- Email: [email protected]
Emergence of carbapenemases in Gram-negative bacteria in Hamburg, Germany
Available online at www.sciencedirect.com
Diagnostic Microbiology and Infectious Disease 71 (2011) 312 – 315 www.elsevier.com/locate/diagmicrobio
Emergence of carbapenemases in Gram-negative bacteria in Hamburg, Germany Moritz Hentschke a,⁎, Veronika Goritzka a , Cristina Belmar Campos a , Philipp Merkel b , Corina Ilchmann c , Heinrich Lellek d , Stefanie Scherpe a , Martin Aepfelbacher a , Holger Rohde a a
Institute of Medical Microbiology, Virology and Hygiene, University Medical Centre Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany b Asklepios Klinik Nord-Heidberg, Anaesthesiology, Tangstedter Landstrasse 400, 22417 Hamburg, Germany c Evangelisches Amalie Sieveking-Krankenhaus, Internal Medicine, Haselkamp 33, 22359 Hamburg, Germany d Bone Marrow Transplantation Unit, University Medical Centre Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany Received 10 March 2011; accepted 25 July 2011
Abstract We analyzed a collection of carbapenem-resistant Gram-negative bacterial isolates and detected VIM-1, VIM-2, and KPC-2 in diverse enterobacterial species and Pseudomonas aeruginosa isolates. Our findings suggest a more widespread dissemination of carbapenemases in Germany than currently appreciated. © 2011 Elsevier Inc. All rights reserved. Keywords: Carbapenemase; Beta-lactamase; Pseudomonas aeruginosa; Enterobacteriaceae
Carbapenems are a class of antibiotics with a broad spectrum of activity against Gram-negative bacteria and are a mainstay of empirical therapy of life-threatening infections. However, the recent emergence of carbapenemases, beta-lactamases with activity against carbapenems, threatens to compromise the use of this class in antibiotic therapy (Queenan and Bush, 2007). Carbapenemases belong to one of 3 beta-lactamase families, class A, class B, or class D. While class A and class D (OXA-type) carbapenemases typically exhibit activity against all betalactam antibiotics, class B metallo-beta-lactamases spare the monobactams, including aztreonam. Due to increasing worldwide spread of carbapenemases in Gram-negative bacteria, we analyzed recently collected carbapenemresistant enterobacterial and Pseudomonas aeruginosa isolates for the presence of these enzymes. Clinical isolates of Enterobacteriaceae resistant to carbapenems have, until recently, very rarely been detected in our diagnostic laboratory. Between June 2010 and January 2011,
⁎ Corresponding author. Tel.: +49-40-74105-8184; fax: +49-40-741053250. E-mail address: [email protected] (M. Hentschke). 0732-8893/$ – see front matter © 2011 Elsevier Inc. All rights reserved. doi:10.1016/j.diagmicrobio.2011.07.011
we detected 10 isolates that were not susceptible to either one or both of the carbapenems imipenem and meropenem. All isolates were analyzed by previously described polymerase chain reaction (PCR) methods for presence of carbapenemases from the classes KPC, VIM, IMP, OXA-48, SIM, GIM, SPM (Dallenne et al., 2010; Ellington et al., 2007), and NDM-1 (NDM-1: CAATATTATGCACCCGGTCG; NDM-1-R: CCTTGCTGTCCTTGATCAGG). Five of the 10 isolates were positive for PCR screening, 2 Klebsiella pneumoniae isolates (positive for VIM and KPC, respectively), 2 Enterobacter cloacae isolates (both positive for VIM), and 1 Citrobacter freundii strain (VIM positive). The full-length carbapenemase genes were amplified, sequenced (KPC-2-F: ATGTCACTGTATCGCCGTC; KPC-2-R: TTACTGCCCGTTGACGCC; VIM-1-F: TTATGGAGCAGCAACGATGT; VIM-1-R: CAAAAGTCCCGCTCCAACGA), and the respective KPC and VIM subtypes of the carbapenemases were determined (Table 1). One K. pneumoniae (EB4), both E. cloacae (EB1 and EB5) strains, and the C. freundii (EB3) isolate carried a VIM-1 encoding gene, while the second K. pneumoniae (EB2) strain harboured blaKPC-2. Two of the strains (EB4-K. pneumoniae and EB5-E. cloacae) were isolated from the same patient from a peripheral hospital. Both were positive to blaVIM-1,
Table 1 Antibiotic susceptibility, sequence type, and carbapenemase of strains described in this study Organism
