Advertisement

Campylobacter jejuni Isolation/Enumeration from Environmental Samples

  • Kelli L. HiettEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1512)

Abstract

Currently, there is no universally accepted standard media or method for the recovery of Campylobacter species. This is likely due to the ubiquity of the organism in nature, the complex sample matrices from which the organism is often recovered, as well as the fragile/viable-but nonculturable state the organism assumes in response to stress. The use of a sterile filter placed upon a nonselective Brucella Agar Blood Plate (BAB), followed by incubation at 37 °C in a hydrogen-containing atmosphere (Campycheck), is one method to recover stressed and emerging Campylobacter spp. from complex environmental matrices; however, this technique does not currently allow for the enumeration of the recovered organisms. Enumeration is performed using serial dilutions of sample homogenate plated onto modified Campy-Cefex media followed by incubation at either 37 °C or 42 °C in a microaerobic atmosphere.

Key words

Campylobacter Cape Town Campycheck VBNC Hydrogen Environmental Zoonotic pathogen 

Supplementary material

Video S1

Campycheck filter placement: Aseptic placement of 0.6 μm filter onto BAB nonselective plate (MP4 24383 kb)

Video S2

Campycheck sample application (MP4 32054 kb)

Video S3

Campycheck filter removal (MP4 31928 kb)

Video S4

Campycheck gas addition using ZipTop bags (MP4 66158 kb)

Video S5

Campycheck gas addition using gas generation system (MP4 48461 kb)

