCU-4 and SKKU-2) were resistant to
polymyxins but susceptible to tigecycline (COL-R/TIG-S), and two
isolates (SKKU-8 and KCU-3) were resistant to tigecycline but
susceptible to polymyxins (COL-S/TIG-R). All six isolates were
resistant to ampicillin-sulbactam. While two isolates (KHU-4 and
KCU-4) were resistant to rifampicin, all the others were susceptible
to rifampicin.
Determination of MIC. In vitro antimicrobial susceptibility testing
was performed by measuring MIC using the broth microdilution
method according to the Clinical and Laboratory Standards
Institute (CLSI) guidelines (CLSI, 2010). Fifteen antimicrobial
agents were tested: imipenem, meropenem, polymyxin B, colistin,
ciprofloxacin, rifampicin, amikacin, cefepime, ceftriaxone, cefoper-
azone-sulbactam, ceftazidime, piperacillin-tazobactam, ampicillin-
sulbactam, tetracycline and tigecycline. Fresh Mueller–Hinton broth
was used for all susceptibility testing. CLSI susceptibility interpre-
tive criteria were used (CLSI, 2010). No breakpoints for rifampicin
and tigecycline are available in the CLSI guidelines; therefore CLSI
criteria recommended for staphylococci were applied to rifam-
picin (resistant ¢4 mg l21), and the criteria of the United States
Food and Drug Administration for Enterobacteriaceae were
used for tigecycline (intermediate 4 mg l21; resistant ¢8 mg l21).

Table 1. Antimicrobial resistance profiles of the A. baumannii isolates used in this study

S, susceptible; R, resistant.

Isolate MIC of antimicrobial agent (mg l”1) (resistance or susceptibility)

Imipenem Meropenem Colistin Polymyxin B Ampicillin- Tigecycline Rifampicin

sulbactam

KRU-A-3 128 (R) .64 (R) 1 (S) 1 (S) 128/64 (R) 2 (S) 2 (S)
KHU-4 128 (R) .64 (R) 1 (S) 1 (S) 128/64 (R) 1 (S) 4 (R)
KCU-4 256 (R) .64 (R) 32 (R) 4 (R) 256/128 (R) 1 (S) 4 (R)
SKKU-2 128 (R) .64 (R) 128 (R) 16 (R) 128/64 (R) 2 (S) 2 (S)
SKKU-8 128 (R) .64 (R) 1 (S) 1 (S) 64/32 (R) 16 (R) 2 (S)
KCU-13 64 (R) .64 (R) 1 (S) 1 (S) 64/32 (R) 16 (R) 2 (S)

