Eur J Clin Microbiol Infect Dis
DOI 10.1007/s10096-014-2125-8
ARTICLE
Pseudomonas aeruginosa in French hospitals between 2001
and 2011: back to susceptibility
C. Slekovec & J. Robert & D. Trystram & J. M. Delarbre & A. Merens &
N. van der Mee-Marquet & C. de Gialluly & Y. Costa & J. Caillon & D. Hocquet &
X. Bertrand & on behalf of the ONERBA
Received: 20 January 2014 / Accepted: 10 April 2014
# Springer-Verlag Berlin Heidelberg 2014
Abstract The European Antimicrobial Resistance Surveil-
lance Network (EARS-Net) reported an increase in the rates
of resistance of Pseudomonas aeruginosa to antimicrobials
between 2008 and 2011 in France. This alarming report was
based on data collected during the harmonisation of
breakpoints by the European Committee on Antimicrobial
Susceptibility Testing (EUCAST) committee. However, these
data were not supported by the findings of other national
surveillance networks. In this study, we assessed the trends in
C. Slekovec : D. Hocquet : X. Bertrand (*)
Service d’Hygiène Hospitalière, Centre Hospitalier Universitaire de
Besançon, 3, Boulevard Fleming, 25030 Besançon Cedex, France
e-mail: xavier.bertrand@univ-fcomte.fr
C. Slekovec : D. Hocquet : X. Bertrand
UMR 6249 Chrono-Environnement, Université de Franche-Comté,
Besançon, France
J. Robert : D. Trystram
Service de Bactériologie-Hygiène, Hôpitaux Universitaires Pitié
Salpêtrière-Charles Foix, AP-HP, Paris, France
J. M. Delarbre
Service de Bactériologie, Centre Hospitalier de Mulhouse,
Mulhouse, France
A. Merens
Laboratoire de bactériologie, Hôpital d’instruction des armées Bégin,
Saint-Mandé, France
N. van der Mee-Marquet : C. de Gialluly
Laboratoire de bactériologie, Centre Hospitalier Universitaire de
Tours, Tours, France
Y. Costa
Laboratoire de bactériologie, Centre Hospitalier de Lagny-Marne la
vallée, Lagny-sur-Marne, France
J. Caillon
Laboratoire de bactériologie, Centre Hospitalier Universitaire de
Nantes, Nantes, France
P. aeruginosa antimicrobial drug resistance at six French hos-
pitals over a longer period of time (2001–2011) and with a
constant definition of resistance. After the exclusion of incom-
plete data and duplicates, we sorted 34,065 isolates into the
antimicrobial resistance patterns defined by the European Cen-
tre for Disease Prevention and Control (ECDC). The proportion
of isolates with a resistant pattern (non-susceptible to one or two
antimicrobial categories), a multidrug-resistant pattern (non-
susceptible to three or four antimicrobial categories) or an
extensively drug-resistant pattern (non-susceptible to five or
six antimicrobial categories) decreased significantly over time.
Logically, the proportion of isolates with a wild-type resistance
pattern has increased significantly over the same period. No
significant changes in the rates of resistance to cephalosporins
and penicillins were observed, whereas carbapenem resistance
rates increased. By contrast, the proportion of isolates resistant
to fluoroquinolones, aminoglycosides and monobactams de-
creased significantly over time. In conclusion, our data do not
confirm the EARS-net data, suggesting instead that antimicro-
bial drug resistance in P. aeruginosa might not have increased
in French hospitals over the last decade.
Introduction
Pseudomonas aeruginosa is a major cause of infection in
hospitalised patients with localised or systemic immune system
defects [1]. Antimicrobial drug resistance in P. aeruginosa is
driven by the extraordinary capacity of this bacterium to de-
velop resistance to almost any antibiotic, through the selection
of mutations in chromosomal genes and the spread of horizon-
tally acquired resistance [2], and it constitutes a growing threat.
