Quorum Sensing-Independent Cellulase-Sensitive Pellicle Formation Is Critical for Colonization of Burkholderia glumae in Rice Plants.

Bacteria form biofilms as a means to adapt to environmental changes for survival. Pellicle is a floating biofilm formed at the air-liquid interface in static culture conditions; however, its functional roles have received relatively little attention compared to solid surface-associated biofilms in gram-negative bacteria. Here we show that the rice pathogen Burkholderia glumae BGR1 forms cellulase-sensitive pellicles in a bis-(3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP)- and flagellum-dependent, but quorum sensing (QS)-independent, manner. Pellicle formation was more favorable at 28°C than at the optimum growth temperature (37°C), and was facilitated by constitutive expression of pelI , a diguanylate cyclase gene from B. glumae , or pleD , the GGDEF response regulator from Agrobacterium tumefaciens . Constitutive expression of pelI or pleD raised the levels of c-di-GMP, facilitated pellicle formation, and suppressed swarming motility in B. glumae . QS-defective mutants of B. glumae formed pellicles, while flagellum-defective mutants did not. Pellicles of B. glumae were sensitive to cellulase but not to proteinase K or DNase I. A gene cluster containing seven genes involved in bacterial cellulose biosynthesis, bcsD , bcsR , bcsQ , bcsA , bcsB , bcsZ , and bcsC , homologous to known genes involved in cellulose biosynthesis in other bacteria, was identified in B. glumae . Mutations in each gene abolished pellicle formation. These results revealed a positive correlation between cellulase-sensitive pellicles and putative cellulose biosynthetic genes. Pellicle-defective mutants did not colonize as successfully as the wild-type strain BGR1 in rice plants, which resulted in a significant reduction in virulence. Our findings show that cellulase-sensitive pellicles produced in a QS-independent manner play important roles in the interactions between rice plants and B. glumae .