Investigation of Biofilm Formation in Yersinia pestis

  • Haihong Fang
Part of the Springer Protocols Handbooks book series (SPH)


The formation of a biofilm, which often grows as an attached extracellular matrix and blocks the flea proventriculus, enhances flea-borne transmission of Yersinia pestis during flea biting. Quantitative investigation of biofilm formation can be used to evaluate Y. pestis transmission in vitro. This chapter reviews methods used to investigate Y. pestis biofilm formation. These methods can be divided into three types based on (i) direct observation of biofilm formation including biofilm colony, crystal violet staining, and Caenorhabditis elegans biofilm assay, (ii) quantitative evaluation of the expression of genes related to biofilm formation by real-time polymerase chain reaction, and (iii) quantification of intracellular cyclic diguanylic acid (c-di-GMP) concentration, which positively correlates with Yersinia biofilm formation.

Key words

Crystal violet staining Biofilm formation c-di-GMP Caenorhabditis elegans 


  1. 1.
    Hobley L, Harkins C, MacPhee CE, Stanley-Wall NR (2015) Giving structure to the biofilm matrix: an overview of individual strategies and emerging common themes. FEMS Microbiol Rev 39(5):649–669CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Hall-Stoodley L, Costerton JW, Stoodley P (2004) Bacterial biofilms: from the natural environment to infectious diseases. Nat Rev Microbiol 2(2):95–108CrossRefPubMedGoogle Scholar
  3. 3.
    Donlan RM (2008) Biofilms on central venous catheters: is eradication possible? Curr Top Microbiol Immunol 322:133–161PubMedGoogle Scholar
  4. 4.
    Hatt JK, Rather PN (2008) Role of bacterial biofilms in urinary tract infections. Curr Top Microbiol Immunol 322:163–192PubMedGoogle Scholar
  5. 5.
    Kouidhi B, Al Qurashi YM, Chaieb K (2015) Drug resistance of bacterial dental biofilm and the potential use of natural compounds as alternative for prevention and treatment. Microb Pathog 80:39–49CrossRefPubMedGoogle Scholar
  6. 6.
    Wu C, Cheng YY, Yin H, Song XN, Li WW, Zhou XX, Zhao LP, Tian LJ, Han JC, Yu HQ (2013) Oxygen promotes biofilm formation of Shewanella putrefaciens CN32 through a diguanylate cyclase and an adhesin. Sci Rep 3:1945CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Yarwood JM, Bartels DJ, Volper EM, Greenberg EP (2004) Quorum sensing in Staphylococcus aureus biofilms. J Bacteriol 186(6):1838–1850CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Yu D, Zhao L, Xue T, Sun B (2012) Staphylococcus aureus autoinducer-2 quorum sensing decreases biofilm formation in an icaR-dependent manner. BMC Microbiol 12:288CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Beenken KE, Dunman PM, McAleese F, Macapagal D, Murphy E, Projan SJ, Blevins JS, Smeltzer MS (2004) Global gene expression in Staphylococcus aureus biofilms. J Bacteriol 186(14):4665–4684CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Vadyvaloo V, Jarrett C, Sturdevant DE, Sebbane F, Hinnebusch BJ (2010) Transit through the flea vector induces a pretransmission innate immunity resistance phenotype in Yersinia pestis. PLoS Pathog 6(2):e1000783CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Zhou D, Yang R (2011) Formation and regulation of Yersinia biofilms. Protein Cell 2(3):173–179CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Sun YC, Koumoutsi A, Darby C (2009) The response regulator PhoP negatively regulates Yersinia pseudotuberculosis and Yersinia pestis biofilms. FEMS Microbiol Lett 290(1):85–90CrossRefPubMedGoogle Scholar
  13. 13.
    Darby C, Hsu JW, Ghori N, Falkow S (2002) Caenorhabditis elegans: plague bacteria biofilm blocks food intake. Nature 417(6886):243–244CrossRefPubMedGoogle Scholar
  14. 14.
    Rempe KA, Hinz AK, Vadyvaloo V (2012) Hfq regulates biofilm gut blockage that facilitates flea-borne transmission of Yersinia pestis. J Bacteriol 194(8):2036–2040CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Fang N, Yang H, Fang H, Liu L, Zhang Y, Wang L, Han Y, Zhou D, Yang R (2015) RcsAB is a major repressor of Yersinia biofilm development through directly acting on hmsCDE, hmsT, and hmsHFRS. Sci Rep 5:9566CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Ren GX, Yan HQ, Zhu H, Guo XP, Sun YC (2014) HmsC, a periplasmic protein, controls biofilm formation via repression of HmsD, a diguanylate cyclase in Yersinia pestis. Environ Microbiol 16(4):1202–1216CrossRefPubMedGoogle Scholar
  17. 17.
    Darby C, Ananth SL, Tan L, Hinnebusch BJ (2005) Identification of gmhA, a Yersinia pestis gene required for flea blockage, by using a Caenorhabditis elegans biofilm system. Infect Immun 73(11):7236–7242CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Nature Singapore Pte Ltd. 2018

Authors and Affiliations

  • Haihong Fang
    • 1
    • 2
  1. 1.State Key Laboratory of Pathogen and BiosecurityBeijing Institute of Microbiology and EpidemiologyBeijingChina
  2. 2.Beijing AcademyBeijingChina

Personalised recommendations