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Background  The synthesis of cellulose is among the most important but poorly understood biochemical processes, especially in bacteria, due to its complexity and high degree of regulation. In this study, we analyzed both the production of cellulose by all known members of the  Rhizobiaceae  and the diversity of  Rhizobium celABC  operon predicted to be involved in cellulose biosynthesis. We also investigated the involvement in cellulose production and biofilm formation of  celC  gene encoding an endoglucanase (CelC2) that is required for canonical symbiotic root hair infection by  Rhizobium leguminosarum  bv. trifolii.    Results  ANU843  celC  mutants lacking (ANU843ΔC2) or overproducing cellulase (ANU843C2 + ) produced greatly increased or reduced amounts of external cellulose micro fibrils, respectively. Calcofluor-stained cellulose micro fibrils were considerably longer when formed by ANU843ΔC2 bacteria rather than by the wild-type strain, in correlation with a significant increase in their flocculation in batch culture. In contrast, neither calcofluor-stained extracellular micro fibrils nor flocculation was detectable in ANU843C2 +  cells. To clarify the role of cellulose synthesis in  Rhizobium  cell aggregation and attachment, we analyzed the ability of these mutants to produce biofilms on different surfaces. Alteration of wild-type CelC2 levels resulted in a reduced ability of bacteria to form biofilms both in abiotic surfaces and  in planta .    Conclusions  Our results support a key role of the CelC2 cellulase in cellulose biosynthesis by modulating the length of the cellulose fibrils that mediate firm adhesion among  Rhizobium  bacteria leading to biofilm formation.  Rhizobium  cellulose is an essential component of the biofilm polysaccharidic matrix architecture and either an excess or a defect of this “building material” seem to collapse the biofilm structure. These results position cellulose hydrolytic enzymes as excellent anti-biofilm candidates.

Role of Rhizobium endoglucanase CelC2 in cellulose biosynthesis and biofilm formation on plant roots and abiotic surfaces