High Level Oxacillin and Vancomycin Resistance and Altered Cell Wall Composition in Staphylococcus aureus Carrying the Staphylococcal mecA and the Enterococcal vanA Gene Complex

Abstract

Recently, for the first time in the history of this bacterial species, methicillin-resistant Staphylococcus aureus (MRSA) carrying the enterococcal vanA gene complex and expressing high level resistance to vancomycin was identified in clinical specimens (CDC (2002) MMWR 51, 565–567). The purpose of our studies was to understand how vanA is expressed in the heterologous background of S. aureus and how it interacts with the mecA-based resistance mechanism, which is also present in these strains and is targeted on cell wall biosynthesis. The vanA-containing staphylococcal plasmid was transferred from the clinical vancomycin-resistant S. aureus (VRSA) strain HIP11714 (CDC (2002) MMWR 51, 565–567) to the methicillin-resistant S. aureus (MRSA) strain COL for which extensive genetic and biochemical information is available on staphylococcal cell wall biochemistry and drug resistance mechanisms. The transconjugant named COLVA showed high and homogeneous resistance to both oxacillin and vancomycin. COLVA grown in vancomycin-containing medium produced an abnormal peptidoglycan: all pentapeptides were replaced by tetrapeptides, and the peptidoglycan contained at least 22 novel muropeptide species that frequently showed a deficit or complete absence of pentaglycine branches. The UDP-MurNAc-pentapeptide, the major component of the cell wall precursor pool in vancomycin-sensitive cells was replaced by UDP-MurNAc-depsipeptide and UDP-MurNAc-tetrapeptide. Transposon inactivation of the β-lactam resistance gene mecA caused complete loss of β-lactam resistance but had no effect on the expression of vancomycin resistance. The two major antibiotic resistance mechanisms encoded by mecA and vanA residing in the same S. aureus appear to use different sets of enzymes for the assembly of cell walls. Recently, for the first time in the history of this bacterial species, methicillin-resistant Staphylococcus aureus (MRSA) carrying the enterococcal vanA gene complex and expressing high level resistance to vancomycin was identified in clinical specimens (CDC (2002) MMWR 51, 565–567). The purpose of our studies was to understand how vanA is expressed in the heterologous background of S. aureus and how it interacts with the mecA-based resistance mechanism, which is also present in these strains and is targeted on cell wall biosynthesis. The vanA-containing staphylococcal plasmid was transferred from the clinical vancomycin-resistant S. aureus (VRSA) strain HIP11714 (CDC (2002) MMWR 51, 565–567) to the methicillin-resistant S. aureus (MRSA) strain COL for which extensive genetic and biochemical information is available on staphylococcal cell wall biochemistry and drug resistance mechanisms. The transconjugant named COLVA showed high and homogeneous resistance to both oxacillin and vancomycin. COLVA grown in vancomycin-containing medium produced an abnormal peptidoglycan: all pentapeptides were replaced by tetrapeptides, and the peptidoglycan contained at least 22 novel muropeptide species that frequently showed a deficit or complete absence of pentaglycine branches. The UDP-MurNAc-pentapeptide, the major component of the cell wall precursor pool in vancomycin-sensitive cells was replaced by UDP-MurNAc-depsipeptide and UDP-MurNAc-tetrapeptide. Transposon inactivation of the β-lactam resistance gene mecA caused complete loss of β-lactam resistance but had no effect on the expression of vancomycin resistance. The two major antibiotic resistance mechanisms encoded by mecA and vanA residing in the same S. aureus appear to use different sets of enzymes for the assembly of cell walls. Until the late 1990s, clinical