First characterization of Tn1546-like structures of vancomycin-resistant Enterococcus faecium Thai isolates.

Introduction: Vancomycin-resistant Enterococcus faecium (VREfm) carrying vanA was first isolated from patient at Siriraj Hospital, Thailand in 2004. Since then, VREfm isolates have been detected increasingly in this 2500-bed university hospital. To understand the epidemiology of vanA VREfm in this setting, the isolates collected during 2004-2013 were characterized.

Methods: A total of 49 vanA VREfm isolates previously confirmed by multiplex PCR were characterized by determining resistance phenotypes to vancomycin, teicoplanin, ampicillin and ciprofloxacin by broth microdilution method. Multilocus sequence typing (MLST) and virulence genes of those isolates were investigated. The Tn1546 structure diversity was studied by long-range overlapping PCR and primer walking sequencing.

Results: Of all isolates studied, 9 sequence types (ST17, ST80, ST78, ST730, ST203, ST18, ST280, ST64, ST323) in clonal complex 17 and a novel ST1051 were revealed. The esp-positive isolates were 73.5%. Of all vanA operons characterized, at least 9 types of Tn1546-like structures were detected. All of vanA determinants contained 5'-end different from the Tn1546 prototype. Approximately 47% of them also carried the insertion sequence IS1251 at the intergenic region between vanS and vanH. Interestingly, another IS (ISEfa4) was found to be inside the sequence of IS1251 in ST17 isolate.

Conclusion: Heterogeneity of vanA VREfm was observed. Nearly all of isolates studied belonged to CC17. One novel ST1051 strain was detected. Isolates in the initial period carried vanA operon similar to the prototype. The diversity of vanA determinants has been increased in the recent isolates. A novel vanA operon structure was detected.