Heterogeneity in the vanB Gene Cluster of Genomically Diverse Clinical Strains of Vancomycin-Resistant Enterococci

Molecular analysis of 17 genomically unrelated clinical VanB-type vancomycin-resistant enterococcus isolates from hospital patients in Germany, Norway, Sweden, the United Kingdom and the United States revealed three subtypes of thevanB gene cluster—vanB1 ,vanB2 , andvanB3 —which was in accordance with previous subtyping of the ligase gene sequence. There was no correlation betweenvanB subtype and levels of vancomycin resistance. All strains studied carried a structurally conservedvanB gene cluster as shown by long-range PCR (long PCR) covering 5,959 bp of the published sequence invanB1 strain V583. Restriction analysis of long PCR amplicons displayed one uniquevanB1 pattern and a secondvanB2 - andvanB3 -specific pattern. ThevanSB -vanYB intergenic sequences with flanking coding regions were identical within eachvanB subtype with one exception. A U.S.vanB2 isolate had a 789-bp enlargement of this region containing a putative open reading frame (ORF) with substantial homology to an ORF in theClostridium perfringens IS1469 insertion element. The molecular heterogeneity within thevanB gene cluster has implications for the selection of PCR primers, as the primers must ensure detection of allvanB subtypes, and is of importance when considering reservoirs and dissemination ofvanB resistance. The molecular identity within thevanB1 and thevanB2 subtype indicates horizontal transmission of both gene clusters between isolates in different geographical areas. Restriction analysis of long PCRvanB amplicons may reveal specific varieties that can be used as epidemiological markers for mobile determinants conferring VanB-type resistance. The finding of three distinctvanB gene clusters should encourage a search for different environmental reservoirs ofvanB resistance determinants.