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Acquisition of extensive drug-resistant Pseudomonas aeruginosa among hospitalized patients: risk factors and resistance mechanisms to carbapenems
Journal of Hospital Infection 79 (2011) 54e58
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Acquisition of extensive drug-resistant Pseudomonas aeruginosa among hospitalized patients: risk factors and resistance mechanisms to carbapenems Y.S. Park a, y, H. Lee b, y, B.S. Chin c, S.H. Han c, S.G. Hong d, S.K. Hong e, H.Y. Kim f, Y. Uh g, H.B. Shin h, E.J. Choo i, S.-H. Han j, W. Song k, S.H. Jeong l, K. Lee l, J.M. Kim c, * a
Department of Internal Medicine, Gachon University Gil Hospital, Incheon, Republic of Korea Department of Laboratory Medicine, Kwandong University College of Medicine, Goyang, Republic of Korea Department of Internal Medicine, Yonsei University College of Medicine, Seoul, Republic of Korea d Department of Laboratory Medicine, Pochon Cha University College of Medicine, Seongnam, Republic of Korea e Department of Internal Medicine, Pochon Cha University College of Medicine, Seongnam, Republic of Korea f Division of Infectious Diseases, Wonju Christian Hospital, Yonsei University Wonju College of Medicine, Wonju, Republic of Korea g Department of Laboratory Medicine, Wonju Christian Hospital, Yonsei University Wonju College of Medicine, Wonju, Republic of Korea h Department of Laboratory Medicine, Soonchunhyang University College of Medicine, Cheonan, Republic of Korea i Department of Internal Medicine, Soonchunhyang University College of Medicine, Cheonan, Republic of Korea j Infection Control Unit, Soonchunhyang University Hospital, Bucheon, Republic of Korea k Department of Laboratory Medicine, Hallym University and Kangnam Sacred Heart Hospital, Seoul, Republic of Korea l Department of Laboratory Medicine and Research Institute of Bacterial Resistance, Yonsei University College of Medicine, Seoul, Republic of Korea b c
a r t i c l e i n f o
s u m m a r y
Article history: Received 24 December 2010 Accepted 3 May 2011 by J.A. Child Available online 20 July 2011
Extensive drug-resistant Pseudomonas aeruginosa (XDRPA) strains, defined as resistant to all available antipseudomonal antibiotics, have been reported recently. This study aimed to investigate the risk factors for XDRPA acquisition by patients and the resistance mechanisms to carbapenems. From June to November 2007, XDRPA isolates were collected from patients in eight tertiary care hospitals. A caseecontrol study was performed to determine factors associated with XDRPA acquisition. EDTA-imipenem disc synergy tests, and polymerase chain reaction amplification and sequencing were performed to detect the presence of metallo-blactamases (MBLs). Risk factor analysis was performed for 33 patients. Mechanical ventilation [odds ratio (OR) 8.2, 95% confidence interval (CI) 1.3e52.2; P ¼ 0.026] and APACHE II score (OR 1.2, 95% CI 1.0e1.3; P ¼ 0.007) were identified as independent risk factors for XDRPA acquisition. Pulsed-field gel electrophoresis of XDRPA identified clonal epidemic isolates co-existing with sporadic isolates. Eight of 43 (19%) XDRPA isolates were shown to produce MBLs; four produced VIM-2 and four produced IMP-6. This study suggests a major role for mechanical ventilation in XDRPA acquisition. Moreover, pulsed-field gel electrophoresis identified a clonal epidemic within hospitals. Taken together, these results suggest that patient-to-patient transmission contributes to XDRPA acquisition in Korea. Ó 2011 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.
Keywords: Carbapenem resistance Extensive drug resistance Pseudomonas aeruginosa Risk factor
Introduction Pseudomonas aeruginosa is one of the main organisms responsible for drug-resistant nosocomial infections, and was the most commonly reported Gram-negative pathogen of nosocomial * Corresponding author. Address: Department of Internal Medicine, Yonsei University College of Medicine, 250 Seongsanno, Seodaemun-gu, Seoul 120-752, Republic of Korea. Tel.: þ82 2 2228 1946; fax: þ82 2 393 6884. E-mail address: [email protected] (J.M. Kim). y These authors contributed equally to this study.
pneumonia from 1987 to 2003.1 The National Nosocomial Infection Surveillance System has reported a significantly increasing trend of resistance.1 Over the past decade, strains with resistance to almost all available antibiotics have emerged, causing major therapeutic problems. Multi-drug-resistant (MDR) strains of P. aeruginosa were initially reported in patients with cystic fibrosis, and numerous cases have subsequently been reported.2 Prevention of the acquisition of MDR P. aeruginosa is essential due to limited therapeutic options and increased mortality.3,4
0195-6701/$ e see front matter Ó 2011 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.jhin.2011.05.014
Y.S. Park et al. / Journal of Hospital Infection 79 (2011) 54e58
Various definitions of antimicrobial resistance have been used in publications dealing with infections caused by Acinetobacter spp. or P. aeruginosa. Multi-drug resistance in P. aeruginosa or Acinetobacter spp. has been variously defined.5 The definition of pandrug resistance is also controversial due to colistin, a ‘rediscovered’ category of antimicrobial agent, and tigecycline, a new drug. Recently, it was proposed that the term ‘extensive drug resistance’ should be used to indicate resistance to all but one or two classes of antimicrobial agents.6 Risk factors for the acquisition of extensive drug-resistant P. aeruginosa (XDRPA) have not been well documented to date. This study aimed to clarify the risk factors for XDRPA acquisition in Korean hospitals, and to document resistance mechanisms to carbapenems. Controls were selected based on improved epidemiological methodology, enabling the assessment of risk factors without the bias of antibiotic usage.7e9 Materials and methods Bacterial strains and antimicrobial susceptibility testing From June to November 2007, XDRPA isolates were collected from clinical specimens at eight tertiary care hospitals (Severance Hospital of Yonsei University College of Medicine, Seoul; Kangnam Sacred Heart Hospital of Hallym University Medical Centre, Seoul; Korea University Anam Hospital, Seoul; Myongji Hospital of Kwandong University College of Medicine, Goyang; Bundang Cha Hospital of Pochon CHA University College of Medicine, Seongnam; Soonchunhyang University Hospital, Bucheon; Yonsei University Wonju Christian Hospital, Wonju; Kosin University Gospel Hospital, Busan). P. aeruginosa isolates that showed resistance or intermediate susceptibility to piperacillin, piperacillin-tazobactam, ceftazidime, cefepime, aztreonam, imipenem, meropenem, amikacin, gentamicin and ciprofloxacin were considered to be XDRPA, and included in the study. Isolates with resistance to colistin were excluded. The minimum inhibitory concentrations (MICs) of antimicrobial agents were determined by agar dilution.10 Caseecontrol study A caseecontrol study was performed to determine factors associated with XDRPA acquisition. The cases were patients from whom XDRPA were isolated in clinical cultures. During the study period, the electronic microbiology database was searched to identify all XDRPA-positive clinical samples obtained from inpatients. Patients who had been hospitalized for <48 h when the sample was collected and those aged 18 years were excluded. The controls were randomly selected adult inpatients in the participating hospitals who did not have XDRPA isolated during their hospital stay. Controls were matched to cases by hospital. Two controls were recruited for each case. Computerized medical, pharmacy and microbiological records were reviewed. Variables investigated as potential risk factors included age, sex, Charlson score,11 length of hospital stay before outcome of interest (time at risk; for case patients, from admission to nosocomial isolation of XDRPA; for controls, complete length of hospital stay), intensive care unit (ICU) stay, surgery, and severity of illness at the time of infection/colonization [calculated by Acute Physiology and Chronic Health Evaluation (APACHE) II score]. The following associated diseases and comorbid conditions were documented: diabetes mellitus, cardiac disease, pulmonary disease, renal disease (creatinine >2.0 mg/dL or on dialysis), neurological disease, malignancy, use of devices before XDRPA isolation (central venous and/or arterial catheter, urinary catheter, mechanical ventilation and nasogastric tube), and exposure to antimicrobials before XDRPA isolation.
