Synthesis and electrochemical detection of a thiazolyl-indole natural product isolated from the nosocomial pathogen Pseudomonas aeruginosa


Pseudomonas aeruginosa is a Gram-negative opportunistic pathogen, capable of surviving in a broad range of natural environments and quickly acquiring resistance. It is associated with hospital-acquired infections, particularly in patients with compromised immunity, and is the primary cause of morbidity and mortality in cystic fibrosis (CF) patients. P. aeruginosa is also of nosocomial importance on dairy farms and veterinary hospitals, where it is a key morbidity factor in bovine mastitis. P. aeruginosa uses a cell-cell communication system consisting of signalling molecules to coordinate bacterial secondary metabolites, biofilm formation, and virulence. Simple and sensitive methods for the detection of biomolecules as indicators of P. aeruginosa infection would be of great clinical importance. Here, we report the synthesis of the P. aeruginosa natural product, barakacin, which was recently isolated from the bovine ruminal strain ZIO. A simple and sensitive electrochemical method was used for barakacin detection using a boron-doped diamond (BDD) and glassy carbon (GC) electrodes, based on cyclic voltammetry (CV) and differential pulse voltammetry (DPV). The influence of electrolyte pH on the peak potential and peak currents was also investigated. At pH 2.0, the peak current was linearly dependent on barakacin concentration (in the range used, 1–10 μM), with correlation coefficients greater than 0.98 on both electrodes. The detection limit (S/N = 3) on the BDD electrode was 100-fold lower than that obtained on the GC electrode. The optimized method using the BDD electrode was extended to bovine (cow feces) and human (sputum of a CF patient) samples. Spiked barakacin was easily detected in these matrices at a limit of 0.5 and 0.05 μM, respectively.

Electrochemical detection of barakacin

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig 6
Fig 7


  1. 1.

    European Centre for Disease Prevention and Control. Annual epidemiological report. Antimicrobial resistance and healthcare-associated infections. ECDC Surveillance report. 2014.

  2. 2.

    Kesarwani M, Hazan R, He J, Que YA, Apidianakis Y, Lesic B, et al. A quorum sensing regulated small volatile molecule reduces acute virulence and promotes chronic infection phenotypes. PLoS Pathog. 2011;7(8):e1002192.

    CAS  Article  Google Scholar 

  3. 3.

    Lepine F, Deziel E, Milot S, Rahme LG. A stable isotope dilution assay for the quantification of the Pseudomonas quinolone signal in Pseudomonas aeruginosa cultures. Biochim Biophys Acta. 2003;1622(1):36–41.

    CAS  Article  Google Scholar 

  4. 4.

    Reen FJ, Mooij MJ, Holcombe LJ, McSweeney CM, McGlacken GP, Morrissey JP, et al. The Pseudomonas quinolone signal (PQS), and its precursor HHQ, modulate interspecies and interkingdom behaviour. FEMS Microbiol Ecol. 2011;77(2):413–28.

    CAS  Article  Google Scholar 

  5. 5.

    Quinn PJ, Markey BK, Leonard F, FitzPatrick E, Fanning S. Concise review of veterinary microbiology. John Wiley & Sons; 2015.

  6. 6.

    Daly M, Power E, Bjorkroth J, Sheehan P, O’Connell A, Colgan M, et al. Molecular analysis of Pseudomonas aeruginosa: epidemiological investigation of mastitis outbreaks in Irish dairy herds. Appl Environ Microbiol. 1999;65(6):2723–9.

    CAS  Google Scholar 

  7. 7.

    Scaccabarozzi L, Leoni L, Ballarini A, Barberio A, Locatelli C, Casula A, et al. Pseudomonas aeruginosa in dairy goats: genotypic and phenotypic comparison of intramammary and environmental isolates. PLoS One. 2015;10(11):e0142973.

    Article  Google Scholar 

  8. 8.

    Donlan RM, Costerton JW. Biofilms: survival mechanisms of clinically relevant microorganisms. Clin Microbiol Rev. 2002;15(2):167–93.

    CAS  Article  Google Scholar 

  9. 9.

    Ortori CA, Dubern JF, Chhabra SR, Camara M, Hardie K, Williams P, et al. Simultaneous quantitative profiling of N-acyl-l-homoserine lactone and 2-alkyl-4(1H)-quinolone families of quorum-sensing signaling molecules using LC-MS/MS. Anal Bioanal Chem. 2011;399(2):839–50.

