Original Article Published on 14 Mar, 2024

Metabolic Profiling Reveals the Importance of Arginine Metabolism via the Arginine Deiminase Pathway in Vancomycin-intermediate S. aureus

Metabolomics of Vancomycin-intermediate S. aureus
Vol. 19 No. 1 : 2024 (204-219)

Tan JK Tan JK
Tan XE Tan XE
Neoh HM Neoh HM

Abstract
Gene mutations located in VraSR and GraSR, two-component regulatory systems have been reported to cause vancomycin resistance in vancomycin-intermediate Staphylococcus aureus (VISA). Subsequent comparative proteomic profiling between vancomycin-susceptible S. aureus (VSSA) and VISAs of the Mu50 genetic lineage revealed up-regulated level of catabolic ornithine carbamoyltransferase (ArcB) in the latter, suggesting a role of arginine catabolism in VISA development. This study aimed to further investigate metabolic pathways associated with VISA development using liquid chromatography mass spectrometry (LCMS)-based untargeted metabolomic profiling. Metabolite profiles were compared among 3 isogenic strains of the Mu50 lineage: Mu50Ω (VSSA), Mu50Ω-vraSm (VISA) and Mu50Ω-vraSm-graRm (VISA). Intracellular levels of α-hydroxyglutaric acid, choline, lysine, malic acid, N-acetylornithine, nicotinamide, and cystathionine were found to be significantly different between VISAs compared to VSSA. VISA cells were found to have lower levels of intracellular citrulline and extracellular arginine, with higher extracellular level of ornithine compared to VSSA. These differences in metabolite levels between VISA and VSSA suggested the importance of arginine metabolism. Activation of the arginine deiminase pathway (ADI) in VISAs of the Mu50 lineage could be further explored as a target for antibiotic development.
Keywords : Arginine deiminase (ADI) pathway, metabolomics, vancomycin-intermediate S. aureus (VISA),
Abstrak
Mutasi gen dalam VraSR dan GraSR, dua sistem pengawalseliaan dua komponen telah dilaporkan menyebabkan rintangan vankomisin dalam Staphylococcus aureus berkerintangan perantaraan vankomisin (VISA). Pemprofilan proteomik perbandingan antara S. aureus rentan vankomisin (VSSA) dan Staphylococcus aureus rintang vankomisin (VISA) daripada keturunan genetik Mu50 mendedahkan pengekspresan yang meningkat untuk protein katabolik ornithine carbamoyltransferase (ArcB); ini mencadangkan peranan katabolisme arginin dalam pembentukan VISA. Kajian ini bertujuan untuk menyiasat dengan lebih lanjut tapak laluan metabolik yang berkaitan dengan pembentukan VISA menggunakan pemprofilan metabolomik tidak bersasar berasaskan "liquid chromatography mass spectrometry" (LCMS). Profil metabolit antara 3 strain isogenik keturunan Mu50 iaitu Mu50Ω (VSSA), Mu50Ω-vraSm (VISA) dan Mu50Ω-vraSm-graRm (VISA) telah dibanding. Paras intrasel bagi asid α-hidroksiglutarik, kolin, lisin, asid malik, N-asetilornithine, nikotinamida dan cystathionine didapati berbeza dengan ketara antara VISA berbanding VSSA. Sel VISA didapati mempunyai tahap citrulline intraselular dan arginin ekstraselular yang lebih rendah, dengan tahap ornithine ekstraselular yang lebih tinggi berbanding VSSA. Perbezaan tahap metabolit antara VISA dan VSSA ini mencadangkan kepentingan metabolisme arginine dalam kerintangan perantaraan kepada vankomisin. Pengaktifan tapak laluan arginine deiminase (ADI) dalam VISA dari keturunan Mu50 boleh diterokai lebih lanjut sebagai sasaran untuk pembangunan antibiotik.
Kata Kunci : Metabolomik, Staphylococcus aureus berkerintangan-perantaraan vankomisin (VISA), tapak laluan arginine deiminase (ADI),

Correspondance Address
Hui-min Neoh. UKM Medical Molecular Biology Institute (UMBI), Universiti Kebangsaan Malaysia, Jalan Yaacob Latif, 56000 Cheras, Kuala Lumpur, Malaysia. Tel: +603-9145-9074 Email: hui-min@ppukm.ukm.edu.my

References
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Chong, J., Soufan, O., Li, C., Caraus, I., Li, S., Bourque, G., Wishart, D.S., Xia, J. 2018. MetaboAnalyst 4.0: Towards more transparent and integrative metabolomics analysis. Nucleic Acids Res 46(W1): W486-94.

