LETTER
The blaOXA-48 gene has been detected in a variety of species and clones of Enterobacteriaceae, circulating mainly throughout the Mediterranean area but progressively disseminating to other geographical areas (1, 2). To our knowledge, we describe here the first cases of OXA-48-producing Enterobacteriaceae in Portugal.
Enterobacter cloacae and Escherichia coli were isolated from the urine of a catheterized patient presenting with pyuria, hematuria, and positive nitrites. The patient was a 74-year-old Caucasian female, admitted at the emergency room of a hospital in Lisbon, Portugal, in March 2013, who was diagnosed with decompensated heart failure and who had been previously treated with the combination amoxicillin-clavulanate, due to infected leg ulcers. In the patient's clinical history, we noted a left nephrectomy for oncocytoma, a nonfunctional right adrenal adenoma, the use of medication for high blood pressure, and several previous admissions for cardiac decompensation.
The two isolates were sent to the National Institute of Health due to their carbapenem resistance, where the genotype was studied to determine the main resistance mechanisms. By the microdilution method, both isolates were found to be nonsusceptible to different classes of antibiotics, with susceptibility to colistin and tigecycline (http://www.eucast.org/clinical_breakpoints/) (Table 1). Conjugation experiments were performed using E. coli J53 (sodium azide resistant) as the recipient, as previously described (3). E. coli J53 Tc17313-1 and E. coli J53 Tc17313-2 transconjugants were resistant to penicillins and ertapenem but remained susceptible to oximino-β-lactams; both transconjugants were additionally nonsusceptible to meropenem and gentamicin, respectively (Table 1).
TABLE 1.
Antimicrobial drugb | MIC (μg/ml) for strain: |
||||
---|---|---|---|---|---|
E. coli J53 | E. cloacae INSRA17313-1 (OXA-48, ACT-34) | E. coli J53 Tc17313-1 (OXA-48) | E. coli INSRA17313-2 (OXA-48, QnrD) | E. coli J53 Tc17313-2 (OXA-48) | |
Amoxicillin | 4 | >256 | >256 | >256 | >256 |
Amoxicillin-clavulanate | 4 | 256 | 64 | 128 | 64 |
Ticarcillin | ≤4 | 1,024 | 128 | 512 | 256 |
Cefuroxime | 2 | 512 | 4 | 8 | 4 |
Cefoxitine | 8 | 512 | 8 | 16 | 8 |
Ceftazidime | ≤0.5 | 64 | ≤0.5 | 4 | ≤0.5 |
Ceftazidime-clavulanate | ≤0.5 | 16 | ≤0.5 | ≤0.5 | ≤0.5 |
Cefotaxime | ≤0.5 | 512 | 2 | 2 | 1 |
Cefotaxime-clavulanate | ≤0.5 | 256 | 1 | 2 | ≤0.5 |
Aztreonam | ≤0.5 | 128 | ≤0.5 | ≤0.5 | ≤0.5 |
Cefepime | ≤1 | 1,024 | ≤1 | 16 | ≤1 |
Imipenem | ≤0.5 | 4 | 1 | 1 | 1 |
Meropenem | ≤0.125 | 8 | 4 | 2 | 2 |
Doripenem | ≤0.125 | 2 | ≤0.125 | ≤0.125 | ≤0.125 |
Ertapenem | ≤0.06 | >64 | 4 | 4 | 2 |
Ciprofloxacin | ≤0.06 | 2 | ≤0.06 | ≤0.06 | ≤0.06 |
Gentamicin | 2 | 32 | 2 | 8 | 8 |
Trimethoprim | ≤2 | >4,096 | ≤2 | 8 | ≤2 |
Colistin | 0.5 | 1 | 0.25 | 0.25 | 0.25 |
Tigecycline | 0.25 | 0.5 | 0.125 | 0.125 | 0.125 |
E. coli J53 Tc17313-1 (harboring OXA-48) and E. coli J53 Tc17313-2 (harboring OXA-48) were transconjugants of E. cloacae INSRA17313-1 (harboring OXA-48 and ACT-34) and E. coli INSRA17313-2 (harboring OXA-48 and QnrD), respectively; E. coli J53 was the recipient strain.
The combinations with clavulanate included that drug at 2 μg/ml.
PCR amplification and sequencing of the most prevalent β-lactamase- and plasmid-mediated quinolone resistance (PMQR)-encoding genes, performed as previously described (3), allowed the identification of the blaOXA-48 gene in both isolates and transconjugants; the chromosomal blaACT-34 variant and the qnrD genes were detected in E. cloacae and E. coli clinical isolates, respectively (Table 1). This is the first description of QnrD-producing E. coli in Europe (4).
By PCR mapping (5), we confirmed the presence of a Tn1999.2 structure containing the blaOXA-48 gene. This composite transposon, located in a conjugative plasmid, encompassed two copies of the insertion sequence IS1999, with IS1R truncating the left-hand copy of the transposase. This Tn1999.2 structure has been described to be associated with higher carbapenem MICs than Tn1999.1 (5, 6). Indeed, the blaOXA-48 gene is usually part of Tn1999 and located in a plasmid (6, 7), although a chromosome-mediated blaOXA-48 gene has recently been found in E. coli (8).
