Skip to main content

Detection of Campylobacter DNA using magnetic nanoparticles coupled with PCR and a colorimetric end-point system

Abstract

Campylobacter is an important food-borne pathogen causing acute gastroenteritis worldwide. Magnetic nanoparticle-based PCR coupled with streptavidin-horseradish peroxidase and a substrate was used for colorimetric detection. Forward primers conjugated to magnetic nanoparticles facilitated separation and concentration of Campylobacter DNA in a sample matrix. After PCR, a green color developed and was observed using the unaided eye, or detected using a spectrophotometer. High specificity and sensitivity of the 100 fg DNA/PCR reaction were achieved in pure culture experiments. The technique was applied for detection of Campylobacter on naturally contaminated chicken skin. All positive results were in agreement with results achieved using a conventional culture method. The magnetic nanoparticle-PCR-enzyme linked gene assay was practical and useful for detection of Campylobacter in complex matrices with PCR-interfering substances.

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

References

  1. 1.

    Butzler JP. Campylobacter from obscurity to celebrity. Clin. Microbiol. Infec. 10: 868–876 (2004)

    Article  Google Scholar 

  2. 2.

    Hughes RA, Rees JH. Clinical and epidemiologic features of Guillain-Barré syndrome. J. Infect. Dis. 176: 92–98 (1997)

    Article  Google Scholar 

  3. 3.

    Hannu T, Mattlia L, Rautelin H, Pelkonen P, Lahdenne P, Siitonen A, Leirisalo-Repo M. Campylobacter-triggered reactive arthritis: A population-based study. Rheumatolog. 41: 312–318 (2002)

    Article  CAS  Google Scholar 

  4. 4.

    Young KT, Davis LM, Dirita VJ. Campylobacter jejuni: Molecular biology and pathogenesis. Nat. Rev. Microbiol. 5: 665–679 (2007)

    Article  CAS  Google Scholar 

  5. 5.

    Vindigni SM, Srijan A, Wongstitwilairoong B, Marcus R, Meek J, Riley PL, Mason C. Prevalence of foodborne microorganisms in retail foods in Thailand. Foodborne Pathog. Dis. 4: 208–215 (2007)

    Article  Google Scholar 

  6. 6.

    Suzuki H, Yamamoto S. Campylobacter contamination in retail poultry meats and by-products in the world: A literature survey. J. Vet. Med. Sci. 71: 255–261 (2009)

    Article  Google Scholar 

  7. 7.

    ISO 10272-1. Microbiology of food and animal feeding stuffs-Horizontal method for detection and numeration of Campylobacter spp.-Part 1: Detection method. International Standard Organization, Geneva, Switzerland (2006)

  8. 8.

    Bang DD, Pedersen K, Madsen M. Development of a PCR assay suitable for Campylobacter spp. mass screening programs in broiler production. J. Rapid Meth. Aut. Mic. 9: 97–113 (2001)

    Article  CAS  Google Scholar 

  9. 9.

    Bang DD, Wedderkopp A, Pedersen K, Madsen M. Rapid PCR using nested primers of the 16S rRNA and the hippuricase (hipO) genes to detect Campylobacter jejuni and Campylobacter coli in environmental samples. Mol. Cell. Probe. 16: 359–369 (2002)

    Article  CAS  Google Scholar 

  10. 10.

    Persson S, Petersen HM, Jespersgaard C, Olsen KEP. Realtime TaqMan polymerase chain reaction-based genus-identification and pyrosequencing based species identification of Campylobacter jejuni, C. coli, C. lari, C. upsaliensis, and C. fetus directly on stool samples. Diagn. Micr. Infec. Dis. 74: 6–10 (2012)

    Article  CAS  Google Scholar 

  11. 11.

    Federighi M, Tholozan J, Cappelier JM, Tissier JP, Jouve JL. Evidence of noncoccoid viable but non-culturable Campylobacter jejuni cells in microcosm water by direct viable count, CTC-DAPI double staining, and scaning electron microscopy. Food Microbiol. 15: 539–550 (1998)

    Article  Google Scholar 

  12. 12.

    Adler M, Wacker R, Niemeyer CM. Sensitivity by combination: Immuno-PCR and related technologies. Analys. 133: 702–718 (2008)

    Article  CAS  Google Scholar 

  13. 13.

    Hong Y, Berrang ME, Liu T, Hofacre CL, Sanchez S, Wang L, Maurer JJ. Rapid detection of Campylobacter coli, C. jejuni, and Salmonella enterica on poultry carcasses by using PCR-enzyme-linked immunosorbent assay. Appl. Environ. Microb. 69: 3492–3499 (2003)

    Article  CAS  Google Scholar 

  14. 14.

    Safarik I, Safarikova M. Magnetic nano-and microparticles in biotechnology. Chem. Paper. 63: 497–505 (2009)

    Article  CAS  Google Scholar 

  15. 15.

    Yu LSL, Unkalis J, Tu SI. Immunomagnetic separation methods for the isolation of Campylobacter jejuni from ground poultry meats. J. Immunol. Method. 256: 11–18 (2001)

    Article  CAS  Google Scholar 

  16. 16.

    Morales-Rayas R, Wolffs PFG, Griffiths MW. Immunocapture and real-time PCR to detect Campylobacter spp. J. Food Protect. 71: 2543–2547 (2008)

    CAS  Google Scholar 

  17. 17.

    Adedayo O, Kirkpatrick BD. Campylobacter jejuni infections: Update on presentation, diagnosis, and management. Hosp. Physicia. 44: 9–15 (2008)

    Google Scholar 

  18. 18.