isolates of Saccharomyces aureus have retained a uniform high sensitivity to vancomycin with minimal inhibitory concentration (MIC) 1The abbreviations used are: MICminimal inhibition concentrationUDP-MurNAc-pentapeptideuridinediphospho-N-acetylmuramyl-l-Ala-d-Glu-l-Lys-d-Ala-d-AlaUDP-MurNAc-tetrapeptideuridine-diphospho-N-acetylmuramyl-l-Ala-d-Glu-l-Lys-d-AlaUDP-MurNAc-depsipeptideuridinediphospho-N-acetylmuramyl-l-Ala-d-Glu-l-Lys-d-Ala-d-LacMurNAcN-acetylmuramic acidGlcNAcN-acetyl-glucosamineLipid IIundecaprenylpyrophosphoryl-N-acetylmuramyl-(peptide)-N-acetylglucosamineMS/MSfragmentation analysis by mass spectrometryLC-MSliquid chromatography (HPLC) coupled to mass spectrometryHPLChigh performance liquid chromatographyPBPpenicillin-binding proteinVRSAvancomycin-resistant S. aureusMRSAmethicillin-resistant S. aureusBHIbrain heart infusion.1The abbreviations used are: MICminimal inhibition concentrationUDP-MurNAc-pentapeptideuridinediphospho-N-acetylmuramyl-l-Ala-d-Glu-l-Lys-d-Ala-d-AlaUDP-MurNAc-tetrapeptideuridine-diphospho-N-acetylmuramyl-l-Ala-d-Glu-l-Lys-d-AlaUDP-MurNAc-depsipeptideuridinediphospho-N-acetylmuramyl-l-Ala-d-Glu-l-Lys-d-Ala-d-LacMurNAcN-acetylmuramic acidGlcNAcN-acetyl-glucosamineLipid IIundecaprenylpyrophosphoryl-N-acetylmuramyl-(peptide)-N-acetylglucosamineMS/MSfragmentation analysis by mass spectrometryLC-MSliquid chromatography (HPLC) coupled to mass spectrometryHPLChigh performance liquid chromatographyPBPpenicillin-binding proteinVRSAvancomycin-resistant S. aureusMRSAmethicillin-resistant S. aureusBHIbrain heart infusion. values in the vicinity of 1 μg/ml. Beginning with the late 1990s, isolates with reduced susceptibility to vancomycin began to be reported from several countries (1Walsh T.R. Howe R.A. Annu. Rev. Microbiol. 2002; 56: 657-675Google Scholar), but the MIC value of these vancomycin intermediate-resistant S. aureus isolates was limited to the range of 8–16 μg/ml of the antibiotic. The first highly vancomycin-resistant S. aureus (VRSA) isolates (MIC over 32 μg/ml) were only detected during the last year in two hospitals in the United States (2CDC MMWR. 2002; 51: 565-567Google Scholar, 3CDC MMWR. 2002; 51: 902Google Scholar). The appearance of such VRSA strains in clinical specimens is of obvious and grave concern, because the spread of VRSA isolates may seriously jeopardize the chemotherapy of multidrug-resistant S. aureus disease and raise the specter of untreatable staphylococcal infections (4Chang S. Sievert D.M. Hageman J.C. Boulton M.L. Tenover F.C. Pouch-Downes F. Shah S. Rudrik J.T. Pupp G.R. Brown W.J. Cardo D. Fridkin S.K. N. Engl. J. Med. 2003; 348: 1342-1347Google Scholar, 5Gonzalez-Zorn B. Courvalin P. Lancet Infect. Dis. 2003; 3: 67-68Google Scholar). minimal inhibition concentration uridinediphospho-N-acetylmuramyl-l-Ala-d-Glu-l-Lys-d-Ala-d-Ala uridine-diphospho-N-acetylmuramyl-l-Ala-d-Glu-l-Lys-d-Ala uridinediphospho-N-acetylmuramyl-l-Ala-d-Glu-l-Lys-d-Ala-d-Lac N-acetylmuramic acid N-acetyl-glucosamine undecaprenylpyrophosphoryl-N-acetylmuramyl-(peptide)-N-acetylglucosamine fragmentation analysis by mass spectrometry liquid chromatography (HPLC) coupled to mass spectrometry high performance liquid chromatography penicillin-binding protein vancomycin-resistant S. aureus methicillin-resistant S. aureus brain heart infusion. minimal inhibition concentration uridinediphospho-N-acetylmuramyl-l-Ala-d-Glu-l-Lys-d-Ala-d-Ala uridine-diphospho-N-acetylmuramyl-l-Ala-d-Glu-l-Lys-d-Ala uridinediphospho-N-acetylmuramyl-l-Ala-d-Glu-l-Lys-d-Ala-d-Lac N-acetylmuramic acid N-acetyl-glucosamine undecaprenylpyrophosphoryl-N-acetylmuramyl-(peptide)-N-acetylglucosamine fragmentation analysis by mass spectrometry liquid chromatography (HPLC) coupled to mass spectrometry high performance liquid chromatography penicillin-binding protein vancomycin-resistant S. aureus methicillin-resistant S. aureus brain heart infusion. The VRSA strain HIP11714 recovered from a dialysis patient in Detroit, Michigan has acquired the vanA gene complex most likely from a vancomycin-resistant Enterococcus faecalis strain coinfecting the diabetic wound and catheter insertion site of the patient (1Walsh T.R. Howe R.A. Annu. Rev. Microbiol. 