BioChip Journal

, Volume 12, Issue 4, pp 287–293 | Cite as

Detection of Bacillus Cereus Using Bioluminescence Assay with Cell Wall-binding Domain Conjugated Magnetic Nanoparticles

  • Chanyong Park
  • Minsuk Kong
  • Ju-Hoon Lee
  • Sangryeol Ryu
  • Sungsu ParkEmail author
Original Article


Bacillus cereus can cause blood infections (i.e., sepsis). Its early detection is very important for treating patients. However, an antibody with high binding affinity to B. cereus is not currently available. Bacteriophage cell wall-binding domain (CBD) has strong and specific binding affinity to B. cereus. Here, we report the improvement in the sensitivity of an ATP bioluminescence assay for B. cereus detection using CBD-conjugated magnetic nanoparticles (CBDMNPs). The assay was able to detect as few as 10 colony forming units (CFU) per mL and 103 CFU per mL in buffer and blood. CBD-MNPs did not show any cross-reactivity with other microorganisms. These results demonstrate the feasibility of the ATP assay for the detection of B. cereus.


Bacillus cereus Cell wall-binding domain Enrichment Magnetic nanoparticles ATP bioluminescence assay 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    Sutherland, A. et al. Development and validation of a novel molecular biomarker diagnostic test for the early detection of sepsis. Crit. Care 15, R149 (2011).CrossRefGoogle Scholar
  2. 2.
    Ikeda, M. et al. Clinical characteristics and antimicrobial susceptibility of Bacillus cereus blood stream infections. Ann. Clin. Microbiol. Antimicrob. 14, 43 (2015).CrossRefGoogle Scholar
  3. 3.
    Goto, M. & Al-Hasan, M. Overall burden of bloodstream infection and nosocomial bloodstream infection in North America and Europe. Clin. Microbiol. Infect. 19, 501–509 (2013).CrossRefGoogle Scholar
  4. 4.
    Angus, D.C. et al. Epidemiology of severe sepsis in the United States: analysis of incidence, outcome, and associated costs of care. Crit. Care Med. 29, 1303–1310 (2001).CrossRefGoogle Scholar
  5. 5.
    Shen, H. et al. Rapid and selective detection of pathogenic bacteria in bloodstream infections with aptamer-based recognition. ACS Appl. Mater. Interfaces 8, 19371–19378 (2016).CrossRefGoogle Scholar
  6. 6.
    Yagupsky, P. & Nolte, F. Quantitative aspects of septicemia. Clin. Microbiol. Rev. 3, 269–279 (1990).CrossRefGoogle Scholar
  7. 7.
    Reier-Nilsen, T., Farstad, T., Nakstad, B., Lauvrak, V. & Steinbakk, M. Comparison of broad range 16S rDNA PCR and conventional blood culture for diagnosis of sepsis in the newborn: a case control study. BMC Pediatr. 9, 5 (2009).CrossRefGoogle Scholar
  8. 8.
    Ahmed, A., Rushworth, J.V., Hirst, N.A. & Millner, P.A. Biosensors for whole-cell bacterial detection. Clin. Microbiol. Rev. 27, 631–646 (2014).CrossRefGoogle Scholar
  9. 9.
    Toh, S.Y., Citartan, M., Gopinath, S.C. & Tang, T.-H. Aptamers as a replacement for antibodies in enzyme-linked immunosorbent assay. Biosens. Bioelectron. 64, 392–403 (2015).CrossRefGoogle Scholar
  10. 10.
    Chen, A. & Yang, S. Replacing antibodies with aptamers in lateral flow immunoassay. Biosens. Bioelectron. 71, 230–242 (2015).CrossRefGoogle Scholar
  11. 11.
    Kong, M. et al. A novel and highly specific phage endolysin cell wall binding domain for detection of Bacillus cereus. Eur. Biophys. J. 44, 437–446 (2015).CrossRefGoogle Scholar
  12. 12.
    Lim, T., Lee, S.Y., Yang, J., Hwang, S.Y. & Ahn, Y. Microfluidic biochips for simple impedimetric detection of thrombin based on label-free DNA aptamers. BioChip J. 11, 109–115 (2017).CrossRefGoogle Scholar
  13. 13.
    Joshi, R. et al. Selection, characterization, and application of DNA aptamers for the capture and detection of Salmonella enterica serovars. Mol. Cell. Probes 23, 20–28 (2009).CrossRefGoogle Scholar
