Transferable vancomycin resistance in a community-associated MRSA lineage

Abstract

We report the case of a patient from Brazil with a bloodstream infection caused by a strain of methicillin-resistant Staphylococcus aureus (MRSA) that was susceptible to vancomycin (designated BR-VSSA) but that acquired the vanA gene cluster during antibiotic therapy and became resistant to vancomycin (designated BR-VRSA). Both strains belong to the sequence type (ST) 8 community-associated genetic lineage that carries the staphylococcal chromosomal cassette mec (SCCmec) type IVa and the S. aureus protein A gene (spa) type t292 and are phylogenetically related to MRSA lineage USA300. A conjugative plasmid of 55,706 bp (pBRZ01) carrying the vanA cluster was identified and readily transferred to other staphylococci. The pBRZ01 plasmid harbors DNA sequences that are typical of the plasmid-associated replication genes rep24 or rep21 described in community-associated MRSA strains from Australia (pWBG745). The presence and dissemination of community-associated MRSA containing vanA could become a serious public health concern.

Figure 1. The Patient’s Clinical Course before and after Isolation of the Vancomycin-Resistant Staphylococcus aureus

The antibiotics used are depicted in colored rectangles, with beta-lactams in blue (cephalexin, cefepime, and piperacillin–tazobactam), clindamycin in green, glycopeptides in pink (vancomycin and teicoplanin), and daptomycin in yellow; the number in each rectangle corresponds to the number of days of treatment with the antibiotic. The drugs are shown in the order in which they were added to therapy. Dosages were as follows: cephalexin, 500 mg every 6 hours, given orally; clindamycin, 450 mg every 8 hours, given orally; cefepime, 1 g every 8 hours, given intravenously; piperacillin–tazobactam, 3.375 g every 6 hours, given intravenously; vancomycin, 1 g every 12 hours, given intravenously; teicoplanin, 400 mg daily, given intravenously; and daptomycin, 6 mg per kilogram of total body weight daily, given intravenously. The final days of hospitalization are not included. SSTI denotes skin and soft-tissue infection, BR-VSSA vancomycin-susceptible S. aureus, BR-VRSA vancomycin-resistant S. aureus, and VREF vancomycin-resistant Enterococcus faecalis.

Figure 2. Results of Pulsed-Field Gel Electrophoresis (PFGE), S1 Nuclease Digestion of Total DNA, Hybridization, and Plasmid Annotation

Panel A shows the results of SmaI digestion of total DNA followed by PFGE. Lane M shows the lambda ladder (with the molecular sizes in kilobases shown at left); lane 1, BR-VSSA; lane 2, BR-VRSA (donor); lane 3, transconjugant number 1 Staphylococcus aureus RN4220-RF; lane 4, transconjugant number 2 S. aureus RN4220-RF; lane 5, S. aureus RN4220-RF (recipient); and lane 6, vancomycin-resistant Enterococcus faecalis (vanA). Panel B shows the results of S1 digestion of the total DNA of staphylococcal strains and vancomycin-resistant E. faecalis followed by PFGE (at left) and hybridization with a VanA probe (at right). In the results at left, lane 1 shows BR-VSSA; lane 2, BR-VRSA (donor); lane 3, transconjugant number 1 S. aureus RN4220-RF; lane 4, transconjugant number 2 S. aureus RN4220-RF; lane 5, S. aureus RN4220RF (recipient); lane 6, vancomycin-resistant E. faecalis; and lane M the lambda ladder. In the results at right, the white and red arrows highlight the positive signal for vanA hybridization in staphylococcal and enterococcal strains, respectively. Panel C shows a schematic representation of the annotation of the BR-VRSA plasmid pBRZ01.

Figure 3. Phylogenetic Comparisons of BR-VRSA and BR-VSSA with Representatives of Other Methicillin-Resistant Staphylococcus aureus (MRSA) Clones

Panel A shows a dendrogram of the pandemic MRSA clones, BR-VRSA and BR-VSSA, generated with PFGE and the use of GelCompar II software, version 6.5 (Applied Maths). Patterns were clustered by means of UPGMA (unweighted pair group method with arithmetic mean), with the use of the Dice similarity coefficient and an optimization of 0.50% and a tolerance of 1.0%. PFGE types, or clusters, were identified on the basis of a similarity of 75% or higher (indicated by the vertical red line). ST denotes sequence type. The phylogenetic tree shown in Panel B is based on whole-genome single-nucleotide polymorphisms (SNPs) and was generated with the use of the maximum-likelihood optimality criterion. Branch lengths are proportional to the number of evolutionary changes (substitutions per site). All nodes have 100% bootstrap support. Sequence types (STs) and clonal complexes (CCs) of the S. aureus strains are indicated.