Open AccessVisualization and characterization of Pseudomonas syringae pv. tomato DC 3000 pellicles
Author(s) -
Farias Gabriela A.,
Olmedilla Adela,
Gallegos MaríaTrinidad
Publication year - 2019
Publication title -
microbial biotechnology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.287
H-Index - 74
ISSN - 1751-7915
DOI - 10.1111/1751-7915.13385
Subject(s) - biofilm , bacterial cellulose , pseudomonas syringae , cellulose , microbiology and biotechnology , intracellular , extracellular , bacteria , chemistry , transposon mutagenesis , biochemistry , biology , mutant , biophysics , pathogen , transposable element , gene , genetics
Summary Cellulose, whose production is controlled by c‐di‐ GMP , is a commonly found exopolysaccharide in bacterial biofilms. Pseudomonas syringae pv. tomato (Pto) DC 3000, a model organism for molecular studies of plant–pathogen interactions, carries the wss ABCDEFGHI operon for the synthesis of acetylated cellulose. The high intracellular levels of the second messenger c‐di‐ GMP induced by the overexpression of the heterologous diguanylate cyclase PleD stimulate cellulose production and enhance air–liquid biofilm (pellicle) formation. To characterize the mechanisms involved in Pto DC 3000 pellicle formation, we studied this process using mutants lacking flagella, biosurfactant or different extracellular matrix components, and compared the pellicles produced in the absence and in the presence of PleD. We have discovered that neither alginate nor the biosurfactant syringafactin are needed for their formation, whereas cellulose and flagella are important but not essential. We have also observed that the high c‐di‐ GMP levels conferred more cohesion to Pto cells within the pellicle and induced the formation of intracellular inclusion bodies and extracellular fibres and vesicles. Since the pellicles were very labile and this greatly hindered their handling and processing for microscopy, we have also developed new methods to collect and process them for scanning and transmission electron microscopy. These techniques open up new perspectives for the analysis of fragile biofilms in other bacterial strains.