Heterogeneity of the vanA gene cluster in clinical isolates of enterococci from the northeastern United States

Abstract

In several strains of Enterococcus faecium isolated in Europe, the cluster of genes encoding high-level resistance to vancomycin (VanA phenotype) resides on a 10.85-kb transposon, Tn1546, or closely related elements. To determine whether Tn1546 was conserved in recent enterococcal isolates from the northeastern United States, seven strains were compared by restriction mapping and DNA hybridization with probes from within the van cluster. Two of the seven strains contained intact Tn1546-like sequences; however, in five of the strains, the organization of the van cluster differed from that of Tn1546. Three of the five strains with variations harbored a novel DNA segment within the van gene cluster. This 1,496-bp segment was similar to IS1165 of Leuconostoc mesenteroides and IS1181 of Staphylococcus aureus and was flanked by 24-and 23-bp imperfect inverted repeats and 8-bp direct repeats. On the basis of these findings, we propose that this element comprises a novel insertion-like sequence, IS1251. Multiple copies of IS1251 were also present at other sites in both resistant and susceptible clinical isolates. Our findings suggest that the van cluster in recent isolates from the northeastern United States differs from that present in the early European VanA phenotype strains.Glycopeptide antibiotics such as vancomycin act by binding to the D-alanyl-D-alanine terminus of peptidoglycan precursors, preventing cell wall synthesis (26). It had been widely assumed that this terminus was ubiquitous among eubacterial species producing peptidoglycan and that vancomycin resistance was therefore unlikely to emerge in the absence of a barrier to vancomycin binding, such as the outer membrane of gram-negative bacteria. Nonetheless, vancomycin-resistant isolates of Enterococcus faecium were identified in France in 1986 (22). Resistant enterococcal strains have subsequently been isolated worldwide, and in 1993 the estimated incidence of glycopeptide resistance among isolates of Enterococcus spp. from intensive care units in the United States was 14% (24).Vancomycin resistance in members of several gram-positive genera is due to the production of peptidoglycan precursors terminating in the depsipeptide D-alanyl-D-lactate, to which vancomycin does not bind (1,17,18,23). In highly resistant (VanA phenotype) E. faecium and Enterococcus faecalis strains, production of this altered precursor is encoded by a plasmid-borne cluster of genes. Insertional mutagenesis experiments with two vancomycin resistance plasmids, pIP816 and pHKK100, have shown that four genes in the cluster are required for resistance : vanR, vanH, vanA, and vanX (3, 4, 15). vanR and vanS act as a sensor-regulatory pair analogous to such previously characterized systems as ompR-envZ and are required for the induction of resistance (3, 29). VanH, a D-specific ␣-keto acid reductase (8), and VanA, a D-alanine-Dalanine ligase of altered specificity (7), act in concert to form the depsipeptide D-alanyl-D-lactate, which is added in place of D-alanyl-D-alanine to the UDP-muram...

Genetic Rearrangements of Tn 1546 -Like Elements in Vancomycin-Resistant Enterococcus faecium Isolates Collected from Hospitalized Patients over a Seven-Year Period

“…The heterogeneity of the Tn1546-like element results from point mutations, deletions, and the integration of insertion sequence (IS) elements (16,17); and among these variations, the presence of IS elements accounts for much of the heterogeneity (3). Such a rearrangement has been shown to play a key role in the evolution of the vanA gene cluster and may modify its transferability (7,8,11). In the present study, we characterized the consequence of Tn1546 in vanA-containing Enterococcus faecium isolates collected from patients over time passed to obtain an understanding of the hospital dissemination of VRE over time.…”

Characterization of Tn 1546 in Vancomycin-Resistant Enterococcus faecium Isolated from Canine Urinary Tract Infections: Evidence of Gene Exchange between Human and Animal Enterococci

“…The concurrent appearance of vanA in both Europe and the United States, despite the lack of avoparcin use in the United States, suggests that the epidemiology and global dissemination of vanA are complex (38,40). Tn1546 is often associated with plasmids, and several reports have emphasized the fact that there are polymorphisms between Tn1546 elements due to insertions, deletions, and point mutations (10,13,14,(36)(37)(38)(39)42).…”

Interruption of the Streptococcus gordonii M5 sspA/sspB intergenic region by an insertion sequence related to IS1167 of Streptococcus pneumoniae

“…This is consistent with the findings of Zhou et al (1995) who reported that IS1167 was similar to a short non-coding region in our previously published nucleotide sequence of the M5 sspB gene (Demuth et al, 1990a (Bennett, 1991). The putative transposase polypeptide is highly similar (85-95% sequence identity) to regions of both ORFl and ORF2 of IS1167 and exhibits 30-4o0/0 sequence identity with transposases encoded by IS1 165 (Johansen & Kibenich, 1992), IS1181 (Derbise et al, 1994), IS1251 (Handwerger et al, 1995) (Chandler & E:ayet, 1993). Whether the lack of these sequences in the ISSgl transposase influences its activity remains to be determined.…”