354 Journal of Medical Microbiology 61

 In vitro combination therapy against A. baumannii

Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 29213,
and Pseudomonas aeruginosa ATCC 27853 were used as control
strains.
Time-kill analysis. Time-kill studies were performed on five
antimicrobial agents (imipenem, colistin, ampicillin-sulbactam, rifam-
picin and tigecycline) and five combinations of these agents (imipenem
plus colistin, imipenem plus ampicillin-sulbactam, colistin plus
rifampicin, colistin plus tigecycline, and tigecycline plus rifampicin)
according to a previously reported method (Petersen et al., 2006).
Time-kill assays were performed in duplicate using concentrations of
0.56 and 16 MIC in both single-agent and combination studies.
Bacterial growth was quantified after 0, 2, 4, 8, 12 and 24 h incubation
at 37 uC by plating 10-fold dilutions on sheep blood agar.
Antimicrobials were considered bactericidal when a ¢3 log10 decrease
in c.f.u. ml21 was reached compared with the initial inocula. Synergy of
the antimicrobial combination was defined as a ¢2 log10 decrease in
c.f.u. ml21 as compared to use of a single agent (Eliopoulos &
Moellering, 1996).
RESULTS
In vitro susceptibilities
The MICs for imipenem, meropenem, colistin, polymy-
xin B, ampicillin-sulbactam, tigecycline and rifampicin of
the six A. baumannii isolates are presented in Table 1.
All isolates were resistant to the carbapenems imipen-
em and meropenem. Additionally, all were resistant to
ciprofloxacin, cefepime, ceftriaxone, cefoperazone-sulbac-
tam, ceftazidime, piperacillin-tazobactam, tetracycline and
ampicillin-sulbactam. While KHU-4 was susceptible to
amikacin (MIC 4 mg l21), the other isolates were resistant
(MICs .128 mg l21).
Single-agent studies
Only imipenem was bactericidal against all six A. baumannii
isolates tested (Table 2); even 0.56 MIC of imipenem
resulted in bactericidal effects against two COL-S/TIG-S
isolates and one COL-R/TIG-S isolate (KCU-4). Colistin
was bactericidal against KCU-4 only (Table 2). Although
16 MIC of colistin initially decreased growth in all A.
baumannii isolates after 2 and 4 h of incubation, regrowth
was observed in five isolates. As observed with the colistin
treatment, ampicillin-sulbactam treatment showed a bacter-
icidal effect on KCU-4 and SKKU-8. Even KCU-4 showed
about 2 log10 regrowth after 24 h of incubation at 16 MIC
of ampicillin-sulbactam, compared to that after 12 h of
incubation with this agent. None of the A. baumannii
isolates used in this study were completely killed by
tigecycline as a single regimen at either 0.56 or 16 MIC.
Rifampicin also had no bactericidal effect against any of the
A. baumannii isolates tested.
Combination studies
Treatment with a combination of 16 MIC imipenem and
colistin exerted bactericidal effects on all six A. baumannii
isolates tested (Table 3, Fig. 1). However, treatment with
0.56 MIC imipenem plus colistin was bactericidal against
only four isolates: the two COL-S/TIG-S isolates, a COL-R/
TIG-S isolate (KCU-4) and a COL-S/TIG-R isolate (SKKU-
8). This combination at 0.56 MIC was not effective
against SKKU-2 and KCU-13.
All A. baumannii isolates were also not detected in
incubations with the combination of 16 MIC imipenem
and ampicillin-sulbactam (Table 3). Compared with 16
imipenem alone, three A. baumannii isolates (KHU-4,
SKKU-2 and KCU-13) were killed earlier by the combina-
tion of 16 imipenem and ampicillin-sulbactam. Imipenem
plus ampicillin-sulbactam at 0.56 MIC displayed bacte-
ricidal activities against all isolates. It is of note that the
combination of 0.56 imipenem and ampicillin-sulbactam
displayed synergistic and bactericidal effects even against
the three isolates (SKKU-2, SKKU-8 and KCU-13) that
were not killed within 24 h by 0.56 imipenem alone.

Table 2. Bactericidal effects of single agents against imipenem-resistant A. baumannii isolates

Resistance* Isolate Bactericidal effectD

Imipenem Colistin Ampicillin- Rifampicin Tigecycline

sulbactam

0.5¾ MIC 1¾ MIC 0.5¾ MIC 1¾ MIC 0.5¾ MIC 1¾ MIC 0.5¾ MIC 1¾ MIC 0.5¾ MIC 1¾ MIC

COL-S/TIG-S KRU-A-3 B B NB NB NB NB NB NB NB NB
KHU-4 B B NB NB NB NB NB NB NB NB
COL-R/TIG-S KCU-4 B B B B NB B NB NB NB NB
SKKU-2 NB B NB NB NB NB NB NB NB NB
COL-S/TIG-R SKKU-8 NB B NB NB NB B NB NB NB NB
KCU-13 NB B NB NB NB NB NB NB NB B

*COL-S, colistin susceptible; COL-R, colistin resistant; TIG-S, tigecycline susceptible; TIG-R, tigecycline resistant.
DB, bactericidal (when ¢3 log10 decrease in c.f.u. ml21 was reached compared with the initial inocula); NB, non-bactericidal.

http://jmm.sgmjournals.org 355
K. R. Peck and others

All tested isolates of bacteria were not detected in

incubations with the combination of 16 MIC colistin and

Tigecycline+Rifampicin

1¾ MIC
rifampicin within 8 h after incubation (Table 3). Although

S
S
S
S
S
S
KRU-A-3 regrew temporarily after 12 h of incubation, it was
eventually eliminated. However, treatment with 0.56 MIC
colistin plus rifampicin was bactericidal against only two
0.5¾ MIC
isolates: KCU-4 and KCU-13. Treatment with a combina-