Indeed, increasing numbers of Ambler class A extended-
spectrum β-lactamases (such as TEM, SHV, PER, GES, VEB
and BEL), class B carbapenemases (metallo-β-lactamases such
as IMP, VIM, SPM and GIM) and class D extended-spectrum

oxacillinases (ES-OXA) are being reported in clinical strains of
P. aeruginosa of various geographic origins [3–8]. The cross-
resistance to multiple antibiotic families can also occur in the
absence of foreign gene acquisition, after the mutation of
regulators of broad-specificity multidrug efflux systems. Mul-
tidrug resistance in P. aeruginosa makes it both difficult and
expensive to treat infections, and can increase morbidity and
mortality [1]. The surveillance of antibiotic resistance trends in
this pathogen is therefore crucial. The 2011 European Antimi-
crobial Resistance Surveillance Network (EARS-Net) report
indicated that, despite the high proportions of resistance ob-
served in European P. aeruginosa isolates in 2011, the overall
situation was stable, with few countries reporting a significant
trend towards an increase or decrease in resistance [9]. This
report indicated that, between 2008 and 2011, resistance rates
in France significantly increased for all the antimicrobial agents
under surveillance: ceftazidime, piperacillin/tazobactam, cipro-
floxacin, aminoglycosides and carbapenems, and combined
resistance. These data were collected during the harmonisation
of breakpoints by the European Committee on Antimicrobial
Susceptibility Testing (EUCAST) committee [10]. The EARS-
Net data were obtained from three French networks of the
ONERBA (French observatory of the epidemiology of bacte-
rial resistance to antibiotics). However, the trend towards
higher resistance was not supported by the data for the other
surveillance networks of the ONERBA, which reported overall
stability [11].
We therefore hypothesised that the discrepancies between
the time trends observed by the EARS-Net and by most of the
other ONERBA networks were probably linked to changes in
the definitions of resistance, which were not taken into ac-
count in the trend analysis. We therefore evaluated time trends
in the antimicrobial drug resistance of P. aeruginosa, within a
sample of French hospitals, over a longer period of time
(2001–2011), using a constant definition of resistance based
on current breakpoints [10].
Materials and methods
Study setting
Six French hospitals collaborating with ONERBA participat-
ed in this study: three university-affiliated hospitals
(Besançon, Paris-La Pitié-Salpétrière, Tours), two general
hospitals (Lagny-sur-Marne and Mulhouse) and one hospital
of the Military Health Service (Saint-Mandé). Five of the six
hospitals provided data for the EARS-Net survey.
Collection of data
All P. aeruginosa isolates were obtained from clinical speci-
mens (screening specimens excluded) and identified by
Eur J Clin Microbiol Infect Dis
routine microbiology methods. Antimicrobial drug suscepti-
bility was determined using the agar diffusion method, ac-
cording to the recommendations of the Antibiogram Commit-
tee of the French Microbiology Society (CA-SFM) [12].
Inhibition zone diameters were extracted retrospectively from
laboratory databases for 12 antimicrobial drugs grouped into
seven antimicrobial drug categories according to ECDC rec-
ommendations [13]: aminoglycosides (gentamicin,
tobramycin, amikacin), carbapenems (imipenem,
meropenem), cephalosporins (ceftazidime, cefepime),
fluoroquinolones (ciprofloxacin), penicillins (ticarcillin–
clavulanic acid, piperacillin–tazobactam), monobactam
(aztreonam) and colistin. As determination of colistin resis-
tance using the disk diffusion method is not relevant [14], we
did not take into account the data for colistin resistance.
Annual numbers of patient-days were collected for each hos-
pital, other than Tours Hospital for 2001–2005 (owing to
missing data), for the calculation of incidence density rates.
Data analysis
Isolates were classified as susceptible or non-susceptible to the
panel of antipseudomonal compounds. We applied 2013
EUCAST guidelines [10] for the interpretation of inhibition
zone diameters for all antipseudomonal agents, except for
aztreonam, for which CA-SFM breakpoints were used [12].
For antimicrobial drug categories for which more than one
agent was tested (e.g. imipenem and meropenem for the
carbapenem category), susceptibility to the least effective
compound was used to classify the entire category. Isolates
were then sorted into four antimicrobial resistance patterns,
according to ECDC recommendations [13]. Wild-type isolates
(WT) were susceptible to all agents; resistant (R) isolates were
non-susceptible to one or two categories of antimicrobial
drugs; multidrug-resistant isolates (MDR) were non-
susceptible to three or four categories of antimicrobial drugs
and extensively drug-resistant isolates (XDR) were non-
susceptible to five or six categories of antimicrobial drugs.