Cui, L., Lian, J.Q., Neoh, H.M., Reyes, E., Hiramatsu, K. 2005. DNA microarray-based identification of genes associated with glycopeptide resistance in Staphylococcus aureus. Antimicrob Agents Chemother 49(8): 3404-13.

Cui, L., Murakami, H., Kuwahara-Arai, K., Hanaki, H., Hiramatsu, K. 2000. Contribution of a thickened cell wall and its glutamine nonamidated component to the vancomycin resistance expressed by Staphylococcus aureus Mu50. Antimicrob Agents Chemother 44(9): 2276-85.

Cui, L., Neoh, H.M., Shoji, M., Hiramatsu, K. 2009. Contribution of vraSR and graSR point mutations to vancomycin resistance in vancomycin-intermediate Staphylococcus aureus. Antimicrob Agents Chemother 53(3): 1231-4.

Dunn, W.B., Broadhurst, D., Begley, P., Zelena, E., Francis-Mcintyre, S., Anderson, N., Brown, M., Knowles, J.D., Halsall, A., Haselden, J. N., Nicholls, A.W., Wilson, I.D., Kell, D.B., Goodacre, R. 2011. Procedures for large-scale metabolic profiling of serum and plasma using gas chromatography and liquid chromatography coupled to mass spectrometry. Nat Protoc 6(7): 1060-83.

Gardner, S.G., Marshall, D.D., Daum, R.S., Powers, R., Somerville, G.A. 2018. Metabolic mitigation of Staphylococcus aureus vancomycin intermediate-level susceptibility. Antimicrob Agents Chemother 62(1): e01608-17.

Gil-De-La-Fuente, A., Godzien, J., Saugar, S., Garcia-Carmona, R., Badran, H., Wishart, D.S., Barbas, C., Otero, A. 2019. CEU Mass Mediator 3.0: A Metabolite Annotation Tool. J Proteome Res 18(2): 797-802.

Guijas, C., Montenegro-Burke, J.R., Domingo-Almenara, X., Palermo, A., Warth, B., Hermann, G., Koellensperger, G., Huan, T., Uritboonthai, W., Aisporna, A.E., Wolan, D.W., Spilker, M.E., Benton, H.P., Siuzdak, G. 2018. METLIN: A technology platform for identifying knowns and unknowns. Anal Chem 90(5): 3156-64.

Hattangady, D.S., Singh, A.K., Muthaiyan, A., Jayaswal, R.K., Gustafson, J.E., Ulanov, A.V., Li, Z., Wilkinson, B.J., Pfeltz, R.F. 2015. Genomic, transcriptomic and metabolomic studies of two well-characterized, laboratory-derived vancomycin-intermediate Staphylococcus aureus strains derived from the same parent strain. Antibiotics 4(1): 76-112.

Hiramatsu, K., Aritaka, N., Hanaki, H., Kawasaki, S., Hosoda, Y., Hori, S., Fukuchi, Y., Kobayashi, I. 1997. Dissemination in Japanese hospitals of strains of Staphylococcus aureus heterogeneously resistant to vancomycin. Lancet 350(9092): 1670-3.

Hiramatsu, K., Kayayama, Y., Matsuo, M., Aiba, Y., Saito, M., Hishinuma, T., Iwamoto, A. 2014. Vancomycin-intermediate resistance in Staphylococcus aureus. JGAR 2(4): 213-24.

Kanehisa, M. 2019. Toward understanding the origin and evolution of cellular organisms. Protein Sci 28(11): 1947-51.

Koehl, J.L., Muthaiyan, A., Jayaswal, R.K., Ehlert, K., Labischinski, H., Wilkinson, B.J. 2004. Cell wall composition and decreased autolytic activity and lysostaphin susceptibility of glycopeptide-intermediate Staphylococcus aureus. Antimicrob Agents Chemother 48(10): 3749-57.

