The endospore-forming rhizobacteria from the Bacillus genus living in association with plant roots are universal symbionts of higher plants, which enhance the adaptive potential of their hosts. These Plant Growth-Promoting Rhizobacteria (PGPR) have numerous traits which allow them to act as biocontrol agents. One of them is suppression of diseases caused by phytopathogens thanks to produced wide range of antimicrobial molecules. Bacillus subtilis is the most outstanding representative with regards to its ability to excrete a variety of antibacterial and antifungal compounds among which are lipopeptides of the surfactin, iturin and fengycin families.
Samples of various plant roots (n=86) were collected from fields in neighbourhood of Gdansk, Poland, and bacteria from its rhizosphere were isolated. Method of isolation was based on resistance of Bacillus endospores to high temperature. In our search for biological agents to control plant pathogens, we obtained 620 bacterial isolates. To confirm their preliminary belonging to Bacillus genus, gram staining, colony morphology, a catalase reaction and nitrate reduction test were assayed.
Moreover, isolates were analyzed according to their ability to biofilm formation. Measurement of biofilm mass was adapted from the protocol published by O'Toole and Kolter (1998) and modified in our laboratory. Studies investigated growth and biofilm production of isolated strains in minimal defined medium (Belitzky medium) with glucose as a sole carbon source, and test was assayed by the ability of cells to pellicle formation in air/liquid interfaces of 96-well plates made of polystyrene. The biofilm was detected by staining with crystal violet. The procedure was performed three times for all strains, and the averages and standard deviations were calculated for all repetitions of the experiment.
The domesticated strain Bacillus subtilis 168 and natural wild-type Bacillus subtilis 3610 formed a biofilm in amount of crystal violet units (CV) 0.95 and 1.80, respectively. Results appeared to detect differences in biofilm-forming ability among investigated strains. Biofilm production at 24 h of incubation varied greatly between different natural isolates, ranging from CV of 0.1 to 2.4.

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