    Josefen MH, Lübek PS, Hansen F, Hoorfar J. Toward an international standard for PCR-based detection of foodborne thermotolerant campylobacters: Interaction of enrichment media and pre-PCR treatment on carcass rinse samples. J. Microbiol. Meth. 58: 39–48 (2004)

    Article  Google Scholar 

  19. 19.

    Walsh MK, Wang X, Weimer BC. Optimizing the immobilization of singlestranded DNA onto glass beads. J. Biochem. Bioph. Meth. 47: 221–231 (2001)

    Article  CAS  Google Scholar 

  20. 20.

    Li H, He Z. Magnetic bead-based DNA hybridization assay with chemiluminescence and chemiluminescent imaging detection. Analys. 134: 800–804 (2009)

    Article  CAS  Google Scholar 

  21. 21.

    Amagliani G, Omicciolio E, Del Campo, A, Bruce IJ, Brandi G, Magnani M. Development of a magnetic capture hybridization-PCR assay for Listeria monocytogenes direct detection in milk samples. J. Appl. Microbiol. 100: 375–383 (2006)

    Article  CAS  Google Scholar 

  22. 22.

    Parkhill J, Wren BW, Mungall K, Ketley JM, Churcher C, Basham D, Chillingwort T, Davies RM, Feltwell T, Holroyd S, Jagels K, Karlyshev AV, Moule S, Pallen MJ, Penn CW, Quail MA, Rajandream MA, Rutherford KM, van Vliet AH, Whitehead S, Barrel BG. The genome sequence of the food-borne pathogen Campylobacter jejuni reveals hypervariable sequences. Natur. 403: 665–668 (2000)

    Article  CAS  Google Scholar 

  23. 23.

    Jangpatarapongsa K, Polpanch D, Yamkamon V, Diththarot Y, Peng-On J, Thiramanas R, Hongseng S, Jootar S, Charoenmak L, Tangboriboonrat P. DNA detection of chronic myelogenous leukemia by magnetic nanoparticles. Analys. 136: 354–358 (2011)

    Article  CAS  Google Scholar 

  24. 24.

    Sails AD, Fox AJ, Bolton FJ, Wareing DRA, Greenway DLA, Borrow R. Development of a PCR ELISA assay for the identification of Campylobacter jejuni and Campylobacter coli. Mol. Cell. Probe. 15: 291–300 (2001)

    Article  CAS  Google Scholar 

  25. 25.

    Grennan B, O’Sullivan NA, Fallon R, Carroll C, Smith T, Glennon M, Maher M. PCR-ELISAs for the detection of Campylobacter jejuni and Campylobacter coli in poultry samples. BioTechnique. 30: 602–610 (2001)

    CAS  Google Scholar 

  26. 26.

    Sails AD, Fox AJ, Bolton FJ, Wareing DRA, Greenway LA. A real-time PCR assay for the detection of Campylobacter jejuni in foods after enrichment culture. Appl. Environ. Microb. 69: 1383–1390 (2003)

    Article  CAS  Google Scholar 

  27. 27.

    Davis MA, Conner DE. Incidence of Campylobacter from raw, retail poultry products. Poultry Sci. 79(Suppl. 1): 54 (2000)

    Article  Google Scholar 

  28. 28.

    Davis MA, Conner DE. Survival of Campylobacter jejuni on poultry skin and meat at varying temperatures. Poultry Sci. 86: 765–767 (2007)

    Article  CAS  Google Scholar 

  29. 29.

    Sasaki Y, Haruna M, Mori T, Kusukawa M, Murakami M, Tsujuyama Y, Ito K, Toyofuku H, Yamada Y. Quantitative estimation of Campylobacter crosscontamination in carcasses and chicken products at an abattoir. Food Contro. 43: 10–17 (2014)

    Article  Google Scholar 

  30. 30.

    Al-soud WA, Jönsson LJ, Rådström P. Identification and characterization of immunoglobulin G in blood as a major inhibitor of diagnostic PCR. J. Clin. Microbiol. 38: 345–350 (2000)

    CAS  Google Scholar 

  31. 31.

    Al-soud WA, Rådström P. Purification and characterization of PCR inhibitory components in blood cells. J. Clin. Microbiol. 39: 485–493 (2001)

    Article  CAS  Google Scholar 

  32. 32.

    Rossen L, Nórskov P, Holmstrøm K, Rasmussen OF. Inhibition of PCR by components of food samples, microbial diagnostic assays and DNA-extraction solutions. Int. J. Food Microbiol. 17: 37–45 (1992)

    Article  CAS  Google Scholar 

  33. 33.

    Thunberg RL, Tran TT, Walderhaung MO. Detection of thermophilic Campylobacter spp. in blood-free enriched samples of inoculated foods by the polymerase chain reaction. J. Food Protect. 63: 299–303 (2000)

    CAS  Google Scholar 

  34. 34.

    Dickinson JH, Nkroll RG, Astori KA. The direct application of the polymerase chain reaction to DNA extracted from foods. Lett. Appl. Microbiol. 20: 212–216 (1995)

    Article  CAS  Google Scholar 

Download references

Author information

Affiliations

Authors

Corresponding author

Correspondence to Wijit Wonglumsom.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Jansaento, W., Jangpatarapongsa, K., Polpanich, D. et al. Detection of Campylobacter DNA using magnetic nanoparticles coupled with PCR and a colorimetric end-point system. Food Sci Biotechnol 25, 193–198 (2016). https://doi.org/10.1007/s10068-016-0029-3

Download citation

Keywords

  • Campylobacter
  • magnetic nanoparticle
  • colorimetric system
  • chicken