Pulso type
Sequence type
Origin
VIM-1
nd
na
KPC-2
nd
ST258
VIM-1
nd
na
VIM-1
nd
ST17
VIM-1
nd
na
VIM-2
1
nd
VIM-2
1
nd
VIM-2
1
ST235
VIM-2
2
nd
VIM-2
2
ST111
VIM-2
2
nd
VIM-2
2
nd
Wound, university hospital Gallbladder, university hospital Urine, university hospital Wound, Peripheral hospital Wound, Peripheral hospital Blood, university hospital Urethra, university hospital Blood, university hospital Urine, university hospital Sputum, university hospital Trachea, university hospital Anal, university hospital
MIC (μg/mL) Imi penem
Mero penem
Erta penem
Dori penem
Tige cycline
Colistin
≥ 32
≥ 32
≥ 32
≥ 32
1
0.5
≥ 32
≥ 32
≥ 32
≥ 32
1
8
≥ 32
8
≥ 32
≥ 32
2
≥ 32
≥ 32
4
≥ 32
Cipro floxacin
MBL Etest
Hodge test
Genta micin
Cefta zidime
Trimethoprim/ sulfamethoxazole
12
2
≥ 256
≥ 32
4 dilution steps
Positive
1
≥ 32
1
≥ 256
≥ 32
Negative
Positive
1
1
2
8
≥ 256
≥ 32
Negative
Positive
≥ 32
0.25
1
≥ 32
1
≥ 256
≥ 32
Negative
Positive
≥ 32
≥ 32
0.5
1
0.032
1
≥ 256
≥ 32
4 dilution steps
Positive
≥ 32
na
≥ 32
na
1
8
≥ 256
16
na
4 dilution steps
nd
≥ 32
≥ 32
na
≥ 32
na
1
≥ 32
≥ 256
16
na
4 dilution steps
nd
≥ 32
≥ 32
na
≥ 32
na
1
8
≥ 256
16
na
4 dilution steps
nd
≥ 32
≥ 32
na
≥ 32
na
1
≥ 32
4
8
na
4 dilution steps
nd
≥ 32
≥ 32
na
≥ 32
na
1
≥ 32
3
16
na
4 dilution steps
nd
≥ 32
≥ 32
na
≥ 32
na
1
≥ 32
8
12
na
4 dilution steps
nd
≥ 32
≥ 32
na
≥ 32
na
1
≥ 32
3
12
na
4 dilution steps
nd
M. Hentschke et al. / Diagnostic Microbiology and Infectious Disease 71 (2011) 312–315
EB1— Enterobacter cloacae EB2— Klebsiella pneumoniae EB3— Citrobacter freundii EB4— Klebsiella pneumoniae EB5— Enterobacter cloacae PA1— Pseudomonas aeruginosa PA2— Pseudomonas aeruginosa PA3— Pseudomonas aeruginosa PA4— Pseudomonas aeruginosa PA5— Pseudomonas aeruginosa PA6— Pseudomonas aeruginosa PA7— Pseudomonas aeruginosa
Enzyme
313
314
M. Hentschke et al. / Diagnostic Microbiology and Infectious Disease 71 (2011) 312–315
which may point towards “in vivo” interspecies transfer of VIM-1 encoding gene. Multilocus sequence typing (MLST) revealed that K. pneumoniae EB2 belongs to sequence type ST258 (http://www.pasteur.fr/recherche/genopole/PF8/mlst/), a well-recognized internationally disseminating clone associated with KPC-2 production (Woodford et al., 2011). In contrast, K. pneumoniae EB4 belongs to ST17, a rarely reported sequence type. All 5 isolates were positive for carbapenemase production in the modified Hodge test, but a combined imipenem/ imipenem-EDTA Etest did not reliably identify strains positive by PCR for metallo-beta-lactamases. The MBL Etest results were negative for VIM-producing K. pneumoniae strain EB4 and C. freundii isolate EB3. All PCRnegative strains were also negative in the modified Hodge test and the MBL Etest. Susceptibility results for imipenem, meropenem, ertapenem, doripenem, and other agents (Table 1) were determined by Etest and interpreted according to EUCAST (http://www. eucast.org). All 5 enterobacterial isolates were either only intermediately susceptible or fully resistant to all tested carbapenems. Furthermore, all isolates were resistant to at least 1 additional class of antibiotics. All strains were fully susceptible to colistin and tigecycline. Carbapenem resistance in P. aeruginosa is very prevalent, but is often associated with porin loss and/or increased efflux (Lister et al., 2009). We therefore also analyzed a collection of 59 imipenem- and/or meropenemresistant P. aeruginosa isolates for the presence of carbapenemases by PCR as described above. The strains were collected between May 2010 and July 2010 with the 2 notable exceptions of PA2 and PA5, which were isolated in January 2008 and December 2009, respectively. VIM-2 was detected in 7 isolates of the 59 P. aeruginosa (primers for amplification of full-length VIM-2: VIM-2-F: AAAGTTATGCCGCACTCACC; VIM-2-R: TGCAACTTCATGTTATGCCG). Analysis by pulsed-field gel electrophoresis (PFGE) demonstrated that the 7 isolates belonged to 2 different pulsotypes—3 strains exhibited the PFGE type 1 and 4 strains the PFGE type 2 (Table 1). One pulsotype was clearly associated