2002; 56: 657-675Google Scholar). HIP11714 was also methicillin-resistant: it carried the heterologous mecA gene, the key component of wide spectrum β-lactam resistance. Clinical aspects of this case (1Walsh T.R. Howe R.A. Annu. Rev. Microbiol. 2002; 56: 657-675Google Scholar), the genetic background of HIP11714 (4Chang S. Sievert D.M. Hageman J.C. Boulton M.L. Tenover F.C. Pouch-Downes F. Shah S. Rudrik J.T. Pupp G.R. Brown W.J. Cardo D. Fridkin S.K. N. Engl. J. Med. 2003; 348: 1342-1347Google Scholar), and the structure of the S. aureus plasmid pLW1043 into which the transposon Tn1546 integrated 2Weigel, L. M., Clewell, D. B., Gill, S. R., Clark, N. C., McDougal, L. K., Flannagan, S. E., Kolonay, J. F., Shetty, J., Killgore, G. E., and Tenover, F. C. (2003) Science302, 1569–1571.2Weigel, L. M., Clewell, D. B., Gill, S. R., Clark, N. C., McDougal, L. K., Flannagan, S. E., Kolonay, J. F., Shetty, J., Killgore, G. E., and Tenover, F. C. (2003) Science302, 1569–1571. are the subject of separate publications. The purpose of the study described here was different: we wanted to examine how two very different antibiotic resistance mechanisms, based either on mecA or on vanA, are expressed in the same S. aureus cell and what impact the enterococcal vanA gene complex may have on the structure of the staphylococcal cell wall. The mechanism of vanA-mediated vancomycin resistance, at least when it is expressed in enterococci, involves the production of a cell wall precursor with abnormal chemical structure (for recent review see Ref. 6Boneca I.G. Chiosis G. Expert Opin. Ther. Targets. 2003; 7: 311-328Google Scholar). The mechanism of mecA-based β-lactam resistance involves the production of a new cell wall biosynthetic enzyme, the penicillin-binding protein 2A (PBP2A) (7Hartman B.J. Tomasz A. J. Bacteriol. 1984; 158: 513-516Google Scholar, 8Reynolds P.E. Brown D.F.J. FEBS Lett. 1985; 192: 28-32Google Scholar, 9Utsui Y. Yokota T. Antimicrob. Agents Chemother. 1985; 28: 397-403Google Scholar). To understand interactions between these two resistance mechanisms each targeting the bacterial cell wall, we transferred the vanA-carrying plasmid from the clinical strain HIP1174 to the MRSA strain COL, in which extensive biochemical and genetic information on cell wall structure (10Boneca I.G. Huang Z.-H. Gage D.A. Tomasz A. J. Biol. Chem. 2000; 275: 9910-9918Google Scholar, 11Ornelas-Soares A. de Lencastre H. de Jonge B. Gage D. Chang Y.-S. Tomasz A. J. Biol. Chem. 1993; 268: 26268-26272Google Scholar, 12Sobral R.G. Ludovice A.M. Gardete S. Tabei K. de Lencastre H. Tomasz A. Microb. Drug Resist. 2003; 9: 231-241Google Scholar) and biosynthesis (13de Jonge B.L. Chang Y.-S. Gage D. Tomasz A. J. Biol. Chem. 1992; 267: 11248-11254Google Scholar, 14Pinho M.G. Filipe S. de Lencastre H. Tomasz A. J. Bacteriol. 2001; 183: 6525-6531Google Scholar, 15Pinho M.G. de Lencastre H. Tomasz A. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 10886-10891Google Scholar) is available. Strain COL does not carry chromosomal or plasmid-born regulatory elements that often control the transcription of mecA in clinical MRSA isolates (16Weller T.M. J. Antimicrob. Chemother. 1999; 43: 15-22Google Scholar, 17Kobayashi T. Zhu Y.F. Nicholls N.J. Lampen J.O. J. Bacteriol. 1987; 169: 3873-3878Google Scholar). Strain COL is one of the fully sequenced MRSA strains (18Ito T. Katayama Y. Hiramatsu K. Antimicrob. Agents Chemother. 