Kuroda, M., Kuroda, H., Oshima, T., Takeuchi, F., Mori, H., Hiramatsu, K. 2003. Two-component system VraSR positively modulates the regulation of cell-wall biosynthesis pathway in Staphylococcus aureus. Mol Microbiol 49(3): 807-21.

Liebeke, M., Meyer, H., Donat, S., Ohlsen, K., Lalk, M. 2010. A metabolomic view of Staphylococcus aureus and its ser/thr kinase and phosphatase deletion mutants: Involvement in cell wall biosynthesis. Chem Biol 17(8): 820-30.

Lindgren, J.K., Thomas, V.C., Olson, M.E., Chaudhari, S.S., Nuxoll, A.S., Schaeffer, C.R., Lindgren, K.E., Jones, J., Zimmerman, M.C., Dunman, P.M., Bayles, K.W. Fey, P.D. 2014. Arginine deiminase in Staphylococcus epidermidis functions to augment biofilm maturation through pH homeostasis. J Bacteriol 196(12): 2277-89.

Makhlin, J., Kofman, T., Borovok, I., Kohler, C., Engelmann, S., Cohen, G. Aharonowitz, Y. 2007. Staphylococcus aureus ArcR controls expression of the arginine deiminase operon. J Bacteriol 189(16): 5976-86.

McCallum, N., Karauzum, H., Getzmann, R., Bischoff, M., Majcherczyk, P., Berger-Bächi, B., Landmann, R. 2006. In vivo survival of teicoplanin-resistant Staphylococcus aureus and fitness cost of teicoplanin resistance. Antimicrob Agents Chemother 50(7): 2352-60.

McGuinness, W.A., Malachowa, N., DeLeo, F.R. 2017. Vancomycin resistance in Staphylococcus aureus. YJBM 90(2): 269-81.

Neoh, H.M., Cui, L., Yuzawa, H., Takeuchi, F., Matsuo, M., Hiramatsu, K. 2008 Mutated response regulator graR is responsible for phenotypic conversion of Staphylococcus aureus from heterogeneous vancomycin-intermediate resistance to vancomycin-intermediate resistance. Antimicrob Agents Chemother 52(1): 45-53.

Sumner, L.W., Amberg, A., Barrett, D., Beale, M.H., Beger, R., Daykin, C.A., Fan, T.W.M., Fiehn, O., Goodacre, R., Griffin, J.L., Hankemeier, T., Hardy, N., Harnly, J., Higashi, R., Kopka, J., Lane, A.N., Lindon, J.C., Marriott, P., Nicholls, A.W., Reily, M.D., Thaden, J.J., Viant, M.R. 2007. Proposed minimum reporting standards for chemical analysis. Metabolomics 3(3): 211-21.

Tan, J.K., Zakaria, S.N.A., Gunasekaran, G., Abdul Sani, N.F., Nasaruddin, M.L., Jaafar, F., Abu Bakar, Z.H., Amir Hamzah, A.I.Z., Nor Aripin, K.N., Mohd Rani, M.D., Noh, N. A., Damanhuri, H.A., Mazlan, M., Makpol, S., Wan Ngah, W.Z. 2023. Metabolomics profiling of age-associated metabolites in Malay population. Oxid Med Cell Longev 2023: 4416410.

Tan, X.E., Neoh, H.M., Looi, M.L., Chin, S.F., Cui, L., Hiramatsu, K., Hussin, S., Jamal, R. 2017. Activated ADI pathway: The initiator of intermediate vancomycin resistance in Staphylococcus aureus. Can J Microbiol 63(3): 260–264.

Tan, X.E., Neoh, H.M., Looi, M.L., Tan, T.L., Hussin, S., Cui, L., Hiramatsu, K., Jamal, R. 2016. Comparative proteomics profiling reveals down-regulation of Staphylococcus aureus virulence in achieving intermediate vancomycin resistance. Malays J Microbiol 12(6): 498-505.

