Abstract
Bacterial exopolysaccharides (EPS) often confer a survival advantage by protecting the cell against abiotic and biotic stresses, including host defensive factors. They are also main components of the extracellular matrix involved in cell–cell recognition, surface adhesion and biofilm formation. Biosynthesis of a growing number of EPS has been reported to be regulated by the ubiquitous second messenger c-di-GMP, which promotes the transition to a biofilm mode of growth in an intimate association with the eukaryotic host. Here we describe a strategy based on the combination of an approach to artificially increase the intracellular level of c-di-GMP in virtually any gram-negative bacteria with a high throughput screening (HTS) for the identification of monosaccharide composition and carbohydrate fingerprinting of novel EPS, or modified variants, that can be involved in host–bacteria interactions.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Whitfield GB, Marmont LS, Howell L (2015) Enzymatic modifications of exopolysaccharides enhance bacterial persistence. Front Microbiol 6:471
Gunn JS, Bakaletz LO, Wozniak DJ (2016) What’s on the outside matters: the role of the extracellular polymeric substance of gram-negative biofilms in evading host immunity and as a target for therapeutic intervention. J Biol Chem 291(24):12538–12546
Killiny N, Martínez RH, Dumenyo CK et al (2013) The exopolysaccharide of Xylella fastidiosa is essential for biofilm formation, plant virulence, and vector transmission. Mol Plant-Microbe Interact 26:1044–1053
Baker P, Hill PJ, Snarr BD et al (2016) Exopolysaccharide biosynthetic glycoside hydrolases can be utilized to disrupt and prevent Pseudomonas aeruginosa biofilms. Sci Adv 2:e1501632
Yu S, Su T, Wu H et al (2015) PslG, a self-produced glycosyl hydrolase, triggers biofilm disassembly by disrupting exopolysaccharide matrix. Cell Res 25:1352–1367
Kawaharada Y, Kelly S, Nielsen MW et al (2015) Receptor-mediated exopolysaccharide perception controls bacterial infection. Nature 523:308–312
Skorupska A, Janczarek M, Marczak M et al (2006) Rhizobial exopolysaccharides: genetic control and symbiotic functions. Microb Cell Factories 5:7
Römling U, Galperin MY, Gomelsky M (2013) Cyclic di-GMP: the first 25 years of a universal bacterial second messenger. Microbiol Mol Biol Rev 77:1–52
Römling U, Gomelsky M, Galperin MY (2005) C-di-GMP: the dawning of a novel bacterial signalling system. Mol Microbiol 57:629–639
Galperin MY, Higdon R, Kolker E (2010) Interplay of heritage and habitat in the distribution of bacterial signal transduction systems. Mol BioSyst 6:721–728
Caly DL, Bellini D, Walsh MA et al (2015) Targeting cyclic di-GMP signalling: a strategy to control biofilm formation? Curr Pharm Des 21:12–24
Liang ZX (2015) The expanding roles of c-di-GMP in the biosynthesis of exopolysaccharides and secondary metabolites. Nat Prod Rep 32:663–683
Schmid J, Sieber V, Rehm B (2015) Bacterial exopolysaccharides: biosynthesis pathways and engineering strategies. Front Microbiol 6:496
Ryan RP, Fouhy Y, Lucey JF et al (2007) Cyclic di-GMP signalling in the virulence and environmental adaptation of Xanthomonas campestris. Mol Microbiol 63:429–442
Chatterjee S, Killiny N, Almeida RP et al (2010) Role of cyclic di-GMP in Xylella fastidiosa biofilm formation, plant virulence, and insect transmission. Mol Plant-Microbe Interact 23:1356–1363
Ross P, Weinhouse H, Aloni Y et al (1987) Regulation of cellulose synthesis in Acetobacter xylinum by cyclic diguanylic acid. Nature 325:279–281
Pérez-Mendoza D, Sanjuán J (2016) Exploiting the commons: cyclic diguanylate regulation of bacterial exopolysaccharide production. Curr Opin Microbiol 30:36–43
Nwodo UU, Green E, Okoh AI (2012) Bacterial exopolysaccharides: functionality and prospects. Int J Mol Sci 13:14002–14015
