|Year : 2014 | Volume
| Issue : 1 | Page : 44-48
Chronological emergence of a class a carbapenemase-producing Enterobacter aerogenes in Taiwan
Zheng-Yi Huang1, Jun-Ren Sun2, Sheng-Ting Hsu2, Ching-Mei Yu2, Cherng-Lih Perng3, Tzong-Shi Chiueh2
1 Department of Pathology, Kaohsiung Armed Forced General Hospital, Kaohsiung; Division of Clinical Pathology, Department of Pathology, Tri-Service General Hospital and National Defense Medical Center, Taipei, Taiwan, Republic of China
2 Division of Clinical Pathology, Department of Pathology, Tri-Service General Hospital and National Defense Medical Center, Taipei, Taiwan, Republic of China
3 Division of Clinical Pathology, Department of Pathology, Tri-Service General Hospital and National Defense Medical Center; Graduate Institute of Pathology, National Defense Medical Center, Taipei, Taiwan, Republic of China
|Date of Submission||11-Apr-2013|
|Date of Decision||01-Jul-2013|
|Date of Acceptance||19-Jul-2013|
|Date of Web Publication||26-Mar-2014|
Division of Clinical Pathology, Department of Pathology, Tri-Service General Hospital, No. 325, Sec. 2, Cheng-Kung Road, Taipei 114, Taiwan
Republic of China
Source of Support: None, Conflict of Interest: None
This study reports the case of a 77-year-old, long-term, bedridden patient, with a nosocomial wound infection caused by a multidrug-resistant strain of Enterobacter aerogenes (E. aerogense). The isolate produced an Ambler-class A carbapenemase, which was demonstrated by the Modified Hodge test (MHT) and a confirmatory inhibition test. However, no known carbapenemase genes were discovered in this isolate by polymerase chain reactions (PCRs) with specific primers. New carbapenemase or other resistant mechanisms could be explored from the isolate of carbapenem-resistant E. aerogense, according to the revised criteria (CLSI, 2012).
Keywords: Carbapenemase, Enterobacter aerogenes, Taiwan
|How to cite this article:|
Huang ZY, Sun JR, Hsu ST, Yu CM, Perng CL, Chiueh TS. Chronological emergence of a class a carbapenemase-producing Enterobacter aerogenes in Taiwan. J Med Sci 2014;34:44-8
|How to cite this URL:|
Huang ZY, Sun JR, Hsu ST, Yu CM, Perng CL, Chiueh TS. Chronological emergence of a class a carbapenemase-producing Enterobacter aerogenes in Taiwan. J Med Sci [serial online] 2014 [cited 2020 Jan 18];34:44-8. Available from: http://www.jmedscindmc.com/text.asp?2014/34/1/44/129394
| Introduction|| |
Enterobacter aerogenes was not an important pathogen until it was reported as one of the common pathogens causing nosocomial infections in recent decades. ,, Strains of this species are naturally resistant to ampicillin, amoxicillin-clavulanate, cefazolin, and cefuroxime, due to the constitutive production of chromosomal AmpC β-lactamases. , In recent studies, the clinical isolates of this species has not only shown the extended-spectrum β-lactamase TEM-24, which results in resistance to β-lactam antibiotics, ,, but also resistance to quinolones, tetracycline, and chloramphenicol, due to reduced drug uptake, by losing porins on the outer membrane. , In the past 10 years, the carbapenem-resistant E. aerogenes has been found in many countries. ,,,, With the emergence of multidrug-resistant E. aerogenes strains, the mortality rate of their infected patients has also increased. 
This study reports the case of a 77-year-old, long-term, bedridden patient having a nosocomial wound infection due to a multidrug-resistant (MDR) strain of E. aerogenes, which was also resistant to carbapenems. The isolate developed a phenotype of the class A carbapenemase after imipenem treatment for three weeks.
