Van c o m y c i n - R e s i s t a n t

E n t e ro c o c c i
Therapeutic Challenges in the 21st Century

William R. Miller, MDa, Barbara E. Murray, MDa,b,

Louis B. Rice, MDc, Cesar A. Arias, MD, MSc, PhDa,b,d,*

KEYWORDS
 Vancomycin resistant enterococcus  Antibiotic resistance  Combination therapy

KEY POINTS
 Multidrug-resistant enterococcal infections continue to be a clinical challenge despite the
advent of new therapeutic agents.
 The genetic plasticity of enterococci underscores the versatile nature of the organisms
and has provided new insights into the mechanisms by which bacteria can become resis-
tant to antibiotics and how commensal organisms evolve to become prominent hospital-
associated opportunistic pathogens.
 Development of resistance to almost all antienterococcal antibiotics currently available in
clinical practice highlights the difficulties facing clinicians in the setting of deep-seated
enterococcal infections.
 Vancomycin-resistant enterococci infections in the 21st century will require the use of new
or innovative therapeutic treatments that involve both old and new antimicrobials.

Disclosure Statement: Dr W.R. Miller has no relevant affiliations or financial disclosures. Dr B.E.
Murray has grant support from Johnson & Johnson, Astellas, Palumed, Intercell and Cubist, and
has served as consultant for Astellas (Theravance), Cubist, Targanta Therapeutics Corporation,
Pfizer, Rib-X, AstraZeneca and Durata Therapeutics. Dr L.B. Rice has served as a consultant for
Astra-Zeneca, Tetraphase Pharmaceuticals, MicuRx, Macrolide Pharmaceuticals and Adenium
Pharmaceuticals. Dr C.A. Arias has received lecture fees, research support and consulting fees
from Theravance, Bayer and Cubist; research support from Forrest Pharmaceuticals and Therav-
ance Inc and has served as speaker for Forest, Theravance, Pfizer, Astra-Zeneca, Cubist, The
Medicines Company and Norvartis.
a
Division of Infectious Diseases, Department of Internal Medicine, University of Texas Medical
School at Houston, 6431 Fannin Street, Houston, TX 77030, USA; b Department of Microbiology
and Molecular Genetics, University of Texas Medical School at Houston, 6431 Fannin Street,
Houston, TX 77030, USA; c Departments of Medicine, Microbiology and Immunology, Warren
Alpert Medical School of Brown University, 593 Eddy Street, Providence, RI 02903, USA;
d
Molecular Genetics and Antimicrobial Resistance Unit, International Center for Microbial Ge-
nomics, Universidad El Bosque, Avenue Cra 9 No. 131 A - 02, Bogotá, Colombia
* Corresponding author. University of Texas Medical School, 6431 Fannin Street, Room MSB
2.112, Houston, TX 77030.
E-mail address: [email protected]

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416 Miller et al

INTRODUCTION: A REPORT FROM THE FRONT LINES

The year 1899 saw the first report of the bacterium Micrococcus zymogenes (thought
today to be Enterococcus faecalis) as a cause of infective endocarditis (IE).1 The pa-
tient, a 37-year-old German man, described a 2-month history of fever with an indo-
lent, yet relentlessly progressive course. The medical staff could only watch
helplessly as he died 18 days later from complications of his illness. One hundred
fifteen years later, a man in his 40s with a hematologic malignancy was found to be
colonized with vancomycin-resistant enterococci (VRE) on admission to the hospital
for treatment of his cancer. During the admission, he became febrile, and Entero-
coccus faecium was isolated from the blood several times.2 The medical team admin-
istered a progressive regimen of antimicrobials, including daptomycin, ampicillin,
gentamicin, quinupristin-dalfopristin, tigecycline, and linezolid; however, after
3 months on therapy, blood cultures remained positive for VRE. Despite the availability
of antibiotics, the eventual outcomes of both patients (separated by medical care that
had evolved for 150 years) were not that different.
As the 21st century dawns, organisms such as multi–drug-resistant (MDR) E fae-
cium present new challenges to clinicians. Medical science races to keep pace
with the spread of resistance determinants and provide clinicians new drugs to com-
bat an ever-changing enemy. The number of antibiotics in the therapeutic armamen-
tarium that are active against enterococci has expanded over the last decade;
however, there are little published clinical data to guide their most effective use.
Below is a profile of the pathogen and its genomic plasticity, the attributes thought
to be associated with its ability to colonize and infect the human host, and its strate-
gies for resisting antimicrobial attack. This review concludes with a synthesis of the
available therapeutic data to guide physicians in selecting the best treatment for these
difficult infections.

PROFILE OF AN OPPORTUNISTIC PATHOGEN

From the Iron Age to the Antibiotic Age
Enterococci are facultative gram-positive cocci that live as commensals of the gastro-
intestinal (GI) tract of animals and humans. Enterococci are rugged and durable, able
to survive in high salt concentrations and elevated temperatures, and able to resist
chemical stress from chlorine and alcohol-based disinfectants.3 Enterococci lack
the cadre of virulence determinants of Staphylococcus aureus or the more pathogenic
streptococci; however, their durability and commensal nature position them to not
only survive but thrive in the modern medical setting. Surveys of organisms respon-
sible for health care–associated infection have found that enterococci are second
only to staphylococci in being isolated from health care–associated infections across
the United States.4
Enterococcal infections, like most human diseases, have a history shaped not only
by bacterial biology but also by the social, economic, and geographic factors that
affect the human host. Whole genome sequencing and analyses of E faecium ge-
nomes have shed some light on the evolutionary history and adaptation of enterococci
with the human host.5,6 This story appears to emerge at the dawn of the Iron Age
(ca. 3000 years ago) with the divergence of 2 genetic lineages (clades) of E faecium.
One, designated clade B, would remain as a commensal of the human gastrointestinal
tract with low potential for infectivity and general lack of antibiotic resistance determi-
nants. The other (clade A) seems to have evolved and adapted to the gastrointestinal
tract of livestock and domesticated animals. This bifurcation was driven, as Lebreton
and colleagues6 hypothesized, by the increasing contact between humans and
 Vancomycin-Resistant Enterococci 417

