Biofilms are structured communities of bacterial cells enclosed in a self-produced polymeric matrix and adherent to an inert or living surface; they have clinical, industrial and environmental impacts. Biofilms that are established by bacteria on plants are found on the surfaces of roots, leaves, seeds and internal vascular tissues where the microbes live in commensal, mutualistic or parasitic/pathogenic associations with their host. The study of the structure of plant-associated biofilms has been considerably helped by the development of techniques using fluorescent markers coupled with confocal scanning laser microscopy as well as scanning electron microscopy. We review several of these techniques as well as some of the research that has dealt with plant-associated biofilms. Our investigations focus on biofilm formation in the early stages of the Rhizobium-legume symbiosis, in which Gram-negative rhizobia provide fixed nitrogen to a host legume, and in return, the legume provides carbon-containing molecules. Because root colonization is an important early step in the establishment of the nitrogen-fixing symbiosis, we looked at Sinorhizobium meliloti attachment and biofilm establishment on the roots of its legume hosts, Medicago sativa L. and Melilotus alba Desr. We also examined biofilm formation by Rhizobium leguminosarum bv. viciae on the roots of Arabidopsis thaliana ( L.) Heynh., a non-legume and non-host. Our ultimate goal is to characterize the rhizobial genes involved in aggregation and attachment to roots because several of these appear to be shared in biofilm formation and rhizobial entry of legume root cells. (c) 2006 The Linnean Society of London, Botanical Journal of the Linnean Society, 2006, 150, 79-88.