Weigel, 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., Tenover, F.C. 2003. Genetic analysis of a high-level vancomycin-resistant isolate of Staphylococcus aureus. Science 302(5650): 1569-71.

Wishart, D.S., Feunang, Y.D., Marcu, A., Guo, A.C., Liang, K., Vázquez-Fresno, R., Sajed, T., Johnson, D., Li, C., Karu, N., Sayeeda, Z., Lo, E., Assempour, N., Berjanskii, M., Singhal, S., Arndt, D., Liang, Y., Badran, H., Grant, J., Serra-Cayuela, A., Liu, Y., Mandal, R., Neveu, V., Pon, A., Knox, C., Wilson, M., Manach, C., Scalbert, A. 2018. HMDB 4.0: The human metabolome database for 2018. Nucleic Acids Res 46(D1): D608-17.
Correspondance Address

Alexander, E.L., Gardete, S., Bar, H.Y., Wells, M.T., Tomasz, A., Rhee, K.Y. 2014. Intermediate-type vancomycin resistance (VISA) in genetically-distinct Staphylococcus aureus isolates is linked to specific, reversible metabolic alterations. PLoS ONE 9(5): e97137.

Chong, J., Soufan, O., Li, C., Caraus, I., Li, S., Bourque, G., Wishart, D.S., Xia, J. 2018. MetaboAnalyst 4.0: Towards more transparent and integrative metabolomics analysis. Nucleic Acids Res 46(W1): W486-94.

Cui, L., Lian, J.Q., Neoh, H.M., Reyes, E., Hiramatsu, K. 2005. DNA microarray-based identification of genes associated with glycopeptide resistance in Staphylococcus aureus. Antimicrob Agents Chemother 49(8): 3404-13.

Cui, L., Murakami, H., Kuwahara-Arai, K., Hanaki, H., Hiramatsu, K. 2000. Contribution of a thickened cell wall and its glutamine nonamidated component to the vancomycin resistance expressed by Staphylococcus aureus Mu50. Antimicrob Agents Chemother 44(9): 2276-85.

Cui, L., Neoh, H.M., Shoji, M., Hiramatsu, K. 2009. Contribution of vraSR and graSR point mutations to vancomycin resistance in vancomycin-intermediate Staphylococcus aureus. Antimicrob Agents Chemother 53(3): 1231-4.

Dunn, W.B., Broadhurst, D., Begley, P., Zelena, E., Francis-Mcintyre, S., Anderson, N., Brown, M., Knowles, J.D., Halsall, A., Haselden, J. N., Nicholls, A.W., Wilson, I.D., Kell, D.B., Goodacre, R. 2011. Procedures for large-scale metabolic profiling of serum and plasma using gas chromatography and liquid chromatography coupled to mass spectrometry. Nat Protoc 6(7): 1060-83.

Gardner, S.G., Marshall, D.D., Daum, R.S., Powers, R., Somerville, G.A. 2018. Metabolic mitigation of Staphylococcus aureus vancomycin intermediate-level susceptibility. Antimicrob Agents Chemother 62(1): e01608-17.

Gil-De-La-Fuente, A., Godzien, J., Saugar, S., Garcia-Carmona, R., Badran, H., Wishart, D.S., Barbas, C., Otero, A. 2019. CEU Mass Mediator 3.0: A Metabolite Annotation Tool. J Proteome Res 18(2): 797-802.

Guijas, C., Montenegro-Burke, J.R., Domingo-Almenara, X., Palermo, A., Warth, B., Hermann, G., Koellensperger, G., Huan, T., Uritboonthai, W., Aisporna, A.E., Wolan, D.W., Spilker, M.E., Benton, H.P., Siuzdak, G. 2018. METLIN: A technology platform for identifying knowns and unknowns. Anal Chem 90(5): 3156-64.

Hattangady, D.S., Singh, A.K., Muthaiyan, A., Jayaswal, R.K., Gustafson, J.E., Ulanov, A.V., Li, Z., Wilkinson, B.J., Pfeltz, R.F. 2015. Genomic, transcriptomic and metabolomic studies of two well-characterized, laboratory-derived vancomycin-intermediate Staphylococcus aureus strains derived from the same parent strain. Antibiotics 4(1): 76-112.