    CAS  Article  Google Scholar 

  10. 10.

    Williams P, Winzer K, Chan WC, Camara M. Look who’s talking: communication and quorum sensing in the bacterial world. Philos T R Soc B. 2007;362(1483):1119–34.

    CAS  Article  Google Scholar 

  11. 11.

    Haussler S, Becker T. The pseudomonas quinolone signal (PQS) balances life and death in Pseudomonas aeruginosa populations. PLoS Pathog. 2008;4(9):e1000166.

    Article  Google Scholar 

  12. 12.

    Govan JR, Deretic V. Microbial pathogenesis in cystic fibrosis: mucoid Pseudomonas aeruginosa and Burkholderia cepacia. Microbiol Rev. 1996;60(3):539–74.

    CAS  Google Scholar 

  13. 13.

    Bala A, Gupta RK, Chhibber S, Harjai K. Detection and quantification of quinolone signalling molecule: a third quorum sensing molecule of Pseudomonas aeruginosa by high performance-thin layer chromatography. J Chromatogr B Analyt Technol Biomed Life Sci. 2013;930:30–5.

    CAS  Article  Google Scholar 

  14. 14.

    McGlacken GP, McSweeney CM, O’Brien T, Lawrence SE, Elcoate CJ, Reen FJ, et al. Synthesis of 3-halo-analogues of HHQ, subsequent cross-coupling and first crystal structure of Pseudomonas quinolone signal (PQS). Tetrahedron Lett. 2010;51(45):5919–21.

    CAS  Article  Google Scholar 

  15. 15.

    Lee J, Wu J, Deng Y, Wang J, Wang C, Wang J, et al. A cell-cell communication signal integrates quorum sensing and stress response. Nat Chem Biol. 2013;9:339–43.

    CAS  Article  Google Scholar 

  16. 16.

    West SE, Zeng L, Kosorok MR, Laxova A, Rock MJ, Splaingard MJ, et al. Respiratory infections with Pseudomonas aeruginosa in children with cystic fibrosis; early detection by serology and assessment of risk factors. JAMA. 2002;287:2958–67.

    Article  Google Scholar 

  17. 17.

    Karpati F, Jonasson J. Polymerase chain reaction for the detection of Pseudomonas aeruginosa, Stenotrophomonas maltophilia and Burkholderia cepacia in sputum of patients with cystic fibrosis. Mol Cell Probes. 1996;10:397–403.

    CAS  Article  Google Scholar 

  18. 18.

    Fletcher MP, Diggle SP, Camara M, Williams P. Biosensor-based assays for PQS, HHQ and related 2-alkyl-4-quinolone quorum sensing signal molecules. Nat Protoc. 2007;2(5):1254–62.

    CAS  Article  Google Scholar 

  19. 19.

    Zhou L, Glennon JD, Luong JHT, Reen FJ, O’Gara F, McSweeney C, et al. Detection of the Pseudomonas quinolone signal (PQS) by cyclic voltammetry and amperometry using a boron doped diamond electrode. Chem Commun. 2011;47(37):10347–9.

    CAS  Article  Google Scholar 

  20. 20.

    Shang F, Muimhneachain EO, Reen FJ, Buzid A, O’Gara F, Luong JHT, et al. One step preparation and electrochemical analysis of IQS, a cell-cell communication signal in the nosocomial pathogen Pseudomonas aeruginosa. Bioorg Med Chem Lett. 2014;24(19):4703–7.

    CAS  Article  Google Scholar 

  21. 21.

    Alatraktchi FA, Breum Andersen S, Krogh Johansen H, Molin S, Svendsen WE. Fast selective detection of pyocyanin using cyclic voltammetry. Sensors. 2016;16(3):1–10.

    Article  Google Scholar 

  22. 22.

    Sismaet HJ, Banerjee A, McNish S, Choi Y, Torralba M, Lucas S, et al. Electrochemical detection of Pseudomonas in wound exudate samples from patients with chronic wounds. Wound Repair Regen. 2016;24(2):366–72.

    Article  Google Scholar 

  23. 23.

    Webster TA, Sismaet HJ, Chan IP, Goluch ED. Electrochemically monitoring the antibiotic susceptibility of Pseudomonas aeruginosa biofilms. Analyst. 2015;140(21):7195–201.

