Skip to main content
SpringerLink
Log in

Single-Step Capture and Targeted Metabolomics of Alkyl-Quinolones in Outer Membrane Vesicles of Pseudomonas aeruginosa

  • Protocol
  • First Online:
Book cover Lipidomics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1609))

Abstract

Outer membrane vesicles (OMVs) are secreted by all Gram-ve pathogens. These nano-scale delivery vehicles contain discrete arrays of prokaryotic pathogenic determinants, including a family of low molecular weight (MW) lipidic quorum signaling alkyl-quinolones (AQs). These are synthesized from β-keto-fatty acids and function like primordial lipidic hormones, which regulate numerous pathogenic factors both inter-species and intra-species. Significantly, AQs can also directly exacerbate pathogenesis by cross-kingdom signaling with the host immune, metabolic, and other systems. In Pseudomonas aeruginosa more than 50 AQs are reported; many with pathogenic mechanisms that are largely unknown. Some of these AQs are exclusively associated with OMVs. Accurate characterization of these OMV-AQs may reveal novel mechanism of diseases and Pseudomonas aeruginosa presents an ideal model. Matrix-free laser desorption/ionization mass spectrometry (LDI-MS) technologies enjoy unique advantages in mass spectrometry (MS)-based imaging and low MW analysis. We report single-step isolation of Pseudomonas aeruginosa OMV on inert ceramic filters and high-resolution mass spectrometry (HRMS) analysis of AQs vesicle in situ.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 119.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 159.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 159.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Kulp A, Kuehn MJ (2010) Biological functions and biogenesis of secreted bacterial outer membrane vesicles. Ann Rev Microbiol 64:163–184. doi:10.1146/annurev.micro.091208.073413

  2. Schwechheimer C, Kuehn MJ (2015a) Outer-membrane vesicles from gram-negative bacteria: biogenesis and functions. Nat Rev Microbiol 13(10):605–619. doi:10.1038/nrmicro3525

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Mashburn LM, Whiteley M (2005) Membrane vesicles traffic signals and facilitate group activities in a prokaryote. Nature 437(7057):422–425. doi:10.1038/nature03925

    Article  CAS  PubMed  Google Scholar 

  4. Schwechheimer C, Kuehn MJ (2015b) Outer-membrane vesicles from gram-negative bacteria: biogenesis and functions. Nat Rev Micro 13(10):605–619. doi:10.1038/nrmicro3525

    Article  CAS  Google Scholar 

  5. Wessel AK, Liew J, Kwon T, Marcotte EM, Whiteley M (2013) Role of Pseudomonas aeruginosa peptidoglycan-associated outer membrane proteins in vesicle formation. J Bacteriol 195(2):213–219. doi:10.1128/JB.01253-12

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Vella BD, Schertzer JW (2015) Understanding and exploiting bacterial outer membrane vesicles. In: Ramos J-L, Goldberg JB, Filloux A (eds) Pseudomonas, New aspects of pseudomonas biology, vol 7. Springer, Dordrecht, pp 217–250. doi:10.1007/978-94-017-9555- 5_9

    Google Scholar 

  7. Roier S, Zingl FG, Cakar F, Durakovic S, Kohl P, Eichmann TO, Klug L, Gadermaier B, Weinzerl K, Prassl R, Lass A, Daum G, Reidl J, Feldman MF, Schild S (2016) A novel mechanism for the biogenesis of outer membrane vesicles in gram-negative bacteria. Nat Comm 7:10515. doi:10.1038/ncomms10515

  8. Chatterjee SN, Chaudhuri K (2012) Gram-negative bacteria: the cell membranes. In: Outer membrane vesicles of bacteria. Springer, Berlin, Heidelberg, pp 15–34. doi:10.1007/978-3-642-30526-9_2

    Chapter  Google Scholar 

  9. Choi CW, Park EC, Yun SH, Lee SY, Lee YG, Hong Y, Park KR, Kim SH, Kim GH, Kim SI (2014) Proteomic characterization of the outer membrane vesicle of pseudomonas putida KT2440. J Proteome Res 13(10):4298–4309. doi:10.1021/pr500411d

  10. Tashiro Y, Sakai R, Toyofuku M, Sawada I, Nakajima-Kambe T, Uchiyama H, Nomura N (2009) Outer membrane machinery and alginate synthesis regulators control membrane vesicle production in Pseudomonas aeruginosa. J Bacteriol 191(24):7509–7519. doi:10.1128/JB.00722-09

  11. Bomberger JM, Maceachran DP, Coutermarsh BA, Ye S, O'Toole GA, Stanton BA (2009a) Long-distance delivery of bacterial virulence factors by Pseudomonas aeruginosa outer membrane vesicles. PLoS Pathog 5(4):e1000382. doi:10.1371/journal.ppat.1000382

  12. Wispelwey B, Hansen EJ, Scheld WM (1989) Haemophilus influenzae outer membrane vesicle-induced blood-brain barrier permeability during experimental meningitis. Infect Immun 57(8):2559–2562