55
Detection of metallo-b-lactamases and sequencing Carbapenemase and metallo-b-lactamase (MBL) production were determined by the modified Hodge test and the double-disc synergy test.12 Polymerase chain reaction (PCR) detection of the blaIMP- and blaVIM-like alleles was performed using IMP-1 and VIM2 primers.13 To determine the type of MBL genes, isolates with blaIMP- and blaVIM-like alleles were used for sequencing. INT-1-F, INT-2-R, IMP-1-F, IMP-1-R, VIM-2-F and VIM-2-R primers were used for blaIMP- and blaVIM-like allele sequencing. The primers used were: IMP-1-F (5’-CAT GGT TTG GTG GTT CTT GT-3’) and IMP-1-R (5’-ATA ATT TGG CGG ACT TTG GC-3’); VIM-2-F (5’-ATG TTC AAA CTT TTG AGT AAG-3’) and VIM-2-R (5’-CTA CTC AAC GAC TGA GCG-3’); and INT-1-F (5’-GGC ATC CAA GCA GCA AG-3’) and INT2-R (5’-AAG CAG ACT TGA CCT GA-3’). The PCR products were subjected to direct sequencing. Both strands of the PCR products were sequenced twice with an automatic sequencer (Model 3730xl; Applied Biosystems, Weiterstadt, Germany). Sequence analysis and comparisons were performed using programs available at the National Center for Biotechnology Information (http://www.ncbi. nlm.nih.gov/). Pulsed-field gel electrophoresis For pulsed-field gel electrophoresis (PFGE) analysis, Xba I-digested genomic DNA of XDRPA was separated using a CHEF-DR II System (Bio-Rad, Hercules, CA, USA).14 The pattern was analysed with Fingerprinting II software (Bio-Rad). Statistical analysis Statistical analyses were performed using Statistical Package for the Social Sciences Version 15 (SPSS Inc., Chicago, IL, USA). Bivariate analyses were performed separately for each of the variables. P-values were calculated using Fisher’s exact test for categorical variables, and Student’s t-test or ManneWhitney U-test for continuous variables. Variables for which the P-value was <0.05 in the bivariate analysis were included in the multi-variate analysis. A forward selection process was used. Risk factors were checked for colinearity by viewing changes in standard errors of multi-variate models. All tests were two-tailed, and a P-value of <0.05 was considered to be significant in the multi-variate model. Results The mechanism of carbapenem resistance was analysed using 43 XDRPA isolates, and full medical records were available for 37 of these patients. Positive cultures within two days of admission necessitated the exclusion of four patients with XDRPA from the risk factor analysis. In total, 33 patients were included in the risk factor analysis. Risk factor analysis Of 33 patients with XDRPA acquisition, 26 were men and seven were women. Their mean age was 65 years (standard deviation 17 years). The most common site from which XDRPA was recovered was the respiratory system (N ¼ 13, 39%), followed by the urinary system (N ¼ 12, 36%). Sixteen patients were receiving care in general wards and 17 were in ICUs. The results of bivariate analyses for baseline demographic and clinical characteristics, associated diseases or comorbid conditions, Charlson score, ICU stay, surgery, time at risk, severity of illness (as calculated by the APACHE II score) and receipt of specific antibiotics are presented in Table I.
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Y.S. Park et al. / Journal of Hospital Infection 79 (2011) 54e58
Table I Bivariate analysis of risk factors for the acquisition of Pseudomonas aeruginosa Risk factor Age in years, mean SD Male sex Associated disease Diabetes mellitus Cardiac disease Pulmonary disease Renal disease Neurological disease Malignancy Charlson comorbidity scale, median (IQR) Related to hospitalization Days at risk, median (IQR) Intensive care unit stay in days, median (IQR) Surgery Device Central venous/arterial catheter Urinary catheter Mechanical ventilation Nasogastric tube Antimicrobial Cephalosporin First generation Second generation Third generation Penicillin with b-lactamase inhibitor Quinolone Aminoglycoside Carbapenem Glycopeptide Septic shock/severe sepsis APACHE II score, median (IQR)
Cases (N ¼ 33)
Controls (N ¼ 66)
65 12 26 (79)
56 19 30 (46)
11 (33) 7 (21) 13 (39) 6 (18) 14 (42) 11 (33) 3 (2e6)
15 (23) 17 (26) 5 (8) 3 (5) 7 (11) 22 (33) 1 (0e3)
OR (95% CI) 4.5 (1.7e11.7) 1.7 0.8 7.9 4.7 6.2 1.0
P-value 0.006 0.002
(0.7e4.3) 0.33 (0.3e2.1) 0.8 (2.5e25.0) <0.001 (1.1e20.0) 0.057 (2.2e17.6) 0.001 (0.4e2.4) 1 0.003
<0.001 <0.001
31 (11e65) 10 (5e21) 9 (3e26) 0 (0e0) 14 (42)
20 (31)
1.7 (0.7e4.0)
19 (58)
12 (18)
6.1 (2.4e15.5) <0.001
24 (73) 15 (46) 16 (49)
19 (29) 3 (4.5) 7 (11)
6.6 (2.6e16.8) <0.001 17.5 (4.6e67.2) <0.001 7.9 (2.8e22.4) <0.001
5 (15) 10 (30) 22 (67) 11 (33)
7 (11) 13 (20) 37 (56) 4 (6)
17 (52) 15 (46) 9 (27) 13 (39) 7 (21)
11 (17) 19 (29) 2 (3) 7 (11) 3 (5)
14 (10e21)
1.5 1.7 1.6 7.8
0.27
(0.4e5.2) (0.7e4.6) (0.7e3.7) (2.2e26.9)
0.52 0.17 0.39 0.001
5.3 (2.1e13.6) 2.1 (0.9e4.9) 12.0 (2.4e59.6) 5.5 (1.9e15.6) 5.6 (1.3e23.2)
0.001 0.12 0.001 0.001 0.03
6 (2e8)
<0.001
Data are N (%) of patients, unless otherwise indicated. OR, odds ratio; CI, confidence interval; SD, standard deviation; IQR, interquartile range; APACHE, Acute Physiology and Chronic Health Evaluation.
Results of the multi-variate analysis are presented in Table II. In the final model, mechanical ventilation [odds ratio (OR) 8.2, 95% confidence interval (CI) 1.3e52.2; P ¼ 0.026] and APACHE II score (OR 1.2, 95% CI 1.0e1.3; P ¼ 0.007) were identified as independent risk factors for XDRPA acquisition. Antimicrobial susceptibility According to the definition of XDRPA, all XDRPA had intermediate susceptibility or were resistant to all tested antimicrobials except colistin. The MICs of imipenem for XDRPA were 8e128 mg/ mL (Table III), and the MICs of all other b-lactams for the isolates were 16 mg/mL. All isolates were susceptible to colistin (MICs 0.5e2 mg/mL).