    CAS  Article  Google Scholar 

  24. 24.

    Webster TA, Sismaet HJ, Conte JL, Chan IP, Goluch ED. Electrochemical detection of Pseudomonas aeruginosa in human fluid samples via pyocyanin. Biosens Bioelectron. 2014;60:265–70.

    CAS  Article  Google Scholar 

  25. 25.

    Webster TA, Sismaet HJ, Sattler AF, Goluch ED. Improved monitoring of P. aeruginosa on agar plates. Anal Methods. 2015;7(17):7150–5.

    CAS  Article  Google Scholar 

  26. 26.

    Sismaet HJ, Webster TA, Goluch ED. Up-regulating pyocyanin production by amino acid addition for early electrochemical identification of Pseudomonas aeruginosa. Analyst. 2014;139(17):4241–6.

    CAS  Article  Google Scholar 

  27. 27.

    Hameed A, Pi H-W, Lin S-Y, Lai W-A, Young L-S, Liu Y-C, et al. Direct electrochemical sensing of phenazine-1-carboxylic acid secreted by Pseudomonas chlororaphis subsp. aureofaciens BCRC 11057T using disposable screen-printed carbon electrode. Electroanalysis. 2016;28(4):846–53.

    CAS  Article  Google Scholar 

  28. 28.

    Bellin DL, Sakhtah H, Rosenstein JK, Levine PM, Thimot J, Emmett K, et al. Integrated circuit-based electrochemical sensor for spatially resolved detection of redox-active metabolites in biofilms. Nat Commun. 2014;5:3256.

    Article  Google Scholar 

  29. 29.

    Bellin DL, Sakhtah H, Zhang Y, Price-Whelan A, Dietrich LE, Shepard KL. Electrochemical camera chip for simultaneous imaging of multiple metabolites in biofilms. Nat Commun. 2016;7:10535.

    CAS  Article  Google Scholar 

  30. 30.

    Bukelman O, Amara N, Mashiach R, Krief P, Meijler MM, Alfonta L. Electrochemical analysis of quorum sensing inhibition. Chem Commun. 2009;20:2836–8.

    Article  Google Scholar 

  31. 31.

    Kim E, Gordonov T, Bentley WE, Payne GF. Amplified and in situ detection of redox-active metabolite using a biobased redox capacitor. Anal Chem. 2013;85(4):2102–8.

    CAS  Article  Google Scholar 

  32. 32.

    Sharp D, Gladstone P, Smith RB, Forsythe S, Davis J. Approaching intelligent infection diagnostics: carbon fibre sensor for electrochemical pyocyanin detection. Bioelectrochemistry. 2010;77(2):114–9.

    CAS  Article  Google Scholar 

  33. 33.

    Webster TA, Goluch ED. Electrochemical detection of pyocyanin in nanochannels with integrated palladium hydride reference electrodes. Lab Chip. 2012;12(24):5195–201.

    CAS  Article  Google Scholar 

  34. 34.

    Webster TA, Sismaet HJ, Goluch ED. Amperometric detection of pyocyanin in nanofluidic channels. Nano LIFE. 2013;03(01):1340011.

    Article  Google Scholar 

  35. 35.

    Seviour T, Doyle LE, Lauw SJL, Hinks J, Rice SA, Nesatyy VJ, et al. Voltammetric profiling of redox-active metabolites expressed by Pseudomonas aeruginosa for diagnostic purposes. Chem Commun. 2015;51(18):3789–92.

    CAS  Article  Google Scholar 

  36. 36.

    Oziat J, Elsen S, Owens RM, Malliaras GG, Mailley P. Electrochemistry provides a simple way to monitor Pseudomonas aeruginosa metabolites. Conf Proc IEEE Eng Med Biol Soc. 2015;7522–5.

  37. 37.

    Zendah I, Shaaban KA, Helmke E, Maier A, Fiebig HH, Laatsch H. Barakacin: a thiazolyl-indole alkaloid isolated from a ruminal Pseudomonas sp. Z Naturforsch B. 2012;67(5):417–20.

    CAS  Article  Google Scholar 

  38. 38.

    Zendah. I. Isolation, Purification and structure elucidation of new secondary metabolites from terrestrial, marine and ruminal microorganisms [doctoral dissertation]. Göttingen: Niedersächsische Staats- und Universitätsbibliothek Göttingen: der Georg-August-Universität zu Göttingen; 2012.