NS
NS
NS
NS
tion of 16 MIC colistin and tigecycline was also bactericidal

S
S
against all A. baumannii isolates (Table 3, Fig. 2). Treatment
with 0.56 MIC colistin plus tigecycline was bactericidal or
synergistic against only four isolates: one COL-S/TIG-S
1¾ MIC
Colistin+Tigecycline

(KRU-A-3), one COL-R/TIG-S (KCU-4), and the two COL-S/

S
S
S
S
S
S
TIG-R isolates. However, SKKU-8 (a COL-S/TIG-R isolate)
showed regrowth after 12 h of incubation with 0.56 colistin
plus tigecycline.
0.5¾ MIC

Treatment with the combination of tigecycline and rifam-

NS

NS
S
S

S
S

picin was the least effective against the A. baumannii

isolates tested (Table 3). Although all isolates reached
undetectable levels in incubations with 16 tigecycline plus
rifampicin, removal took longer (12–24 h) than for the
1¾ MIC
Colistin+Rifampicin

other combinations. The 0.56 MIC of tigecycline and

Synergistic effectD

S
S
S
S
S
S
Table 3. Synergistic effects of antimicrobial combinations against imipenem-resistant A. baumannii isolates

rifampicin was synergistic against only two COL-S/TIG-R

isolates. Therefore, only two COL-S/TIG-R isolates were
DS, synergistic (when ¢2 log10 decrease in c.f.u. ml21 as compared to use of a single agent); NS, non-synergistic.
*COL-S, colistin-susceptible; COL-R, colistin-resistant; TIG-S, tigecycline-susceptible; TIG-R, tigecycline-resistant.

not detected by the combination of 0.56 tigecycline and

0.5¾ MIC

rifampicin within 24 h of incubation.

NS
NS

NS
NS
S

DISCUSSION
A. baumannii infections have traditionally been treated with
Imipenem+Ampicillin-

1¾ MIC

broad-spectrum cephalosporins, b-lactams and b-lactamase

S
S
S
S
S
S

inhibitors, and carbapenems (Munoz-Price & Weinstein,

sulbactam

2008). However, the emergence of and subsequent increase

in MDR A. baumannii isolates, including carbapenem-
0.5¾ MIC

resistant isolates, have limited the treatment options. Thus,

S
S
S
S
S
S

treatment with polymyxins such as polymyxin B and colistin,

which had previously been abandoned due to problems of
nephrotoxicity and neurotoxicity, is being used for these
infections (Peleg et al., 2008; Li et al., 2006b). In addition,
1¾ MIC
Imipenem+Colistin

tigecycline, a new glycylcycline, has been introduced to

S
S
S
S
S
S

treat MDR Gram-negative bacterial infections including A.

baumannii (Peleg et al., 2008). However, studies have
reported development of resistance to colistin or tigecycline
0.5¾ MIC

during the treatment (Hawley et al., 2008; Li et al., 2006a;

NS
NS
NS

Owen et al., 2007; Peleg et al., 2007). Monotherapy with

S
S
S

colistin may be problematic for the treatment of colistin-

heteroresistant A. baumannii infections (Owen et al., 2007).
Even extreme drug-resistant (XDR) A. baumannii isolates,
KRU-A-3

displaying resistance to all antimicrobials, including

SKKU-2
SKKU-8
KCU-13
Isolate

KHU-4
KCU-4

polymyxins and tigecycline, have emerged (Doi et al., 2009;

Park et al., 2009d). Thus, combination therapy has been
recommended not only to combat MDR A. baumannii
infections but also to inhibit or reduce the emergence of
COL-R/TIG-S

COL-S/TIG-R
COL-S/TIG-S

resistance during treatment.