with the bone marrow transplantation unit (PFGE type 1), while isolates from the other pulsotype (PFGE type 2) were recovered from diverse wards of the university hospital. MLST was performed on 1 random isolate of each pulsotype. PA3 from PFGE type 1 exhibited sequence type ST235, and PA5 from PFGE type 2 belongs to ST111 (http://pubmlst.org/paeruginosa/). Both sequence types have previously been associated with international spread, production of VIM metallo-betalactamases, and nosocomial infections (Cholley et al., 2011; Woodford et al., 2011). All 7 VIM-2–positive isolates were resistant to most anti-P. antibiotics. The MBL imipenem/imipenem-EDTA Etest result was positive for all 7 isolates. Reporting of carbapenemases in Enterobacteriaceae from Germany has been scarce and generally involved
single isolates (Gottig et al., 2010; Pfeifer et al., 2011; Weile et al., 2007, 2009) or clonal outbreaks (Castanheira et al., 2004; Wendt et al., 2010). Although carbapenem resistance in P. aeruginosa from Germany is not uncommon, carbapenemases have been found very rarely and VIM-2–positive strains have been reported from only 1 individual isolation (Henrichfreise et al., 2005) and 1 outbreak (Valenza et al., 2010). Here, we describe several carbapenemases that render multiple, unrelated Gramnegative bacterial isolates resistant to carbapenems. Surprisingly, these were detected during a relatively short period. Therefore, while Germany is currently believed to have a very low prevalence of carbapenemases, our findings challenge this perception. The perceived low prevalence in the past may have resulted from inadequate detection or underreporting. The genetic basis of carbapenem resistance in the carbapenemase-negative enterobacterial isolates currently remains unclear. A combination of membrane impermeability and expression of ESBL or AmpC-beta-lactamases may be an underlying mechanism. Yet, the recently described NDM-1 pandemic, which had remained undetected in Germany for several years (Gottig et al., 2010), highlights the importance of this analysis. References Castanheira M, Toleman MA, Jones RN, Schmidt FJ, Walsh TR (2004) Molecular characterization of a beta-lactamase gene, blaGIM-1, encoding a new subclass of metallo-beta-lactamase. Antimicrob Agents Chemother 48:4654–4661. Cholley P, Thouverez M, Hocquet D, van der Mee-Marquet N, Talon D, Bertrand X (2011) The majority of multi-drug resistant Pseudomonas aeruginosa isolates from hospitals in eastern France belongs to a few clonal types. J Clin Microbiol May 18. [Epub ahead of print]. Dallenne C, Da Costa A, Decre D, Favier C, Arlet G (2010) Development of a set of multiplex PCR assays for the detection of genes encoding important beta-lactamases in Enterobacteriaceae. J Antimicrob Chemother 65:490–495. Ellington MJ, Kistler J, Livermore DM, Woodford N (2007) Multiplex PCR for rapid detection of genes encoding acquired metallo-beta-lactamases. J Antimicrob Chemother 59:321–322. Gottig S, Pfeifer Y, Wichelhaus TA, Zacharowski K, Bingold T, Averhoff B, Brandt C, Kempf VA (2010) Global spread of New Delhi metallobeta-lactamase 1. Lancet Infect Dis 10:828–829. Henrichfreise B, Wiegand I, Sherwood KJ, Wiedemann B (2005) Detection of VIM-2 metallo-beta-lactamase in Pseudomonas aeruginosa from Germany. Antimicrob Agents Chemother 49:1668–1669. Lister PD, Wolter DJ, Hanson ND (2009) Antibacterial-resistant Pseudomonas aeruginosa: clinical impact and complex regulation of chromosomally encoded resistance mechanisms. Clin Microbiol Rev 22: 582–610. Pfeifer Y, Witte W, Holfelder M, Busch J, Nordmann P, Poirel L (2011) NDM-1-producing Escherichia coli in Germany. Antimicrob Agents Chemother 55:1318–1319. Queenan AM, Bush K (2007) Carbapenemases: the versatile betalactamases. Clin Microbiol Rev 20:440–458. Valenza G, Joseph B, Elias J, Claus H, Oesterlein A, Engelhardt K, Turnwald D, Frosch M, Abele-Horn M, Schoen C (2010) First survey of metallo-beta-lactamases in clinical isolates of Pseudomonas aeruginosa in a German university hospital. Antimicrob Agents Chemother 54:3493–3497.