domesticated animals owing to the spread of animal husbandry for agricultural and
economic gain, the specialization of diets that resulted, and the changing patterns
of hygiene that accompanied urbanization. Interestingly, the animal clade would go
on to diverge again, this time a mere 75 years ago, at about the time antibiotics
were introduced in clinical medicine. This occurrence gave rise to a hospital-
associated lineage (designated clade A1), which seems to have a better ability to infect
humans, disseminate in the hospital environment, and acquire a plethora of antibiotic
resistance determinants, the hallmarks of the current VRE epidemic.
A cardinal feature of the members of the A1 clade is the remarkable adaptability of
their genome. This plasticity stems from the large repertoire of mobile genetic ele-
ments capable of transferring pieces of DNA as small as a single gene to large path-
ogenicity islands and chromosomal fragments.7 In both E faecalis and E faecium, the
transmissibility of mobile elements and recruitment of a large number of antibiotic
resistance determinants seem to be associated in part with a deficiency in the CRISPR
(clustered regularly interspaced short palindromic repeats)–associated system (cas).8
CRISPRs and the cas genes are present in a diverse number of archaea and bacteria.9
Together, they encode an adaptive immunity against phages, plasmids, and other mo-
bile genetic elements. Proteins encoded by the cas genes can process and integrate
small sequences of foreign DNA between the clustered repeats, which serve as a
template to create small RNAs that recognize and bind incoming foreign nucleic
acid, targeting them for destruction by further components of the cas system. With
an apparent limitation in the ability to degrade incoming DNA, it is thought the ge-
nomes of the A1 clade were primed to evolve and respond to the selective pressures
of the hospital environment. This ability, when coupled with intrinsic resistance to
several broad-spectrum antibiotics (eg, cephalosporins) and increases in the numbers
of chronically and critically ill patients, set the stage for the emergence and dissemi-
nation of this multidrug-resistant clade.

Enterococcal Infections: A Tale of Two Species

From the late 1970s and into the early 1980s, E faecalis recorded a 3-fold surge in rates
of nosocomial bacteremia, whereas rates of community-associated infection remained
flat.10 Although these isolates largely retained their sensitivity to ampicillin, their emer-
gence would serve as a harbinger of the challenge to come. In the late 1970s, clinicians
noted an increase in the minimum inhibitory concentration (MIC) of ampicillin from 4 to
64 mg/mL in isolates of E faecium, and over the next decade an alarming phenotype
characterized by high-level resistance to ampicillin (>128 mg/mL) began to emerge.11,12
This resistance depends on the chromosomally encoded penicillin-binding protein
5 (PBP5), a monofunctional transpeptidase capable of cross-linking peptidoglycan
when all other PBPs are inactivated.13 It was later recognized that differences in up
to 5% of the amino acids between the PBP5 of resistant strains, compared with
ampicillin-sensitive strains (PBP5-S), correlated with both the resistance phenotype
and origin of the strain.14 Most often, PBP5-resistance strains were found in the
hospital-associated clade, whereas PBP5 ampicillin-sensitive strains predominated
in community-associated strains, reflecting the shift from clade B to A1 within the hos-
pital environment. At about the same time, the glycopeptide vancomycin entered wide-
spread use against methicillin-resistant S aureus, setting the stage for the emergence of
vancomycin resistance in enterococci.
The glycopeptide antibiotic vancomycin binds to the last 2 residues of the penta-
peptide moiety of peptidoglycan (the terminal D-Ala-D-Ala), thereby inhibiting
transglycosylation and transpeptidation reactions, leading to an arrest in cell wall syn-
thesis.15 Two species of enterococci, Enterococcus gallinarum and Enterococcus
418 Miller et al

casseliflavus, possess related chromosomally encoded gene clusters designated

vanC1 and vanC2 that can confer intrinsic low-level resistance to vancomycin (MICs
of up to 2–32 mg/mL) via a change in the terminal precursor’s residues to D-Ala-D-
Ser.16 In contrast, high-level resistance to vancomycin (MICs >64 mg/mL) is mediated
by the production of pentadepsipeptides (ending in D-Ala-D-Lac) and destruction of
the normal D-Ala-D-Ala ending precursors. The enzymatic machinery is encoded by
the vanA gene cluster frequently found on transposons within plasmids (eg, Tn1546)
and less often the vanB cluster integrated into the chromosome on Tn5382.17,18 Of
note, 7 other van clusters have been described so far, of which, vanD and vanM
also allow synthesis of peptidoglycan ending in D-Ala-D-Lac and confer high levels
of resistance to vancomycin.17,19 The recent source of the genes implicated in vanco-
mycin resistance in enterococci is likely environmental, as the genus Paenibacillus has
a cluster nearly identical to that of vanA found in enterococci.20
Clinically relevant strains of VRE were first described from England in 1988.21 It was
later postulated that VRE in Europe arose first in livestock owing to the association of
the emergence of VRE with the use of avoparcin, a glycopeptide antibiotic used as a
growth promoter, a finding that led to the ban of this practice in 1997.22 In the United
States, vancomycin resistance in E faecium arose in clinical isolates and quickly came
to dominate the landscape of enterococcal infections, with vancomycin resistance re-
ported in more than 80% of E faecium and 8% to 9% of E faecalis.4

MASTER OF SURVIVAL
Host Colonization—Preamble to Infection
The first step toward infection with VRE appears to involve colonization of the host
gastrointestinal tract. The normal flora of individuals in the United States consists of
a diverse mixture of bacterial taxa, including Enterobacteriaceae, anaerobes, and
gram-positive organisms, such as vancomycin-sensitive enterococci. However, in
health care settings, VRE are often found colonizing the intestines of hospitalized pa-
tients (particularly the critically ill or patients with multiple comorbidities). This finding is
reflected in the risk factors for hospital acquisition of VRE including prolonged stay in a
hospital or other long-term care facility, solid organ or bone marrow transplantation,
physical proximity to other patients with VRE, use of antibiotics, hemodialysis, and
indwelling hardware such as urinary catheters.23 A study of VRE colonization in pa-
tients from 2 intensive care units found, using multivariate analysis, that occupation
of a room by a previous patient with VRE was an independent predictor of subsequent
patients becoming colonized, an effect that persisted for 2 weeks.24 Investigations
conducted in outpatient dialysis units found up to 18% of patients colonized, with
15% acquiring VRE over a 6-month period. Further, evaluation of the chairs and
gowns of staff at a dialysis unit found rates of VRE contamination of 58% and 30%,
respectively.25
Once established as members of the gastrointestinal tract flora, VRE can take
advantage of perturbations of microbial diversity to dominate the bacterial populations
in the gut. Antibiotics play a large role in this transition, via direct inhibitory effects on
the microbes themselves and the host’s innate immune response. One consequence
of therapy with broad-spectrum antimicrobials is the collateral damage they inflict on
otherwise benign commensal flora, providing an opening for microbes resistant to
these agents, such as VRE, to expand and fill the void. A study of 146 patients who
were admitted to the intensive care unit with negative VRE surveillance cultures and
who received treatment with either piperacillin-tazobactam or cefepime showed sub-
sequent colonization rates of 26.4% and 31.1%, respectively.26
 Vancomycin-Resistant Enterococci 419

