International Spread and Persistence of TEM-24 Is Caused by the Confluence of Highly Penetrating Enterobacteriaceae Clones and an IncA/C 2 Plasmid Containing Tn 1696 ::Tn 1 and IS 5075 -Tn 21

Abstract

ABSTRACT TEM-24 remains one of the most widespread TEM-type extended-spectrum β-lactamases (ESBLs) among Enterobacteriaceae . To analyze the reasons influencing its spread and persistence, a multilevel population genetics study was carried out on 28 representative TEM-24 producers from Belgium, France, Portugal, and Spain (13 Enterobacter aerogenes isolates, 6 Escherichia coli isolates, 6 Klebsiella pneumoniae isolates, 2 Proteus mirabilis isolates, and 1 Klebsiella oxytoca isolate, from 1998 to 2004). Clonal relatedness (XbaI pulsed-field gel electrophoresis [PFGE] and E. coli phylogroups) and antibiotic susceptibility were determined by standard procedures. Plasmid analysis included determination of the incompatibility group (by PCR, hybridization, and/or sequencing) and comparison of restriction fragment length polymorphism (RFLP) patterns. Characterization of genetic elements conferring antibiotic resistance included integrons (classes 1, 2, and 3) and transposons (Tn 3 , Tn 21 , and Tn 402 ). Similar PFGE patterns were identified among E. aerogenes , K. pneumoniae , and P. mirabilis isolates, while E. coli strains were diverse (phylogenetic groups A, B2, and D). Highly related 180-kb IncA/C 2 plasmids conferring resistance to kanamycin, tobramycin, chloramphenicol, trimethoprim, and sulfonamides were identified. Each plasmid contained defective In0-Tn 402 ( dfrA1 - aadA1 , aacA4 , or aacA4 - aacC1 - orfE - aadA2 - cmlA1 ) and In4-Tn 402 ( aacA4 or dfrA1 - aadA1 ) variants. These integrons were located within Tn 21 , Tn 1696 , or hybrids of these transposons, with IS 5075 interrupting their IR tnp and IR mer . In all cases, bla TEM-24 was part of an IS 5075 -ΔTn 1 transposon within tnp 1696 , mimicking other genetic elements containing bla TEM-2 and bla TEM-3 variants. The international dissemination of TEM-24 is fuelled by an IncA/C 2 plasmid acquired by different enterobacterial clones which seem to evolve by gaining diverse genetic elements. This work highlights the risks of a confluence between highly penetrating clones and highly promiscuous plasmids in the spread of antibiotic resistance, and it contributes to the elucidation of the origin and evolution of TEM-2 ESBL derivatives.