Duplicates were defined on the basis of patient identity, iso-
lation time and antimicrobial resistance pattern. We retained
only one isolate per pattern, per patient and per year. We
determined the proportion of non-susceptible isolates for each
antimicrobial class and the crude incidence of antimicrobial
resistance drug patterns per 1,000 patient-days. A subanalysis
was performed with isolates obtained from blood cultures.
Statistical analysis
We used non-parametric Spearman rank tests to analyse the
trends in terms of proportions. The rho (ρ) value (Spearman’s
rank correlation coefficient) obtained in this test lies between
−1 and +1. Values close to −1 or +1 indicate a negative or a
positive correlation respectively. Values close to 0 indicate an
Eur J Clin Microbiol Infect Dis
Fig. 1 Trends in the proportions (%) of the different antibiotic resistance
patterns defined by the European Centre for Disease Prevention and
Control (ECDC) for P. aeruginosa isolated at six French hospitals be-
tween 2001 and 2011 (n=34,065). Dashed grey line wild-type (WT)
absence of correlation. We used a negative binomial regres-
sion test to analyse trends in incidence. The regression coef-
ficient (b) obtained expresses the expected change in the
logarithm of the expected number of counts of the response
variable for a one-unit change in the predictive variable.
Probability of 0.05 or below was considered significant. All
statistical analyses were performed using Stata version 10.0
software (Stata, College Station, TX, USA).
Results
We analysed the resistance data for 58,810 P. aeruginosa
isolates collected during the 11-year surveillance period at
the six participating hospitals. After the exclusion of incom-
plete data and duplicates, 34,065 isolates (2,404 of which
Fig. 2 Trends in the crude incidence (per 1,000 patient-days) of the
different antibiotic resistance patterns defined by the ECDC for
P. aeruginosa isolates from six French hospitals between 2001 and
2011 (n=34,065). Dashed grey line wild-type (WT) resistance pattern,
= 0.944, p<0.001
WT
= -0.874, p<0.001
R
= -0.916, p=0.001
MDR
= -0.601 p=0.039
XDR
resistance pattern, solid grey line resistant (R) pattern, dashed black line
multidrug-resistant (MDR) pattern, solid black line extensively drug-
resistant (XDR) pattern. The ρ value is Spearman’s rank correlation
coefficient
were from blood cultures) were considered for analysis
(2,763 to 3,382 isolates per year).
The proportion of isolates that showed R, MDR and XDR
resistance patterns decreased significantly over time (Fig. 1).
By contrast, the proportion of isolates with a WT resistance
pattern increased significantly over time. The trends of the
incidence of resistance patterns confirmed the trends of the
proportions (Fig. 2). However, incidence trends of the MDR
and XDR patterns did not reach the threshold for statistical
significance. Of note, the global incidence of P. aeruginosa
infections per 1,000 patient-days was stable during the survey
(Spearman’s ρ=0.004, p=0.189). Identical trends were ob-
served for bloodstream isolates of P. aeruginosa (data not
shown).
The proportions of isolates resistant to fluoroquinolones,
aminoglycosides and monobactams decreased significantly
b = 0.038, p<0.001
WT
b = -0.019, p=0.001
b = -0.011, p=0.063
MDR
b = -0.023, p=0.077
XDR
solid grey line resistant (R) pattern, dashed black line multidrug-resistant
(MDR) pattern, solid black line extensively drug-resistant (XDR) pattern.
The regression coefficient (b) was calculated in the negative binomial
regression test

Fig. 3 Trends in the antibiotic
resistance (%) of the different
antibiotics for P. aeruginosa
isolated at six French hospitals
between 2001 and 2011 (n=
34,065). Form the top to the
bottom (proportion in 2011): thin
solid grey line monobactam
resistance proportion, dashed
grey line fluoroquinolone
resistance proportion, dotted
black line carbapenem resistance
proportion, bold solid black line
penicillin resistance proportion,
bold solid grey line
aminoglycoside resistance
proportion, dashed black line
cephalosporin resistance
proportion
over time, whereas the proportion of isolates resistant to
carbapenems increased, and those of isolates resistant to pen-
icillins and cephalosporins remain stable (Fig. 3).