M. Hentschke et al. / Diagnostic Microbiology and Infectious Disease 71 (2011) 312–315 Weile J, Ohler S, Schonthal S, Knabbe C (2009) A VIM-1 metallo-betalactamase-producing Klebsiella pneumoniae clinical isolate in an acute hospital in Germany. Int J Antimicrob Agents 33:389–391. Weile J, Rahmig H, Gfroer S, Schroeppel K, Knabbe C, Susa M (2007) First detection of a VIM-1 metallo-beta-lactamase in a carbapenem-resistant Citrobacter freundii clinical isolate in an acute hospital in Germany. Scand J Infect Dis 39:264–266.
315
Wendt C, Schutt S, Dalpke AH, Konrad M, Mieth M, Trierweiler-Hauke B, Weigand MA, Zimmermann S, Biehler K, Jonas D (2010) First outbreak of Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae in Germany. Eur J Clin Microbiol Infect Dis 29:563–570. Woodford N, Turton JF, Livermore DM (2011) Multiresistant Gramnegative bacteria: the role of high-risk clones in the dissemination of antibiotic resistance. FEMS Microbiol Rev Feb 9. [Epub ahead of print].
Diagnostic Microbiology and Infectious Disease 71 (2011) 312 – 315 www.elsevier.com/locate/diagmicrobio
Emergence of carbapenemases in Gram-negative bacteria in Hamburg, Germany Moritz Hentschke a,⁎, Veronika Goritzka a , Cristina Belmar Campos a , Philipp Merkel b , Corina Ilchmann c , Heinrich Lellek d , Stefanie Scherpe a , Martin Aepfelbacher a , Holger Rohde a a
Institute of Medical Microbiology, Virology and Hygiene, University Medical Centre Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany b Asklepios Klinik Nord-Heidberg, Anaesthesiology, Tangstedter Landstrasse 400, 22417 Hamburg, Germany c Evangelisches Amalie Sieveking-Krankenhaus, Internal Medicine, Haselkamp 33, 22359 Hamburg, Germany d Bone Marrow Transplantation Unit, University Medical Centre Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany Received 10 March 2011; accepted 25 July 2011
Abstract We analyzed a collection of carbapenem-resistant Gram-negative bacterial isolates and detected VIM-1, VIM-2, and KPC-2 in diverse enterobacterial species and Pseudomonas aeruginosa isolates. Our findings suggest a more widespread dissemination of carbapenemases in Germany than currently appreciated. © 2011 Elsevier Inc. All rights reserved. Keywords: Carbapenemase; Beta-lactamase; Pseudomonas aeruginosa; Enterobacteriaceae
Carbapenems are a class of antibiotics with a broad spectrum of activity against Gram-negative bacteria and are a mainstay of empirical therapy of life-threatening infections. However, the recent emergence of carbapenemases, beta-lactamases with activity against carbapenems, threatens to compromise the use of this class in antibiotic therapy (Queenan and Bush, 2007). Carbapenemases belong to one of 3 beta-lactamase families, class A, class B, or class D. While class A and class D (OXA-type) carbapenemases typically exhibit activity against all betalactam antibiotics, class B metallo-beta-lactamases spare the monobactams, including aztreonam. Due to increasing worldwide spread of carbapenemases in Gram-negative bacteria, we analyzed recently collected carbapenemresistant enterobacterial and Pseudomonas aeruginosa isolates for the presence of these enzymes. Clinical isolates of Enterobacteriaceae resistant to carbapenems have, until recently, very rarely been detected in our diagnostic laboratory. Between June 2010 and January 2011,
⁎ Corresponding author. Tel.: +49-40-74105-8184; fax: +49-40-741053250. E-mail address: [email protected] (M. Hentschke). 0732-8893/$ – see front matter © 2011 Elsevier Inc. All rights reserved. doi:10.1016/j.diagmicrobio.2011.07.011