Table II Multi-variate analysis of risk factors for the acquisition of extensive drug-resistant Pseudomonas aeruginosa Risk factor Mechanical ventilation APACHE II score
Adjusted OR (95% CI)
P-value
8.2 (1.3e52.2) 1.2 (1.0e1.3)
0.026 0.007
OR, odds ratio; CI, confidence interval; APACHE, Acute Physiology and Chronic Health Evaluation. Adjusted for pulmonary disease, time at risk, urinary catheter, penicillin with b-lactamase inhibitor, quinolone and carbapenem usage.
Table III Antimicrobial susceptibility of extensive drug-resistant Pseudomonas aeruginosa P. aeruginosa (N ¼ 43)
Antimicrobial agent
Piperacillin Piperacillin/tazobactam Ceftazidime Cefepime Aztreonam Imipenem Meropenem Amikacin Gentamicin Ciprofloxacin Colistin
MIC range (mg/L)
MIC50 (mg/L)
MIC90 (mg/L)
128e>256 128e>256 16e>128 32e>128 16e>128 8e>128 8e>128 64e>128 32e>128 8e>128 0.5e2
256 256 64 >128 128 16 32 >128 >128 32 1
>256 >256 >128 >128 >128 64 >128 >128 >128 64 2
MIC, minimum inhibitory concentration; MIC50 and MIC90, MICs for 50% and 90% of strains, respectively.
Mechanism of carbapenem resistance Of the 43 XDRPA isolates, 11 were positive for the modified Hodge test. Among them, eight (19%) were shown to produce MBLs by the EDTA-imipenem disk synergy test and PCR; four produced VIM-2 and four produced IMP-6. PFGE profile PFGE of Xba I-digested DNA of XDRPA revealed clonal epidemic XDRPA isolates co-existing with sporadic isolates (Figure 1). Among the XDRPA isolates from the same hospital, PFGE patterns were genetically related (Dice coefficient >80%); three of four from Hospital HA, three of six from Hospital HE, and five of six from Hospital HH. Discussion The important nosocomial pathogens P. aeruginosa and Acinetobacter spp. were categorized as MDR Gram-negative bacilli on a recent report by the Infectious Diseases Society of America.15 Unfortunately, no antibiotics (except tigecycline) have been developed specifically for MDR Gram-negative bacilli. There is a growing number of reported cases of infections caused by Gramnegative organisms for which no adequate therapeutic options exist.16 Thus, strategies to prevent the nosocomial emergence and spread of MDR Gram-negative bacilli are essential. Elucidation of the risk factors associated with the acquisition of antimicrobial-resistant P. aeruginosa by patients has been an area of active research, prompting numerous reports. Length of hospital stay, prior admission history, haemodialysis, prior antimicrobial usage and invasive therapeutic interventions are all known risk factors for the acquisition of imipenem-resistant P. aeruginosa.17e21 However, these analyses of the acquisition of MDR P. aeruginosa have some limitations due to heterogeneity of the MDR definition and the selection of control groups between studies. A few descriptive studies have been reported for pandrug-resistant P. aeruginosa.16,22 However, risk factors for the acquisition of P. aeruginosa resistant to all b-lactam antimicrobials, aminoglycosides and fluoroquinolones have not been well described to date. Epidemiological studies using DNA fingerprinting have shown that the acquisition of MDR P. aeruginosa may occur by selection of drug resistance by pre-existing sensitive strains and by cross-infection with resistant strains.23e25 The relative contributions of antimicrobial selective pressure and transmission between patients on the emergence of MDR P. aeruginosa are not yet known. In the present study, independent risk factors for XDRPA acquisition were mechanical ventilation and APACHE II score. Mechanical ventilation
100
90
80
70
60
50
40
Y.S. Park et al. / Journal of Hospital Infection 79 (2011) 54e58
Isolate no.
57
MBL gene PFGE type
..HE6 .A VIM-2 ..HF8 .B ..HD1 .C1 ..HG5 .C2 ..HE1, HE3, HE4 .C3 ..HG1 .D1 ..HG3 .D2 ..HG6, HG11 .E1 ..HG12 .E2 ..HG2 VIM-2 .F ..HG9 .G ..HG10 .H ..HE5 .I ..HG4 .J1 ..HG7 .J2 ..HA1 .K1 ..HA3, HA4 .K2 ..HA2, HB1 VIM-2 (HA2) .K3 ..HE2 IMP-6 .L1 ..HG8 .L2 ..HH1, HH2 .M1 ..HH3 .M2 ..HH4, HH5 .M3 ..HF1 .N ..HH6 .O ..HF5 VIM-2 .P1 ..HF7 IMP-6 .P2 ..HF4 .P3 ..HF10 .P4 ..HF3 .Q ..HC1 .R ..HF9 IMP-6 .S ..HF2 .T1 ..HF6 IMP-6 .T2 Figure 1. Pulsed-field gel electrophoresis (PFGE) dendrogram of extensive drug-resistant Pseudomonas aeruginosa. Ten of 12 isolates from Hospital HF were available for analysis. The strains were clustered by the unweighted pair group method with arithmetic averages. The scale indicates the percentage of genetic similarity. Isolate numbers indicate hospital and isolate numbers. MBL, metallo-b-lactamase.
is one of the most commonly implicated risk factors for the acquisition of MDR P. aeruginosa.26 Surprisingly, antimicrobial selection was not documented as a risk factor. Although an epidemiological study and outbreak investigation were not performed, there may be cross-transmission between patients receiving respiratory care in the same hospital, because mechanical ventilation was documented as a risk factor and clonality was observed on PFGE. This finding suggests that, during the study period, patientto-patient transmission contributed more to XDRPA acquisition than antimicrobial selective pressure. The definition of pandrug resistance in Gram-negative bacilli has been complicated by the rediscovery of colistin and the development of tigecycline. Recently, Falagas and Karageorgopoulos proposed that the term ‘extensive drug resistance’ should designate resistance to all but one or two classes of antimicrobial agents.6 In the present study, extensive drug resistance was defined as decreased susceptibility to all available antimicrobials for P. aeruginosa except colistin. The controls were selected at random from all inpatients who did not have XDRPA isolates, because control patients infected with the antimicrobial-susceptible organism would not only exaggerate
the association between antimicrobial exposure and patients with XDRPA, but fail to represent the base population. Resistance mechanisms that are expressed frequently in nosocomial strains of P. aeruginosa include b-lactamases, alterations in cell wall channels (porins) and efflux pumps. This bacterium can become resistant to quinolones through mutations in the genes gyrA and parC, and to aminoglycosides by expressing aminoglycoside-modifying enzymes. This study focused on the mechanisms of carbapenem resistance of XDRPA, because the acquisition of b-lactamases by P. aeruginosa is a major clinical concern.27 For P. aeruginosa, carbapenem resistance attributed to b-lactamases is due to MBLs in general. In Korea, VIM-2-like enzymes represented the majority of MBLs in P. aeruginosa, and 9% of imipenem-resistant P. aeruginosa isolated between 1995 and 1999 in a tertiary care hospital were found to produce VIM-2.14 The present study found that 9% of XDRPA carried VIM-2 genes and another 9% carried IMP-6 genes. In Korea, the IMP-type b-lactamases have rarely been reported in P. aeruginosa, except for a recent outbreak of IMP-1-carrying strains.28 In summary, this study supports a major role for mechanical ventilation in XDRPA acquisition. Moreover, PFGE revealed clonal
