Annals of Biomedical Engineering

, Volume 42, Issue 11, pp 2314–2321 | Cite as

Rapid Antimicrobial Susceptibility Testing with Electrokinetics Enhanced Biosensors for Diagnosis of Acute Bacterial Infections

  • Tingting Liu
  • Yi Lu
  • Vincent Gau
  • Joseph C. Liao
  • Pak Kin WongEmail author


Rapid pathogen detection and antimicrobial susceptibility testing (AST) are required in diagnosis of acute bacterial infections to determine the appropriate antibiotic treatment. Molecular approaches for AST are often based on the detection of known antibiotic resistance genes. Phenotypic culture analysis requires several days from sample collection to result reporting. Toward rapid diagnosis of bacterial infection in non-traditional healthcare settings, we have developed a rapid AST approach that combines phenotypic culture of bacterial pathogens in physiological samples and electrochemical sensing of bacterial 16S rRNA. The assay determines the susceptibility of pathogens by detecting bacterial growth under various antibiotic conditions. AC electrokinetic fluid motion and Joule heating induced temperature elevation are optimized to enhance the sensor signal and minimize the matrix effect, which improve the overall sensitivity of the assay. The electrokinetics enhanced biosensor directly detects the bacterial pathogens in blood culture without prior purification. Rapid determination of the antibiotic resistance profile of Escherichia coli clinical isolates is demonstrated.


Electrokinetic enhancement Point-of-care Antimicrobial susceptibility testing Acute bacterial infections Electrochemical sensing 



This work was supported by NIH Health Director’s New Innovator Award (1DP2OD007161-01) and NIAID (1U01AI082457-01 and 2R44AI088756-03).


