Abstract
Three species of the β-Proteobacterial genus Herbaspirillum are able to fix nitrogen in endophytic associations with such important agricultural crops as maize, rice, sorghum, sugar-cane and wheat. In addition, Herbaspirillum rubrisubalbicans causes the mottled-stripe disease in susceptible sugar-cane cultivars as well as the red-stripe disease in some sorghum cultivars. The xylem of these cultivars exhibited a massive colonisation of mucus-producing bacteria leading to blocking the vessels. A cluster of eight genes (bcs) are involved in cellulose synthesis in Herbaspirillum rubrisubalbicans. Mutation of bcsZ, that encodes a 1,4-endoglucanase, impaired the exopolysaccharide production, the ability to form early biofilm and colonize sorghum when compared to the wild-type strain M1. This mutation also impaired the ability of Herbaspirillum rubrisubalbicans M1 to cause the red-stripe disease in Sorghum bicolor. We show cellulose synthesis is involved in the biofilm formation and as a consequence significantly modulates bacterial-plant interactions, indicating the importance of cellulose biosynthesis in this process.
Highlights
Nitrogen fixation, solubilisation of phosphorus, production of phyto-hormones and siderophores are some of the mechanisms that plant growth promoting rhizobacteria (PGPR) use to modulate plant growth[1]
To confirm the presence of cellulose structures in the H. rubrisubalbicans M1 bacterial biofilm, we perform a microscopy assay during the biofilm formation in glass fibres
The cells were stained with calcofluor a fluorescent dye[30] that binds β(14)-linked glucans. In these experiments biofilm formation started with H. rubrisubalbicans M1 aggregating on the glass fibres eventually forming clumps that were intensively stained by calcofluor (Fig. 1a)
Summary
Production of Cellulose by H. rubrisubalbicans Strains. To confirm the presence of cellulose structures in the H. rubrisubalbicans M1 bacterial biofilm, we perform a microscopy assay during the biofilm formation in glass fibres. A treatment of H. rubrisubalbicans with cellulase before inoculation of maize led to 10-fold decrease in bacterial attachment of the wild-type strain to root-cells (from 3 × 106 to 5 × 105 CFU). The mutant strain TRT1 showed similar results, except for root volume which was significantly smaller than those colonized by the wild-type strain (Fig. S7). These results suggest that the bcsZ gene is not essential for plant growth promotion in maize. The results show that cellulose synthesis is important for cell aggregation, biofilm formation, colonisation of gramineae roots and for development of the red-stripe disease in sorghum by H. rubrisubalbicans
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