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epidemics within hospitals. Taken together, these results suggest that patient-to-patient transmission contributes to XDRPA acquisition in Korea. Carbapenem resistance of XDRPA is mainly due to a non-carbapenemase mechanism, and 19% of XDRPA produce MBLs. Conflict of interest statement None declared. Funding sources This work was partly supported by the National Institute of Health, Korea Centres for Disease Control and Prevention, Ministry of Health and Welfare, Republic of Korea (Grant No. 2007-S3-G-004). Acknowledgements The authors wish to thank the following for collecting isolates, reviewing medical records and data management: Kkot Sil Lee (Myungji Hospital of Kwandong University College of Medicine, Goyang); Kyoung Ho Roh (Konyang University Hospital, Daejeon); Il Kwon Bae (Kosin University Gospel Hospital, Busan); and Shin Young Park, Ji-Hea Kang, Sae Bom Kim and Jin Hee Kim (Gil Hospital of Gachon University of Medicine and Science, Incheon). The authors also thank Younghee Suh for screening the MBL-producing isolates. References 1. Gaynes R, Edwards JR. Overview of nosocomial infections caused by Gramnegative bacilli. Clin Infect Dis 2005;41:848e854. 2. Aris RM, Gilligan PH, Neuringer IP, Gott KK, Rea J, Yankaskas JR. The effects of panresistant bacteria in cystic fibrosis patients on lung transplant outcome. Am J Respir Crit Care Med 1997;155:1699e1704. 3. Aloush V, Navon-Venezia S, Seigman-Igra Y, Cabili S, Carmeli Y. Multidrugresistant Pseudomonas aeruginosa: risk factors and clinical impact. Antimicrob Agents Chemother 2006;50:43e48. 4. Giske CG, Monnet DL, Cars O, Carmeli Y. Clinical and economic impact of common multidrug-resistant Gram-negative bacilli. Antimicrob Agents Chemother 2008;52:813e821. 5. Paterson DL. The epidemiological profile of infections with multidrug-resistant Pseudomonas aeruginosa and Acinetobacter species. Clin Infect Dis 2006; 43(Suppl. 2):S43eS48. 6. Falagas ME, Karageorgopoulos DE. Pandrug resistance (PDR), extensive drug resistance (XDR), and multidrug resistance (MDR) among Gram-negative bacilli: need for international harmonization in terminology. Clin Infect Dis 2008;46:1121e1122, author reply 1122. 7. Harris AD, Karchmer TB, Carmeli Y, Samore MH. Methodological principles of caseecontrol studies that analyzed risk factors for antibiotic resistance: a systematic review. Clin Infect Dis 2001;32:1055e1061. 8. Harris AD, Samore MH, Lipsitch M, Kaye KS, Perencevich E, Carmeli Y. Controlgroup selection importance in studies of antimicrobial resistance: examples applied to Pseudomonas aeruginosa, Enterococci, and Escherichia coli. Clin Infect Dis 2002;34:1558e1563.
9. Kaye KS, Harris AD, Samore M, Carmeli Y. The caseecaseecontrol study design: addressing the limitations of risk factor studies for antimicrobial resistance. Infect Control Hosp Epidemiol 2005;26:346e351. 10. Clinical and Laboratory Standard Institute. Performance standards for antimicrobial susceptibility testing; seventeenth informational supplement. M100-S17. Wayne, PA: CLSI; 2007. 11. Charlson ME, Pompei P, Ales KL, MacKenzie CR. A new method of classifying prognostic comorbidity in longitudinal studies: development and validation. J Chron Dis 1987;40:373e383. 12. Lee K, Chong Y, Shin HB, Kim YA, Yong D, Yum JH. Modified Hodge and EDTA-disk synergy tests to screen metallo-beta-lactamase-producing strains of Pseudomonas and Acinetobacter species. Clin Microbiol Infect 2001;7:88e91. 13. Lee K, Yum JH, Yong D, et al. Novel acquired metallo-beta-lactamase gene, bla(SIM-1), in a class 1 integron from Acinetobacter baumannii clinical isolates from Korea. Antimicrob Agents Chemother 2005;49:4485e4491. 14. Lee K, Lim JB, Yum JH, et al. bla(VIM-2) cassette-containing novel integrons in metallo-beta-lactamase-producing Pseudomonas aeruginosa and Pseudomonas putida isolates disseminated in a Korean hospital. Antimicrob Agents Chemother 2002;46:1053e1058. 15. Talbot GH, Bradley J, Edwards Jr JE, Gilbert D, Scheld M, Bartlett JG. Bad bugs need drugs: an update on the development pipeline from the antimicrobial availability task force of the infectious diseases society of America. Clin Infect Dis 2006;42:657e668. 16. Falagas ME, Bliziotis IA, Kasiakou SK, Samonis G, Athanassopoulou P, Michalopoulos A. Outcome of infections due to pandrug-resistant (PDR) Gramnegative bacteria. BMC Infect Dis 2005;5:24. 17. Tam VH, Chang KT, LaRocco MT, et al. Prevalence, mechanisms, and risk factors of carbapenem resistance in bloodstream isolates of Pseudomonas aeruginosa. Diagn Microbiol Infect Dis 2007;58:309e314. 18. Fortaleza CM, Freire MP, Filho Dde C, de Carvalho Ramos M. Risk factors for recovery of imipenem- or ceftazidime-resistant Pseudomonas aeruginosa among patients admitted to a teaching hospital in Brazil. Infect Control Hosp Epidemiol 2006;27:901e906. 19. Ozkurt Z, Ertek M, Erol S, Altoparlak U, Akcay MN. The risk factors for acquisition of imipenem-resistant Pseudomonas aeruginosa in the burn unit. Burns 2005;31:870e873. 20. Zavascki AP, Cruz RP, Goldani LZ. Risk factors for imipenem-resistant Pseudomonas aeruginosa: a comparative analysis of two caseecontrol studies in hospitalized patients. J Hosp Infect 2005;59:96e101. 21. Harris AD, Smith D, Johnson JA, Bradham DD, Roghmann MC. Risk factors for imipenem-resistant Pseudomonas aeruginosa among hospitalized patients. Clin Infect Dis 2002;34:340e345. 22. Wang CY, Jerng JS, Chen KY, et al. Pandrug-resistant Pseudomonas aeruginosa among hospitalised patients: clinical features, risk factors and outcomes. Clin Microbiol Infect 2006;12:63e68. 23. Ferreira AC, Gobara S, Costa SE, et al. Emergence of resistance in Pseudomonas aeruginosa and Acinetobacter species after the use of antimicrobials for burned patients. Infect Control Hosp Epidemiol 2004;25:868e872. 24. Crespo MP, Woodford N, Sinclair A, et al. Outbreak of carbapenem-resistant Pseudomonas aeruginosa producing VIM-8, a novel metallo-beta-lactamase, in a tertiary care center in Cali, Colombia. J Clin Microbiol 2004;42:5094e5101. 25. Nouer SA, Nucci M, de-Oliveira MP, Pellegrino FL, Moreira BM. Risk factors for acquisition of multidrug-resistant Pseudomonas aeruginosa producing SPM metallo-beta-lactamase. Antimicrob Agents Chemother 2005;49:3663e3667. 26. Falagas ME, Kopterides P. Risk factors for the isolation of multi-drug-resistant Acinetobacter baumannii and Pseudomonas aeruginosa: a systematic review of the literature. J Hosp Infect 2006;64:7e15. 27. Bonomo RA, Szabo D. Mechanisms of multidrug resistance in Acinetobacter species and Pseudomonas aeruginosa. Clin Infect Dis 2006;43(Suppl. 2):S49eS56. 28. Nho SO, Jin JS, Kim JW, et al. Dissemination of the blaIMP-1 and blaVIM-2 metallo-beta-lactamase genes among genetically unrelated Pseudomonas aeruginosa isolates in a South Korean hospital. Int J Antimicrob Agents 2008;31: 586e588.