Hiramatsu, K., Aritaka, N., Hanaki, H., Kawasaki, S., Hosoda, Y., Hori, S., Fukuchi, Y., Kobayashi, I. 1997. Dissemination in Japanese hospitals of strains of Staphylococcus aureus heterogeneously resistant to vancomycin. Lancet 350(9092): 1670-3.

Hiramatsu, K., Kayayama, Y., Matsuo, M., Aiba, Y., Saito, M., Hishinuma, T., Iwamoto, A. 2014. Vancomycin-intermediate resistance in Staphylococcus aureus. JGAR 2(4): 213-24.

Kanehisa, M. 2019. Toward understanding the origin and evolution of cellular organisms. Protein Sci 28(11): 1947-51.

Koehl, J.L., Muthaiyan, A., Jayaswal, R.K., Ehlert, K., Labischinski, H., Wilkinson, B.J. 2004. Cell wall composition and decreased autolytic activity and lysostaphin susceptibility of glycopeptide-intermediate Staphylococcus aureus. Antimicrob Agents Chemother 48(10): 3749-57.

Kuroda, M., Kuroda, H., Oshima, T., Takeuchi, F., Mori, H., Hiramatsu, K. 2003. Two-component system VraSR positively modulates the regulation of cell-wall biosynthesis pathway in Staphylococcus aureus. Mol Microbiol 49(3): 807-21.

Liebeke, M., Meyer, H., Donat, S., Ohlsen, K., Lalk, M. 2010. A metabolomic view of Staphylococcus aureus and its ser/thr kinase and phosphatase deletion mutants: Involvement in cell wall biosynthesis. Chem Biol 17(8): 820-30.

Lindgren, J.K., Thomas, V.C., Olson, M.E., Chaudhari, S.S., Nuxoll, A.S., Schaeffer, C.R., Lindgren, K.E., Jones, J., Zimmerman, M.C., Dunman, P.M., Bayles, K.W. Fey, P.D. 2014. Arginine deiminase in Staphylococcus epidermidis functions to augment biofilm maturation through pH homeostasis. J Bacteriol 196(12): 2277-89.

Makhlin, J., Kofman, T., Borovok, I., Kohler, C., Engelmann, S., Cohen, G. Aharonowitz, Y. 2007. Staphylococcus aureus ArcR controls expression of the arginine deiminase operon. J Bacteriol 189(16): 5976-86.

McCallum, N., Karauzum, H., Getzmann, R., Bischoff, M., Majcherczyk, P., Berger-Bächi, B., Landmann, R. 2006. In vivo survival of teicoplanin-resistant Staphylococcus aureus and fitness cost of teicoplanin resistance. Antimicrob Agents Chemother 50(7): 2352-60.

McGuinness, W.A., Malachowa, N., DeLeo, F.R. 2017. Vancomycin resistance in Staphylococcus aureus. YJBM 90(2): 269-81.

Neoh, H.M., Cui, L., Yuzawa, H., Takeuchi, F., Matsuo, M., Hiramatsu, K. 2008 Mutated response regulator graR is responsible for phenotypic conversion of Staphylococcus aureus from heterogeneous vancomycin-intermediate resistance to vancomycin-intermediate resistance. Antimicrob Agents Chemother 52(1): 45-53.

Sumner, L.W., Amberg, A., Barrett, D., Beale, M.H., Beger, R., Daykin, C.A., Fan, T.W.M., Fiehn, O., Goodacre, R., Griffin, J.L., Hankemeier, T., Hardy, N., Harnly, J., Higashi, R., Kopka, J., Lane, A.N., Lindon, J.C., Marriott, P., Nicholls, A.W., Reily, M.D., Thaden, J.J., Viant, M.R. 2007. Proposed minimum reporting standards for chemical analysis. Metabolomics 3(3): 211-21.

Tan, J.K., Zakaria, S.N.A., Gunasekaran, G., Abdul Sani, N.F., Nasaruddin, M.L., Jaafar, F., Abu Bakar, Z.H., Amir Hamzah, A.I.Z., Nor Aripin, K.N., Mohd Rani, M.D., Noh, N. A., Damanhuri, H.A., Mazlan, M., Makpol, S., Wan Ngah, W.Z. 2023. Metabolomics profiling of age-associated metabolites in Malay population. Oxid Med Cell Longev 2023: 4416410.