This is the first report of carbapenem-resistant E. aerogenes that demonstrates a phenotype of class A carbapenemase, in Taiwan.
| Case Report|| |
A 77-year-old male has had a history of old cerebral vascular accident, with right hemiparesis, stupor consciousness, hypertension, and type-2 diabetes mellitus with nephropathy, for years. He had no recent traveling history. Regular hemodialysis was initiated a year ago. He was admitted to create an arteriovenous shunt over his left arm. The operation was performed on hospitalization day 2. Subsequently, the patient presented with shortness of breath, fever, and dyspnea with leukocytosis. Aspiration pneumonia was diagnosed on hospitalization day 4. Arterial blood gas analysis revealed a pH of 7.71, PaCO 2 of 15.9 mmHg, and PaO 2 of 63.1 mmHg. After an emergent endotracheal tube insertion, he was transferred to the Intensive Care Unit (ICU) on hospitalization day 9. Isolates of E. aerogenes were obtained from two sets of sputum culture, and they were found to be sensitive to amikacin, piperacillin, third-generation cephalosporin, imipenem, ertapenem, and intermittent to ciprofloxacin on hospitalization day 5. Imipenem-sensitive Pseudomonas aeruginosa (P. aeruginosa) was isolated from his sputum on hospitalization day 17. The patient received tracheostomy on hospitalization day 28 because of ventilator dependence. In his infection course [Table 1], despite administering the serial anti-microbial agent, prescriptions including amoxicillin, piperacillin, first- and fourth-generation cephalosporin, vancomycin, and ciprofloxacin, the P. aeruginosa-related pneumonia persisted. The antibiograms of the P. aeruginosa isolate remained unchanged until an imipenem-resistant P. aeruginosa was noted on hospitalization day 81. A few weeks later, a 5 × 8 cm 2 pressure sore was noted over his sacral region, owing to his being bedridden for a long period of time. Mixed infections of P. aeruginosa and E. aerogenes were found in the wound culture on hospitalization day 107. Not only did the E. aerogenes isolate remain resistant to carbapenems, it also became resistant to carbapenems. According to the antibiograms of the two imipenem-resistant strains, the patient was then treated with amikacin (1000 mg, q.d.) and cefepime (1000 mg, q.d.). Although his wound infection was controlled, the nosocomial pneumonia caused by the imipenem-resistant P. aeruginosa persisted. At present, he is still hospitalized in a Respiratory Care Center for controlling the nosocomial pneumonia.
|Table 1: Chronological emergence of carbapenem-resistant phenotypes for Pseudomonas aeruginosa and Enterobacter aerogenes during hospitalization|
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| Laboratory Result|| |
Both imipenem-resistant P. aeruginosa and E. aerogenes were isolated from the wound swab on hospitalization day 107. The Vitek-2 ID 32 GN and AST-N044 systems (bioMerieux Vitek, Marcy-l'Etoile, France) were used for species identification and the antimicrobial susceptibility test. All antibiotics, except amikacin, showed high-level minimum inhibitory concentration (MIC) values. Both disk diffusion and microdilution methods were also performed and revealed compatible results with the MIC values of Vitek-2 (MIC: amikacin <4 μg/ml, cefepime: 8 μg/ml, imipenem >8 μg/ml, ertapenem >4 μg/ml, meropenem: 8 μg/ml, tigecycline: 2 μg/ml, colistin: 0.25 μg/ml, polymyxin B: 0.5 μg/ml). Carbapenem susceptibilities for Enterobacteriaceae are interpreted according to the new Clinical and Laboratory Standards Institute (CLSI) criteria, 2012. The Modified Hodge test was performed to detect whether carbapenemase was produced by the two imipenem-resistant isolates.  A strong positive result was revealed for the isolate of E. aerogenes, but not for the isolate of P. aeruginosa [Figure 1]a. We used a carbapenemase-inhibitor-impregnated agar to test and classify the carbapenem-resistant strains. , Aminophenylboronic acid (APBA) was employed to inhibit class A carbapenemase. Class B metallo-carbapanemase was suppressed by ethylenediaminetetraacetic acid (EDTA), and 6-pyridinedicarboxylic acid (DPA) and cloxacillin were utilized to inhibit β-lactamase. The size differences of the inhibition zone on the meropenem disks between the control and inhibitor of the APBA-impregnated Mueller-Hinton-agar were measured with a cut-off value of 4 mm, while those between the control and inhibitors of EDTA, DPA, and cloxacillin were measured with a cut-off value of 5 mm. The positive result was noted only on the meropenem disk with APBA [Figure 1]b. The E. aerogenes isolate was therefore phenotypically typed as a strain-producing Ambler class A carbapenemase. However, the isolate of imipenem-resistant P. aeruginosa showed a negative result in the Modified Hodge test.