  1. 1.
    An, G., G. Nieman, and Y. Vodovotz. Toward computational identification of multiscale “tipping points” in acute inflammation and multiple organ failure. Ann. Biomed. Eng. 40:2414–2424, 2012.PubMedCrossRefGoogle Scholar
  2. 2.
    Campbell, J., and J. A. Washington. Evaluation of the necessity for routine terminal subcultures of previously negative blood cultures. J. Clin. Microbiol. 12:576–578, 1980.PubMedPubMedCentralGoogle Scholar
  3. 3.
    Castellanos, A., A. Ramos, A. Gonzalez, N. G. Green, and H. Morgan. Electrohydrodynamics and dielectrophoresis in microsystems: scaling laws. J. Phys. D-Appl. Phys. 36:2584–2597, 2003.CrossRefGoogle Scholar
  4. 4.
    Chen, C. H., Y. Lu, M. L. Y. Sin, K. E. Mach, D. D. Zhang, V. Gau, J. C. Liao, and P. K. Wong. Antimicrobial susceptibility testing using high surface-to-volume ratio microchannels. Anal. Chem. 82:1012–1019, 2010.PubMedCrossRefPubMedCentralGoogle Scholar
  5. 5.
    Chiu, M. L., W. Lawi, S. T. Snyder, P. K. Wong, J. C. Liao, and V. Gau. Matrix effects—a challenge toward automation of molecular analysis. J. Assoc. Lab. Autom. 15:233–242, 2010.CrossRefGoogle Scholar
  6. 6.
    Daum, R. S., T. Ito, K. Hiramatsu, F. Hussain, K. Mongkolrattanothai, M. Jamklang, and S. Boyle-Wang. A novel methicillin-resistance cassette in community-acquired methicillin-resistant staphylococcus aureus isolates of diverse genetic backgrounds. J. Infect. Dis. 186:1344–1347, 2002.PubMedCrossRefGoogle Scholar
  7. 7.
    Davies, J., and D. Davies. Origins and evolution of antibiotic resistance. Microbiol. Mol. Biol. Rev. 74:417–+, 2010.Google Scholar
  8. 8.
    Erickson, D., X. Z. Liu, R. Venditti, D. Q. Li, and U. J. Krull. Electrokinetically based approach for single-nucleotide polymorphism discrimination using a microfluidic device. Anal. Chem. 77:4000–4007, 2005.PubMedCrossRefGoogle Scholar
  9. 9.
    Green, N. G., A. Ramos, A. Gonzalez, A. Castellanos, and H. Morgan. Electrothermally induced fluid flow on microelectrodes. J. Electrostat. 53:71–87, 2001.CrossRefGoogle Scholar
  10. 10.
    Hansen, W. L., J. Beuving, A. Verbon, and P. F. Wolffs. One-day workflow scheme for bacterial pathogen detection and antimicrobial resistance testing from blood cultures. J. Vis. Exp. 2012. doi: 10.3791/3254.
  11. 11.
    Hawkey, P. M. Pre-clinical experience with daptomycin. J. Antimicrob. Chemother. 62:7–14, 2008.CrossRefGoogle Scholar
  12. 12.
    Herold, B. C., L. C. Immergluck, M. C. Maranan, D. S. Lauderdale, R. E. Gaskin, S. Boyle-Vavra, C. D. Leitch, and R. S. Daum. Community-acquired methicillin-resistant staphylococcus aureus in children with no identified predisposing risk. Jama-Journal of the American Medical Association. 279:593–598, 1998.CrossRefGoogle Scholar
  13. 13.
    Huttner, B., and S. Harbarth. Antibiotics are not automatic anymore—the French national campaign to cut antibiotic overuse. Plos Med. 6, 2009.Google Scholar
  14. 14.
    Ingber, D. E. From cellular mechanotransduction to biologically inspired engineering. Ann. Biomed. Eng. 38:1148–1161, 2010.PubMedCrossRefPubMedCentralGoogle Scholar
  15. 15.
    Ingham, C. J., M. van den Ende, D. Pijnenburg, P. C. Wever, and P. M. Schneeberger. Growth and multiplexed analysis of microorganisms on a subdivided, highly porous, inorganic chip manufactured from anopore. Appl. Environ. Microbiol. 71:8978–8981, 2005.PubMedCrossRefPubMedCentralGoogle Scholar
  16. 16.
    Kuper, K. M., D. M. Boles, J. E. Mohr, and A. Wanger. Antimicrobial susceptibility testing: a primer for clinicians. Pharmacotherapy. 29:1326–1343, 2009.PubMedCrossRefGoogle Scholar
  17. 17.
    Lawi, W., C. Wiita, S. T. Snyder, F. Wei, D. Wong, P. K. Wong, J. C. Liao, D. Haake, and V. Gau. A microfluidic cartridge system for multiplexed clinical analysis. J. Assoc. Lab. Autom. 14:407–412, 2009.Google Scholar
  18. 18.
    Levy, S. B., and B. Marshall. Antibacterial resistance worldwide: causes, challenges and responses. Nat. Med. 10:S122–S129, 2004.PubMedCrossRefGoogle Scholar
  19. 19.
    Liu, T., M. L. Y. Sin, J. D. Pyne, V. Gau, J. C. Liao, and P. K. Wong. Electrokinetic stringency control in self-assembled monolayer-based biosensors for multiplex urinary tract infection diagnosis. Nanomed. Nanotechnol. Biol. Med. 10:159–166, 2014.CrossRefGoogle Scholar
  20. 20.
    Louie, M., and F. R. Cockerill. Susceptibility testing—phenotypic and genotypic tests for bacteria and mycobacteria. Infect. Dis. Clin. North Am. 15:1205–+, 2001.Google Scholar
  21. 21.
    Lu, Y., J. Gao, D. D. Zhang, V. Gau, J. C. Liao, and P. K. Wong. Single cell antimicrobial susceptibility testing by confined microchannels and electrokinetic loading. Anal. Chem. 85:3971–3976, 2013.PubMedCrossRefPubMedCentralGoogle Scholar
  22. 22.