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Acquisition of extensive drug-resistant Pseudomonas aeruginosa among hospitalized patients: risk factors and resistance mechanisms to carbapenems Y.S. Park a, y, H. Lee b, y, B.S. Chin c, S.H. Han c, S.G. Hong d, S.K. Hong e, H.Y. Kim f, Y. Uh g, H.B. Shin h, E.J. Choo i, S.-H. Han j, W. Song k, S.H. Jeong l, K. Lee l, J.M. Kim c, * a
Department of Internal Medicine, Gachon University Gil Hospital, Incheon, Republic of Korea Department of Laboratory Medicine, Kwandong University College of Medicine, Goyang, Republic of Korea Department of Internal Medicine, Yonsei University College of Medicine, Seoul, Republic of Korea d Department of Laboratory Medicine, Pochon Cha University College of Medicine, Seongnam, Republic of Korea e Department of Internal Medicine, Pochon Cha University College of Medicine, Seongnam, Republic of Korea f Division of Infectious Diseases, Wonju Christian Hospital, Yonsei University Wonju College of Medicine, Wonju, Republic of Korea g Department of Laboratory Medicine, Wonju Christian Hospital, Yonsei University Wonju College of Medicine, Wonju, Republic of Korea h Department of Laboratory Medicine, Soonchunhyang University College of Medicine, Cheonan, Republic of Korea i Department of Internal Medicine, Soonchunhyang University College of Medicine, Cheonan, Republic of Korea j Infection Control Unit, Soonchunhyang University Hospital, Bucheon, Republic of Korea k Department of Laboratory Medicine, Hallym University and Kangnam Sacred Heart Hospital, Seoul, Republic of Korea l Department of Laboratory Medicine and Research Institute of Bacterial Resistance, Yonsei University College of Medicine, Seoul, Republic of Korea b c
a r t i c l e i n f o
s u m m a r y
Article history: Received 24 December 2010 Accepted 3 May 2011 by J.A. Child Available online 20 July 2011
Extensive drug-resistant Pseudomonas aeruginosa (XDRPA) strains, defined as resistant to all available antipseudomonal antibiotics, have been reported recently. This study aimed to investigate the risk factors for XDRPA acquisition by patients and the resistance mechanisms to carbapenems. From June to November 2007, XDRPA isolates were collected from patients in eight tertiary care hospitals. A caseecontrol study was performed to determine factors associated with XDRPA acquisition. EDTA-imipenem disc synergy tests, and polymerase chain reaction amplification and sequencing were performed to detect the presence of metallo-blactamases (MBLs). Risk factor analysis was performed for 33 patients. Mechanical ventilation [odds ratio (OR) 8.2, 95% confidence interval (CI) 1.3e52.2; P ¼ 0.026] and APACHE II score (OR 1.2, 95% CI 1.0e1.3; P ¼ 0.007) were identified as independent risk factors for XDRPA acquisition. Pulsed-field gel electrophoresis of XDRPA identified clonal epidemic isolates co-existing with sporadic isolates. Eight of 43 (19%) XDRPA isolates were shown to produce MBLs; four produced VIM-2 and four produced IMP-6. This study suggests a major role for mechanical ventilation in XDRPA acquisition. Moreover, pulsed-field gel electrophoresis identified a clonal epidemic within hospitals. Taken together, these results suggest that patient-to-patient transmission contributes to XDRPA acquisition in Korea. Ó 2011 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.
Keywords: Carbapenem resistance Extensive drug resistance Pseudomonas aeruginosa Risk factor
Introduction Pseudomonas aeruginosa is one of the main organisms responsible for drug-resistant nosocomial infections, and was the most commonly reported Gram-negative pathogen of nosocomial * Corresponding author. Address: Department of Internal Medicine, Yonsei University College of Medicine, 250 Seongsanno, Seodaemun-gu, Seoul 120-752, Republic of Korea. Tel.: þ82 2 2228 1946; fax: þ82 2 393 6884. E-mail address: [email protected] (J.M. Kim). y These authors contributed equally to this study.
pneumonia from 1987 to 2003.1 The National Nosocomial Infection Surveillance System has reported a significantly increasing trend of resistance.1 Over the past decade, strains with resistance to almost all available antibiotics have emerged, causing major therapeutic problems. Multi-drug-resistant (MDR) strains of P. aeruginosa were initially reported in patients with cystic fibrosis, and numerous cases have subsequently been reported.2 Prevention of the acquisition of MDR P. aeruginosa is essential due to limited therapeutic options and increased mortality.3,4
0195-6701/$ e see front matter Ó 2011 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.jhin.2011.05.014
Y.S. Park et al. / Journal of Hospital Infection 79 (2011) 54e58
Various definitions of antimicrobial resistance have been used in publications dealing with infections caused by Acinetobacter spp. or P. aeruginosa. Multi-drug resistance in P. aeruginosa or Acinetobacter spp. has been variously defined.5 The definition of pandrug resistance is also controversial due to colistin, a ‘rediscovered’ category of antimicrobial agent, and tigecycline, a new drug. Recently, it was proposed that the term ‘extensive drug resistance’ should be used to indicate resistance to all but one or two classes of antimicrobial agents.6 Risk factors for the acquisition of extensive drug-resistant P. aeruginosa (XDRPA) have not been well documented to date. This study aimed to clarify the risk factors for XDRPA acquisition in Korean hospitals, and to document resistance mechanisms to carbapenems. Controls were selected based on improved epidemiological methodology, enabling the assessment of risk factors without the bias of antibiotic usage.7e9 Materials and methods Bacterial strains and antimicrobial susceptibility testing From June to November 2007, XDRPA isolates were collected from clinical specimens at eight tertiary care hospitals (Severance Hospital of Yonsei University College of Medicine, Seoul; Kangnam Sacred Heart Hospital of Hallym University Medical Centre, Seoul; Korea University Anam Hospital, Seoul; Myongji Hospital of Kwandong University College of Medicine, Goyang; Bundang Cha Hospital of Pochon CHA University College of Medicine, Seongnam; Soonchunhyang University Hospital, Bucheon; Yonsei University Wonju Christian Hospital, Wonju; Kosin University Gospel Hospital, Busan). P. aeruginosa isolates that showed resistance or intermediate susceptibility to piperacillin, piperacillin-tazobactam, ceftazidime, cefepime, aztreonam, imipenem, meropenem, amikacin, gentamicin and ciprofloxacin were considered to be XDRPA, and included in the study. Isolates with resistance to colistin were excluded. The minimum inhibitory concentrations (MICs) of antimicrobial agents were determined by agar dilution.10 Caseecontrol study A caseecontrol study was performed to determine factors associated with XDRPA acquisition. The cases were patients from whom XDRPA were isolated in clinical cultures. During the study period, the electronic microbiology database was searched to identify all XDRPA-positive clinical samples obtained from inpatients. Patients who had been hospitalized for <48 h when the sample was collected and those aged 18 years were excluded. The controls were randomly selected adult inpatients in the participating hospitals who did not have XDRPA isolated during their hospital stay. Controls were matched to cases by hospital. Two controls were recruited for each case. Computerized medical, pharmacy and microbiological records were reviewed. Variables investigated as potential risk factors included age, sex, Charlson score,11 length of hospital stay before outcome of interest (time at risk; for case patients, from admission to nosocomial isolation of XDRPA; for controls, complete length of hospital stay), intensive care unit (ICU) stay, surgery, and severity of illness at the time of infection/colonization [calculated by Acute Physiology and Chronic Health Evaluation (APACHE) II score]. The following associated diseases and comorbid conditions were documented: diabetes mellitus, cardiac disease, pulmonary disease, renal disease (creatinine >2.0 mg/dL or on dialysis), neurological disease, malignancy, use of devices before XDRPA isolation (central venous and/or arterial catheter, urinary catheter, mechanical ventilation and nasogastric tube), and exposure to antimicrobials before XDRPA isolation.
