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M E T H O D S I N M O L E C U L A R M E D I C I N E
 TM
Humana Press
 AntibioticResistance
Edited by 
Stephen H. Gillespie
 Methods and Protocols
 
Multiplex PCR Detection of VRE3 
1
Multiplex Polymerase Chain Reaction Detectionof
vanA
,
vanB 
,
vanC-1
,and
vanC-2/3 
Genesin
Enterococci 
Robin Patel, Jim R. Uhl, and Franklin R. Cockerill, III1. Introduction
Resistance to the glycopeptide antibiotic vancomycin in enterococci, is phe-notypically and genotypically heterogeneous. Three glycopeptide resistancephenotypes, VanA, VanB, and VanC, account for most glycopeptide resistancein enterococci; they can be distinguished on the basis of the level and induc-ibility of resistance to vancomycin and teicoplanin.VanA type glycopeptide resistance is characterized by acquired induc-ible resistance to both vancomycin and teicoplanin and has been describedfor
Enterococcus faecalis
,
Enterococcus faecium
,
Enterococcus gallinarum
,
 Enterococcus casseliflavus
,
Enterococcus durans
,
Enterococcus mundtii
,
 Enterococcus raffinosus
, and
 Enterococcus avium
(
Table 1
)
(1)
. VanA is themost completely understood type of vancomycin resistance. It is mediated bytransposon Tn
1546 
or related elements. Tn
1546 
was originally described on aplasmid from an
 E. faecium
isolate. It consists of a series of genes encoding 9polypeptides that can be assigned to different functional groups: Transpositionfunctions (ORF1 and ORF2), regulation of vancomycin resistance genes (VanRand VanS), synthesis of the depsipeptide, D-alanyl-D-lactate which whenincorporated into the pentapeptide peptidoglycan precursor form a pentapep-tide peptidoglycan precursor to which neither vancomycin nor teicoplanin willbind (VanH and VanA), and hydrolysis of normal peptidoglycan (VanX andVanY); the function of VanZ is unknown. The
vanR, vanS, vanH, vanA
, and
vanX 
genes are necessary and sufficient for the inducible expression of resis-tance to glycopeptides. VanY and VanZ are accessory peptides and are notrequired for resistance. Genetic heterogeneity has been described in
vanA
gene
3 
From:
Methods in Molecular Medicine, vol. 48: Antibiotic Resistance Methods and Protocols 
Edited by: S. H. Gillespie © Humana Press Inc., Totowa, NJ
 
4Patel, Uhl, and Cockerill 
clusters of vancomycin resistant enterococci (VRE). The
vanA
gene cluster hasbeen found on the chromosome as well as on plasmids.VanB type glycopeptide resistance is characterized by acquired inducibleresistance to various concentrations of vancomycin but not to teicoplaninand has been described in
 E. faecalis
and
E. faecium
(
Table 1
). The
vanB
gene cluster, as described in an
 E. faecalis
isolate, has homology to the
vanA
gene cluster but has been less well studied. It appears to be located onthe chromosome.VanC type glycopeptide resistance is a less well characterized type of van-comycin resistance. VanC type glycopeptide resistance is characterized by lowlevel vancomycin resistance but teicoplanin susceptibility and has beendescribed as an intrinsic property of 
 E. gallinarum
,
E. casseliflavus
, and
 Enterococcus flavescens
(
Table 1
)
(2–4)
. The VanC phenotype is felt to bechromosomally encoded and expressed constitutively, although recent datasuggest that vancomycin resistance may be inducible in at least some strains of 
 E. gallinarum
. Pentapeptide peptidoglycan precursors in strains with VanCvancomycin resistance terminate in the D-alanyl-D-serine rather than in D-alanyl-D-alanine. The genes encoding for the synthesis of the depsipeptide D-alanyl-D-serine are referred to as
vanC-1
(in
 E. gallinarum
),
vanC-2
(in
 E. casseliflavus
)and
vanC-3
(in
 E. flavescens).
Table 1Vancomycin Resistant Enterococci
VancomycinTeicoplaninMICMICBacterialPhenotypeGenotype(
µ
g/mL)(
µ
g/mL)ExpressionTransferspeciesVanA
vanA
64–100016–512Inducible+
E. faecium E. faecalis E. avium E. gallinarum E. durans E. mundtii E. casseliflavus E. raffinosus
VanB
vanB
4–10000.5–1Inducible+
E. faecium E. faecalis
VanC
vanC-1
2–320.5–1Constitutive/–
E. gallinarum
InducibleVanC
vanC-2
2–320.5–1Constitutive–
E. casseliflavus
VanC
vanC-3
2–320.5–1Constitutive–
E. flavescens
of 00

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dawdshanu

Antibiotic Resistance

reply04 / 15 / 2010
dhanisidik

mikrobiologi kedokteran

reply01 / 29 / 2010
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