Disruption of the gram-negative intestinal flora by broad-spectrum antibiotics also

has profound effects on the host innate immune response and alters the ability to con-
trol VRE. These pathways rely on signals from the toll-like receptor family of proteins,
which recognize conserved pathogen-associated molecular patterns such as lipo-
polysaccharides. These signals, in turn, activate the MyD88-dependent signaling
cascade resulting in the ultimate production of a variety of antimicrobial peptides.
The peptide RegIIIg, a lectin with potent anti–gram-positive activity, was shown to
play an important role in controlling VRE colonization in mice.27 Elimination of the
normal intestinal anaerobic and gram-negative flora via the administration of metroni-
dazole, neomycin, and vancomycin resulted in down regulation of the production of
RegIIIg by the murine intestinal epithelial cells. In the absence of this antimicrobial
peptide, mice were unable to clear VRE from the intestine. Supplementing the mouse
with oral lipopolysaccharide (a component of the gram-negative cell envelope), but not
lipoteichoic acid (a component of the gram-positive cell wall), restored the production
of RegIIIg and allowed the mice to control the growth of VRE. In the human gastroin-
testinal tract, RegIIIa is thought to play a similar role.28 By analogy, in the hospital
setting, the administration of broad-spectrum antibiotics to patients can eliminate
gram-negative organisms in the gastrointestinal tract, much as was shown in the
mouse model. Without gram-negative bacteria, it seems likely that human production
of RegIIIa would also decrease, allowing overgrowth of resistant colonizers such as
VRE. Once established in the gut, VRE pose a risk of invasive infection in vulnerable
patients. Clinical studies in recipients of hematopoietic stem cell transplants showed
a clear progression from colonization to dominance and subsequent bacteremia in
patients in whom dominance of VRE in their gastrointestinal tracts occurred.29

Virulence Factors
The shift from commensal to pathogen, while aided by resistance to antimicrobials,
also depends on the ability of an organism to adhere to or invade tissues and evade
host defenses. Although enterococci lack a robust repertoire of secreted toxins like
those produced by the staphylococci or streptococci, a cadre of virulence determi-
nants have been reported to help them to adhere to tissues and form biofilms.
Among the best studied of these determinants are proteins that harbor the LPxTG
motif (which is the protein signature for attachment to the cell wall peptidoglycan).
Several are found as a feature of the core genome of enterococci, including the collagen
adhesins of E faecalis (Ace) and E faecium (Acm). Adherence to extracellular matrix mol-
ecules such as collagen and laminin would be an important first step in establishing an
infection, and both Ace and Acm are thought to be involved in this process. The role of
Ace and Acm in supporting infection was shown in a rat model of IE, as deletion mutants
in both species were found to be attenuated as compared to the wild type.30,31 In addi-
tion, antibodies against Ace provided protection against developing IE in this same
model.31 Further, in community-associated isolates of E faecium, the acm allele is often
a pseudogene, which does not express a functional product, resulting in a phenotype
that is unable to bind collagen.32 Another major group of genes in the core genome
encode the enterococcal pili. These multiprotein structures are long filamentous protru-
sions from the cell surface that are also shown to promote adherence to the matrix pro-
teins fibrinogen and collagen and support biofilm formation.33 The E faecalis Ebp pilus
operon consists of 3 structural genes and a pilus-specific sortase; the individual sub-
units contain the LPxTG recognition motif that allows the pilus sortase to assemble
the structural proteins into a complete pilus before it is anchored to the cell wall via a
housekeeping sortase.34,35 In E faecium, deletion of genes from a homologous operon,
known as Ebpfm, impaired bacterial adhesion and biofilm formation and resulted in
420 Miller et al

impaired pathogenic potential compared with the wild type in a mixed infection model of
murine urinary tract infection.36
Several other proteins possessing the LPxTG motif are acquired determinants and
not present in all enterococcal isolates. Aggregation substance proteins are a family of
plasmid-encoded surface adhesins that mediate adhesion to molecules of the extra-
cellular matrix and promote clumping, biofilm formation, and high-frequency transfer
of plasmid DNA.37 In animal models of IE, E faecalis strains overexpressing aggrega-
tion substance were found to have larger vegetations with increased bacterial loads
compared with mutants lacking the gene.38 Esp (enterococcal surface protein) and
Espfm (its homologue in E faecium) have been implicated in biofilm formation and in
the pathogenesis of IE.39,40 This determinant is often found within a pathogenicity
island that seems to be associated with hospital-acquired isolates.39
Another important subset of cell envelope–located virulence factors contains the
WxL motif, a signature sequence that is conserved among the low G 1 C gram-
positive bacteria and binds to peptidoglycan.41 ElrA (enterococcal leucine-rich
repeat-containing protein A) is one of multiple WxL proteins in E faecalis and is located
in a polycistronic operon; previous characterizations of a homologue in Listeria spp.
showed its involvement in inducing uptake of the bacteria into nonphagocytic cells.42
Deletion of the elrA gene attenuated lethality in a mouse peritonitis model and
decreased the strain’s ability to infect macrophages and induce an interleukin-6–
mediated inflammatory response. In silico analysis of the E faecium genome identified
6 genes encoding potential WxL-containing proteins. The genes are organized in 3 op-
erons, termed WxL locus A, B, and C. Using electron microscopy and enzyme-linked
immunosorbent assays, locus A and C were found to be antigenic and expressed on
the cell surface at much higher levels in clinical strains (as opposed to community-
associated lineages). In addition, WxL proteins from locus A were able to bind collagen
and fibronectin and deletion of all 3 loci-attenuated virulence in a mixed infection rat
endocarditis model.43
Glycolipids and polysaccharides are also important components of the entero-
coccal cell envelope. Lipoteichoic acid is an antigenic component of gram-positive
cell walls, and antibodies directed against this epitope were shown to enhance opso-
nization and killing of enterococci by complement and leukocytes.44 Some isolates of
E faecalis, particularly members of hospital-associated clades, possess a capsular
polysaccharide gene locus that allows the bacteria to synthesize a polysaccharide
shield for the antigenic lipoteichoic acid, enabling evasion of antibody-mediated
opsonization.45 Other polysaccharides are found shown to be important in the pro-
duction of biofilm, which mediates persistence on environmental surfaces and re-
duces the effectiveness of many antimicrobials. Enterococcal polysaccharide
antigen is the product of a 16-gene locus named epa that produces a rhamnose-
containing polysaccharide important for the formation of biofilm.46 It is present in all
E faecalis, and antibodies directed against this antigen can be found in most samples
of sera from patients with deep-seated E faecalis infections.47 In animal models, dele-
tion of this locus from the laboratory strain E faecalis OG1RF led to attenuation in both
a mouse peritonitis and murine urinary tract infection model.48 In addition to poly-
saccharides, glycolipids play a role in enterococcal pathogenesis. Deficiency of the
glycolipid a-diglucosyldiacylglycerol has been described to reduce both biofilm for-
mation and adherence to host cells in E faecalis, and this defect was associated
with faster sterilization of blood cultures compared with a wild type strain in mouse
model of bacteremia.49
Secreted virulence factors also play a role in the pathogenesis of experimental
enterococcal infection. Approximately 30% of E faecalis produce the toxin cytolysin
 Vancomycin-Resistant Enterococci 421