55
Detection of metallo-b-lactamases and sequencing Carbapenemase and metallo-b-lactamase (MBL) production were determined by the modified Hodge test and the double-disc synergy test.12 Polymerase chain reaction (PCR) detection of the blaIMP- and blaVIM-like alleles was performed using IMP-1 and VIM2 primers.13 To determine the type of MBL genes, isolates with blaIMP- and blaVIM-like alleles were used for sequencing. INT-1-F, INT-2-R, IMP-1-F, IMP-1-R, VIM-2-F and VIM-2-R primers were used for blaIMP- and blaVIM-like allele sequencing. The primers used were: IMP-1-F (5’-CAT GGT TTG GTG GTT CTT GT-3’) and IMP-1-R (5’-ATA ATT TGG CGG ACT TTG GC-3’); VIM-2-F (5’-ATG TTC AAA CTT TTG AGT AAG-3’) and VIM-2-R (5’-CTA CTC AAC GAC TGA GCG-3’); and INT-1-F (5’-GGC ATC CAA GCA GCA AG-3’) and INT2-R (5’-AAG CAG ACT TGA CCT GA-3’). The PCR products were subjected to direct sequencing. Both strands of the PCR products were sequenced twice with an automatic sequencer (Model 3730xl; Applied Biosystems, Weiterstadt, Germany). Sequence analysis and comparisons were performed using programs available at the National Center for Biotechnology Information (http://www.ncbi. nlm.nih.gov/). Pulsed-field gel electrophoresis For pulsed-field gel electrophoresis (PFGE) analysis, Xba I-digested genomic DNA of XDRPA was separated using a CHEF-DR II System (Bio-Rad, Hercules, CA, USA).14 The pattern was analysed with Fingerprinting II software (Bio-Rad). Statistical analysis Statistical analyses were performed using Statistical Package for the Social Sciences Version 15 (SPSS Inc., Chicago, IL, USA). Bivariate analyses were performed separately for each of the variables. P-values were calculated using Fisher’s exact test for categorical variables, and Student’s t-test or ManneWhitney U-test for continuous variables. Variables for which the P-value was <0.05 in the bivariate analysis were included in the multi-variate analysis. A forward selection process was used. Risk factors were checked for colinearity by viewing changes in standard errors of multi-variate models. All tests were two-tailed, and a P-value of <0.05 was considered to be significant in the multi-variate model. Results The mechanism of carbapenem resistance was analysed using 43 XDRPA isolates, and full medical records were available for 37 of these patients. Positive cultures within two days of admission necessitated the exclusion of four patients with XDRPA from the risk factor analysis. In total, 33 patients were included in the risk factor analysis. Risk factor analysis Of 33 patients with XDRPA acquisition, 26 were men and seven were women. Their mean age was 65 years (standard deviation 17 years). The most common site from which XDRPA was recovered was the respiratory system (N ¼ 13, 39%), followed by the urinary system (N ¼ 12, 36%). Sixteen patients were receiving care in general wards and 17 were in ICUs. The results of bivariate analyses for baseline demographic and clinical characteristics, associated diseases or comorbid conditions, Charlson score, ICU stay, surgery, time at risk, severity of illness (as calculated by the APACHE II score) and receipt of specific antibiotics are presented in Table I.
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Y.S. Park et al. / Journal of Hospital Infection 79 (2011) 54e58
Table I Bivariate analysis of risk factors for the acquisition of Pseudomonas aeruginosa Risk factor Age in years, mean SD Male sex Associated disease Diabetes mellitus Cardiac disease Pulmonary disease Renal disease Neurological disease Malignancy Charlson comorbidity scale, median (IQR) Related to hospitalization Days at risk, median (IQR) Intensive care unit stay in days, median (IQR) Surgery Device Central venous/arterial catheter Urinary catheter Mechanical ventilation Nasogastric tube Antimicrobial Cephalosporin First generation Second generation Third generation Penicillin with b-lactamase inhibitor Quinolone Aminoglycoside Carbapenem Glycopeptide Septic shock/severe sepsis APACHE II score, median (IQR)
Cases (N ¼ 33)
Controls (N ¼ 66)
65 12 26 (79)
56 19 30 (46)
11 (33) 7 (21) 13 (39) 6 (18) 14 (42) 11 (33) 3 (2e6)
15 (23) 17 (26) 5 (8) 3 (5) 7 (11) 22 (33) 1 (0e3)
OR (95% CI) 4.5 (1.7e11.7) 1.7 0.8 7.9 4.7 6.2 1.0
P-value 0.006 0.002
(0.7e4.3) 0.33 (0.3e2.1) 0.8 (2.5e25.0) <0.001 (1.1e20.0) 0.057 (2.2e17.6) 0.001 (0.4e2.4) 1 0.003
<0.001 <0.001
31 (11e65) 10 (5e21) 9 (3e26) 0 (0e0) 14 (42)
20 (31)
1.7 (0.7e4.0)
19 (58)
12 (18)
6.1 (2.4e15.5) <0.001
24 (73) 15 (46) 16 (49)
19 (29) 3 (4.5) 7 (11)
6.6 (2.6e16.8) <0.001 17.5 (4.6e67.2) <0.001 7.9 (2.8e22.4) <0.001
5 (15) 10 (30) 22 (67) 11 (33)
7 (11) 13 (20) 37 (56) 4 (6)
17 (52) 15 (46) 9 (27) 13 (39) 7 (21)
11 (17) 19 (29) 2 (3) 7 (11) 3 (5)
14 (10e21)
1.5 1.7 1.6 7.8
0.27
(0.4e5.2) (0.7e4.6) (0.7e3.7) (2.2e26.9)
0.52 0.17 0.39 0.001
5.3 (2.1e13.6) 2.1 (0.9e4.9) 12.0 (2.4e59.6) 5.5 (1.9e15.6) 5.6 (1.3e23.2)
0.001 0.12 0.001 0.001 0.03
6 (2e8)
<0.001
Data are N (%) of patients, unless otherwise indicated. OR, odds ratio; CI, confidence interval; SD, standard deviation; IQR, interquartile range; APACHE, Acute Physiology and Chronic Health Evaluation.
Results of the multi-variate analysis are presented in Table II. In the final model, mechanical ventilation [odds ratio (OR) 8.2, 95% confidence interval (CI) 1.3e52.2; P ¼ 0.026] and APACHE II score (OR 1.2, 95% CI 1.0e1.3; P ¼ 0.007) were identified as independent risk factors for XDRPA acquisition. Antimicrobial susceptibility According to the definition of XDRPA, all XDRPA had intermediate susceptibility or were resistant to all tested antimicrobials except colistin. The MICs of imipenem for XDRPA were 8e128 mg/ mL (Table III), and the MICs of all other b-lactams for the isolates were 16 mg/mL. All isolates were susceptible to colistin (MICs 0.5e2 mg/mL).