Tan, X.E., Neoh, H.M., Looi, M.L., Chin, S.F., Cui, L., Hiramatsu, K., Hussin, S., Jamal, R. 2017. Activated ADI pathway: The initiator of intermediate vancomycin resistance in Staphylococcus aureus. Can J Microbiol 63(3): 260–264.

Tan, X.E., Neoh, H.M., Looi, M.L., Tan, T.L., Hussin, S., Cui, L., Hiramatsu, K., Jamal, R. 2016. Comparative proteomics profiling reveals down-regulation of Staphylococcus aureus virulence in achieving intermediate vancomycin resistance. Malays J Microbiol 12(6): 498-505.

Weigel, 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., Tenover, F.C. 2003. Genetic analysis of a high-level vancomycin-resistant isolate of Staphylococcus aureus. Science 302(5650): 1569-71.

Wishart, D.S., Feunang, Y.D., Marcu, A., Guo, A.C., Liang, K., Vázquez-Fresno, R., Sajed, T., Johnson, D., Li, C., Karu, N., Sayeeda, Z., Lo, E., Assempour, N., Berjanskii, M., Singhal, S., Arndt, D., Liang, Y., Badran, H., Grant, J., Serra-Cayuela, A., Liu, Y., Mandal, R., Neveu, V., Pon, A., Knox, C., Wilson, M., Manach, C., Scalbert, A. 2018. HMDB 4.0: The human metabolome database for 2018. Nucleic Acids Res 46(D1): D608-17.



Alexander, E.L., Gardete, S., Bar, H.Y., Wells, M.T., Tomasz, A., Rhee, K.Y. 2014. Intermediate-type vancomycin resistance (VISA) in genetically-distinct Staphylococcus aureus isolates is linked to specific, reversible metabolic alterations. PLoS ONE 9(5): e97137.

Chong, J., Soufan, O., Li, C., Caraus, I., Li, S., Bourque, G., Wishart, D.S., Xia, J. 2018. MetaboAnalyst 4.0: Towards more transparent and integrative metabolomics analysis. Nucleic Acids Res 46(W1): W486-94.

Cui, L., Lian, J.Q., Neoh, H.M., Reyes, E., Hiramatsu, K. 2005. DNA microarray-based identification of genes associated with glycopeptide resistance in Staphylococcus aureus. Antimicrob Agents Chemother 49(8): 3404-13.

Cui, L., Murakami, H., Kuwahara-Arai, K., Hanaki, H., Hiramatsu, K. 2000. Contribution of a thickened cell wall and its glutamine nonamidated component to the vancomycin resistance expressed by Staphylococcus aureus Mu50. Antimicrob Agents Chemother 44(9): 2276-85.

Cui, L., Neoh, H.M., Shoji, M., Hiramatsu, K. 2009. Contribution of vraSR and graSR point mutations to vancomycin resistance in vancomycin-intermediate Staphylococcus aureus. Antimicrob Agents Chemother 53(3): 1231-4.

Dunn, W.B., Broadhurst, D., Begley, P., Zelena, E., Francis-Mcintyre, S., Anderson, N., Brown, M., Knowles, J.D., Halsall, A., Haselden, J. N., Nicholls, A.W., Wilson, I.D., Kell, D.B., Goodacre, R. 2011. Procedures for large-scale metabolic profiling of serum and plasma using gas chromatography and liquid chromatography coupled to mass spectrometry. Nat Protoc 6(7): 1060-83.

Gardner, S.G., Marshall, D.D., Daum, R.S., Powers, R., Somerville, G.A. 2018. Metabolic mitigation of Staphylococcus aureus vancomycin intermediate-level susceptibility. Antimicrob Agents Chemother 62(1): e01608-17.

Gil-De-La-Fuente, A., Godzien, J., Saugar, S., Garcia-Carmona, R., Badran, H., Wishart, D.S., Barbas, C., Otero, A. 2019. CEU Mass Mediator 3.0: A Metabolite Annotation Tool. J Proteome Res 18(2): 797-802.