Resistance*

Not a few studies have been performed on the in vitro

activities of combination therapies against A. baumannii
infections. Tripodi et al. (2007) reported synergistic effects

356 Journal of Medical Microbiology 61

 In vitro combination therapy against A. baumannii

12 KRU-A-3 (COL-S/TIG-S) 12 KHU-4 (COL-S/TIG-S)

10 10

log(c.f.u. ml 1)

log(c.f.u. ml 1)
_

_
8 8
6 6
4 4
2 2

4 8 12 16 20 24 4 8 12 16 20 24
Time (h) Time (h)
12 12
KCU-4 (COL-R/TIG-S) SKKU-2 (COL-R/TIG-S)
10 10
log(c.f.u. ml 1)

log(c.f.u. ml 1)
_

_
8 8
6 6
4 4
2 2

4 8 12 16 20 24 4 8 12 16 20 24
Time (h) Time (h)
12 SKKU-8 (COL-S/TIG-R) 12 KCU-13 (COL-S/TIG-R)
10 10
log(c.f.u. ml 1)

log(c.f.u. ml 1)
_

_
8 8
6 6
4 4
2 2

4 8 12 16 20 24 4 8 12 16 20 24
Time (h) Time (h)

Fig. 1. Effects of imipenem (IMP), colistin (COL), and a combination of these agents on the viability of six imipenem-resistant A.
baumannii isolates. COL-S, colistin-susceptible; COL-R, colistin-resistant; TIG-S, tigecycline-susceptible; TIG-R, tigecycline-
resistant. e, 0.5¾ MIC IMP; h, 0.5¾ MIC COL; #, 0.5¾ MIC IMP+COL; X, 1¾ MIC IMP; &, 1¾ MIC COL; $, 1¾ MIC
IMP+COL. The in vitro time-kill experiments were duplicated; mean values are plotted. In most duplicate experiments, similar
time-kill results were obtained.

for combinations of rifampicin plus imipenem or ampicil-
lin-sulbactam against MDR A. baumannii isolates. Colistin
plus minocycline, tigecycline plus amikacin, carbapenems
(imipenem and meropenem) plus polymyxins (polymyxin
B and colistin), imipenem plus tigecycline, colistin plus
vancomycin, colistin plus teicoplanin, and rifampicin plus
sulbactam have also been reported to have synergistic effects
(Hornsey & Wareham, 2011; Moland et al., 2008; Pachón-
Ibáñez et al., 2006; Gordon et al., 2010; Pankey & Ashcraft,
2009; Sopirala et al., 2010; Tan et al., 2007). In addition,
imipenem plus sulbactam and colistin plus rifampicin
combinations were effective in vitro against carbapenem-
resistant A. baumannii isolates (Song et al., 2007). However,
other in vitro time-kill studies demonstrated that tigecy-
cline is ineffective when used in combination with
polymyxin B, minocycline, imipenem, levofloxacin, ampi-
cillin-sulbactam and rifampicin against carbapenem-non-
susceptible A. baumannii isolates (Moland et al., 2008;
Scheetz et al., 2007). Overall, many in vitro and a few in vivo
time-kill studies have indicated that antibiotic combination
therapy is effective against infections caused by imipenem-
resistant A. baumannii, although the specific regimens that
were shown to be effective differ in each study. To our
knowledge, few studies except that of Vila-Farres et al.
(2011) have been performed on colistin-resistant A.
baumannii isolates.
In the present study, the effectiveness of five antimicrobial
agents singly and in combinations against imipenem-
resistant, colistin-resistant or tigecycline-resistant A.
baumannii blood isolates were evaluated using an in vitro
time-kill analysis. In single-agent studies, only imipenem
exhibited consistent bactericidal activity against all A.
baumannii isolates tested. Although a bacteridical effect of
16 imipenem might be anticipated, even 0.56 MIC
imipenem was bactericidal against some A. baumannii
isolates. The other agents used singly yielded inconsis-
tent results against A. baumannii even at 16 MIC. In
particular, colistin or tigecycline alone was not always
bactericidal even against colistin-susceptible or tigecycline-
susceptible isolates, respectively (Table 2). This suggests
that either colistin or tigecycline monotherapy is not
likely to be effective against infections with carbapenem-
resistant A. baumannii isolates, irrespective of antimicro-
bial resistance.