Table II Multi-variate analysis of risk factors for the acquisition of extensive drug-resistant Pseudomonas aeruginosa Risk factor Mechanical ventilation APACHE II score
Adjusted OR (95% CI)
P-value
8.2 (1.3e52.2) 1.2 (1.0e1.3)
0.026 0.007
OR, odds ratio; CI, confidence interval; APACHE, Acute Physiology and Chronic Health Evaluation. Adjusted for pulmonary disease, time at risk, urinary catheter, penicillin with b-lactamase inhibitor, quinolone and carbapenem usage.
Table III Antimicrobial susceptibility of extensive drug-resistant Pseudomonas aeruginosa P. aeruginosa (N ¼ 43)
Antimicrobial agent
Piperacillin Piperacillin/tazobactam Ceftazidime Cefepime Aztreonam Imipenem Meropenem Amikacin Gentamicin Ciprofloxacin Colistin
MIC range (mg/L)
MIC50 (mg/L)
MIC90 (mg/L)
128e>256 128e>256 16e>128 32e>128 16e>128 8e>128 8e>128 64e>128 32e>128 8e>128 0.5e2
256 256 64 >128 128 16 32 >128 >128 32 1
>256 >256 >128 >128 >128 64 >128 >128 >128 64 2
MIC, minimum inhibitory concentration; MIC50 and MIC90, MICs for 50% and 90% of strains, respectively.
Mechanism of carbapenem resistance Of the 43 XDRPA isolates, 11 were positive for the modified Hodge test. Among them, eight (19%) were shown to produce MBLs by the EDTA-imipenem disk synergy test and PCR; four produced VIM-2 and four produced IMP-6. PFGE profile PFGE of Xba I-digested DNA of XDRPA revealed clonal epidemic XDRPA isolates co-existing with sporadic isolates (Figure 1). Among the XDRPA isolates from the same hospital, PFGE patterns were genetically related (Dice coefficient >80%); three of four from Hospital HA, three of six from Hospital HE, and five of six from Hospital HH. Discussion The important nosocomial pathogens P. aeruginosa and Acinetobacter spp. were categorized as MDR Gram-negative bacilli on a recent report by the Infectious Diseases Society of America.15 Unfortunately, no antibiotics (except tigecycline) have been developed specifically for MDR Gram-negative bacilli. There is a growing number of reported cases of infections caused by Gramnegative organisms for which no adequate therapeutic options exist.16 Thus, strategies to prevent the nosocomial emergence and spread of MDR Gram-negative bacilli are essential. Elucidation of the risk factors associated with the acquisition of antimicrobial-resistant P. aeruginosa by patients has been an area of active research, prompting numerous reports. Length of hospital stay, prior admission history, haemodialysis, prior antimicrobial usage and invasive therapeutic interventions are all known risk factors for the acquisition of imipenem-resistant P. aeruginosa.17e21 However, these analyses of the acquisition of MDR P. aeruginosa have some limitations due to heterogeneity of the MDR definition and the selection of control groups between studies. A few descriptive studies have been reported for pandrug-resistant P. aeruginosa.16,22 However, risk factors for the acquisition of P. aeruginosa resistant to all b-lactam antimicrobials, aminoglycosides and fluoroquinolones have not been well described to date. Epidemiological studies using DNA fingerprinting have shown that the acquisition of MDR P. aeruginosa may occur by selection of drug resistance by pre-existing sensitive strains and by cross-infection with resistant strains.23e25 The relative contributions of antimicrobial selective pressure and transmission between patients on the emergence of MDR P. aeruginosa are not yet known. In the present study, independent risk factors for XDRPA acquisition were mechanical ventilation and APACHE II score. Mechanical ventilation
100
90
80
70
60
50
40
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Isolate no.
57
MBL gene PFGE type
..HE6 .A VIM-2 ..HF8 .B ..HD1 .C1 ..HG5 .C2 ..HE1, HE3, HE4 .C3 ..HG1 .D1 ..HG3 .D2 ..HG6, HG11 .E1 ..HG12 .E2 ..HG2 VIM-2 .F ..HG9 .G ..HG10 .H ..HE5 .I ..HG4 .J1 ..HG7 .J2 ..HA1 .K1 ..HA3, HA4 .K2 ..HA2, HB1 VIM-2 (HA2) .K3 ..HE2 IMP-6 .L1 ..HG8 .L2 ..HH1, HH2 .M1 ..HH3 .M2 ..HH4, HH5 .M3 ..HF1 .N ..HH6 .O ..HF5 VIM-2 .P1 ..HF7 IMP-6 .P2 ..HF4 .P3 ..HF10 .P4 ..HF3 .Q ..HC1 .R ..HF9 IMP-6 .S ..HF2 .T1 ..HF6 IMP-6 .T2 Figure 1. Pulsed-field gel electrophoresis (PFGE) dendrogram of extensive drug-resistant Pseudomonas aeruginosa. Ten of 12 isolates from Hospital HF were available for analysis. The strains were clustered by the unweighted pair group method with arithmetic averages. The scale indicates the percentage of genetic similarity. Isolate numbers indicate hospital and isolate numbers. MBL, metallo-b-lactamase.
is one of the most commonly implicated risk factors for the acquisition of MDR P. aeruginosa.26 Surprisingly, antimicrobial selection was not documented as a risk factor. Although an epidemiological study and outbreak investigation were not performed, there may be cross-transmission between patients receiving respiratory care in the same hospital, because mechanical ventilation was documented as a risk factor and clonality was observed on PFGE. This finding suggests that, during the study period, patientto-patient transmission contributed more to XDRPA acquisition than antimicrobial selective pressure. The definition of pandrug resistance in Gram-negative bacilli has been complicated by the rediscovery of colistin and the development of tigecycline. Recently, Falagas and Karageorgopoulos proposed that the term ‘extensive drug resistance’ should designate resistance to all but one or two classes of antimicrobial agents.6 In the present study, extensive drug resistance was defined as decreased susceptibility to all available antimicrobials for P. aeruginosa except colistin. The controls were selected at random from all inpatients who did not have XDRPA isolates, because control patients infected with the antimicrobial-susceptible organism would not only exaggerate
the association between antimicrobial exposure and patients with XDRPA, but fail to represent the base population. Resistance mechanisms that are expressed frequently in nosocomial strains of P. aeruginosa include b-lactamases, alterations in cell wall channels (porins) and efflux pumps. This bacterium can become resistant to quinolones through mutations in the genes gyrA and parC, and to aminoglycosides by expressing aminoglycoside-modifying enzymes. This study focused on the mechanisms of carbapenem resistance of XDRPA, because the acquisition of b-lactamases by P. aeruginosa is a major clinical concern.27 For P. aeruginosa, carbapenem resistance attributed to b-lactamases is due to MBLs in general. In Korea, VIM-2-like enzymes represented the majority of MBLs in P. aeruginosa, and 9% of imipenem-resistant P. aeruginosa isolated between 1995 and 1999 in a tertiary care hospital were found to produce VIM-2.14 The present study found that 9% of XDRPA carried VIM-2 genes and another 9% carried IMP-6 genes. In Korea, the IMP-type b-lactamases have rarely been reported in P. aeruginosa, except for a recent outbreak of IMP-1-carrying strains.28 In summary, this study supports a major role for mechanical ventilation in XDRPA acquisition. Moreover, PFGE revealed clonal