we detected 10 isolates that were not susceptible to either one or both of the carbapenems imipenem and meropenem. All isolates were analyzed by previously described polymerase chain reaction (PCR) methods for presence of carbapenemases from the classes KPC, VIM, IMP, OXA-48, SIM, GIM, SPM (Dallenne et al., 2010; Ellington et al., 2007), and NDM-1 (NDM-1: CAATATTATGCACCCGGTCG; NDM-1-R: CCTTGCTGTCCTTGATCAGG). Five of the 10 isolates were positive for PCR screening, 2 Klebsiella pneumoniae isolates (positive for VIM and KPC, respectively), 2 Enterobacter cloacae isolates (both positive for VIM), and 1 Citrobacter freundii strain (VIM positive). The full-length carbapenemase genes were amplified, sequenced (KPC-2-F: ATGTCACTGTATCGCCGTC; KPC-2-R: TTACTGCCCGTTGACGCC; VIM-1-F: TTATGGAGCAGCAACGATGT; VIM-1-R: CAAAAGTCCCGCTCCAACGA), and the respective KPC and VIM subtypes of the carbapenemases were determined (Table 1). One K. pneumoniae (EB4), both E. cloacae (EB1 and EB5) strains, and the C. freundii (EB3) isolate carried a VIM-1 encoding gene, while the second K. pneumoniae (EB2) strain harboured blaKPC-2. Two of the strains (EB4-K. pneumoniae and EB5-E. cloacae) were isolated from the same patient from a peripheral hospital. Both were positive to blaVIM-1,
Table 1 Antibiotic susceptibility, sequence type, and carbapenemase of strains described in this study Organism
Pulso type
Sequence type
Origin
VIM-1
nd
na
KPC-2
nd
ST258
VIM-1
nd
na
VIM-1
nd
ST17
VIM-1
nd
na
VIM-2
1
nd
VIM-2
1
nd
VIM-2
1
ST235
VIM-2
2
nd
VIM-2
2
ST111
VIM-2
2
nd
VIM-2
2
nd
Wound, university hospital Gallbladder, university hospital Urine, university hospital Wound, Peripheral hospital Wound, Peripheral hospital Blood, university hospital Urethra, university hospital Blood, university hospital Urine, university hospital Sputum, university hospital Trachea, university hospital Anal, university hospital
MIC (μg/mL) Imi penem
Mero penem
Erta penem
Dori penem
Tige cycline
Colistin
≥ 32
≥ 32
≥ 32
≥ 32
1
0.5
≥ 32
≥ 32
≥ 32
≥ 32
1
8
≥ 32
8
≥ 32
≥ 32
2
≥ 32
≥ 32
4
≥ 32
Cipro floxacin
MBL Etest
Hodge test
Genta micin
Cefta zidime
Trimethoprim/ sulfamethoxazole
12
2
≥ 256
≥ 32
4 dilution steps
Positive
1
≥ 32
1
≥ 256
≥ 32
Negative
Positive
1
1
2
8
≥ 256
≥ 32
Negative
Positive
≥ 32
0.25
1
≥ 32
1
≥ 256
≥ 32
Negative
Positive
≥ 32
≥ 32
0.5
1
0.032
1
≥ 256
≥ 32
4 dilution steps
Positive
≥ 32
na
≥ 32
na
1
8
≥ 256
16
na
4 dilution steps
nd
≥ 32
≥ 32
na
≥ 32
na
1
≥ 32
≥ 256
16
na
4 dilution steps
nd
≥ 32
≥ 32
na
≥ 32
na
1
8
≥ 256
16
na
4 dilution steps
nd
≥ 32
≥ 32
na
≥ 32
na
1
≥ 32
4
8
na
4 dilution steps
nd
≥ 32
≥ 32
na
≥ 32
na
1
≥ 32
3
16
na
4 dilution steps
nd
≥ 32
≥ 32
na
≥ 32
na
1
≥ 32
8
12
na
4 dilution steps
nd
≥ 32
≥ 32
na
≥ 32
na
1
≥ 32
3
12
na
4 dilution steps
nd
M. Hentschke et al. / Diagnostic Microbiology and Infectious Disease 71 (2011) 312–315
EB1— Enterobacter cloacae EB2— Klebsiella pneumoniae EB3— Citrobacter freundii EB4— Klebsiella pneumoniae EB5— Enterobacter cloacae PA1— Pseudomonas aeruginosa PA2— Pseudomonas aeruginosa PA3— Pseudomonas aeruginosa PA4— Pseudomonas aeruginosa PA5— Pseudomonas aeruginosa PA6— Pseudomonas aeruginosa PA7— Pseudomonas aeruginosa
Enzyme
313
314
M. Hentschke et al. / Diagnostic Microbiology and Infectious Disease 71 (2011) 312–315