    Mach, K. E., R. Mohan, E. J. Baron, M. C. Shih, V. Gau, P. K. Wong, and J. C. Liao. A biosensor platform for rapid antimicrobial susceptibility testing directly from clinical samples. J. Urol. 185:148–153, 2011.PubMedCrossRefPubMedCentralGoogle Scholar
  23. 23.
    Mach, K. E., P. K. Wong, and J. C. Liao. Biosensor diagnosis of urinary tract infections: a path to better treatment? Trends Pharmacol. Sci. 32:330–336, 2011.PubMedCrossRefPubMedCentralGoogle Scholar
  24. 24.
    Murray, P. R. Determination of the optimum incubation period of blood culture broths for the detection of clinically significant septicemia. J. Clin. Microbiol. 21:481–485, 1985.PubMedPubMedCentralGoogle Scholar
  25. 25.
    Ouyang, M. X., R. Mohan, Y. Lu, T. T. Liu, K. E. Mach, M. L. Y. Sin, M. McComb, J. Joshi, V. Gau, P. K. Wong, and J. C. Liao. An ac electrokinetics facilitated biosensor cassette for rapid pathogen identification. Analyst. 138:3660–3666, 2013.PubMedCrossRefPubMedCentralGoogle Scholar
  26. 26.
    Pfaller, M. A., R. N. Jones, and P. Coll. Amer. Performance accuracy of antibacterial and antifungal susceptibility test methods—report from the college of American pathologists microbiology surveys program (2001–2003). Arch. Pathol. Lab. Med. 130:767–778, 2006.PubMedGoogle Scholar
  27. 27.
    Pinner, R. W., S. M. Teutsch, L. Simonsen, L. A. Klug, J. M. Graber, M. J. Clarke, and R. L. Berkelman. Trends in infectious diseases mortality in the United States. JAMA 275:189–193, 1996.PubMedCrossRefGoogle Scholar
  28. 28.
    Ramos, A., H. Morgan, N. G. Green, and A. Castellanos. Ac electrokinetics: a review of forces in microelectrode structures. J. Phys. D-Appl. Phys. 31:2338–2353, 1998.CrossRefGoogle Scholar
  29. 29.
    Riahi, R., K. E. Mach, R. Mohan, J. C. Liao, and P. K. Wong. Molecular detection of bacterial pathogens using microparticle enhanced double-stranded DNA probes. Anal. Chem. 83:6349–6354, 2011.PubMedCrossRefPubMedCentralGoogle Scholar
  30. 30.
    Sezonov, G., D. Joseleau-Petit, and R. D’Ari. Escherichia coli physiology in luria-bertani broth. J. Bacteriol. 189:8746–8749, 2007.PubMedCrossRefPubMedCentralGoogle Scholar
  31. 31.
    Sin, M. L. Y., V. Gau, J. C. Liao, and P. K. Wong. Electrothermal fluid manipulation of high-conductivity samples for laboratory automation applications. J. Assoc. Lab. Autom. 15:426–432, 2010.CrossRefGoogle Scholar
  32. 32.
    Sin, M. L. Y., T. T. Liu, J. D. Pyne, V. Gau, J. C. Liao, and P. K. Wong. In situ electrokinetic enhancement for self-assembled-monolayer-based electrochemical biosensing. Anal. Chem. 84:2702–2707, 2012.PubMedCrossRefPubMedCentralGoogle Scholar
  33. 33.
    Sin, M. L., K. E. Mach, P. K. Wong, and J. C. Liao. Advances and challenges in biosensor-based diagnosis of infectious diseases. Expert Review of Molecular Diagnostics. 14:225–244, 2014.PubMedCrossRefPubMedCentralGoogle Scholar
  34. 34.
    Tondo, E. C., M. C. M. Guimarpes, J. A. P. Henriques, and M. A. Z. Ayub. Assessing and analysing contamination of a dairy products processing plant by staphylococcus aureus using antibiotic resistance and PFGE. Can. J. Microbiol. 46:1108–1114, 2000.PubMedCrossRefGoogle Scholar
  35. 35.
    von Lilienfeld-Toal, M., L. E. Lehmann, A. D. Raadts, C. Hahn-Ast, K. S. Orlopp, G. Marklein, I. Purr, G. Cook, A. Hoeft, A. Glasmacher, and F. Stuber. Utility of a commercially available multiplex real-time PCR assay to detect bacterial and fungal pathogens in febrile neutropenia. J. Clin. Microbiol. 47:2405–2410, 2009.CrossRefGoogle Scholar
  36. 36.
    Waldeisen, J. R., T. Wang, D. Mitra, and L. P. Lee. A real-time pcr antibiogram for drug-resistant sepsis. Plos One 6, 2011.Google Scholar
  37. 37.
    Yanagihara, K., Y. Kitagawa, M. Tomonaga, K. Tsukasaki, S. Kohno, M. Seki, H. Sugimoto, T. Shimazu, O. Tasaki, A. Matsushima, Y. Ikeda, S. Okamoto, N. Aikawa, S. Hori, H. Obara, A. Ishizaka, N. Hasegawa, J. Takeda, S. Kamihira, K. Sugahara, S. Asari, M. Murata, Y. Kobayashi, H. Ginba, Y. Sumiyama, and M. Kitajima. Evaluation of pathogen detection from clinical samples by real-time polymerase chain reaction using a sepsis pathogen DNA detection kit. Crit. Care. 14, 20.Google Scholar
  38. 38.
    Zuker, M. Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res. 31:3406–3415, 2003.PubMedCrossRefPubMedCentralGoogle Scholar

Copyright information

© Biomedical Engineering Society 2014

Authors and Affiliations

  • Tingting Liu
    • 1
  • Yi Lu
    • 1
  • Vincent Gau
    • 2
  • Joseph C. Liao
    • 3
    • 4
  • Pak Kin Wong
    • 1
    Email author
  1. 1.Department of Aerospace and Mechanical EngineeringThe University of ArizonaTucsonUSA
  2. 2.GeneFluidics Inc.IrwindaleUSA
  3. 3.Department of UrologyStanford UniversityStanfordUSA
  4. 4.Veterans Affairs Palo Alto Health Care SystemPalo AltoUSA

Personalised recommendations