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epidemics within hospitals. Taken together, these results suggest that patient-to-patient transmission contributes to XDRPA acquisition in Korea. Carbapenem resistance of XDRPA is mainly due to a non-carbapenemase mechanism, and 19% of XDRPA produce MBLs. Conflict of interest statement None declared. Funding sources This work was partly supported by the National Institute of Health, Korea Centres for Disease Control and Prevention, Ministry of Health and Welfare, Republic of Korea (Grant No. 2007-S3-G-004). Acknowledgements The authors wish to thank the following for collecting isolates, reviewing medical records and data management: Kkot Sil Lee (Myungji Hospital of Kwandong University College of Medicine, Goyang); Kyoung Ho Roh (Konyang University Hospital, Daejeon); Il Kwon Bae (Kosin University Gospel Hospital, Busan); and Shin Young Park, Ji-Hea Kang, Sae Bom Kim and Jin Hee Kim (Gil Hospital of Gachon University of Medicine and Science, Incheon). The authors also thank Younghee Suh for screening the MBL-producing isolates. References 1. Gaynes R, Edwards JR. Overview of nosocomial infections caused by Gramnegative bacilli. Clin Infect Dis 2005;41:848e854. 2. Aris RM, Gilligan PH, Neuringer IP, Gott KK, Rea J, Yankaskas JR. The effects of panresistant bacteria in cystic fibrosis patients on lung transplant outcome. Am J Respir Crit Care Med 1997;155:1699e1704. 3. Aloush V, Navon-Venezia S, Seigman-Igra Y, Cabili S, Carmeli Y. Multidrugresistant Pseudomonas aeruginosa: risk factors and clinical impact. Antimicrob Agents Chemother 2006;50:43e48. 4. Giske CG, Monnet DL, Cars O, Carmeli Y. Clinical and economic impact of common multidrug-resistant Gram-negative bacilli. Antimicrob Agents Chemother 2008;52:813e821. 5. Paterson DL. The epidemiological profile of infections with multidrug-resistant Pseudomonas aeruginosa and Acinetobacter species. Clin Infect Dis 2006; 43(Suppl. 2):S43eS48. 6. Falagas ME, Karageorgopoulos DE. Pandrug resistance (PDR), extensive drug resistance (XDR), and multidrug resistance (MDR) among Gram-negative bacilli: need for international harmonization in terminology. Clin Infect Dis 2008;46:1121e1122, author reply 1122. 7. Harris AD, Karchmer TB, Carmeli Y, Samore MH. Methodological principles of caseecontrol studies that analyzed risk factors for antibiotic resistance: a systematic review. Clin Infect Dis 2001;32:1055e1061. 8. Harris AD, Samore MH, Lipsitch M, Kaye KS, Perencevich E, Carmeli Y. Controlgroup selection importance in studies of antimicrobial resistance: examples applied to Pseudomonas aeruginosa, Enterococci, and Escherichia coli. Clin Infect Dis 2002;34:1558e1563.
9. Kaye KS, Harris AD, Samore M, Carmeli Y. The caseecaseecontrol study design: addressing the limitations of risk factor studies for antimicrobial resistance. Infect Control Hosp Epidemiol 2005;26:346e351. 10. Clinical and Laboratory Standard Institute. Performance standards for antimicrobial susceptibility testing; seventeenth informational supplement. M100-S17. Wayne, PA: CLSI; 2007. 11. Charlson ME, Pompei P, Ales KL, MacKenzie CR. A new method of classifying prognostic comorbidity in longitudinal studies: development and validation. J Chron Dis 1987;40:373e383. 12. Lee K, Chong Y, Shin HB, Kim YA, Yong D, Yum JH. Modified Hodge and EDTA-disk synergy tests to screen metallo-beta-lactamase-producing strains of Pseudomonas and Acinetobacter species. Clin Microbiol Infect 2001;7:88e91. 13. Lee K, Yum JH, Yong D, et al. Novel acquired metallo-beta-lactamase gene, bla(SIM-1), in a class 1 integron from Acinetobacter baumannii clinical isolates from Korea. Antimicrob Agents Chemother 2005;49:4485e4491. 14. Lee K, Lim JB, Yum JH, et al. bla(VIM-2) cassette-containing novel integrons in metallo-beta-lactamase-producing Pseudomonas aeruginosa and Pseudomonas putida isolates disseminated in a Korean hospital. Antimicrob Agents Chemother 2002;46:1053e1058. 15. Talbot GH, Bradley J, Edwards Jr JE, Gilbert D, Scheld M, Bartlett JG. Bad bugs need drugs: an update on the development pipeline from the antimicrobial availability task force of the infectious diseases society of America. Clin Infect Dis 2006;42:657e668. 16. Falagas ME, Bliziotis IA, Kasiakou SK, Samonis G, Athanassopoulou P, Michalopoulos A. Outcome of infections due to pandrug-resistant (PDR) Gramnegative bacteria. BMC Infect Dis 2005;5:24. 17. Tam VH, Chang KT, LaRocco MT, et al. Prevalence, mechanisms, and risk factors of carbapenem resistance in bloodstream isolates of Pseudomonas aeruginosa. Diagn Microbiol Infect Dis 2007;58:309e314. 18. Fortaleza CM, Freire MP, Filho Dde C, de Carvalho Ramos M. Risk factors for recovery of imipenem- or ceftazidime-resistant Pseudomonas aeruginosa among patients admitted to a teaching hospital in Brazil. Infect Control Hosp Epidemiol 2006;27:901e906. 19. Ozkurt Z, Ertek M, Erol S, Altoparlak U, Akcay MN. The risk factors for acquisition of imipenem-resistant Pseudomonas aeruginosa in the burn unit. Burns 2005;31:870e873. 20. Zavascki AP, Cruz RP, Goldani LZ. Risk factors for imipenem-resistant Pseudomonas aeruginosa: a comparative analysis of two caseecontrol studies in hospitalized patients. J Hosp Infect 2005;59:96e101. 21. Harris AD, Smith D, Johnson JA, Bradham DD, Roghmann MC. Risk factors for imipenem-resistant Pseudomonas aeruginosa among hospitalized patients. Clin Infect Dis 2002;34:340e345. 22. Wang CY, Jerng JS, Chen KY, et al. Pandrug-resistant Pseudomonas aeruginosa among hospitalised patients: clinical features, risk factors and outcomes. Clin Microbiol Infect 2006;12:63e68. 23. Ferreira AC, Gobara S, Costa SE, et al. Emergence of resistance in Pseudomonas aeruginosa and Acinetobacter species after the use of antimicrobials for burned patients. Infect Control Hosp Epidemiol 2004;25:868e872. 24. Crespo MP, Woodford N, Sinclair A, et al. Outbreak of carbapenem-resistant Pseudomonas aeruginosa producing VIM-8, a novel metallo-beta-lactamase, in a tertiary care center in Cali, Colombia. J Clin Microbiol 2004;42:5094e5101. 25. Nouer SA, Nucci M, de-Oliveira MP, Pellegrino FL, Moreira BM. Risk factors for acquisition of multidrug-resistant Pseudomonas aeruginosa producing SPM metallo-beta-lactamase. Antimicrob Agents Chemother 2005;49:3663e3667. 26. Falagas ME, Kopterides P. Risk factors for the isolation of multi-drug-resistant Acinetobacter baumannii and Pseudomonas aeruginosa: a systematic review of the literature. J Hosp Infect 2006;64:7e15. 27. Bonomo RA, Szabo D. Mechanisms of multidrug resistance in Acinetobacter species and Pseudomonas aeruginosa. Clin Infect Dis 2006;43(Suppl. 2):S49eS56. 28. Nho SO, Jin JS, Kim JW, et al. Dissemination of the blaIMP-1 and blaVIM-2 metallo-beta-lactamase genes among genetically unrelated Pseudomonas aeruginosa isolates in a South Korean hospital. Int J Antimicrob Agents 2008;31: 586e588.