1999; 43: 1449-1458Google Scholar) in which a large body of genetic information is also available concerning genetic determinants, auxiliary genes (19de Lencastre H. Tomasz A. Antimicrob. Agents Chemother. 1994; 38: 2590-2598Google Scholar, 20de Lencastre H. Wu S.W. Pinho M.G. Ludovice A.M. Filipe S.R. Gardete S. Sobral R. Gill S. Chung M. Tomasz A. Microb. Drug Resist. 1999; 5: 163-175Google Scholar, 21Ray S.S. Bonanno J.B. Rajashankar K.R. Pinho M. He G. de Lencastre H. Tomasz A. Burley S.K. Structure. 2002; 10: 1499-1508Google Scholar, 22Ray S.S. Bonanno J.B. Chen H. de Lencastre H. Wu S. Tomasz A. Burley S.K. Proteins Struct. Funct. Genet. 2003; 50: 170-173Google Scholar), that are essential for the optimal expression of β-lactam resistance. These genetic determinants may provide useful tools for the dissection of the vanA- and mecA-mediated resistance mechanisms. Construction of Strain COLVA by Filter Mating—In the filter mating experiments the recipient used was the MRSA strain COL, and the donor was the clinical VRSA isolate HIP11714, which carried transposon Tn1546 on pLW1043. of donor and recipient grown in brain heart Detroit, were in a to 1 in of the donor a filter by of the on at for the were into and on μg/ml vancomycin μg/ml for the of the was by two on the and was transferred to the at the the and of the drug resistance was and the transconjugant was named strain and aureus strains were grown in at with was by the at with an peptidoglycan and cell wall precursor were on grown in the of or μg/ml vancomycin and also in μg/ml these experiments the grown in with the were into medium the same antibiotic resistance level and of were by the the by the and by the of analysis A. S. H. Antimicrob. Agents Chemother. Scholar). and of wall peptidoglycan was and by described (10Boneca I.G. Huang Z.-H. Gage D.A. Tomasz A. J. Biol. Chem. 2000; 275: 9910-9918Google Scholar, 11Ornelas-Soares A. de Lencastre H. de Jonge B. Gage D. Chang Y.-S. Tomasz A. J. Biol. Chem. 1993; 268: 26268-26272Google Scholar, Jonge B.L. Chang Y.-S. Gage D. Tomasz A. J. Biol. Chem. 1992; 267: 11248-11254Google Scholar). The by the were reduced by and by high performance liquid chromatography (HPLC) with a described (13de Jonge B.L. Chang Y.-S. Gage D. Tomasz A. J. Biol. Chem. 1992; 267: 11248-11254Google Scholar). The of were and by (13de Jonge B.L. Chang Y.-S. Gage D. Tomasz A. J. Biol. Chem. 1992; 267: 11248-11254Google Scholar), and analysis was of and by of the were by a described D. B. J. J. Bacteriol. Scholar) with K. Tomasz A. J. Bacteriol. Scholar). strains COL and COLVA grown in μg/ml or was to the the cells to the of cell wall experiments in our have that an of the of in the of cell Proc. Natl. Acad. Sci. U. S. A. Scholar, N. Y. Acad. Sci. Scholar) be used of vancomycin D. L. D. D. J. J. D.M. J. Bacteriol. Scholar) for the of the cell wall precursor was with a were to a The was at a of with a of in to in in The of was by was The were detected by at was used to by coupled to mass T.R. Scholar, J. S.K. Chem. 1993; Scholar). The was for the and it was reduced to the To the of and the was the of the was to by of were by and in was at a of into a mass in the analysis Zhu J. 3: Scholar, T.R. Huang Chem. Scholar) of was at a of with the produced to the and of the The was to an of analysis a Scholar, Chem. Scholar) with a at of to and in Strain COLVA grown either in or in with μg/ml vancomycin were for susceptibility to oxacillin and vancomycin by the The grown in showed high level resistance to oxacillin μg/ml) and the grown in vancomycin showed vancomycin resistance μg/ml) of the of Strain COLVA in and of recovered from the peptidoglycan of strain COL was with that of strain COLVA with both strains grown in The two were that in strain COLVA to of the muropeptide of strain COL (13de Jonge B.L. Chang Y.