None of the plasmids carried by the clinical isolates and transconjugants belonged to an incompatibility group identified by a PCR-based replicon typing method (9). However, we detected, as previously described (10), the presence of the repA, traU, and parA genes, suggesting that those plasmids had an IncL/IncM-pOXA-48a-like backbone (10). The OXA-48-producing E. coli isolate belonged to the ST1429 lineage, based on multilocus sequence typing (http://mlst.warwick.ac.uk/mlst/dbs/Ecoli). This sequence type was previously reported in Sweden in 2004 in an E. coli isolate that did not produce any carbapenemase (http://mlst.warwick.ac.uk/mlst/dbs/Ecoli).
In conclusion, this study shows the presence of OXA-48-producing Enterobacteriaceae in Portugal. Although we could not link our patient with a history of travel to countries of endemicity and no other case was identified in that hospital, we highlight the potential of interspecies dissemination of blaOXA-48 gene-harboring plasmids and, consequently, the importance of concerted actions to manage carbapenem resistance.
Nucleotide sequence accession number.
The new blaACT nucleotide sequence was submitted to the EMBL nucleotide sequence database as blaACT-34 under accession number HG975300.
ACKNOWLEDGMENT
The authors thank Fundação para a Ciência e a Tecnologia (FCT) for project grant PEst-OE/AGR/UI0211/2011-2014, Strategic Project UI211-2011-2014. V. Manageiro was supported by grant SFRH/BPD/77486/2011 from FCT, Lisbon, Portugal.
Footnotes
Published ahead of print 22 September 2014
REFERENCES
- 1.Poirel L, Potron A, Nordmann P. 2012. OXA-48-like carbapenemases: the phantom menace. J. Antimicrob. Chemother. 67:1597–1606. 10.1093/jac/dks121. [DOI] [PubMed] [Google Scholar]
- 2.Patel G, Bonomo RA. 2013. “Stormy waters ahead”: global emergence of carbapenemases. Front. Microbiol. 4:48. 10.3389/fmicb.2013.00048. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 3.Manageiro V, Ferreira E, Caniça M, Manaia CM. 2014. GES-5 among the β-lactamases detected in ubiquitous bacteria isolated from aquatic environment samples. FEMS Microbiol. Lett. 351:64–69. 10.1111/1574-6968.12340. [DOI] [PubMed] [Google Scholar]
- 4.Guillard T, Grillon A, de Champs C, Cartier C, Madoux J, Berçot B, Lebreil AL, Lozniewski A, Riahi J, Vernet-Garnier V, Cambau E. 2014. Mobile insertion cassette elements found in small non-transmissible plasmids in Proteeae may explain qnrD mobilization. PLoS One 9:e87801. 10.1371/journal.pone.0087801. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 5.Giani T, Conte V, Di Pilato V, Aschbacher R, Weber C, Larcher C, Rossolini GM. 2012. Escherichia coli from Italy producing OXA-48 carbapenemase encoded by a novel Tn1999 transposon derivative. Antimicrob. Agents Chemother. 56:2211–2213. 10.1128/AAC.00035-12. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6.Carrër A, Poirel L, Yilmaz M, Akan OA, Feriha C, Cuzon G, Matar G, Honderlick P, Nordmann P. 2010. Spread of OXA-48-encoding plasmid in Turkey and beyond. Antimicrob. Agents Chemother. 54:1369–1373. 10.1128/AAC.01312-09. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 7.Aubert D, Naas T, Héritier C, Poirel L, Nordmann P. 2006. Functional characterization of IS1999, an IS4 family element involved in mobilization and expression of β-lactam resistance genes. J. Bacteriol. 188:6506–6514. 10.1128/JB.00375-06. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 8.Beyrouthy R, Robin F, Cougnoux A, Dalmasso G, Darfeuille-Michaud A, Mallat H, Dabboussi F, Hamzé M, Bonnet R. 2013. Chromosome-mediated OXA-48 carbapenemase in highly virulent Escherichia coli. J. Antimicrob. Chemother. 68:1558–1561. 10.1093/jac/dkt051. [DOI] [PubMed] [Google Scholar]
- 9.Carattoli A, Bertini A, Villa L, Falbo V, Hopkins KL, Threlfall EJ. 2005. Identification of plasmids by PCR-based replicon typing. J. Microbiol. Methods 63:219–228. 10.1016/j.mimet.2005.03.018. [DOI] [PubMed] [Google Scholar]
- 10.Poirel L, Bonnin RA, Nordmann P. 2012. Genetic features of the widespread plasmid coding for the carbapenemase OXA-48. Antimicrob. Agents Chemother. 56:559–562. 10.1128/AAC.05289-11. [DOI] [PMC free article] [PubMed] [Google Scholar]