|Figure 1: Modified Hodge Test and (a) confirmatory inhibition test (b) for the Enterobacter aerogenes. 1: E. coli ATCC 25922 as a reporter strain; 2: K. pneumonia ATCC BAA 1760 as the negative control; 3: Enterobacter aerogenes isolate showed a positive result; 4: Pseudomonas aeruginosa isolate showed a negative result; APB: aminophenylboronic acid, DPA: 6-pyridinedicarboxylic acid, CLX: cloxacillin sodium salt monohydrate, EDTA: ethylenediaminetetraacetic acid|
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To verify the class A carbapenemase genes, PCRs were performed with primers specific for the Ambler class A (blaKPC, blaGES, blaSME, blaNMC-A ), B (blaIMP, blaVIM, blaGIM, blaSIM, blaSPM ), or D (blaOXA ) genes, as in the previous studies. ,,,, However, both isolates revealed no meaningful carbapenemase gene bands on the electrophoresis gel.
| Discussion|| |
The imipenem-resistant mechanism of E. aerogenes has been well investigated in Western countries. , However, there is still no report about the emergence of imipenem-resistant E. aerogenes in East Asia. Transport of the carbapenemase gene via plasmids has been reported among different species.  From our antibiotic-sensitive vigilance system, it is noted that the imipenem-resistant rate of P. aeruginosa has increased from 9% (2009) to 24% (2011). There has been no E. aerogenes resistant to imipenem in our record, before 2011. So far, the imipenem-resistant rate of E. aerogenes has been about 1.2% (2/169). Although imipenem-resistant isolates of P. aeruginosa and E. aerogenes had been obtained from the same wound culture concomitantly on hospital day 107, the chronological transfer of plasmids between the two isolates has been ruled out because of their different results in the modified Hodge tests. The carbapenem resistance of these two isolates may be due to the different mechanisms involved.
According to publications in the past two decades, E. aerogenes develops imipenem-resistance in three main mechanisms [Table 2]. They are, (I) reduction in drug uptake due to the loss of porin in the outer membrane, ,, (II) increase in production of carbapenemase, ,, and (III) combination with loss of porin in the outer membrane and production of extended spectrum beta-lactamase (ESBL). , The mechanism of porin loss on the outer membrane has been investigated clearly. ,,,, E. aerogenes is shown to be capable of adapting rapidly to its permeability by regulating the expression of porin, in the previous studies. Long-term use of imipenem has been shown to be associated with in vivo development of porin-deficient mutants.  Moreover, the carbapenem resistance of E. aerogense has been found to be reversible after discontinuing the prescription of carbapenems. , Moreover, Mallea et al. have also described clinical E. aerogenes strains presenting complex resistant strategies associated with β-lactamase production, impermeability, and active efflux pumps. 
|Table 2: Literature review for carbapenem-resistant Enterobacter aerogenes|
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Long-term use of carbapenems against the ESBL- and AmpC-producing E. aerogenes could lead to the emergence of carbapenem resistance. Several β-lactamases, including, Ambler class A (CTX-M-3, CTX-M-14, TEM-1, and SHV-12) and Ambler class D (DHA-1) β-lactamases were reported to result in carbapenem resistance. , Moreover, the specific carbapenemases (Ambler classes A and B) encoded by plasmids were also identified from the strains of E. aerogenes in some Western countries. , The Ambler class A (blaKPC, blaGES ), B (blaIMP, blaVIM, blaGIm, blaSIM, blaSPM, ), and D (blaOXA ) genes were not found in this isolate. Hence, further investigation to detect whether the E. aerogenes isolate contains an unknown carbapenemase is needed.
