Background: Primary screening for VRE with PCR directed against vanA allowed identification of vanA+ samples from which VRE could not be isolated when selective culture methods were used. From such a sample a vancomycin-susceptible, vanA+ Enterococcus faecium, Efm-V1511, was isolated, when vancomycin selection was not used during culture. Similar isolates with variable susceptibility to vancomycin were obtained in the following months. Objectives: To characterize Efm-V1511 and investigate the causes of variable susceptibility to vancomycin. Methods: All strains were sequenced using Illumina technology. Plasmids containing vanA were reconstructed by scaffolding to known plasmids or plasmids were sequenced using Oxford Nanopore MinION. Derived structures were verified by PCR and sequencing. Furthermore, selected vanA+ vancomycin-susceptible isolates were passaged in the presence of vancomycin and vancomycin-resistant variants obtained were sequenced. Results: Efm-V1511 belonged to ST1421 and contained a 49 696 bp plasmid pHVH-V1511 carrying a Tn1546-derived genetic element. Within this element vanX was truncated by a 252 bp 3' deletion explaining the susceptibility of Efm-V1511. Between March 2016 and April 2017, 48 isolates containing pHVH-V1511 were identified. All were ST1421. In isolates resistant to vancomycin, resistance could be attributed to changes in ddl disrupting gene function sometimes accompanied by changes in vanS, increased pHVH-V1511 copy number or the existence of an additional vanA-containing plasmid encoding a functional vanX. Conclusions: E. faecium carrying pHVH-V1511 is capable of nosocomial transmission and may develop clinical resistance to vancomycin. Strains may not be detected using standard culture methods for VRE.
Background: Primary screening for VRE with PCR directed against vanA allowed identification of vanA+ samples from which VRE could not be isolated when selective culture methods were used. From such a sample a vancomycin-susceptible, vanA+ Enterococcus faecium, Efm-V1511, was isolated, when vancomycin selection was not used during culture. Similar isolates with variable susceptibility to vancomycin were obtained in the following months. Objectives: To characterize Efm-V1511 and investigate the causes of variable susceptibility to vancomycin. Methods: All strains were sequenced using Illumina technology. Plasmids containing vanA were reconstructed by scaffolding to known plasmids or plasmids were sequenced using Oxford Nanopore MinION. Derived structures were verified by PCR and sequencing. Furthermore, selected vanA+ vancomycin-susceptible isolates were passaged in the presence of vancomycin and vancomycin-resistant variants obtained were sequenced. Results: Efm-V1511 belonged to ST1421 and contained a 49 696 bp plasmid pHVH-V1511 carrying a Tn1546-derived genetic element. Within this element vanX was truncated by a 252 bp 3' deletion explaining the susceptibility of Efm-V1511. Between March 2016 and April 2017, 48 isolates containing pHVH-V1511 were identified. All were ST1421. In isolates resistant to vancomycin, resistance could be attributed to changes in ddl disrupting gene function sometimes accompanied by changes in vanS, increased pHVH-V1511 copy number or the existence of an additional vanA-containing plasmid encoding a functional vanX. Conclusions: E. faecium carrying pHVH-V1511 is capable of nosocomial transmission and may develop clinical resistance to vancomycin. Strains may not be detected using standard culture methods for VRE.
Authors: Lauren M Petersen; Isabella W Martin; Wayne E Moschetti; Colleen M Kershaw; Gregory J Tsongalis Journal: J Clin Microbiol Date: 2019-12-23 Impact factor: 5.948