Guijas, C., Montenegro-Burke, J.R., Domingo-Almenara, X., Palermo, A., Warth, B., Hermann, G., Koellensperger, G., Huan, T., Uritboonthai, W., Aisporna, A.E., Wolan, D.W., Spilker, M.E., Benton, H.P., Siuzdak, G. 2018. METLIN: A technology platform for identifying knowns and unknowns. Anal Chem 90(5): 3156-64.

Hattangady, D.S., Singh, A.K., Muthaiyan, A., Jayaswal, R.K., Gustafson, J.E., Ulanov, A.V., Li, Z., Wilkinson, B.J., Pfeltz, R.F. 2015. Genomic, transcriptomic and metabolomic studies of two well-characterized, laboratory-derived vancomycin-intermediate Staphylococcus aureus strains derived from the same parent strain. Antibiotics 4(1): 76-112.

Hiramatsu, K., Aritaka, N., Hanaki, H., Kawasaki, S., Hosoda, Y., Hori, S., Fukuchi, Y., Kobayashi, I. 1997. Dissemination in Japanese hospitals of strains of Staphylococcus aureus heterogeneously resistant to vancomycin. Lancet 350(9092): 1670-3.

Hiramatsu, K., Kayayama, Y., Matsuo, M., Aiba, Y., Saito, M., Hishinuma, T., Iwamoto, A. 2014. Vancomycin-intermediate resistance in Staphylococcus aureus. JGAR 2(4): 213-24.

Kanehisa, M. 2019. Toward understanding the origin and evolution of cellular organisms. Protein Sci 28(11): 1947-51.

Koehl, J.L., Muthaiyan, A., Jayaswal, R.K., Ehlert, K., Labischinski, H., Wilkinson, B.J. 2004. Cell wall composition and decreased autolytic activity and lysostaphin susceptibility of glycopeptide-intermediate Staphylococcus aureus. Antimicrob Agents Chemother 48(10): 3749-57.

Kuroda, M., Kuroda, H., Oshima, T., Takeuchi, F., Mori, H., Hiramatsu, K. 2003. Two-component system VraSR positively modulates the regulation of cell-wall biosynthesis pathway in Staphylococcus aureus. Mol Microbiol 49(3): 807-21.

Liebeke, M., Meyer, H., Donat, S., Ohlsen, K., Lalk, M. 2010. A metabolomic view of Staphylococcus aureus and its ser/thr kinase and phosphatase deletion mutants: Involvement in cell wall biosynthesis. Chem Biol 17(8): 820-30.

Lindgren, J.K., Thomas, V.C., Olson, M.E., Chaudhari, S.S., Nuxoll, A.S., Schaeffer, C.R., Lindgren, K.E., Jones, J., Zimmerman, M.C., Dunman, P.M., Bayles, K.W. Fey, P.D. 2014. Arginine deiminase in Staphylococcus epidermidis functions to augment biofilm maturation through pH homeostasis. J Bacteriol 196(12): 2277-89.

Makhlin, J., Kofman, T., Borovok, I., Kohler, C., Engelmann, S., Cohen, G. Aharonowitz, Y. 2007. Staphylococcus aureus ArcR controls expression of the arginine deiminase operon. J Bacteriol 189(16): 5976-86.

McCallum, N., Karauzum, H., Getzmann, R., Bischoff, M., Majcherczyk, P., Berger-Bächi, B., Landmann, R. 2006. In vivo survival of teicoplanin-resistant Staphylococcus aureus and fitness cost of teicoplanin resistance. Antimicrob Agents Chemother 50(7): 2352-60.

McGuinness, W.A., Malachowa, N., DeLeo, F.R. 2017. Vancomycin resistance in Staphylococcus aureus. YJBM 90(2): 269-81.

Neoh, H.M., Cui, L., Yuzawa, H., Takeuchi, F., Matsuo, M., Hiramatsu, K. 2008 Mutated response regulator graR is responsible for phenotypic conversion of Staphylococcus aureus from heterogeneous vancomycin-intermediate resistance to vancomycin-intermediate resistance. Antimicrob Agents Chemother 52(1): 45-53.

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