(Cyl), which is encoded by an 8-gene operon carried on plasmids or chromosomal

pathogenicity islands.50,51 The toxin is a pore-forming multimer of 2 subunits, CylLL
and CylLS, which are activated by CylA, a serine protease. The spectrum of activity
includes other gram-positive organisms and erythrocytes of humans, horses, and
rabbits but not sheep or cows (a specificity thought to be conferred by membrane
levels of phosphatidylcholine). Clinical data from a retrospective series of 190 patients
found that those with Cyl-producing isolates had a 5-fold increased risk of mortality
at 3 weeks as compared to Cyl negative isolates.52 However, a subsequent larger
prospective study of 398 cases of E faecalis bacteremia failed to confirm this asso-
ciation.53 Two enterococcal proteases, gelatinase (GelE) and extracellular serine pro-
tease, mediate virulence under the control of the Fsr quorum sensor, a 2-component
signaling system that bears homology to agr (accessory gene regulator) in S aureus.54
GelE is able to digest host matrix proteins and also activates autolysin, which pro-
motes biofilm formation caused by cell lysis and release of DNA into the extracellular
space.55 In a rabbit model of endocarditis, GelE deletion mutants were associated
with decreased vegetation colony counts and increased migration of inflammatory
cells (such as neutrophils) to the site of infection.56 Similarly, in a rat model of IE, E fae-
calis with a disruption of GelE was attenuated and had a lower IE induction rate than
the wild type.57 Finally, a variety of other proteins involved in stress response and
neutralization of reactive oxygen species has been associated with enterococcal viru-
lence, presumably allowing the bacteria to survive the oxidative burst of human poly-
morphonuclear leukocytes and macrophages.58

Antibiotic Resistance—Beyond Vancomycin

The flexibility of the enterococcal genetic repertoire has given rise to a variety of resis-
tance strategies as new antimicrobial compounds are deployed. A comprehensive
look at these strategies has recently been described in depth elsewhere.59,60 This sec-
tion discusses the mechanisms mediating resistance to newer agents used to treat
serious VRE infections.

Daptomycin
Daptomycin (DAP) is a lipopeptide antibiotic with in vitro bactericidal activity against
both E faecalis and E faecium, including VRE.61 DAP consists of a cyclic polypeptide
core with a lipid tail that facilitates insertion into the bacterial membrane in a calcium-
dependent manner, a characteristic shared with cationic peptides of the innate im-
mune response.62 The precise mechanism by which the antibiotic mediates cell death
is not known, but DAP-treated bacteria rapidly lose membrane physicochemical prop-
erties with disruption of the ionic gradient used to drive many biosynthetic pathways.63
It is postulated that DAP forms a multimeric pore structure in the presence of phos-
phatidylglycerol, with 2 tetramers aligned opposite each other on the inner and outer
leaflet of the membrane.64 Enrichment of the membrane with cardiolipin (CL), with its
4 bulky fatty acyl chains, stabilizes the membrane to local perturbations caused by the
DAP insertion. This CL-DAP interaction seems to decrease the ability of the antibiotic
to transition from the outer to the inner leaflet, preventing tetramer alignment and pore
formation. Both E faecalis and E faecium have adopted strategies to avoid DAP-
mediated membrane damage; however, the underlying mechanism seems to be spe-
cies specific.
In E faecalis, diversion of the antibiotic from critical membrane locations such as the
dividing septum is associated with daptomycin resistance (DAP-R). This resistance
seems to be associated with redistribution of CL microdomains, which are suggested
to act as decoys to sequester the antibiotic away from important septal areas and
422 Miller et al

might also protect the membrane at these locations (the diversion hypothesis).65
Changes associated with this redistribution were described in isolates both in vitro
and in vivo and implicated genes involved in the cell envelope stress response (the
3-component regulatory system, LiaFSR) and phospholipid synthesis.66,67 In contrast,
E faecium does not exhibit the characteristic redistribution of cardiolipin microdo-
mains seen in E faecalis. Instead, E faecium seems to repel the DAP molecule from
the cell surface, changes more akin to the proposed repulsion strategy for DAP resis-
tance described in staphylococci. In this scenario, the positively charged calcium-
DAP complex is repelled from the cell surface owing to electrostatic changes (as
the cell envelope becomes more positively charged).68 Interestingly, despite the
mechanistic differences in DAP resistance between E faecalis and E faecium, similar
genes in both species (including liaFSR, see below) are involved.
The lia operon (for Lipid II Interacting Antibiotics) belongs to a family of 3-component
signaling systems composed of a sensor histidine kinase (LiaS), its cognate response
regulator (LiaR), and a putative transmembrane negative regulator protein (LiaF). The
LiaFSR system is conserved in all gram-positive organisms of clinical relevance.69
In the presence of cell envelope stress, the system seems to be activated either by
phosphorylation of LiaR (via LiaS) or by mutations in LiaR that mimic phosphoryla-
tion.70 In E faecalis, a mutation in liaF (resulting in a deletion of an isoleucine residue
at position 177) seemed to activate the system leading to an increase in DAP MIC
from 1 to 4 mg/mL. More importantly, this single change in LiaF resulted in loss of
DAP bactericidal activity in vitro.71 This increase in MIC and loss of bactericidal activity
at a supposedly susceptible MIC questions the current DAP breakpoint for entero-
cocci (4 mg/mL, which is 4 times higher that than for S aureus). DAP-susceptible
E faecium isolates with DAP MICs close to the susceptibility breakpoint (3–4 mg/mL)
often harbor mutations associated with DAP resistance (mostly in liaFSR),72 and,
like E faecalis, these isolates are tolerant (ie, even at 5x, MIC DAP is not bactericidal)
to DAP in vitro.73 In a recent report, a patient with recalcitrant E faecium bacteremia
treated with DAP experienced failure with this antibiotic, and the initial bloodstream
isolate harbored alterations in LiaRS, supporting the notion that changes in LiaFSR
are clinically associated with DAP therapeutic failure.2 Of note, mutations associated
with DAP resistance have been described in isolates from patients who had never
been exposed to DAP, and it is postulated that, in addition to spread from other pa-
tients, naturally occurring antimicrobial peptides (like those produced by the innate im-
mune system) may trigger cell envelope adaptive responses similar to those observed
with DAP.74,75
The yycFG operon is involved in cell wall homeostasis and is highly conserved among
the low G 1 C gram-positive bacteria. The yycFG operon is known to be active in modi-
fication of peptidoglycan synthesis, modulating autolysin expression (including pcsB
in streptococci), and mutations in this operon are found in vancomycin-intermediate
S aureus and in vancomycin-resistant S aureus strains.76 In addition, the yycFG operon
has also been found to have indirect effects on fatty acid metabolism and membrane
fluidity. The histidine kinase (YycG) of this 2-component system from S aureus is found
to associate with the lipid bilayer via a transmembrane domain and responds to stress
via a conserved sensing sequence known as the PAS domain.77,78 Upon activation, the
sensor kinase auto phosphorylates before passing the phosphate to its cognate
response regulator (YycF) to induce transcription of the regulon. This system was asso-
ciated with DAP-R in a clinical strain-pair of E faecium that developed resistance to ther-
apy.79 Changes found in the PAS domain were postulated to alter the sensing activity of
the YycFG system, although it is not understood precisely how this effect is mediated.
Further, in a genomic survey of resistant isolates of E faecium, changes in the yycFG
 Vancomycin-Resistant Enterococci 423

