Evaluation of fluorescently labeled lectins for noninvasive localization of extracellular polymeric substances in Sphingomonas biofilms

Three strains of Sphingomonas were grown as biofilms and tested for binding of five fluorescently labeled lectins (Con A-type IV-TRITC or -Cy5, Pha-E- TRITC, PNA-TRITC, UEA 1-TRITC, and WGA-Texas red). Only ConA and WGA were s ignificantly bound by the biofilms. Binding of the live lectins to artifici al biofilms made of the commercially available Sphinogomonas extracellular polysaccharides was similar to binding to living biofilms, Staining of the living and artificial biofilms by ConA might be explained as binding of the lectin to the terminal mannosyl and terminal glucosyl residues in the poly saccharides secreted by Sphingomonas as well as to the terminal mannosyl re sidue in glycosphingolipids. Staining of the biofilms by WGA could only be explained as binding to the Sphingomonas glycosphingolipid membrane, bindin g to the cell wall, or nonspecific binding. Glycoconjugation of ConA and WG A with the target sugars glucose and N-acetylglucosamine, respectively, was used as a method for evaluation of the specificity of the lectin; towards Sphingomonas biofilms and Sphingomonas polysaccharides. Our results show th at the binding of lectins to biofilms does not necessarily prove the presen ce of specific target sugars in the extracellular polymeric substances (EPS ) in biofilms, The lectins may bind to non-EPS targets or adhere nonspecifi cally to components of the biofilm matrix.