-S. Gage D. Tomasz A. J. Biol. Chem. 1992; 267: 11248-11254Google Scholar) was often a from the with in COLVA grown in the of vancomycin these novel the muropeptide species of the peptidoglycan of COLVA in the of oxacillin μg/ml) caused a of all but one of the novel and the peptidoglycan to a of strain COL grown in the of β-lactam (13de Jonge B.L. Chang Y.-S. Gage D. Tomasz A. J. Biol. Chem. 1992; 267: 11248-11254Google Scholar) of the in Strain COLVA in novel muropeptide that in the cells were and to mass the and for the 22 new muropeptide species The chemical from the are in The most and novel of these are the complete of pentapeptides by and the deficit complete of pentaglycine in in muropeptide and in the donor of were in that we for of of the acid and the of and and The of muropeptide in grown different is in and acid of cell wall from vancomycin-resistant strain are in The in the were from and of the reduced were from acid of the reduced with structure was for the muropeptide are in The in the were from and of the reduced were from acid of the reduced structure was for the muropeptide in a new of major in the peptidoglycan of strain COL and vancomycin-resistant strain of of muropeptide species are expressed the of recovered from the of muropeptide species are expressed the of recovered from the in a new of the the of the cell wall precursor pool in strains COL and COLVA grown either in medium or in with These are the same that were for effect on the of the peptidoglycan in strains COL and COLVA grown in the with mass analysis identified in the precursor pool that were and were present in in both strains and in first and component was only detected in strain and were identified the cell wall muropeptide and the of B. D. J. Scholar), the to of the precursor was identified in both COL and COLVA a of the carrying a on the of the The precursor pool of COLVA not also contained an component which had time and mass of the described in vancomycin-resistant S. J. J. Bacteriol. 1992; Scholar, S. M. Courvalin P. Scholar). different were when the cell wall precursor pool was from COLVA grown in vancomycin-containing medium to be present the major component in the grown was and was replaced by two new major and several were also analysis of identified it the had a time of on the and a mass that by 1 mass from that of the The of the cell wall precursor pool in strain COLVA grown in with μg/ml oxacillin was from the precursor of COLVA grown in the of of the UDP-MurNAc-depsipeptide The of in the cell wall precursor pool in COL and COLVA grown different is in of cell wall precursor pool in the strain COL strain of cells were for the last of to μg/ml cells were for the last of to μg/ml cells were for the last of to μg/ml and and were integrated To cells were for the last of to μg/ml and were integrated in a new of the of UDP-MurNAc-depsipeptide by analysis T.R. Scholar, J. S.K. Chem. 1993; Scholar) in the showed that the at time has a mass of the value of for To provide a for the analysis of the was of the at two at and that to the loss of and from the precursor and The at to the structure The first at the in this from the of acid between and To the of the fragmentation of the at was by the precursor at the Scholar, Chem. Scholar) with a high of The fragmentation from the a of at and to the loss of and of were also to the fragmentation from the were and to the and The at and be to the and The last two fragmentation of from both the and the loss of and from the described the appearance at and The at may be the of the loss of and from the was A. N. A. Tabei K. D. D. J. M. J. Biol. Chem. 2002; Scholar). the provide a that the from the COLVA strain of S. aureus grown in the of vancomycin of the of the cell wall precursor of the of by of of COLVA grown in was at on two