operon were the second most commonly found changes associated with DAP-R, with
changes in LiaFSR encountered most often.73
The contribution of alterations in enzymes involved in phospholipid metabolism to
the DAP-resistance phenotype has not been fully elucidated, but such changes
seem to act synergistically with those related to the cell envelope stress response
mediated by LiaFSR (or other regulatory systems, see earlier discussion). Alterations
involving enzymes such as cardiolipin synthase (Cls) appear to occur after changes in
LiaFSR, potentiating the resistance phenotype.80 Davlieva and colleagues81 recently
found that substitutions in one of the phospholipase domains of Cls increased the cat-
alytic activity of the enzyme, albeit marginally. Experimental data have also found that
when additional copies of Cls harboring changes found in DAP-R strains were intro-
duced in trans on a plasmid into the laboratory strain of E faecalis OG1RF, they
increased the DAP MICs.67 These findings suggest that Cls substitutions are associ-
ated with a gain in function of the enzyme; however, how this change is linked to redis-
tribution of CL microdomains is unknown.

Oxazolidinones
Linezolid and the recently approved tedizolid are the clinically available oxazolidi-
nones. These compounds bind to the A site of ribosomes and disrupt the docking
of the aminoacyl-transfer RNA, inhibiting the delivery of peptides and the subsequent
elongation of the polypeptide chain.82,83 Tedizolid differs from linezolid in that the
former harbors a fourth D-ring constituent and also has a hydroxymethyl group on
the oxazolidinone ring. These modifications greatly increase the binding affinity of tedi-
zolid for its target compared with linezolid, with MICs 4- to 8-fold lower than those of
linezolid.84 A common mechanism that mediates resistance to both compounds is
target modification, namely mutations in the 23S rRNA. The substitutions G2505A
and G2576U are the most common changes observed in resistant clinical isolates.
As enterococci possess multiple copies of the 23S rRNA gene, the number of mutated
alleles correlates directly with the level of resistance.85 The number of mutated copies
has been associated with the length of drug exposure and is accelerated by recombi-
nation of mutant alleles with wild type alleles within the enterococcal genome.86 This
gene-dosage effect was demonstrated in vitro with a laboratory strain of E faecalis and
its recombination-deficient mutant exposed to linezolid. The time to first G2576U
mutation was similar for both strains, as expected for a spontaneous mutation rate,
but the wild type developed subsequent mutations 6 passages sooner than the
recombination deficient strain, indicating interallelic recombination within the cell
played a role in the propagation of resistant alleles.87
The ribosomal proteins L3 and L4, which are part of the peptidyl-transferase center,
have also been associated with resistance to oxazolidinones in both enterococci and
staphylococci.88,89 It was found in Streptococcus pneumoniae that changes in L3 not
only confer oxazolidinone resistance, but also seem to have compensatory effects
to balance the fitness cost associated with mutations in the 23S rRNA.90 In a genetic
survey to characterize linezolid-resistant versus susceptible Staphylococcus epider-
midis clinical isolates, mutations in 23S rRNA genes were always accompanied by
changes in L3, a finding the authors speculated may be caused by its compensatory
nature.91 Further, whole genome sequencing of 21 S epidermidis bloodstream isolates
resistant to linezolid found 19 isolates harbored coexisting mutations in 23S rRNA and
L3.92 Whether these findings are generalizable to other gram-positive cocci, including
enterococci, is not known.
Two transmissible oxazolidinone resistance determinants have been described to
date, cfr and optrA. Cfr (for chloramphenicol-florfenicol resistance) is a methylase
424 Miller et al

that modifies the adenine nucleotide at position 2503 of the 23S rRNA. It has been
found in clinical isolates of E faecalis and in other clinically relevant gram-positive or-
ganisms such as staphylococci.93,94 It has been postulated that cfr has spread from
animal strains to humans, as this gene seems to be widespread in bacterial isolates
recovered from livestock (where florfenicol use is widespread) and is usually found
on plasmids or other mobile genetic elements that can readily transfer between bac-
terial cells.95 Of note, the binding of tedizolid to the ribosome does not seem to be
affected by Cfr-mediated methylation of the 23S rRNA, and this compound retains
activity in vitro against enterococcal strains that carry the cfr gene.96 The optrA
gene encodes a putative ABC transporter associated with elevated MICs to the oxa-
zolidinones in the absence of known 23S rRNA mutations or the gene cfr and has been
recently identified in both E faecalis and E faecium from animal and human sources.97

