Letter | Published:

Staphylococcus aureus inactivates daptomycin by releasing membrane phospholipids

Nature Microbiology volume 2, Article number: 16194 (2016) | Download Citation


Daptomycin is a bactericidal antibiotic of last resort for serious infections caused by methicillin-resistant Staphylococcus aureus (MRSA)1,2. Although resistance is rare, treatment failure can occur in more than 20% of cases3,4 and so there is a pressing need to identify and mitigate factors that contribute to poor therapeutic outcomes. Here, we show that loss of the Agr quorum-sensing system, which frequently occurs in clinical isolates, enhances S. aureus survival during daptomycin treatment. Wild-type S. aureus was killed rapidly by daptomycin, but Agr-defective mutants survived antibiotic exposure by releasing membrane phospholipids, which bound and inactivated the antibiotic. Although wild-type bacteria also released phospholipid in response to daptomycin, Agr-triggered secretion of small cytolytic toxins, known as phenol soluble modulins, prevented antibiotic inactivation. Phospholipid shedding by S. aureus occurred via an active process and was inhibited by the β-lactam antibiotic oxacillin, which slowed inactivation of daptomycin and enhanced bacterial killing. In conclusion, S. aureus possesses a transient defence mechanism that protects against daptomycin, which can be compromised by Agr-triggered toxin production or an existing therapeutic antibiotic.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.


  1. 1.

    , , & Daptomycin: a lipopeptide antibiotic for the treatment of serious Gram-positive infections. J. Antimicrob. Chemother. 55, 283–288 (2005).

  2. 2.

    , & Serious infections caused by methicillin-resistant Staphylococcus aureus. Clin. Infect. Dis. 51, S183–S197 (2010).

  3. 3.

    et al. Insights and clinical perspectives of daptomycin resistance in Staphylococcus aureus: a review of the available evidence. Int. J. Antimicrob. Agents. 46, 278–289 (2015).

  4. 4.

    et al. Evaluation of effectiveness and safety of high-dose daptomycin: results from patients included in the European Cubicin® outcomes registry and experience. Adv. Ther. 12, 1192–1205 (2015).

  5. 5.

    et al. Synthesis of staphylococcal virulence factors is controlled by a regulatory RNA molecule. EMBO J. 12, 3967–3975 (1993).

  6. 6.

    & Quorum sensing in Staphylococci. Annu. Rev. Genet. 42, 541–564 (2008).

  7. 7.

    et al. Accessory gene regulator (agr) locus in geographically diverse Staphylococcus aureus isolates with reduced susceptibility to vancomycin. Antimicrob. Agents Chemother. 46, 1492–1502 (2002).

  8. 8.

    et al. Persistent bacteremia due to methicillin-resistant Staphylococcus aureus infection is associated with agr dysfunction and low-level in vitro resistance to thrombin-induced platelet microbicidal protein. J. Infect. Dis. 190, 1140–1149 (2004).

  9. 9.

    et al. Tracking the in vivo evolution of multidrug resistance in Staphylococcus aureus by whole-genome sequencing. Proc. Natl Acad. Sci. USA 104, 9451–9456 (2007).

  10. 10.

    et al. agr function in clinical Staphylococcus aureus isolates. Microbiology 154, 2265–2274 (2008).

  11. 11.

    et al. Evolutionary trade-offs underlie the multi-faceted virulence of Staphylococcus aureus. PLoS Biol. 13, e1002229 (2015).

  12. 12.

    et al. Increased mortality with accessory gene regulator (agr) dysfunction in Staphylococcus aureus among bacteremic patients. Antimicrob. Agents Chemother. 55, 1082–1087 (2011).

  13. 13.

    et al. Antibiotic-mediated selection of quorum-sensing-negative Staphylococcus aureus. mBio. 3, e00459 (2013).

  14. 14.

    , , & What role does the quorum-sensing accessory gene regulator system play during Staphylococcus aureus bacteremia? Trends Microbiol. 22, 676–685 (2014).

  15. 15.

    , & Activity of daptomycin alone and in combination with rifampin and gentamicin against Staphylococcus aureus assessed by time–kill methodology. Antimicrob. Agents Chemother. 51, 1504–1507 (2007).

  16. 16.

    et al. Daptomycin is effective in treatment of experimental endocarditis due to methicillin-resistant and glycopeptide-intermediate Staphylococcus aureus. Antimicrob. Agents Chemother. 52, 2538–2543 (2008).

  17. 17.

    & Daptomycin-nonsusceptible Staphylococcus aureus: the role of combination therapy with daptomycin and gentamicin. Genes 6, 1256–1267 (2015).

  18. 18.

    et al. Abscess formation and alpha-hemolysin induced toxicity in a mouse model of Staphylococcus aureus peritoneal infection. Infect. Immun. 80, 3721–3732 (2012).

  19. 19.

    et al. Rapid bactericidal activity of daptomycin against methicillin-resistant and methicillin-susceptible Staphylococcus aureus peritonitis in mice as measured with bioluminescent bacteria. Antimicrob. Agents Chemother. 51, 1787–1794 (2007).

  20. 20.

    , & Scanning electron microscopy of Staphylococcus aureus and Enterococcus faecalis exposed to daptomycin. J. Med. Microbiol. 30, 45–49 (1989).

  21. 21.

    , , , & Daptomycin exerts bactericidal activity without lysis of Staphylococcus aureus. Antimicrob. Agents Chemother. 52, 2223–2225 (2008).

  22. 22.

    , , & Oligomerization of daptomycin on membranes. Biochim. Biophys. Acta. 1808, 1154–1160 (2011).

  23. 23.

    , , & Interaction of daptomycin with lipid bilayers: a lipid extracting effect. Biochemistry 53, 5384–5392 (2014).

  24. 24.

    & Correlation of cell membrane lipid profiles with daptomycin resistance in methicillin-resistant Staphylococcus aureus. Antimicrob. Agents Chemother. 57, 1082–1085 (2013).

  25. 25.

    , , , & Inhibition of daptomycin by pulmonary surfactant: in vitro modeling and clinical impact. J. Infect. Dis. 191, 2149–2152 (2005).

  26. 26.

    & Contribution of bacterial outer membrane vesicles to innate bacterial defense. BMC. Microbiol. 11, 258 (2011).

  27. 27.

    et al. Identification of novel cytolytic peptides as key virulence determinants for community-associated MRSA. Nat. Med. 13, 1510–1514 (2007).

  28. 28.

    et al. RNAIII-independent target gene control by the agr quorum-sensing system: insight into the evolution of virulence regulation in Staphylococcus aureus. Mol. Cell 32, 150–158 (2008).

  29. 29.

    & Synergy of daptomycin with oxacillin and other β-lactams against methicillin-resistant Staphylococcus aureus. Antimicrob. Agents Chemother. 48, 2871–2875 (2004).

  30. 30.

    et al. Penicillin binding protein 1 is important in the compensatory response of Staphylococcus aureus to daptomycin-induced membrane damage and is a potential target for β-lactam–daptomycin synergy. Antimicrob. Agents Chemother. 60, 451–458 (2015).

  31. 31.

    , , , & The Agr quorum-sensing system regulates fibronectin binding but not hemolysis in the absence of a functional electron transport chain. Infect. Immun. 82, 4337–4347 (2014).

  32. 32.

    , , , & Transforming the untransformable: application of direct transformation to manipulate genetically Staphylococcus aureus and Staphylococcus epidermidis. mBio. 3, e00277 (2012).

  33. 33.

    et al. Systematic mutational analysis of the LytTR DNA binding domain of Staphylococcus aureus virulence gene transcription factor AgrA. Nucleic Acids Res. 42, 12523–12536 (2014).

  34. 34.

    , & Transcriptional downregulation of agr expression in Staphylococcus aureus during growth in human serum can be overcome by constitutively active mutant forms of the sensor kinase AgrC. FEMS Microbiol. Lett. 349, 153–162 (2013).

  35. 35.

    et al. in Manual of Clinical Microbiology Ch. 118, 1526–1543 (ASM Press, 1999).

  36. 36.

    , & Agar and broth dilution methods to determine the minimal inhibitory concentration (MIC) of antimicrobial substances. Nat. Protoc. 3, 163–175 (2008).

  37. 37.

    , , & In vitro activity of daptomycin against methicillin-resistant Staphylococcus aureus, including heterogeneously glycopeptide-resistant strains. Antimicrob. Agents Chemother. 50, 3189–3191 (2006).

  38. 38.

    , & Daptomycin bactericidal activity and correlation between disk and broth microdilution method results in testing of Staphylococcus aureus strains with decreased susceptibility to vancomycin. Antimicrob. Agents Chemother. 50, 2330–2336 (2006).

  39. 39.

    et al. Bactericidal activity of gentamicin against Enterococcus faecalis in vitro and in vivo. Antimicrob. Agents Chemother. 44, 2077–2080 (2000).

  40. 40.

    et al. Pharmacodynamics of daptomycin in a murine thigh model of Staphylococcus aureus infection. Antimicrob. Agents Chemother. 45, 845–851 (2001).

  41. 41.

    , & In vivo pharmacodynamic activity of daptomycin. Antimicrob. Agents Chemother. 48, 63–68 (2004).

  42. 42.

    The pharmacokinetic and pharmacodynamic properties of vancomycin. Clin. Infect. Dis. 42, S35–S39 (2006).

  43. 43.

    et al. Lack of bactericidal antagonism or synergism in vitro between oxacillin and vancomycin against methicillin-susceptible strains of Staphylococcus aureus. Antimicrob. Agents Chemother. 54, 773–777 (2010).

  44. 44.

    & Pharmacodynamics, population dynamics, and the evolution of persistence in Staphylococcus aureus. PLoS Genet. 9, e1003123 (2013).

  45. 45.

    , , & G*Power 3: a flexible statistical power analysis program for the social, behavioral, and biomedical sciences. Behav. Res. Methods 39, 175–191 (2007).

  46. 46.

    et al. Bacterial division. Mechanical crack propagation drives millisecond daughter cell separation in Staphylococcus aureus. Science 348, 574–578 (2015).

  47. 47.

    & A rapid method of total lipid extraction and purification. Can. J. Biochem. Physiol. 37, 911–917 (1959).

  48. 48.

    , , & Polar lipid and isoprenoid quinone composition in the classification of Staphylococcus. J. Gen. Microbiol. 130, 2427–2437 (1984).

  49. 49.

    et al. In vitro susceptibility of Staphylococcus aureus to thrombin-induced platelet microbicidal protein-1 (tPMP-1) is influenced by cell membrane phospholipid composition and asymmetry. Microbiology 153, 1187–1197 (2007).

  50. 50.

    , & Quantitative analysis of phospholipids by thin-layer chromatography and phosphorus analysis of spots. Lipids 1, 85–86 (1966).

Download references


The authors thank the following for providing bacterial strains, phage or reagents: J.M. Van Dijl (University Medical Center Groningen), M. Otto (NIH, Bethesda), R. Massey (University of Bath), M. Horsburgh (University of Liverpool), T. Foster (Trinity College Dublin) and the Network on Antimicrobial Resistance in Staphylococcus aureus (NARSA) Program under NIAID/NIH contract no. HHSN272200700055C. L. Haigh (Imperial College) is thanked for analysing modified daptomycin samples. A. Nobbs (University of Bristol) is acknowledged for helpful discussions and comments on the manuscript. A.M.E. acknowledges funding from the Department of Medicine, Imperial College. S.W. acknowledges funding from the BBSRC and Wellcome Trust. K.L.P. is supported by a PhD studentship from the Faculty of Medicine, Imperial College London. S.H. is supported by a scholarship from the Inlaks Shivdasani Foundation. T.B.C. is a Sir Henry Dale Fellow jointly funded by the Wellcome Trust and Royal Society (grant no. 107660/Z/15/Z).

Author information


  1. MRC Centre for Molecular Bacteriology and Infection, Imperial College London, Armstrong Road, London SW7 2AZ, UK

    • Vera Pader
    • , Sanika Hakim
    • , Kimberley L. Painter
    • , Sivaramesh Wigneshweraraj
    • , Thomas B. Clarke
    •  & Andrew M. Edwards


  1. Search for Vera Pader in:

  2. Search for Sanika Hakim in:

  3. Search for Kimberley L. Painter in:

  4. Search for Sivaramesh Wigneshweraraj in:

  5. Search for Thomas B. Clarke in:

  6. Search for Andrew M. Edwards in:


V.P., S.W., T.B.C. and A.M.E. designed the experiments. V.P., S.H., T.B.C. and A.M.E. performed experiments. K.L.P. generated and characterized mutants. V.P., S.H., T.B.C. and A.M.E. analysed data. All authors contributed to the writing of the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Andrew M. Edwards.

Supplementary information

PDF files

  1. 1.

    Supplementary information

    Supplementary Tables 1–4, Supplementary Figures 1–22, Supplementary Discussion and Supplementary References

About this article

Publication history






Further reading