Discussion
Our data for trends in resistance to fluoroquinolones, amino-
glycosides and monobactams are consistent with the reported
decrease in resistance to fluoroquinolone and gentamicin in
Canada [15], the USA [16] and Europe [17]. The decline in
resistance to fluoroquinolones and aminoglycosides between
2001 and 2011 presumably resulted from a decrease in the use
of these compounds observed during this period in French
hospitals [18], but we did not collect data on the antimicrobial
consumption of the six participating hospitals to confirm this
hypothesis.
However, our data are not consistent with those of the
EARS-Net for all antimicrobial classes, with the exception
of carbapenems. This Europe-wide surveillance network re-
ported increasing proportions of P. aeruginosa isolates resis-
tant to all the antimicrobial agents tested (ceftazidime,
piperacillin/tazobactam, ciprofloxacin, aminoglycosides and
carbapenems) and increasing proportions of isolates with
combined resistance, between 2008 and 2011, in France [9].
In the 2011 report, the EARS-Net coordinators pointed out
“that the use of microbiological breakpoints may change over
time, when breakpoint protocols are updated or changed. As
data on quantitative measures (i.e. zone diameters in disk
diffusion tests or MIC values) are not provided by all partic-
ipating laboratories, only the reported S, I, and R results are
considered for the analyses”. The criteria defining susceptibil-
ity to ceftazidime and cefepime in the French recommenda-
tions were updated in 2008 and those for susceptibility to
Eur J Clin Microbiol Infect Dis
ciprofloxacin were updated in 2010. The EARS-Net data
showed an increase in the proportion of isolates non-
susceptible to ceftazidime from 8 % in 2008 to 17 % in
2009. Our results suggest that this increase might be more
likely to reflect a change in the breakpoints used to define
susceptibility than a real change in P. aeruginosa antimicro-
bial drug resistance rates. Moreover, caution is required more
generally when considering the trends reported by the EARS-
Net. For instance, although “trend figures” indicated an in-
crease in resistance to fluoroquinolones, aminoglycosides and
combined resistance, an asterisk was used to indicate “signif-
icant trends in the overall data that were not supported by data
from laboratories consistently reporting for all 4 years”.
In contrast to these discrepancies, the increase in the pro-
portion of isolates resistant to carbapenems that we observed
between 2001 and 2011 is consistent with that reported by the
EARS-Net. This concordance probably reflects the lack of
change in the breakpoints used to interpret EARS-net data for
carbapenems. The growing proportion of isolates displaying
carbapenem resistance undoubtedly reflects the increase in
carbapenem use in French hospitals during this period [18].
Our study is undoubtedly less representative than the
French nationwide EARS-Net survey. However, we had the
advantage of collecting raw resistance data and we were able
apply the same criteria for their interpretation throughout the
period concerned. Moreover, we included approximately
3,000 non-duplicate isolates per year, whereas the EARS-net
surveillance included about 1,000 isolates per year.
In conclusion, our study suggests that, with the exception
of carbapenem resistance, resistance of P. aeruginosa to anti-
microbial drugs is generally decreasing. This observation
contrasts with the large number of reports of the occurrence
[19–22] or international spread of multidrug-resistant
P. aeruginosa, producing extended-spectrum β-lactamases
Eur J Clin Microbiol Infect Dis
(mostly metallo-β-lactamase) [8, 23]. These emerging mech-
anisms did not begin to affect the antimicrobial drug resistance
of P. aeruginosa in France until 2011, but careful monitoring
of resistance trends is required. Furthermore, antibiotic man-
agement policies focusing on carbapenem use should be im-
plemented in French hospitals, particularly in the face of
recent increases in the incidence of multidrug-resistant
Enterobacteriaceae.
Conflicts of interest The authors declare that they have no conflict of
interest.
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