  39. 39.

    El Sayed MT, Ahmed KM, Mahmoud K, Hilgeroth A. Synthesis, cytostatic evaluation and structure activity relationships of novel bis-indolylmethanes and their corresponding tetrahydroindolocarbazoles. Eur J Med Chem. 2015;90:845–59.

    Article  Google Scholar 

  40. 40.

    Luong JHT, Male KB, Glennon JD. Boron-doped diamond electrode: synthesis, characterization, functionalization and analytical applications. Analyst. 2009;134(10):1965–79.

    CAS  Article  Google Scholar 

  41. 41.

    Shang F, Liu Y, Hrapovic S, Glennon JD, Luong JHT. Selective detection of dopamine using a combined permselective film of electropolymerized (poly-tyramine and poly-pyrrole-1-propionic acid) on a boron-doped diamond electrode. Analyst. 2009;134(3):519–27.

    CAS  Article  Google Scholar 

  42. 42.

    Filipe OMS, Brett CMA. Characterization of carbon film electrodes for electroanalysis by electrochemical impedance. Electroanalysis. 2004;16(12):994–1001.

    CAS  Article  Google Scholar 

  43. 43.

    Bannerman DD, Chockalingam A, Paape MJ, Hope JC. The bovine innate immune response during experimentally-induced Pseudomonas aeruginosa mastitis. Vet Immunol Immunopathol. 2005;107(3–4):201–15.

    CAS  Article  Google Scholar 

  44. 44.

    Shaheen M, Tantary H, Nabi S. A. treatise on bovine mastitis: disease and disease economics, etiological basis, risk factors, impact on human health, therapeutic management. Prevention and control strategy. J Adv Dairy Res. 2016;4(150):1–10.

    Google Scholar 

Download references


This research was financially supported by SFI/EI Technology Innovation Development Award (TIDA) (SFI/12/TIDA/B2405). FOG acknowledges the grants awarded by the European Commission (FP7-PEOPLE-2013-ITN, 607786; FP7-KBBE-2012-6, CP-TP-312184; FP7-KBBE-2012-6, 311975; OCEAN 2011–2, 287589; Marie Curie 256596; EU-634486); Science Foundation Ireland (SSPC-2, 12/RC/2275; 13/TIDA/B2625; 12/TIDA/B2411; 12/TIDA/B2405; 14/TIDA/2438); the Department of Agriculture and Food (FIRM/RSF/CoFoRD; FIRM 08/RDC/629; FIRM 1/F009/MabS; FIRM 13/F/516); the Irish Research Council for Science, Engineering and Technology (PD/2011/2414; GOIPG/2014/647); the Health Research Board/Irish Thoracic Society (MRCG-2014-6); the Marine Institute (Beaufort award C2CRA 2007/082); and Teagasc (Walsh Fellowship 2013). JDG thanks Science Foundation Ireland (08/SRC/B1412) for research funding of the Irish Separation Science Cluster (ISSC) under the Strategic Research Cluster Programme. GMG acknowledges supports by Science Foundation Ireland (SFI/12/IP/1315, SFI/09/RFP/CHS2353, and SSPC2 12/RC/2275), the Irish Research Council (GOIPG/2013/336), and the UCC for a Strategic Research Fund Ph.D. Studentship.

Author information



Corresponding authors

Correspondence to Jeremy D. Glennon or Gerard P. McGlacken.

Ethics declarations

Sputum samples were collected from paediatric patients attending the CF clinic at Cork University Hospital, Ireland. Ethical approval was granted by the Clinical Research Ethics Committee (CREC) for sputum collection and samples were handled according to the approved guidelines. Written informed consent from all patients/guardians was obtained for acquisition and analysis outlined in this study.

Conflict of interest

The authors declare that they have no competing interests.

Electronic supplementary material

Below is the link to the electronic supplementary material.


(PDF 843 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Buzid, A., Muimhneacháin, E.Ó., Reen, F.J. et al. Synthesis and electrochemical detection of a thiazolyl-indole natural product isolated from the nosocomial pathogen Pseudomonas aeruginosa . Anal Bioanal Chem 408, 6361–6367 (2016).

Download citation


  • Natural product
  • Biomarker
  • Pseudomonas aeruginosa
  • Electrochemical detection
  • Bovine
  • Cystic fibrosis