which may point towards “in vivo” interspecies transfer of VIM-1 encoding gene. Multilocus sequence typing (MLST) revealed that K. pneumoniae EB2 belongs to sequence type ST258 (http://www.pasteur.fr/recherche/genopole/PF8/mlst/), a well-recognized internationally disseminating clone associated with KPC-2 production (Woodford et al., 2011). In contrast, K. pneumoniae EB4 belongs to ST17, a rarely reported sequence type. All 5 isolates were positive for carbapenemase production in the modified Hodge test, but a combined imipenem/ imipenem-EDTA Etest did not reliably identify strains positive by PCR for metallo-beta-lactamases. The MBL Etest results were negative for VIM-producing K. pneumoniae strain EB4 and C. freundii isolate EB3. All PCRnegative strains were also negative in the modified Hodge test and the MBL Etest. Susceptibility results for imipenem, meropenem, ertapenem, doripenem, and other agents (Table 1) were determined by Etest and interpreted according to EUCAST (http://www. eucast.org). All 5 enterobacterial isolates were either only intermediately susceptible or fully resistant to all tested carbapenems. Furthermore, all isolates were resistant to at least 1 additional class of antibiotics. All strains were fully susceptible to colistin and tigecycline. Carbapenem resistance in P. aeruginosa is very prevalent, but is often associated with porin loss and/or increased efflux (Lister et al., 2009). We therefore also analyzed a collection of 59 imipenem- and/or meropenemresistant P. aeruginosa isolates for the presence of carbapenemases by PCR as described above. The strains were collected between May 2010 and July 2010 with the 2 notable exceptions of PA2 and PA5, which were isolated in January 2008 and December 2009, respectively. VIM-2 was detected in 7 isolates of the 59 P. aeruginosa (primers for amplification of full-length VIM-2: VIM-2-F: AAAGTTATGCCGCACTCACC; VIM-2-R: TGCAACTTCATGTTATGCCG). Analysis by pulsed-field gel electrophoresis (PFGE) demonstrated that the 7 isolates belonged to 2 different pulsotypes—3 strains exhibited the PFGE type 1 and 4 strains the PFGE type 2 (Table 1). One pulsotype was clearly associated with the bone marrow transplantation unit (PFGE type 1), while isolates from the other pulsotype (PFGE type 2) were recovered from diverse wards of the university hospital. MLST was performed on 1 random isolate of each pulsotype. PA3 from PFGE type 1 exhibited sequence type ST235, and PA5 from PFGE type 2 belongs to ST111 (http://pubmlst.org/paeruginosa/). Both sequence types have previously been associated with international spread, production of VIM metallo-betalactamases, and nosocomial infections (Cholley et al., 2011; Woodford et al., 2011). All 7 VIM-2–positive isolates were resistant to most anti-P. antibiotics. The MBL imipenem/imipenem-EDTA Etest result was positive for all 7 isolates. Reporting of carbapenemases in Enterobacteriaceae from Germany has been scarce and generally involved
single isolates (Gottig et al., 2010; Pfeifer et al., 2011; Weile et al., 2007, 2009) or clonal outbreaks (Castanheira et al., 2004; Wendt et al., 2010). Although carbapenem resistance in P. aeruginosa from Germany is not uncommon, carbapenemases have been found very rarely and VIM-2–positive strains have been reported from only 1 individual isolation (Henrichfreise et al., 2005) and 1 outbreak (Valenza et al., 2010). Here, we describe several carbapenemases that render multiple, unrelated Gramnegative bacterial isolates resistant to carbapenems. Surprisingly, these were detected during a relatively short period. Therefore, while Germany is currently believed to have a very low prevalence of carbapenemases, our findings challenge this