According to CLSI 2012, the carbapenem-resistant MIC (μg/mL) breakpoints for Enterobacteriaceae shifted from ≥16 μg/ml to ≥4 μg/ml. After applying the revised carbapenem interpretation criteria for Enterobacteriaceae, more strains of imipenem-resistant E. aerogenes were isolated, but none of them produced carbapenemase. This is the first isolate of imipenem-resistant E. aerogenes producing the Ambler class A carbapenemase in Asia. In order to sensitively isolate carbapenem-resistant E. aerogenes, the revised interpretation criteria should be implemented. Moreover, further investigation on the carbapenemase genes from those isolates or other resistant mechanisms should also be conducted.
| Acknowledgments|| |
This study was supported by grants DOD-92-54 from the Department of Defense, Taiwan, Republic of China, and TSGH-C95-40 from the Tri-Service General Hospital.
| Disclosure|| |
All authors declare that they have no competing financial interests.
| References|| |
|1.||Gaston MA. Enterobacter: An emerging nosocomial pathogen. J Hosp Infect 1988;11:197-208. |
|2.||Pitout JD, Moland ES, Sanders CC, Thomson KS, Fitzsimmons SR. Beta-lactamases and detection of beta-lactam resistance in Enterobacter spp. Antimicrob Agents Chemother 1997;41:35-9. |
|3.||Sanders WE Jr., Sanders CC. Enterobacter spp.: Pathogens poised to flourish at the turn of the century. Clin Microbiol Rev 1997;10:220-41. |
|4.||Marchandin H, Godreuil S, Darbas H, Jean-Pierre H, Jumas-Bilak E, Chanal C, et al. Extended-spectrum beta-lactamase TEM-24 in an Aeromonas clinical strain: Acquisition from the prevalent Enterobacter aerogenes clone in France.. Antimicrob Agents Chemother 2003;47:3994-5. |
|5.||Arpin C, Coze C, Rogues AM, Gachie JP, Bebear C, Quentin C. Epidemiological study of an outbreak due to multidrug-resistant Enterobacter aerogenes in a medical intensive care unit. J Clin Microbiol 1996;34:2163-9. |
|6.||Charrel RN, Pages JM, De Micco P, Mallea M. Prevalence of outer membrane porin alteration in beta-lactam-antibiotic-resistant Enterobacter aerogenes. Antimicrob Agents Chemother 1996;40:2854-8. |
|7.||Yigit H, Anderson GJ, Biddle JW, Steward CD, Rasheed JK, Valera LL, et al. Carbapenem resistance in a clinical isolate of Enterobacter aerogenes is associated with decreased expression of OmpF and OmpC porin analogs. Antimicrob Agents Chemother 2002;46:3817-22. |
|8.||Bornet C, Davin-Regli A, Bosi C, Pages JM, Bollet C. Imipenem resistance of Enterobacter aerogenes mediated by outer membrane permeability. J Clin Microbiol 2000;38:1048-52. |
|9.||Yang Q, Wang H, Sun H, Chen H, Xu Y, Chen M. Phenotypic and genotypic characterization of Enterobacteriaceae with decreased susceptibility to carbapenems: Results from large hospital-based surveillance studies in China. Antimicrob Agents Chemother 2010;54:573-7. |
|10.||Biendo M, Canarelli B, Thomas D, Rousseau F, Hamdad F, Adjide C, et al. Successive emergence of extended-spectrum beta-lactamase-producing and carbapenemase-producing Enterobacter aerogenes isolates in a university hospital. J Clin Microbiol 2008;46:1037-44. |