sets of one contained only of vancomycin in the range of μg/ml to μg/ml. the of vancomycin the medium also contained oxacillin at a concentration of μg/ml. The of bacterial was at for cells on the μg/ml and the of on the μg/ml vancomycin. The of oxacillin reduced the vancomycin MIC value from to μg/ml in of the and a of the of by of to the one in was to the effect of of vancomycin on the oxacillin resistance of strain of μg/ml vancomycin in the oxacillin reduced the oxacillin MIC of strain COLVA from to μg/ml in of the and the in the of of mecA on and of Strain mecA P. Tomasz A. Antimicrob. Agents Chemother. Scholar) was into strain COLVA to The of showed that has retained high level resistance to vancomycin but resistance to oxacillin was reduced to μg/ml Strain was grown in μg/ml and cell and precursor pool and The cell wall precursor pool showed a very to that of COLVA grown in vancomycin-containing medium The of the cell wall peptidoglycan is in is that COLVA with an mecA produced a muropeptide that was very to the muropeptide of strain COLVA that carried an are and mechanism of involves inhibition of bacterial that cell wall vancomycin to the of the cell wall precursor and it from for cell wall The mechanisms of β-lactam and resistance the of of these a with for (7Hartman B.J. Tomasz A. J. Bacteriol. 1984; 158: 513-516Google Scholar, 8Reynolds P.E. Brown D.F.J. FEBS Lett. 1985; 192: 28-32Google Scholar, 9Utsui Y. Yokota T. Antimicrob. Agents Chemother. 1985; 28: 397-403Google Scholar). a cell wall precursor with the of which is only by the antibiotic I.G. Chiosis G. Expert Opin. Ther. Targets. 2003; 7: 311-328Google Scholar) Strain COLVA both of drug resistance, and the purpose of the studies described here was to examine how these two mechanisms, one based on the mecA and the on the vanA gene are expressed in the same staphylococcal The of and analysis that strain COLVA carrying both mecA and vanA of homogeneous of that are to high of either oxacillin μg/ml) or vancomycin of the cell wall precursor pool of COLVA is with the expression of the vanA gene complex in the of vancomycin. The UDP-MurNAc-pentapeptide, which was the component of the pool of both strains COL and COLVA grown in medium was in and was replaced by two new in COLVA grown in vancomycin-containing These two were identified the and of These and the appearance of and are to the chemical of the cell wall described in vancomycin-resistant D. L. J. Antimicrob. Agents Chemother. 1992; Scholar, M. F. P. Courvalin P. Microbiol. Scholar, M. F. L. P. Courvalin P. Microbiol. Scholar). at the of peptidoglycan from COL and COLVA that the of S. aureus wall were the of high from the with were in COL, in COLVA grown in and in COLVA grown in the of vancomycin. The of and was also in COL and and in COLVA grown in and in COLVA grown in vancomycin-containing these in of the of the muropeptide of strains COL and COLVA several major The of COLVA grown in a of that are from strain COL grown the same These the muropeptide species in the peptidoglycan of COLVA grown in the of vancomycin. of the new muropeptide species by mass spectrometry showed that the from both the and also from the muropeptide were also in the pentaglycine which are the of the cell wall of COL and S. aureus strains (13de Jonge B.L. Chang Y.-S. Gage D. Tomasz A. J. Biol. Chem. 1992; 267: 11248-11254Google Scholar). The in that the deficit in the pentaglycine was in and that may have the during the of muropeptide and and in of not in strain COL was the of of the acid or the of that we the of an and for and strain COLVA was of expressing high level resistance to both oxacillin and these two had and on the expression of the μg/ml oxacillin reduced the vancomycin MIC value of strain COLVA from to μg/ml. μg/ml vancomycin reduced the oxacillin MIC of COLVA from to μg/ml. both the homogeneous were also to The experiments in and 1 provide into in the mechanism of cell wall in expressing either the β-lactam or the vancomycin resistance has that in strain COL the assembly of peptidoglycan in the of oxacillin is by two the of and the of M.G. Filipe S. de Lencastre H. Tomasz A. J. Bacteriol. 