Relevance of hot spots in the evolution and transmission of Tn1546 in glycopeptide-resistant Enterococcus faecium (GREF) from broiler origin.
OBJECTIVE
Glycopeptide-resistant enterococci are still present within the broiler sector, despite the EU ban of avoparcin more than a decade ago. In the present study, we have developed a rapid method for screening the flanking regions at the integration point of Tn1546 in glycopeptide-resistant Enterococcus faecium isolated from broiler farms.


METHODS
Total DNA was digested, ligated and amplified using primers from inside Tn1546. The resulting amplicons were purified and sequenced. Two new primers were designed based on obtained sequences.


RESULTS
Two main insertion points have been repeatedly found in isolates from the UK (n = 150). The first insertion point revealed that 25 isolates harboured Tn1546 positioned in a sequence with 96% homology to a streptomycin adenyltransferase gene (AY604739) from a Staphylococcus intermedius plasmid. At this insertion point, a direct repeat (GTCCT) was duplicated as previously described, indicating transposition at the target site. Furthermore, this 'hot spot' was also detected in isolates from Norway (2/8) and Denmark (17/20). The second insertion point detected in 45 isolates from the UK revealed integration into an Inc18-like plasmid, most likely by a process of target site recombination.


CONCLUSIONS
The presence of a common insertion point for isolates from different geographical areas could suggest the insertion of Tn1546 by transposition in a plasmid-specific site, followed by genetic rearrangement both inside the transposon and in the flanking regions.
Garcia-Migura
Lourdes
L
National Food Institute, DTU, Antimicrobial Resistance Unit, Bülowsvej 27, DK-1790 Copenhagen V, Denmark. lgmi@food.dtu.dk
Hasman
Henrik
H
Svendsen
Christina
C
Jensen
Lars Bogø
LB
D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001483:Base Sequence; D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D003718:Denmark; D024881:Drug Resistance, Bacterial; D016984:Enterococcus faecium / Q000187:drug effects* / Q000235:genetics* / Q000302:isolation & purification; D023061:Gene Order; D005798:Genes, Bacterial; D006020:Glycopeptides / Q000494:pharmacology*; D008969:Molecular Sequence Data; D009664:Norway; D009713:Nucleotidyltransferases / Q000235:genetics; D010957:Plasmids; D011200:Poultry / Q000382:microbiology*; D011995:Recombination, Genetic; D012091:Repetitive Sequences, Nucleic Acid; D017422:Sequence Analysis, DNA; D006113:United Kingdom
D000900:Anti-Bacterial Agents; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D006020:Glycopeptides; D009713:Nucleotidyltransferases; C022678:streptomycin 3''-adenylyltransferase