JAC Advance Access originally published online on July 29, 2003
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Journal of Antimicrobial Chemotherapy (2003) 52, 528-529
© 2003 The British Society for Antimicrobial Chemotherapy
Correspondence |
Detection of CTX-M-15 extended-spectrum ß-lactamase in the United Kingdom
Antibiotic Resistance Monitoring and Reference Laboratory, Specialist and Reference Microbiology Division, Health Protection Agency—Colindale, London NW9 5HT, UK
Keywords: ESBLs, CTX-M ß-lactamases, Escherichia coli, cefpodoxime
Sir,
Extended-spectrum ß-lactamases (ESBLs) are a major source of resistance to oxyimino-cephalosporins in Gram-negative bacteria. Most ESBLs are mutants of the classical TEM and SHV enzymes, but other classes are emerging. Most importantly, these non-TEM/SHV ESBLs include the CTX-M family, which includes at least 27 alleles, divided between four major sequence sub-types (see http://www.lahey.org/studies/other.stm#table1). Some or all of the CTX-M enzymes have evolved via the genetic escape and mutation of the chromosomal ß-lactamases of Kluyvera spp.1 CTX-M enzymes predominantly attack cefotaxime, and most are only weakly active against ceftazidime. They are most prevalent in South America, but are appearing in Europe, with major outbreaks in Poland and Spain. Recently, CTX-M-type ß-lactamases have been reported for the first time in the UK,2,3 with a major outbreak around Birmingham.2 We sought to investigate their presence among selected ESBL-producing isolates of Enterobacteriaceae collected in a survey of bacteria from hospitalized patients in the UK and Ireland.4
Twenty-six hospitals in the UK and Ireland collected up to 200 consecutive clinically-significant isolates from separate inpatients during a multicentre survey of inpatient isolates, conducted in 2001.4 Disc testing was carried out at the hospital laboratories, and isolates with key resistances, including to third-generation cephalosporins, were sent to the Antibiotic Resistance Monitoring and Reference Laboratory for MIC determinations by the BSAC agar dilution method. Isolates of Enterobacteriaceae with cefotaxime MICs at least eight-fold higher than ceftazidime MICs (except Serratia spp. where this phenotype is AmpC-associated) were screened for blaCTX-M genes by PCR using universal primers.5 PCR products were sequenced on both strands to determine which CTX-M sub-type was present. Sub-type-specific primers5,6 were then used to amplify and sequence the entire gene. blaCTX-M-positive isolates were compared by pulsed-field gel electrophoresis (PFGE) of XbaI-digested DNA.
Among 122 cephalosporin-resistant isolates of Enterobacteriaceae collected, seven (6%) demonstrated cefotaxime MICs at least eight-fold greater than those of ceftazidime. These were five isolates of Escherichia coli, and one isolate each of Enterobacter sp. and Klebsiella sp. Three of the E. coli isolates, with cefotaxime MICs 
256 mg/L and ceftazidime MICs 32 mg/L, yielded a 544-bp amplicon with universal blaCTX-M primers, as also did one E. coli isolate highly resistant to both cefotaxime and ceftazidime (MICs 128 mg/L). The last isolate was screened on the basis that, with MICs above the upper limits tested, it might fulfil the inclusion criteria. Sequencing of entire blaCTX-M alleles revealed that all four E. coli isolates produced CTX-M-15 (also known as UOE-1) enzyme, which differs from CTX-M-3 only by an Asp-240 
Gly substitution.6 The four CTX-M-15-producing isolates were collected at three hospitals, in London (two isolates), Newcastle-upon-Tyne and Belfast. All four producers had unique PFGE profiles, including the two isolates from the hospital in London (Figure 1), indicating that they had acquired the resistance gene independently.
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CTX-M-15 is one of the CTX-M types with greatest relative activity against ceftazidime, perhaps explaining why the isolates were clearly resistant to this drug as well as cefotaxime. Nevertheless, the high-level resistance to ceftazidime seen in one of the E. coli isolates (MIC > 128 mg/L) suggests the presence of other mechanisms in addition to CTX-M-15.
In conclusion, CTX-M-15 enzyme has appeared in distinct strains of E. coli from at least three centres in the UK. It has previously been reported from Japan, Bulgaria and Poland, India and France, whilst its close relative, CTX-M-3, is more widely distributed. Previously, CTX-M-9 and a CTX-M-25-like enzyme were reported in isolates of Klebsiella spp. in the UK,2,3 but these and CTX-M-15 each belong to distinct and different CTX-M sub-types. Thus the CTX-M-producers reported in the UK result from independent events in which Enterobacteriaceae have acquired three different blaCTX-M alleles. Many laboratories in the UK screen for ESBL production using only ceftazidime, then carrying out a cephalosporin-clavulanate synergy test on resistant isolates. The emergence of CTX-M enzymes suggests that this practice may lead to under-reporting. Diagnostic laboratories should screen for ESBLs either with both cefotaxime and ceftazidime or, alternatively, with cefpodoxime; they should still perform clavulanate synergy tests on resistant isolates.
Acknowledgements
We thank Jane Turton for her assistance with PFGE. This work was supported by Wyeth UK, to whom we are grateful.
Footnotes
* Corresponding author. Tel: +44-20-8200-4400; Fax: +44-20-8358-3292; E-mail: neil.woodford@hpa.org.uk 
References
1
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Poirel, L., Kampfer, P. & Nordmann, P. (2002). Chromosome-encoded Ambler class A ß-lactamase of Kluyvera georgiana, a probable progenitor of a subgroup of CTX-M extended-spectrum ß-lactamases. Antimicrobial Agents and Chemotherapy 46, 4038–40.
2
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Brenwald, N. P., Jevons, G., Andrews, J. M. et al. (2003). An outbreak of a CTX-M-type ß-lactamase-producing Klebsiella pneumoniae: the importance of using cefpodoxime to detect extended-spectrum ß-lactamases. Journal of Antimicrobial Chemotherapy 51, 195–6.
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Alobwede, I., M’Zali, F. H., Livermore, D. M. et al. (2003). CTX-M extended-spectrum ß-lactamase arrives in the UK. Journal of Antimicrobial Chemotherapy 51, 470–1.
4 . Livermore, D. M., Mushtaq, S., James, D. et al. (2003). In-vitro activity of piperacillin/tazobactam and other broad-spectrum antibiotics against bacteria from hospitalised patients in the British Isles. International Journal of Antimicrobial Agents, in press.
5 . Saladin, M., Cao, V. T., Lambert, T. et al. (2002). Diversity of CTX-M ß-lactamases and their promoter regions from Enterobacteriaceae isolated in three Parisian hospitals. FEMS Microbiology Letters 209, 161–8.[Web of Science][Medline]
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