Lipoglycopeptides
The lipoglycopeptides are a class of compounds based on the modified glycopeptide
core to which a lipophilic side chain has been added. Several lipoglycopeptides are
now in clinical use and include telavancin, dalbavancin, and oritavancin. These com-
pounds, like vancomycin, bind to peptidoglycan precursors ending in D-Ala-D-Ala.
However, unlike vancomycin, they harbor hydrophobic moieties that permit insertion
into the cell membrane, which is likely to affect membrane homeostasis in addition to
their effect on cell wall synthesis.98,99 This double mechanism of action increases the
in vitro activity against enterococci. Despite the improvement in activity, telavancin
and dalbavancin still exhibit high MICs against VRE (4–8 and 16–32 mg/mL, respec-
tively, although several-fold lower than those of vancomycin) that preclude their use
in clinical settings against these organisms.100 Indeed, telavancin and dalbavancin
do not bind peptidoglycan precursors ending in D-Ala-D-Lac.100
More promising is oritavancin, which demonstrates in vitro activity against VRE medi-
ated by both vanA and vanB gene clusters. This enhanced activity seems to be derived
from its mechanism of binding to peptidoglycan precursors. Nuclear magnetic reso-
nance spectroscopy data have shown that oritavancin has more interactions with the
peptidoglycan precursors than vancomycin, spanning the third amino acid of the pen-
tadepsipeptides (L-lysine) and amino acids present in the cross bridge of both staphy-
lococci and enterococci, mitigating the effect of the D-Ala to D-Lac substitution.101 In
addition, the lipophilic 4’-chlorobiphenylmethyl side chain has greater binding affinity
for lipid II, both concentrating the antibiotic near the site of active synthesis and disrupt-
ing transglycosylation.102 This improved binding to the cell wall assembly apparatus
coupled with its membrane effects mediate potent bactericidal activity against both
growing cells and biofilms. As this compound has recently entered clinical practice,
the degree and mechanisms of resistance are not fully characterized. Arthur and col-
leagues103 found that high-level expression of the vanA gene cluster and an increase
in D-Lac-ending precursors was associated with 16-fold higher MICs to oritavan-
cin. Additionally, expression of the vanZ gene of the same cluster increased oritavancin
MIC by 4-fold, although the mechanism is not known.103 Thus, the role of oritavancin in
the treatment of VRE infections remains to be established.

Streptogramins
Streptogramins are a class of compounds that inhibit protein synthesis by targeting
the 50S subunit of the bacterial ribosome. Quinupristin-dalfopristin (Q/D) is a mixture
of pristinamycin derivatives, streptogramin A (dalfopristin) and B (quinupristin), which
exhibit in vitro bactericidal activity owing to the synergism between the 2 compounds.
The binding of dalfopristin has been reported to induce a conformational change in the
 Vancomycin-Resistant Enterococci 425

ribosome that unmasks a high affinity binding site for quinupristin, leading to irrevers-
ible inhibition of the ribosome complex.104 Importantly, Q/D is effective against
E faecium but not E faecalis, as the latter possesses a chromosomal gene named
lsa (for lincosamide and streptogramin A resistance), which provides all E faecalis
with intrinsic resistance to streptogramin A and lincosamides (eg, clindamycin). This
gene encodes a putative protein with an adenosine triphosphate–binding cassette
motif of transporter proteins but not the transmembrane region that would be ex-
pected for an efflux pump, and its exact mechanism of action remains unknown.105
Reduced susceptibility to Q/D in E faecium is mediated by several mechanisms.
First, modification of dalfopristin via the acetyltransferases VatD and VatE reduces
binding affinity owing to steric hindrance, disrupting the bactericidal synergy of the
2 compounds.106 Enzymatic cleavage of the ring structure of quinupristin by the lac-
tonases VgbA and VgbB, originally described in staphylococci, leads to inactivation of
the compound.107 The ermB gene involved in macrolide resistance is a ribosomal
methyltransferase, which also affects the binding of quinupristin. It confers a pheno-
type, MLSB (for Macrolide, Lincosamide, Streptogramin B), that results in resistance
to these antibiotics and is widespread in enterococci.108 Dalfopristin, a streptogramin
A, remains active in vitro in the presence of the erm gene, but as a bacteriostatic
agent.106 In vivo, the presence of erm may compromise the activity of Q/D, as this
compound exhibited decreased activity against MLSB enterococci in a rat endocardi-
tis model.109
Efflux of Q/D from the bacterial cell also plays a role in resistance, and some resis-
tant isolates of E faecium have been found to contain a mutation in the eatA gene (for
Enterococcus ABC Transporter) that was shown to confer resistance to Q/D when
introduced into susceptible strains.110

Glycylcyclines
Tigecycline is a glycylcycline antibiotic and a synthetic derivative of minocycline with
activity against both gram-negative and gram-positive bacteria, including methicillin-
resistant S aureus and VRE. Similar to the tetracycline class of antimicrobials, tigecy-
cline binds to the 16S rRNA of the 30S subunit of the ribosome and inhibits the docking
of the aminoacyl-transfer RNA.111 Unlike the tetracycline class of antibiotics, tigecy-
cline MICs are not affected by typical tetracycline resistance determinants such as
efflux pumps (tetK, tetL) or ribosomal protection proteins (tetO, tetM, tetS).112 Resis-
tance in E faecalis has been described in 2 case reports of patients with intra-
abdominal infections treated with tigecycline and in antimicrobial surveys.113–115
Decreased susceptibility in E faecium was recently described in both clinical isolates
and strains passaged in vitro. Using genomic analysis, a mutation in the S10 protein, a
constituent of the 30S ribosomal subunit, was found in 4 of the 5 isolates with
decreased susceptibility.116 Analysis of resistant strains found that the changes in
S10 are clustered in an extended loop of the protein that lies in close proximity to
the tigecycline binding site of the 16S rRNA, possibly interfering with access to or
binding of its target.117 Even modest increases in the tigecycline MIC are problematic
in serious infections such as bacteremia or endocarditis because of the low achievable
serum concentration of this antibiotic.

COMBINED ARMS—THERAPEUTIC STRATEGIES FOR THE TREATMENT OF

VANCOMYCIN-RESISTANT ENTEROCOCCI

Managing deep-seated enterococcal infections has always been a clinical challenge.

Even with the advent of penicillin, clinicians noted that IE caused by enterococci was
426 Miller et al

