Abstract
The acidic exopolysaccharide (EPS) produced by Rhizobium leguminosarum bv. trifolii is required for the establishment of effective symbiosis with compatible host plants (Trifolium spp.). In the rhizobium-legume interaction, early stages of root infection and nodule development have been well studied from a genetic standpoint. However, factors important for colonization of several surfaces by rhizobia, including soil particles and roots, have not yet been thoroughly investigated. The aim of this study was establishing of environmental factors affecting production of EPS by R. leguminosarum bv. trifolii strain 24.2 and the role of this polysaccharide in bacterial surface properties and attachment ability. Besides the wild-type strain, its derivatives differing in the level of EPS produced were used to these analyses. The ability of attachment to abiotic and biotic surfaces of these strains were established using CFU counting experiments. Three-dimensional structure and other parameters of biofilms formed were characterized in confocal laser scanning microscopy. Electrokinetic (zeta) potential of rhizobial cells were determined using Laser Doppler Velocimetry. It was evidenced that the ability of R. leguminosarum bv. trifolii to produce EPS significantly affected bacterial attachment and biofilm formation on both abiotic and biotic surfaces. In addition, the presence of this polysaccharide influenced the zeta potential of rhizobial cells. Mutant strains having a mutation in genes involved in EPS synthesis were significantly impaired in attachment, whereas strains overproducing this polysaccharide showed higher adhesion efficiency to all of the tested materials. EPS facilitated attachment of bacterial cells to the tested surfaces most probably due to hydrophobic interactions and heterogeneity of the envelope surface. EPS produced by R. leguminosarum bv. trifolii plays a significant role in attachment and biofilm formation to both abiotic and biotic surfaces as well as bacterial surface properties.
Highlights
Rhizobium leguminosarum bv. trifolii is a nitrogenfixing bacterium that can either exist as a free-living organism in the soil or live in symbiotic association with clover (Trifolium spp.) (Fagerli and Svenning 2005; Duodu et al 2009)
EPS-deficient mutant strains of R. leguminosarum bvs. trifolii and viciae, and Sinorhizobium meliloti induce formation of empty or only partially infected nodulelike structures on roots of compatible host plants that are ineffective in nitrogen fixation (Ivashina et al 1994; Cheng and Walker 1998; Workum et al 1998; Janczarek and Rachwał 2013)
Extracellular polysaccharide secreted by R. leguminosarum bv. trifolii into the environment plays an essential role in the symbiotic interaction of this bacterium with its host plant, i.e. the clover (Trifolium pratense)
Summary
Rhizobium leguminosarum bv. trifolii is a nitrogenfixing bacterium that can either exist as a free-living organism in the soil or live in symbiotic association with clover (Trifolium spp.) (Fagerli and Svenning 2005; Duodu et al 2009). Besides flavonoids and Nod factors, rhizobial acidic exopolysaccharides (EPS) play a significant role in the establishment of an effective symbiosis especially with legumes that form indeterminate-type nodules (e.g. clover, pea, vetch, and alfalfa) (Rolfe et al 1996; Becker and Pühler 1998; Janczarek and Urbanik-Sypniewska 2013). This polysaccharide is indispensable for protection against host plant defence reactions and initiation and elongation of infection threads, special tubular structures via which rhizobia colonize root nodules (Cheng and Walker 1998; Workum et al 1998; Jaszek et al 2014).
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