Different IncI1 plasmids from Escherichia coli carry ISEcp1-blaCTX-M-15 associated with different Tn2-derived elements
Introduction
The CTX-M extended spectrum β-lactamases (ESBL), particularly CTX-M-15, have become dominant worldwide in recent years (D'Andrea et al., 2013). blaCTX-M-3, the apparent progenitor of blaCTX-M-15, appears to have been captured from the Kluyvera ascorbata chromosome by the insertion sequence ISEcp1 (Rodríguez et al., 2004). ISEcp1 is bounded by 14-bp inverted repeats (IRL and IRR) but uses IRL in conjunction with alternative sequences resembling these IR to mobilise adjacent regions, creating 5-bp direct repeats (DR) of flanking sequence on transposition and ISEcp1 also provides a promoter for expression of adjacent genes (Poirel et al, 2003, Poirel et al, 2005).
blaCTX-M-15 has generally been found 48 bp beyond the IRR end of ISEcp1 in a 2.971-kb transposition unit containing a 1.315-kb fragment of the K. ascorbata chromosome that also includes a partial open reading frame, designated orf477Δ (Fig. 1A). In IncF plasmids, which commonly carry blaCTX-M-15 in Escherichia coli, this transposition unit is inserted in the transposon Tn2, which carries blaTEM-1b (Bailey et al, 2011, Partridge, Hall, 2005). Tn2 also includes tnpA (transposase) and tnpR (resolvase) genes and a resolution (res) site and is flanked by 38-bp inverted repeats, designated IRtnp and IRTEM here. In these IncF plasmids, the IRtnp end of Tn2 is truncated by IS26 (Fig. 1B) and this structure has been found as part of large multiresistance regions (MRR) that also include different combinations of the blaOXA-30, aac(3)-IIe (Partridge, 2011), aac(6′)-Ib-cr and tetA(A) genes and sometimes a class 1 integron and other resistance genes (Boyd et al, 2004, Partridge et al, 2011b, Woodford et al, 2009). blaCTX-M-15 is often associated with E. coli multi-locus sequence type (ST) 131, particularly when carried on IncF plasmids, and with clonal complex CC405 (D'Andrea et al, 2013, Naseer, Sundsfjord, 2011).
blaCTX-M-15 has also been found on IncI1 plasmids, but generally with fewer additional resistance genes than in IncF plasmids (e.g. Hopkins et al., 2006) and a few IncI1 plasmids carrying blaCTX-M-15 have now been completely sequenced. A survey demonstrating the dominance of blaCTX-M genes in clinical isolates from western Sydney, Australia in 2005–2007 revealed that five E. coli with four different pulsed-field gel electrophoresis (PFGE) profiles carried blaCTX-M-15 on an IncI1 plasmid (Zong et al., 2008). Here, we have completely sequenced three of these plasmids to compare the blaCTX-M-15 contexts with those on IncF plasmids and the plasmid backbones with other IncI1 plasmids.
Section snippets
Bacterial strains
Five E. coli (JIE113, JIE139, JIE174, JIE236 and JIE242) carrying blaCTX-M-15 and an IncI1 replicon isolated in June 2006–January 2007 from urine samples from different patients were from a larger collection of Enterobacteriaceae clinical isolates referred for microbiological testing at Westmead Hospital, Sydney, Australia (Zong et al., 2008). As the PFGE fingerprints of JIE236 and JIE242 were identical (Zong et al., 2008), only JIE242 was studied further. Multi-locus sequencing typing (MLST)
blaCTX-M-15 is carried by different strains and different IncI1 plasmids
By MLST, JIE113 is ST448, both JIE139 and JIE242 are ST69, and JIE174 is the novel ST2495. This contrasts with the isolates from the same collection with blaCTX-M-15 on IncF or IncX4 plasmids, which were either ST131 (n = 8) or ST405 (n = 4) (Partridge et al, 2011a, Partridge et al, 2011b), although ST2495 differs from ST405 by only 3 nucleotide changes in a single allele, gyrB. ST69 is part of E. coli “clonal group A” (CgA), originally identified in a community outbreak of urinary tract
Conclusions
Comparison of the plasmids analysed here with the well-characterised R64 and R621a builds on the conclusions of Takahashi et al. (2011) about a conserved overall organisation of IncI1 plasmids with various insertions, deletions and substitutions that may influence compatibility and other functions. The ST31 and ST37 sequences also suggest that, unlike the case of ST2 plasmids carrying blaCMY-2 (Tagg et al., 2014), plasmids assigned to at least some other pMLST groups include backbones that are
Acknowledgments
Z.Z. was supported by an Endeavour International postgraduate Research Scholarship from the Australian Government Department of Education, Science and Training. Plasmid sequencing was funded by a Research Grant from the Australian Society for Antimicrobials. This work was supported by grants G1001021, G1002076 and G1046886 from the Australian National Health and Medical Research Council. We thank Carola Venturini for preparing plasmid DNA, Joey Lai for library preparation and Michael Roper and
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Communicated by Julian Rood.
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Permanent address: Department of Biology and Wildlife Diseases, Faculty of Veterinary Hygiene and Ecology, University of Veterinary and Pharmaceutical Sciences Brno, Brno, Czech Republic; CEITEC VFU, University of Veterinary and Pharmaceutical Sciences Brno, Brno, Czech Republic.