much more difficult to treat than that caused by streptococci, with mortality rates of
60% with penicillin monotherapy in IE.10 The discovery that aminoglycosides, when
added to b-lactams (or vancomycin), resulted in better patient outcomes and
in vitro synergism with bactericidal activity, led to this combination becoming the stan-
dard of care for enterococcal IE with cure rates increasing to 88%.118 However, the
spread of resistance determinants, including high-level resistance to ampicillin, ami-
noglycosides, and vancomycin led to a new era in which the cell wall synthesis inhib-
itor/aminoglycoside combination is obsolete in serious VRE infections.
In this scenario, the only compound with current US Food and Drug Administration
(FDA) approval for treatment of VRE infections is linezolid (Q/D previously carried such
an indication, but this approval has since been withdrawn). However, linezolid has the
limitation that it is a bacteriostatic antibiotic (less desirable in endovascular infections)
and that important hematologic and neurologic toxicity may emerge during prolonged
therapy (ie, in the case of IE). The only reliably in vitro bactericidal options against
multidrug-resistant VRE are DAP and oritavancin, although neither carry an FDA indi-
cation for VRE. The latter antibiotic (oritavancin) is only approved for soft tissue infec-
tions, and its use (including the dosing regimen) for deep seated infections remains to
be established. As a result, DAP (despite also lacking an indication for VRE) has
become almost the go-to antibiotic for severe VRE infections, including IE and other
endovascular infections. The reason is that DAP generally seems to retain bactericidal
activity against susceptible VRE strains, at least in vitro, and the toxicity profile is
favorable in infections that require prolonged administration.119 However, robust clin-
ical evidence to support the use of DAP over linezolid (an FDA-approved drug) is lack-
ing. Recent meta-analyses have tried to address this issue and attempted to
summarize the available clinical data on the treatment of VRE bacteremia with linezolid
compared with DAP.120–122 In the analysis conducted by Whang and colleagues,120
there was no statistical difference in clinical success between DAP and linezolid,
although there was a trend favoring the latter. In the studies by Balli and colleagues,121
and Chuang and colleagues,122 linezolid compared favorably with DAP with regard to
infection-related mortality (odds ratio, 3.61; 95% confidence interval [CI], 1.42–9.2)
and pooled all-cause mortality (odds ratio, 1.43; 95% CI, 1.09–1.86), respectively. In
contrast, a recent observational study of the largest cohort of patients with VRE
bacteremia yet published provided important insights into this question.123 Using
the Veterans Affairs medical record database, 644 patients with VRE bacteremia
treated exclusively with DAP or linezolid were identified. In this study, linezolid was
associated with a significantly higher risk of treatment failure compared with DAP
(risk ratio, 1.15; 95% CI, 1.02–1.3), which was defined as a composite endpoint of
30-day all-cause mortality, microbiologic failure, and 60-day recurrence of VRE
bacteremia. Although retrospective and observational in nature, the use of predefined
clinically relevant outcomes and propensity analysis strengthen the latter study con-
clusions. Despite these efforts, the lack of prospective randomized, controlled trials
precludes definitive conclusions regarding optimal therapy for VRE bacteremia.
An important consideration when using DAP is that the approved dose seems to
be suboptimal for the treatment of severe VRE infections. In a pharmacokinetic/phar-
macodynamic model using simulated endocardial vegetations, DAP used at both
6 mg/kg and 10 mg/kg resulted in bactericidal activity against VR E faecium at 8 hours,
but by 72 hours, bacteria from vegetations treated with 6 mg/kg showed regrowth.124
Further analysis of DAP at doses of 6, 8, 10, and 12 mg/kg against 2 clinical strains of
E faecium and one of E faecalis in the same pharmacokinetic/pharmacodynamic
model showed sustained reductions in colony count at 96 hours only in the groups
receiving 10 and 12 mg/kg.125 Importantly, the E faecalis strain exposed to 6 and
 Vancomycin-Resistant Enterococci 427

8 mg/kg, but not 12 mg/kg, developed reduced susceptibility to DAP with a change in
MIC from 0.5 mg/mL to 16 mg/mL. These experiments strongly suggest that DAP con-
centrations at the site of infection are likely to be of paramount importance for activity,
and the consideration of higher doses in serious infections is warranted.

Dual b-lactam Combinations

Recent data originating in Spain indicate that the combination of ampicillin and ceftri-
axone is equivalent to the standard b-lactam plus aminoglycoside combination with
less toxicity in E faecalis IE.126 The use of the double b-lactam combination was first
explored in vitro by showing that differential inhibition of the enterococcal PBPs by
cefotaxime and amoxicillin resulted in synergistic bactericidal effect.127 In a non-
randomized observational study of E faecalis IE involving 246 patients across multiple
centers in Spain and Italy, there were no significant differences in mortality during
treatment at 3-month follow-up or in relapses between standard combination therapy
of b-lactam plus aminoglycosides versus the dual b-lactam combination.126 However,
this study was not designed to assess noninferiority, and further experience is needed
before abandoning aminoglycosides for susceptible isolates. In vitro data suggest a
similar rationale with the use of ampicillin plus imipenem in E faecium, as this combi-
nation was able to reduce bacterial counts by 5 log when compared against single
agents in a rabbit model of endocarditis.128 It was also successfully used to cure a
patient with IE caused by E faecalis with high-level resistance to aminoglycosides
(HLRAg).129

Daptomycin Combinations
Combinations of DAP plus b-lactams are among the most promising and potent com-
binations against multidrug-resistant enterococcal isolates. Observations that resis-
tance to cationic antimicrobial peptides (such as DAP and the human cathelicidin
LL-37) sensitized various organisms (including staphylococci and enterococci) to
b-lactam antibiotics emerged from the use of the combination of DAP and b-lactams
as salvage therapy for recalcitrant enterococcal infections.130,131 Subsequent investi-
gations found in vitro synergistic effects between DAP plus b-lactams including ampi-
cillin, ceftaroline, and ertapenem.132–134 There is a lack of concrete clinical data as to
which agent provides the greatest benefit when added to the regimen, but case re-
ports most commonly feature the use of ampicillin or ceftaroline. Of note, the genetic
pathway to DAP resistance seems to influence the activity of the DAP/b-lactam com-
bination. Two recent publications73,135 testing the combination of DAP-ampicillin and
DAP-ceftaroline in vitro suggested that the presence of liaFSR mutations is indicative
of synergistic activity with these combinations. In contrast, synergism was not
observed in isolates that harbor mutations in yycFG (another regulatory system
involved in cell wall homeostasis). Experimental data in vitro have also shown that
the concomitant use of DAP and b-lactams (in this case amoxicillin and ampicillin)
delayed or prevented the emergence of DAP resistance in both E faecalis and E fae-
cium.136 This finding suggests that combination therapy with DAP plus b-lactams may
play a role in preventing emergence of DAP resistance, prolonging the usefulness
of this antibiotic in complicated VRE infections. Clinical data are urgently needed to
support the use of such a combination against enterococci.
In addition to b-lactams, DAP has been combined with a variety of other antimicro-
bial agents in challenging VRE infections. DAP plus tigecycline has been evaluated
both in vitro and in vivo against VRE, with synergism shown via the checkerboard
method, in time-kill curves, and in a mouse-wound model of enterococcal infection.137
There are several case reports of MDR E faecium IE also treated with this same
428 Miller et al

combination.138 The use of DAP plus aminoglycosides (gentamicin or streptomycin) in

the absence of HLRAg has also been reported139 and was found to be synergistic in an
animal model of endocarditis.140 There are some conflicting laboratory data on the use
of rifampin with DAP. One study found synergy between DAP and rifampin in 75% of
the 24 isolates of linezolid-resistant VRE tested by time-kill curve, without any evi-
dence of antagonism.141 A second study, however, found that rifampin antagonized
the action of both DAP and linezolid in time-kill studies against VRE, although this
was apparent only at a high inoculum (1 x 109 colony-forming units per milliliter).142
Further studies are needed to determine the role of DAP plus rifampin combinations,
although mutations in the rpoB gene conferring rifampin resistance have been asso-
ciated with DAP nonsusceptibility in other gram-positive organisms.143 Fosfomycin,
an agent currently available only in an oral formulation in the United States, is concen-
trated in the urine and may have a role in providing synergy with DAP against MDR
VRE urinary tract infections.144