Rühmann B, Schmid J, Sieber V (2015) Methods to identify the unexplored diversity of microbial exopolysaccharides. Front Microbiol 6:565
Bylund J, Burgess LA, Cescutti P et al (2006) Exopolysaccharides from Burkholderia cenocepacia inhibit neutrophil chemotaxis and scavenge reactive oxygen species. J Biol Chem 281:2526–2532
Schmid J, Sieber V (2015) Enzymatic transformations involved in the biosynthesis of microbial exo-polysaccharides based on the assembly of repeat units. Chembiochem 16:1141–1147
Wilson RP, Winter SE, Spees AM et al (2011) The Vi capsular polysaccharide prevents complement receptor 3-mediated clearance of Salmonella enterica serotype Typhi. Infect Immun 79:830–837
Marshall JM, Gunn JS (2015) The O-antigen capsule of Salmonella enterica serovar Typhimurium facilitates serum resistance and surface expression of FliC. Infect Immun 83:3946–3959
Pérez-Mendoza D, Coulthurst SJ, Sanjuán J et al (2011) N-Acetylglucosamine-dependent biofilm formation in Pectobacterium atrosepticum is cryptic and activated by elevated c-di-GMP levels. Microbiology 157:3340–3348
Pérez-Mendoza D, Rodríguez-Carvajal MA, Romero-Jiménez L et al (2015) Novel mixed-linkage beta-glucan activated by c-di-GMP in Sinorhizobium meliloti. Proc Natl Acad Sci U S A 112:E757–E765
Rühmann B, Schmid J, Sieber V (2015) High throughput exopolysaccharide screening platform: from strain cultivation to monosaccharide composition and carbohydrate fingerprinting in one day. Carbohydr Polym 122:212–220
Romero-Jiménez L, Rodríguez-Carbonell D, Gallegos MT et al (2015) Mini-Tn7 vectors for stable expression of diguanylate cyclase PleD* in Gram-negative bacteria. BMC Microbiol 15:190
Waddell CS, Craig NL (1989) Tn7 transposition: recognition of the attTn7 target sequence. Proc Natl Acad Sci U S A 86:3958–3962
Bao Y, Lies DP, Fu H et al (1991) An improved Tn7-based system for the single-copy insertion of cloned genes into chromosomes of gram-negative bacteria. Gene 109:167–168
Peters JE, Craig NL (2001) Tn7: smarter than we thought. Nat Rev Mol Cell Biol 2:806–814
Choi KH, Schweizer HP (2006) mini-Tn7 insertion in bacteria with single attTn7 sites: example Pseudomonas aeruginosa. Nat Protocol 1:153–161
Rühmann B, Schmid J, Sieber V (2016) Automated modular high throughput exopolysaccharide screening platform coupled with highly sensitive carbohydrate fingerprint analysis. J Vis Exp 110:e53249
Demarre G, Guerout AM, Matsumoto-Mashimo C et al (2005) A new family of mobilizable suicide plasmids based on broad host range R388 plasmid (IncW) and RP4 plasmid (IncP alpha) conjugative machineries and their cognate Escherichia coli host strains. Res Microbiol 156:245–255
Blatny JM, Brautaset T, Winther-Larsen HC et al (1997) Construction and use of a versatile set of broad-host-range cloning and expression vectors based on the RK2 replicon. Appl Environ Microbiol 63:370–379
Pérez-Mendoza D, Aragón IM, Prada-Ramírez HA et al (2014) Responses to elevated c-di-GMP levels in mutualistic and pathogenic plant-interacting bacteria. PLoS One 9:e91645
Rühmann B, Schmid J, Sieber V (2014) Fast carbohydrate analysis via liquid chromatography coupled with ultra violet and electrospray ionization ion trap detection in 96-well format. J Chromatogr A 1350:44–50
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2018 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Schmid, J., Rühmann, B., Sieber, V., Romero-Jiménez, L., Sanjuán, J., Pérez-Mendoza, D. (2018). Screening of c-di-GMP-Regulated Exopolysaccharides in Host Interacting Bacteria. In: Medina, C., López-Baena, F. (eds) Host-Pathogen Interactions. Methods in Molecular Biology, vol 1734. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7604-1_21
Download citation
DOI: https://doi.org/10.1007/978-1-4939-7604-1_21
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-7603-4
Online ISBN: 978-1-4939-7604-1
eBook Packages: Springer Protocols