http://jmm.sgmjournals.org 357
K. R. Peck and others

12 KRU-A-3 (COL-S/TIG-S) 12 KHU-4 (COL-S/TIG-S)

10 10

log(c.f.u. ml 1)

log(c.f.u. ml 1)
_

_
8 8
6 6
4 4
2 2

4 8 12 16 20 24 4 8 12 16 20 24
Time (h) Time (h)
12 KCU-4 (COL-R/TIG-S) 12 SKKU-2 (COL-R/TIG-S)
10 10
log(c.f.u. ml 1)

log(c.f.u. ml 1)
_

_
8 8
6 6
4 4
2 2

4 8 12 16 20 24 4 8 12 16 20 24
Time (h) Time (h)
12 SKKU-8 (COL-S/TIG-R) 12 KCU-13 (COL-S/TIG-R)
10 10
log(c.f.u. ml 1)

log(c.f.u. ml 1)
_

_
8 8
6 6
4 4
2 2

4 8 12 16 20 24 4 8 12 16 20 24
Time (h) Time (h)

Fig. 2. Effects of colistin (COL), tigecycline (TIG) and a combination of these agents on the viability of six imipenem-resistant A.
baumannii isolates. COL-S, colistin-susceptible; COL-R, colistin-resistant; TIG-S, tigecycline-susceptible; TIG-R, tigecycline-
resistant. e, 0.5¾ MIC COL; h, 0.5¾ MIC TIG; #, 0.5¾ MIC COL+TIG; X, 1¾ MIC COL; &, 1¾ MIC TIG; $, 1¾ MIC
COL+TIG. The in vitro time-kill experiments were duplicated; mean values are plotted. In most duplicate experiments, similar
time-kill results were obtained.

In contrast to the single-agent experiments, combination
regimens displayed excellent bactericidal activities. All
imipenem-resistant A. baumannii isolates tested were not
detected in incubations with all five combinations of
antimicrobial agents at 16 MIC. Although treatment
with combinations of antimicrobial agents at 0.56 MIC
was effective against some isolates, the effects were not
consistent. Only 0.56 MIC imipenem plus ampicillin-
sulbactam displayed bactericidal activity against all imipe-
nem-resistant A. baumannii isolates. As both imipenem and
ampicillin-sulbactam target the bacterial cell wall, their
combination would be expected to kill the bacteria more
rapidly, which may not have clinical implications. Excluding
the combination imipenem and ampicillin-sulbactam, 0.56
MIC colistin plus tigecycline was the most effective, showing
synergistic or bactericidal effects against four A. baumannii
isolates. These results may indicate that increasing the
dosage of antimicrobial agents sufficiently is required to
achieve a bactericidal effect against resistant A. baumannii.
However, high dosage of antimicrobial agents may lead to
the further emergence of resistance and may increase the
toxic effects of those agents. Thus, further investigation of
the doses of combination agents that are sufficient to kill
bacteria and to prevent the development of resistance is
required.
Although the imipenem-resistant A. baumannii isolates
examined in this study belonged to different PFGE types,
they have similar imipenem resistance mechanisms, blaOXA-51
and blaOXA-23, and all of them belonged to European clone II,
an internationally disseminated clone (data not shown).
While blaOXA-23 is the main mechanism of imipenem
resistance in A. baumannii, especially in Korea (Park et al.,
2009d; Kim et al., 2010), A. baumannii isolates possessing
blaIMP and blaVIM are also important contributors to
imipenem resistance, but such isolates were not included in
this study. Our study did not analyse the colistin and
tigecycline resistance mechanisms for the A. baumannii
isolates tested. Thus, our results may not be generalizable
to all imipenem-resistant, colistin-resistant or tigecycline-
resistant A. baumannii isolates. However, our data suggest
that some antimicrobial combinations may be effective for
combating imipenem-resistant A. baumannii infections,
including those due to colistin-resistant or tigecycline-
resistant bacteria.

358 Journal of Medical Microbiology 61


ACKNOWLEDGEMENTS
This study was supported by a grant from the Korea Health 21 R&D
Project, Ministry of Health, Welfare, and Family Affairs, Republic of
Korea (grant no. A102065).
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