Abstract
Invasive methicillin-resistant Staphylococcus aureus (MRSA) infections are often characterized by recalcitrance to antimicrobial therapy, which is a function not only of widespread antimicrobial resistance among clinical isolates, but also the capacity to form biofilms. Biofilms consist of ordered populations of bacterial colonies encased in a polysaccharide and/or proteinaceous matrix. This unique physiologic adaptation limits penetration of antimicrobial molecules and innate immune effectors to the infectious focus, increasing the likelihood of treatment failure and progression to chronic infection. Investigation of mechanisms of biofilm formation and dispersal, as well as the physiologic adaptations to the biofilm lifestyle, is therefore critical to developing new therapies to combat MRSA infections. In this chapter, we describe two in vitro methods for the investigation of staphylococcal biofilm formation, a microtiter plate-based assay of biofilm formation under static conditions and a flow cell-based assay of biofilm formation under fluid shear. We also detail an in vivo murine model of catheter-associated biofilm formation that is amenable to imaging and microbiologic analyses. Special consideration is given to the conditions necessary to support biofilm formation by clinical isolates of S. aureus.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Lewis K (2001) Riddle of biofilm resistance. Antimicrob Agents Chemother 45:999–1007
Keren I, Kaldalu N, Spoering A et al (2004) Persister cells and tolerance to antimicrobials. FEMS Microbiol Lett 230:13–18
Patti JM, Allen BL, McGavin MJ et al (1994) MSCRAMM-mediated adherence of microorganisms to host tissues. Annu Rev Microbiol 48:585–617
Sillanpaa J, Xu Y, Nallapareddy SR et al (2004) A family of putative MSCRAMMs from Enterococcus faecalis. Microbiology 150:2069–2078
Beenken KE, Blevins JS, Smeltzer MS (2003) Mutation of sarA in Staphylococcus aureus limits biofilm formation. Infect Immun 71:4206–4211
Cafiso V, Bertuccio T, Santagati M et al (2004) Presence of the ica operon in clinical isolates of Staphylococcus epidermidis and its role in biofilm production. Clin Microbiol Infect 10:1081–1088
Fitzpatrick F, Humphreys H, O’Gara JP (2005) The genetics of staphylococcal biofilm formation-will a greater understanding of pathogenesis lead to better management of device-related infection? Clin Microbiol Infect 11:967–973
Cramton SE, Gerke C, Schnell NF et al (1999) The intracellular adhesin (ica) locus is present in Staphylococcus aureus and is required for biofilm formation. Infect Immun 67:5427–5433
Beenken KE, Dunman PM, McAleese F et al (2004) Global gene expression in Staphylococcus aureus biofilms. J Bacteriol 186:4665–4684
Fitzpatrick F, Humpreys H, O’Gara JP (2005) Evidence for icaADBC-independent biofilm development mechanism in methicillin-resistant Staphylococcus aureus clinical isolates. J Clin Microbiol 43:1973–1976
O’Neill E, Pozzi C, Houston P et al (2007) Association between methicillin susceptibility and biofilm regulation in Staphylococcus aureus isolates from device-related infections. J Clin Microbiol 45:1379–1388
Rice KC, Mann EE, Endres JL et al (2007) The cidA murein hydrolase regulator contributes to DNA release and biofilm development in Staphylococcus aureus. Proc Natl Acad Sci USA 104:8113–8118
Fluckiger U, Ulrich M, Steinhuber A et al (2005) Biofilm formation, icaADBC transcription, and polysaccharide intercellular adhesin synthesis by staphylococci in a device-related infection model. Infect Immun 73:1811–1819
McKenney D, Pouliot KL, Wang Y et al (1999) Broadly protective vaccine for Staphylococcus aureus based on an in vivo-expressed antigen. Science 284:1523–1527
Yao Y, Sturdevandt DE, Otto M (2005) Genome wide analysis of gene expression in Staphylococcus epidermidis biofilms: insights into the pathophysiology of S. epidermidis biofilms and the role of phenol-soluble modulins in formation of biofilms. J Infect Dis 191:289–298
Yarwood JM, Bartels DJ, Volper EM et al (2004) Quorum sensing in Staphylococcus aureus biofilms. J Bacteriol 186:1838–1850
Vuong C, Gerke C, Somerville GA et al (2003) Quorum-sensing control of biofilm factors in Staphylococcus epidermidis. J Infect Dis 188:706–718
Schwartz K, Syed AK, Stephenson RE et al (2012) Functional amyloids composed of phenol soluble modulins stabilize Staphylococcus aureus biofilms. PLoS Pathog 8:e1002744
Boles BR, Thoendel M, Roth AJ et al (2010) Identification of genes involved in polysaccharide-independent Staphylococcus aureus biofilm formation. PLoS One 5:e10146
Mrak LN, Zielenska AK, Beenken KE et al (2012) saeRS and sarA act synergistically to repress protease production and promote biofilm formation in Staphylococcus aureus. PLoS One 7:e38453
Zielenska AK, Beenken KE, Mrak LN et al (2012) sarA-mediated repression of protease production plays a key role in the pathogenesis of Staphylococcus aureus USA300 isolates. Mol Microbiol 86:1183–1196
Kolodkin-Gal I, Romero D, Cao S et al (2010) D-amino acids trigger biofilm disassembly. Science 328:627–629
Christensen GD, Simpson WA, Bisno AL et al (1983) Experimental foreign body infections in mice challenged with slime-producing Staphylococcus epidermidis. Infect Immun 40:407–410
Rupp ME, Ulphani JS, Fey PD et al (1999) Characterization of Staphylococcus epidermidis polysaccharide intercellular adhesin/hemagglutinin in the pathogenesis of intravascular catheter-associated infection in a rat model. Infect Immun 67:2656–2659
Kadurugamuwa JL, Sin L, Albert E et al (2003) Direct continuous method for monitoring biofilm infection in a mouse model. Infect Immun 71:882–890
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2014 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Cassat, J.E., Smeltzer, M.S., Lee, C.Y. (2014). Investigation of Biofilm Formation in Clinical Isolates of Staphylococcus aureus . In: Ji, Y. (eds) Methicillin-Resistant Staphylococcus Aureus (MRSA) Protocols. Methods in Molecular Biology, vol 1085. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-664-1_12
Download citation
DOI: https://doi.org/10.1007/978-1-62703-664-1_12
Published:
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-62703-663-4
Online ISBN: 978-1-62703-664-1
eBook Packages: Springer Protocols