Other Combinations
Linezolid is a bacteriostatic agent, and combination therapy to enhance activity or pre-
vent resistance would be of benefit, especially in deep-seated infections such as IE, or
if source control is not feasible. However, linezolid seems not to have a synergistic
effect when added to other antibiotics. For example, one study found that 99.2% of
1380 organism-drug combinations failed to show synergism with linezolid using the
checkerboard broth microdilution method.145 The combination of linezolid plus imipe-
nem or doxycycline did display synergism, but each in only one of 23 VRE isolates.
A second in vitro analysis determined that linezolid plus either doxycycline or Q/D
(tested isolates were sensitive to both) provided a synergistic effect.146 Therefore,
limited data suggest that linezolid combinations are unlikely to be reliable for treatment
of deep-seated VRE infections.
Q/D showed clinical response in 68.8% of patients infected with VRE in a prospec-
tive, noncomparative, multicenter study in which no other effective agents were avail-
able.147 A series of 56 patients with VRE infection at a major oncology center treated
with Q/D plus minocycline showed a similar response rate (68%).148 In vivo data from a
rabbit model of endocarditis showed that Q/D combined with imipenem or levofloxa-
cin resulted in greater reductions in vegetation colony counts than did Q/D alone.149
The combination of Q/D plus doxycycline was shown in vitro via the checkerboard
method to achieve synergism in 36% of E faecium clinical isolates.150 Clinically,
case reports of Q/D plus doxycycline and rifampin or Q/D plus high-dose ampicillin
(24 g/d) have also been successful in treating enterococcal IE.138

Therapeutic Approach
A framework for approaching complicated enterococcal infections is presented in
Fig. 1. For E faecalis isolates susceptible to ampicillin, this agent plus either an amino-
glycoside (if no HLRAg) or ceftriaxone (HLRAg) is still the treatment of choice. For pa-
tients with penicillin allergy, vancomycin may be substituted for ampicillin if the isolate
is not VRE, although vancomycin plus aminoglycosides may increase nephrotoxicity.
E faecium are often resistant to ampicillin, vancomycin, and the aminoglycosides
(gentamicin and streptomycin). Thus, in severe infections caused by these organisms,
DAP and linezolid are likely to be the drugs of choice. We favor DAP (particularly in
recalcitrant and endovascular infections) over linezolid because of the inherent bacte-
ricidal activity of the former. An important caveat is that particular attention should be
paid to both the DAP MIC of the isolate and the DAP dosage. In the case of MICs near
the breakpoint (eg, 3–4 mg/mL), and for deep-seated infections (endovascular or
 Vancomycin-Resistant Enterococci 429

E faecalis E faecium

Amp + CRO DAP-S DAP-T DAP-R

or DAP MIC ≤ 2 DAP MIC 3–4 DAP MIC >4
Amp + AG
___________ DAP 10–12 DAP 10–12 Linezolid
May use mg/kg mg/kg or
Amp/Sulbactam consider and DAP plus β-
For rare isolates β-lactam β-lactam lactam
with Amp-R due or Consider and
to produc on of Other ac ve adding another
β-lactamase a Other ac ve a
agent for agent(s)
a
deep-seated agent for with in vitro
infec ons not deep-seated ac vity
responding to infec ons
ini al therapy

Fig. 1. Suggested approach to complicated VRE infections. For ampicillin-sensitive E faecalis,

a combination of ampicillin plus either gentamicin or streptomycin, if no HLRAg, or ampi-
cillin plus ceftriaxone if HLRAg is present, is the preferred therapy. For E faecium, the au-
thors prefer DAP for its bactericidal action. For deep-seated infections or isolates with
DAP MIC 3 to 4 mg/mL (DAP tolerant) we suggest the addition of a b-lactam for synergism.
For isolates with DAP MIC greater than 4 linezolid may be used; however, if treating a deep-
seated or endovascular infection, consider the addition of DAP plus a b-lactam or at least
one other agent with demonstrated in vitro activity (eg, tigecycline, AG, or oritavancin). AG,
aminoglycoside; Amp, ampicillin; Amp-R, ampicillin resistant; CRO, ceftriaxone; DAP-R dap-
tomycin resistant; DAP-S, daptomycin susceptible; DAP-T, daptomycin tolerant.a Other active
agents (confirm with in vitro testing): (1) tigecycline, (2) linezolid, (3) oritavancin, (4) Q/D (E
faecium only), (5) AG (gentamicin or streptomycin) if no HLRAg.

source control not achievable), we favor the use of combination therapy (DAP plus
ampicillin or ceftaroline), using DAP at high doses (10–12 mg/kg). Additional active
agents may be required if bacteremia persists. In the case of resistance to both
DAP and linezolid, DAP plus b-lactams, Q/D, or oritavancin (if available) may be con-
siderations, in combination with at least 1 other active agent (eg, tigecycline).

SUMMARY—NEW MILLENNIUM, NEW STRATEGIES

Through the first decade of the 21st century, the emergence and spread of antibiotic
resistance determinants is presenting a new clinical challenge to physicians. Antibiotics
previously used for their reliability or low toxicity are being supplanted by newer agents
active against MDR organisms; however, therapeutic decisions must be made at the
bedside with limited clinical data available on how best to use these new drugs. Further-
more, the advances and expansion of the health care system have conspired to create
an environment that allows the transmission and proliferation of MDR organisms such
as VRE. There is concern that improper use of these new agents in this setting will
compromise their effectiveness or possibly promote the development of resistance.
430 Miller et al

As we tackle the spread of resistant bacteria, we must be judicious in our use of an-
tibiotics, the tools that make modern medicine possible. Novel combination therapies,
with old agents and new, may help take back the health care setting from MDR organ-
isms such as VRE. Strategies such as the use of daptomycin plus b-lactams offer the
potential for synergism against a bacterium that is difficult to kill. Analogous to com-
bination therapy in human immunodeficiency virus infection, such regimens may also
delay or preclude the development of resistance in VRE infections. Clinical data are
urgently needed to establish the effectiveness of such therapy and limit unnecessary
increases in complexity of regimens, health care costs, and side effects associated
with delivery.

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