|11.||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;7:88-91. |
|12.||Cohen Stuart J, Leverstein-Van Hall MA. Guideline for phenotypic screening and confirmation of carbapenemases in Enterobacteriaceae. Int J Antimicrob Agents 2010;36:205-10. |
|13.||Tsakris A, Kristo I, Poulou A, Themeli-Digalaki K, Ikonomidis A, Petropoulou D, et al. Evaluation of boronic acid disk tests for differentiating KPC-possessing Klebsiella pneumoniae isolates in the clinical laboratory. J Clin Microbiol 2009;47:362-7. |
|14.||Naas T, Nordmann P. Analysis of a carbapenem-hydrolyzing class A beta-lactamase from Enterobacter cloacae and of its LysR-type regulatory protein. Proc Natl Acad Sci U S A 1994;91:7693-7. |
|15.||Naas T, Vandel L, Sougakoff W, Livermore DM, Nordmann P. Cloning and sequence analysis of the gene for a carbapenem-hydrolyzing class A beta-lactamase, Sme-1, from Serratia marcescens S6. Antimicrob Agents Chemother 1994;38:1262-70. |
|16.||Ellington MJ, Kistler J, Livermore DM, Woodford N. Multiplex PCR for rapid detection of genes encoding acquired metallo-beta-lactamases. J Antimicrob Chemother 2007;59:321-2. |
|17.||Dallenne C, Da Costa A, Decre D, Favier C, Arlet G. Development of a set of multiplex PCR assays for the detection of genes encoding important beta-lactamases in Enterobacteriaceae. J Antimicrob Chemother 2010;65:490-5. |
|18.||Poirel L, Walsh TR, Cuvillier V, Nordmann P. Multiplex PCR for detection of acquired carbapenemase genes. Diagn Microbiol Infect Dis 2011;70:119-23. |
|19.||Vrioni G, Daniil I, Voulgari E, Ranellou K, Koumaki V, Ghirardi S, et al. Comparative Evaluation of a Prototype Chromogenic Medium (ChromID CARBA) for detecting carbapenemase-producing enterobacteriaceae in surveillance rectal swabs. J Clin Microbiol 2012;50:1841-6. |
|20.||Chen Y, Zhou Z, Jiang Y, Yu Y. Emergence of NDM-1-producing Acinetobacter baumannii in China. J Antimicrob Chemother 2011;66:1255-9. |
|21.||Thiolas A, Bollet C, La Scola B, Raoult D, Pages JM. Successive emergence of Enterobacter aerogenes strains resistant to imipenem and colistin in a patient. Antimicrob Agents Chemother 2005;49:1354-8. |
|22.||Bosi C, Davin-Regli A, Bornet C, Mallea M, Pages JM, Bollet C. Most Enterobacter aerogenes strains in France belong to a prevalent clone. J Clin Microbiol 1999;37:2165-9. |
|23.||Arnold RS, Thom KA, Sharma S, Phillips M, Kristie Johnson J, Morgan DJ. Emergence of Klebsiella pneumoniae carbapenemase-producing bacteria. South Med J 2011;104:40-5. |
|24.||Chen YG, Zhang Y, Yu YS, Qu TT, Wei ZQ, Shen P, et al. In vivo development of carbapenem resistance in clinical isolates of Enterobacter aerogenes producing multiple beta-lactamases. Int J Antimicrob Agents 2008;32:302-7. |
|25.||Mallea M, Chevalier J, Bornet C, Eyraud A, Davin-Regli A, Bollet C, et al. Porin alteration and active efflux: Two in vivo drug resistance strategies used by Enterobacter aerogenes. Microbiology 1998;144:3003-9. |
[Table 1], [Table 2]