2001; 183: 6525-6531Google Scholar, 15Pinho M.G. de Lencastre H. Tomasz A. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 10886-10891Google Scholar). The inhibition of expression of vancomycin resistance by oxacillin in strain COLVA that the which is the only that in the of oxacillin (13de Jonge B.L. Chang Y.-S. Gage D. Tomasz A. J. Biol. Chem. 1992; 267: 11248-11254Google Scholar), is not to the cell wall The inhibition of vancomycin resistance by in has in a was that which has to is to wall S. D. J. L. Microbiol. Lett. 1992; Scholar). the of UDP-MurNAc-depsipeptide precursor and in the peptidoglycan of strain COLVA be by one of the highly oxacillin is with the of peptidoglycan of COLVA grown in the of for the muropeptide all the were the of S. aureus grown in the of (13de Jonge B.L. Chang Y.-S. Gage D. Tomasz A. J. Biol. Chem. 1992; 267: 11248-11254Google Scholar). for this from the inactivation of mecA in strain which had no on the vancomycin MIC it resistance to oxacillin The sensitivity of the to of vancomycin may be by the same of the vanA gene complex by vancomycin to the of the cell wall precursor in and the cell wall precursor pool with the abnormal in abnormal cell wall precursor to be an for which is the only available for wall in the of oxacillin in the The of to is of that the high of sensitivity of this protein to of the structure of the of cell wall Jonge B.L. Chang N. Gage D. Antimicrob. Agents Chemother. Scholar, Jonge B.L. Gage D. N. Antimicrob. Agents Chemother. 2002; Scholar). that the vanA gene complex be fully expressed in S. aureus with several of the mechanism that were identified in vancomycin-resistant M. R. Antimicrob. Agents Chemother. 2001; Scholar). The novel of our study was that we the in cell wall from the of wall precursor pool all the to the of the studies no in the of the cell of and vancomycin-resistant Jonge B.L. Gage D. S. Drug Resist. Scholar, D. D. D. D. J. L. J. Scholar). a of the cell wall structure of and vancomycin-resistant faecalis has the complete absence of or from the peptidoglycan of the A. N. A. Tabei K. D. D. J. M. J. Biol. Chem. 2002; Scholar, Jonge B.L. S. Gage D. Antimicrob. Agents Chemother. Scholar). was Jonge B.L. S. Gage D. Antimicrob. Agents Chemother. Scholar) that the may be of a the of which the from the peptidoglycan of the The complete of pentapeptides by in the peptidoglycan of COLVA grown in vancomycin-containing medium may be by a the of both and abnormal in COLVA grown in medium that the have a high of for the of acid The absence or complete pentaglycine in muropeptide of grown in the of vancomycin that the is not an for the and S. B. Antimicrob. Agents Chemother. 2003; Scholar). that such abnormal carrying pentaglycine may in the biosynthetic and muropeptide into the pentaglycine are also in the donor that in pentaglycine are not in the was that the of to pentaglycine in the cell wall of S. aureus H. H. B. J. Biol. Chem. Scholar). to be the deficit in pentaglycine in vancomycin S. aureus also of and the of such The vancomycin resistance of the strain of COLVA in which mecA was that the key protein of β-lactam resistance, does not in the expression of vancomycin resistance but is to the biosynthesis of cell in the of β-lactam of the muropeptide be by of the of S. The two major antibiotic resistance mechanisms encoded by mecA and vanA residing in the same S. aureus appear to use different sets of enzymes for the assembly of cell walls. for in of the COLVA