References

  1. 1.
    Agunos A, Waddell L, Leger D et al (2014) A systematic review characterizing on-farm sources of Campylobacter spp. for broiler chickens. PLoS One 9(8):e104905. doi: 10.1371/journal.pone.0104905 CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Silley P (2003) Campylobacter and fluoroquinolones: a bias data set? Environ Microbiol 5(4):219–230. doi: 10.1046/j.1462-2920.2003.00425.x CrossRefPubMedGoogle Scholar
  3. 3.
    Hofreuter D (2014) Defining the metabolic requirements for the growth and colonization capacity of Campylobacter jejuni. Front Cell Infect Nicrobiol 4:137. doi: 10.3389/fcimb.2014.00137 Google Scholar
  4. 4.
    Ugarte-Ruiz M, Gómez-Barrero S, Porrero MC et al (2012) Evaluation of four protocols for the detection and isolation of thermophilic Campylobacter from different matrices. J Appl Microbiol 113(1):200–208. doi: 10.1111/j.1365-2672.2012.05323.x CrossRefPubMedGoogle Scholar
  5. 5.
    Corry JE, Post DE, Colin P et al (1995) Culture media for the isolation of campylobacters. Int J Food Microbiol 26(1):43–76CrossRefPubMedGoogle Scholar
  6. 6.
    Gharst G, Oyarzabal OA, Hussain SK (2013) Review of current methodologies to isolate and identify Campylobacter spp. from foods. J Microbiol Methods 95(1):84–92. doi: 10.1016/j.mimet.2013.07.014 CrossRefPubMedGoogle Scholar
  7. 7.
    Duarte A, Botteldoorn N, Coucke W et al (2015) Effect of exposure to stress conditions on propidium monoazide (PMA)-qPCR based Campylobacter enumeration in broiler carcass rinses. Food Microbiol 48:182–190. doi: 10.1016/j.fm.2014.12.011 CrossRefPubMedGoogle Scholar
  8. 8.
    Kim SA, Lee YM, Hwang IG et al (2009) Eight enrichment broths for the isolation of Campylobacter jejuni from inoculated suspensions and ground pork. Lett Appl Microbiol 49(5):620–626. doi: 10.1111/j.1472-765X.2009.02714.x CrossRefPubMedGoogle Scholar
  9. 9.
    Tangvatcharin P, Chanthachum S, Kopaiboon P et al (2005) Comparison of methods for the isolation of thermotolerant Campylobacter from poultry. J Food Prot 68(3):616–620CrossRefPubMedGoogle Scholar
  10. 10.
    Williams LK, Sait LC, Cogan TA et al (2012) Enrichment culture can bias the isolation of Campylobacter subtypes. Epidemiol Infect 140(7):1227–1235. doi: 10.1017/s0950268811001877 CrossRefPubMedGoogle Scholar
  11. 11.
    de Boer P, Rahaoui H, Leer RJ et al (2015) Real-time PCR detection of Campylobacter spp.: a comparison to classic culturing and enrichment. Food Microbiol 51:96–100. doi: 10.1016/j.fm.2015.05.006 CrossRefPubMedGoogle Scholar
  12. 12.
    Magajna B, Schraft H (2015) Evaluation of propidium monoazide and quantitative PCR to quantify viable Campylobacter jejuni biofilm and planktonic cells in log phase and in a viable but nonculturable state. J Food Prot 78(7):1303–1311. doi: 10.4315/0362-028x.jfp-14-583 CrossRefPubMedGoogle Scholar
  13. 13.
    Jokinen CC, Koot JM, Carrillo CD et al (2012) An enhanced technique combining pre-enrichment and passive filtration increases the isolation efficiency of Campylobacter jejuni and Campylobacter coli from water and animal fecal samples. J Microbiol Methods 91(3):506–513. doi: 10.1016/j.mimet.2012.09.005 CrossRefPubMedGoogle Scholar
  14. 14.
    Simmons M, Hiett KL, Stern NJ et al (2008) Comparison of poultry exudate and carcass rinse sampling methods for the recovery of Campylobacter spp. subtypes demonstrates unique subtypes recovered from exudate. J Microbiol Methods 74(2-3):89–93. doi: 10.1016/j.mimet.2008.03.007 CrossRefPubMedGoogle Scholar
  15. 15.
    Acke E, McGill K, Golden O et al (2009) A comparison of different culture methods for the recovery of Campylobacter species from pets. Zoonoses Public Health 56(9-10):490–495. doi: 10.1111/j.1863-2378.2008.01205.x CrossRefPubMedGoogle Scholar
  16. 16.
    Koziel M, Corcoran GD, Sleator RD et al (2014) Detection and molecular analysis of Campylobacter ureolyticus in domestic animals. Gut Pathogens 6:9. doi: 10.1186/1757-4749-6-9 CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Fitzgerald C, Tu ZC, Patrick M et al (2014) Campylobacter fetus subsp. testudinum subsp. nov. isolated from humans and reptiles. Int J Syst Evol Microbiol 64(Pt 9):2944–2948. doi: 10.1099/ijs.0.057778-0
  18. 18.
    Chaban B, Ngeleka M, Hill JE (2010) Detection and quantification of 14 Campylobacter species in pet dogs reveals an increase in species richness in feces of diarrheic animals. BMC Microbiol 10:73. doi: 10.1186/1471-2180-10-73 CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Kaakoush NO, Mitchell HM, Man SM (2014) Role of emerging Campylobacter species in inflammatory bowel diseases. Inflamm Bowel Dis 20(11):2189–2197. doi: 10.1097/mib.0000000000000074 CrossRefPubMedGoogle Scholar
  20. 20.
    Casanova C, Schweiger A, von Steiger N et al (2015) Campylobacter concisus pseudo-outbreak caused by improved culture conditions. J Clin Microbiol 53(2):660–662. doi: 10.1128/jcm.02608-14 CrossRefPubMedGoogle Scholar
  21. 21.
    Lastovica AJ, le Roux E (2000) Efficient isolation of campylobacteria from stools. J Clin Microbiol 38(7):2798–2799PubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2017

Authors and Affiliations

  1. 1.Poultry Microbiological Safety and Processing Research Unit, United States National Poultry Research Center, Agricultural Research ServiceU.S. Department of AgricultureAthensUSA

Personalised recommendations