    CAS  PubMed  PubMed Central  Google Scholar 

  13. Bomberger JM, MacEachran DP, Coutermarsh BA, Ye SY, O'Toole GA, Stanton BA (2009b) Long-distance delivery of bacterial virulence factors by Pseudomonas aeruginosa outer membrane vesicles. PLoS Pathog 5(4):e1000382. doi:10.1371/journal.ppat.1000382

  14. Kulkarni HM, Jagannadham MV (2014) Biogenesis and multifaceted roles of outer membrane vesicles from gram-negative bacteria. Microbiology 160(Pt 10):2109–2121. doi:10.1099/mic.0.079400-0

    Article  CAS  PubMed  Google Scholar 

  15. Kuehn MJ, Kesty NC (2005) Bacterial outer membrane vesicles and the host-pathogen interaction. Genes Dev 19(22):2645–2655. doi:10.1101/gad.1299905

    Article  CAS  PubMed  Google Scholar 

  16. Schertzer JW, Whiteley M (2013) Bacterial outer membrane vesicles in trafficking, communication and the host-pathogen interaction. J Mol Microbiol Biotechnol 23(1–2):118–130. doi:10.1159/000346770

    Article  CAS  PubMed  Google Scholar 

  17. Dean CR, Franklund CV, Retief JD, Coyne MJ Jr, Hatano K, Evans DJ, Pier GB, Goldberg JB (1999) Characterization of the serogroup O11 O-antigen locus of Pseudomonas aeruginosa PA103. J Bacteriol 181(14):4275–4284

    Google Scholar 

  18. Dubern JF, Diggle SP (2008) Quorum sensing by 2-alkyl-4-quinolones in Pseudomonas aeruginosa and other bacterial species. Mol Biosyst 4(9):882–888. doi:10.1039/b803796p

  19. Camara M, Williams P, Barrett D, Halliday N, Knox A, Smyth A, Fogarty A, Barr H, Forrester D (2016) Alkyl quinolones as biomarkers of pseudomonas aeruginosa infection and uses thereof. US Patent 20,160,131,648

    Google Scholar 

  20. Lepine F, Milot S, Deziel E, He J, Rahme LG (2004) Electrospray/mass spectrometric identification and analysis of 4-hydroxy-2-alkylquinolines (HAQs) produced by Pseudomonas aeruginosa. J Am Soc Mass Spectrom 15(6):862–869. doi:10.1016/j.jasms.2004.02.012

  21. Inaba T, Oura H, Morinaga K, Toyofuku M, Nomura N (2015) The pseudomonas quinolone signal inhibits biofilm development of Streptococcus mutans. Microbes Environ 30(2):189–191. doi:10.1264/jsme2.ME14140

  22. Liu YC, Chan KG, Chang CY (2015) Modulation of host biology by Pseudomonas aeruginosa quorum sensing signal molecules: messengers or traitors. Front Microbiol 6:1226. doi:10.3389/fmicb.2015.01226

  23. Kim K, Kim YU, Koh BH, Hwang SS, Kim SH, Lepine F, Cho YH, Lee GR (2010) HHQ and PQS, two Pseudomonas aeruginosa quorum-sensing molecules, down-regulate the innate immune responses through the nuclear factor-kappaB pathway. Immunology 129(4):578–588. doi:10.1111/j.1365-2567.2009.03160.x

  24. Kaparakis-Liaskos M, Ferrero RL (2015) Immune modulation by bacterial outer membrane vesicles. Nat Rev Immunol 15(6):375–387. doi:10.1038/nri3837

    Article  CAS  PubMed  Google Scholar 

  25. Legendre C, Reen FJ, Mooij MJ, McGlacken GP, Adams C, O'Gara F (2012) Pseudomonas aeruginosa alkyl quinolones repress hypoxia-inducible factor 1 (HIF-1) signaling through HIF-1alpha degradation. Infect Immun 80(11):3985–3992. doi:10.1128/IAI.00554-12

  26. Huse H, Whiteley M (2011) 4-Quinolones: smart phones of the microbial world. Chem Rev 111(1):152–159. doi:10.1021/cr100063u

    Article  CAS  PubMed  Google Scholar 

  27. Collier DN, Anderson L, McKnight SL, Noah TL, Knowles M, Boucher R, Schwab U, Gilligan P, Pesci EC (2002) A bacterial cell to cell signal in the lungs of cystic fibrosis patients. FEMS Microbiol Lett 215(1):41–46

    Article  CAS  PubMed  Google Scholar 

  28. Gruber JD, Chen W, Parnham S, Beauchesne K, Moeller P, Flume PA, Zhang YM (2016) The role of 2,4-dihydroxyquinoline (DHQ) in Pseudomonas aeruginosa pathogenicity. PeerJ 4:e1495. doi:10.7717/peerj.1495