perception. The perceived low prevalence in the past may have resulted from inadequate detection or underreporting. The genetic basis of carbapenem resistance in the carbapenemase-negative enterobacterial isolates currently remains unclear. A combination of membrane impermeability and expression of ESBL or AmpC-beta-lactamases may be an underlying mechanism. Yet, the recently described NDM-1 pandemic, which had remained undetected in Germany for several years (Gottig et al., 2010), highlights the importance of this analysis. References Castanheira M, Toleman MA, Jones RN, Schmidt FJ, Walsh TR (2004) Molecular characterization of a beta-lactamase gene, blaGIM-1, encoding a new subclass of metallo-beta-lactamase. Antimicrob Agents Chemother 48:4654–4661. Cholley P, Thouverez M, Hocquet D, van der Mee-Marquet N, Talon D, Bertrand X (2011) The majority of multi-drug resistant Pseudomonas aeruginosa isolates from hospitals in eastern France belongs to a few clonal types. J Clin Microbiol May 18. [Epub ahead of print]. Dallenne C, Da Costa A, Decre D, Favier C, Arlet G (2010) Development of a set of multiplex PCR assays for the detection of genes encoding important beta-lactamases in Enterobacteriaceae. J Antimicrob Chemother 65:490–495. Ellington MJ, Kistler J, Livermore DM, Woodford N (2007) Multiplex PCR for rapid detection of genes encoding acquired metallo-beta-lactamases. J Antimicrob Chemother 59:321–322. Gottig S, Pfeifer Y, Wichelhaus TA, Zacharowski K, Bingold T, Averhoff B, Brandt C, Kempf VA (2010) Global spread of New Delhi metallobeta-lactamase 1. Lancet Infect Dis 10:828–829. Henrichfreise B, Wiegand I, Sherwood KJ, Wiedemann B (2005) Detection of VIM-2 metallo-beta-lactamase in Pseudomonas aeruginosa from Germany. Antimicrob Agents Chemother 49:1668–1669. Lister PD, Wolter DJ, Hanson ND (2009) Antibacterial-resistant Pseudomonas aeruginosa: clinical impact and complex regulation of chromosomally encoded resistance mechanisms. Clin Microbiol Rev 22: 582–610. Pfeifer Y, Witte W, Holfelder M, Busch J, Nordmann P, Poirel L (2011) NDM-1-producing Escherichia coli in Germany. Antimicrob Agents Chemother 55:1318–1319. Queenan AM, Bush K (2007) Carbapenemases: the versatile betalactamases. Clin Microbiol Rev 20:440–458. Valenza G, Joseph B, Elias J, Claus H, Oesterlein A, Engelhardt K, Turnwald D, Frosch M, Abele-Horn M, Schoen C (2010) First survey of metallo-beta-lactamases in clinical isolates of Pseudomonas aeruginosa in a German university hospital. Antimicrob Agents Chemother 54:3493–3497.
M. Hentschke et al. / Diagnostic Microbiology and Infectious Disease 71 (2011) 312–315 Weile J, Ohler S, Schonthal S, Knabbe C (2009) A VIM-1 metallo-betalactamase-producing Klebsiella pneumoniae clinical isolate in an acute hospital in Germany. Int J Antimicrob Agents 33:389–391. Weile J, Rahmig H, Gfroer S, Schroeppel K, Knabbe C, Susa M (2007) First detection of a VIM-1 metallo-beta-lactamase in a carbapenem-resistant Citrobacter freundii clinical isolate in an acute hospital in Germany. Scand J Infect Dis 39:264–266.
315
Wendt C, Schutt S, Dalpke AH, Konrad M, Mieth M, Trierweiler-Hauke B, Weigand MA, Zimmermann S, Biehler K, Jonas D (2010) First outbreak of Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae in Germany. Eur J Clin Microbiol Infect Dis 29:563–570. Woodford N, Turton JF, Livermore DM (2011) Multiresistant Gramnegative bacteria: the role of high-risk clones in the dissemination of antibiotic resistance. FEMS Microbiol Rev Feb 9. [Epub ahead of print].