  14. 14.
    Kretzer, J.W. et al. Use of high-affinity cell wall-binding domains of bacteriophage endolysins for immobilization and separation of bacterial cells. Appl. Environ. Microbiol. 73, 1992–2000 (2007).CrossRefGoogle Scholar
  15. 15.
    Kong, M., Shin, J.H., Heu, S., Park, J.-K. & Ryu, S. Lateral flow assay-based bacterial detection using engineered cell wall binding domains of a phage endolysin. Biosens. Bioelectron. 96, 173–177 (2017).CrossRefGoogle Scholar
  16. 16.
    Molin, O., Nilsson, L. & Anséhn, S. Rapid detection of bacterial growth in blood cultures by bioluminescent assay of bacterial ATP. J. Clin. Microbiol. 18, 521–525 (1983).Google Scholar
  17. 17.
    Nilsson, L., Molin, Ö. & Ånséhn, S. Bioluminescent assay of bacterial ATP for rapid detection of bacterial growth in clinical blood cultures. Luminescence 3, 101–104 (1989).Google Scholar
  18. 18.
    Park, C. et al. 3D-printed microfluidic magnetic preconcentrator for the detection of bacterial pathogen using an ATP luminometer and antibody-conjugated magnetic nanoparticles. J. Microbiol. Methods 132, 128–133 (2017).Google Scholar
  19. 19.
    Arroyo, M.G. et al. Effectiveness of ATP bioluminescence assay for presumptive identification of microorganisms in hospital water sources. BMC Infect. Dis. 17, 458 (2017).CrossRefGoogle Scholar
  20. 20.
    Wright, D., Chapman, P. & Siddons, C. Immunomagnetic separation as a sensitive method for isolating Escherichia coli O157 from food samples. Epidemiol. Infect. 113, 31–39 (1994).CrossRefGoogle Scholar
  21. 21.
    Skjerve, E. & Olsvik, Ø. Immunomagnetic separation of Salmonella from foods. Int. J. Food Microbiol. 14, 11–17 (1991).CrossRefGoogle Scholar
  22. 22.
    Aydin, M. et al. Rapid and Sensitive Detection of Escherichia coli O157:H7 in Milk and Ground Beef Using Magnetic Bead–Based Immunoassay Coupled with Tyramide Signal Amplification. J. Food Prot. 77, 100–105 (2014).CrossRefGoogle Scholar
  23. 23.
    Lee, J., Park, C., Kim, Y. & Park, S. Signal enhancement in ATP bioluminescence to detect bacterial pathogens via heat treatment. BioChip J. 11, 287–293 (2017).CrossRefGoogle Scholar
  24. 24.
    Ganesh, I. et al. An integrated microfluidic PCR system with immunomagnetic nanoparticles for the detection of bacterial pathogens. Biomed. Microdevices 18, 116 (2016).CrossRefGoogle Scholar
  25. 25.
    Kong, M., Kim, M. & Ryu, S. Complete genome sequence of Bacillus cereus bacteriophage PBC1. J. Virol. 86, 6379–6380 (2012).CrossRefGoogle Scholar
  26. 26.
    Pal, S., Alocilja, E.C. & Downes, F.P. Nanowire labeled direct-charge transfer biosensor for detecting Bacillus species. Biosens. Bioelectron. 22, 2329–2336 (2007).CrossRefGoogle Scholar
  27. 27.
    Oda, M. et al. Role of sphingomyelinase in infectious diseases caused by Bacillus cereus. PLoS ONE 7, e38054 (2012).CrossRefGoogle Scholar
  28. 28.
    Mastronardi, C., Yang, L., Halpenny, M., Toye, B. & Ramírez-Arcos, S. Evaluation of the sterility testing process of hematopoietic stem cells at Canadian Blood Services. Transfusion 52, 1778–1784 (2012).CrossRefGoogle Scholar

Copyright information

© The Korean BioChip Society and Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Department of Biomedical EngineeringSungkyunkwan UniversitySuwonRepublic of Korea
  2. 2.Department of Agricultural BiotechnologySeoul National UniversitySeoulRepublic of Korea
  3. 3.Department of Food Science and BiotechnologyKyungHee UniversityYonginRepublic of Korea
  4. 4.School of Mechanical EngineeringSungkyunkwan UniversitySuwonRepublic of Korea

Personalised recommendations