  29. Bala A, Chhibber S, Harjai K (2014) Pseudomonas quinolone signalling system: a component of quorum sensing cascade is a crucial player in the acute urinary tract infection caused by Pseudomonas aeruginosa. Int J Med Microbiol 304(8):1199–1208. doi:10.1016/j.ijmm.2014.08.013

  30. Palmer GC, Schertzer JW, Mashburn-Warren L, Whiteley M (2011) Quantifying Pseudomonas aeruginosa quinolones and examining their interactions with lipids. Methods Mol Biol 692:207–217. doi:10.1007/978-1-60761-971-0_15

  31. Diggle SP, Fletcher MP, Camara M, Williams P (2011) Detection of 2-alkyl-4-quinolones using biosensors. Methods Mol Biol 692:21–30. doi:10.1007/978-1-60761-971-0_2

    Article  CAS  PubMed  Google Scholar 

  32. Choi DS, Kim DK, Choi SJ, Lee J, Choi JP, Rho S, Park SH, Kim YK, Hwang D, Gho YS (2011) Proteomic analysis of outer membrane vesicles derived from Pseudomonas aeruginosa. Proteomics 11(16):3424–3429

    Google Scholar 

  33. Bala A, Gupta RK, Chhibber S, Harjai K (2013) Detection and quantification of quinolone signalling molecule: a third quorum sensing molecule of Pseudomonas aeruginosa by high performance-thin layer chromatography. J Chromatogr B Anal Technol Biomed Life Sci 930:30–35. doi:10.1016/j.jchromb.2013.04.027

  34. Chutkan H, Macdonald I, Manning A, Kuehn MJ (2013) Quantitative and qualitative preparations of bacterial outer membrane vesicles. Methods Mol Biol 966:259–272. doi:10.1007/978-1-62703-245-2_16

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Baig NF, Dunham SJ, Morales-Soto N, Shrout JD, Sweedler JV, Bohn PW (2015) Multimodal chemical imaging of molecular messengers in emerging Pseudomonas aeruginosa bacterial communities. Analyst 140(19):6544–6552. doi:10.1039/c5an01149c

  36. Peterson DS (2007) Matrix-free methods for laser desorption/ionization mass spectrometry. Mass Spectrom Rev 26(1):19–34. doi:10.1002/mas.20104

    Article  CAS  PubMed  Google Scholar 

  37. Coffinier Y, Boukherroub R (2014) Porous Silicon-Based Mass Spectrometry. In: Canham L (ed) Handbook of Porous Silicon. Springer International Publishing, Cham, pp 869–885. doi:10.1007/978-3-319-05744-6_88

    Google Scholar 

  38. Kusano M, Kawabata S, Tamura Y, Mizoguchi D, Murouchi M, Kawasaki H, Arakawa R, Tanaka K (2014) Laser Desorption/Ionization Mass Spectrometry (LDI-MS) of lipids with iron oxide nanoparticle-coated targets. Mass Spectrom (Tokyo) 3(1):A0026. doi:10.5702/massspectrometry.A0026

    Article  Google Scholar 

  39. Ghosh D, Panchagnula V, Dhaware D (2016) Selective detection and analysis of small molecules. EU Patent EP 2676287 A2.

    Google Scholar 

  40. Pluháček T, Lemr K, Ghosh D, Milde D, Novák J, Havlíček V (2016) Characterization of microbial siderophores by mass spectrometry. Mass Spectrom Rev 35(1):35–47

    Article  PubMed  Google Scholar 

  41. Strohalm M, Kavan D, Novak P, Volny M, Havlicek V (2010) mMass 3: a cross-platform software environment for precise analysis of mass spectrometric data. Anal Chem 82(11):4648–4651

    Article  CAS  PubMed  Google Scholar 

  42. Kadurugamuwa JL, Beveridge TJ (1995) Virulence factors are released from Pseudomonas aeruginosa in association with membrane vesicles during normal growth and exposure to gentamicin: a novel mechanism of enzyme secretion. J Bacteriol 177(14):3998–4008

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Special Center for Molecular Medicine, Jawaharlal Nehru University, New Delhi, 110067, India

    Pallavi Lahiri & Dipankar Ghosh

Authors
  1. Pallavi Lahiri
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Dipankar Ghosh
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Dipankar Ghosh .

Editor information

Editors and Affiliations

  1. University of Miami, Miami, Florida, USA

    Sanjoy K. Bhattacharya

Rights and permissions

Reprints and permissions

Copyright information

© 2017 Springer Science+Business Media LLC

About this protocol

Cite this protocol

Lahiri, P., Ghosh, D. (2017). Single-Step Capture and Targeted Metabolomics of Alkyl-Quinolones in Outer Membrane Vesicles of Pseudomonas aeruginosa . In: Bhattacharya, S. (eds) Lipidomics. Methods in Molecular Biology, vol 1609. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6996-8_15

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-6996-8_15

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6995-1

  • Online ISBN: 978-1-4939-6996-8

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation