Phenotypic Characterization and Transformation Attempts Reveal Peculiar Traits of Subspecies Strain De Donno.

subsp. strain De Donno has been recently identified as the causal agent of a severe disease affecting olive trees in a wide area of the Apulia Region (Italy). While insights on the genetics and epidemiology of this virulent strain have been gained, its phenotypic and biological traits remained to be explored. We investigated in vitro behavior of the strain and compare its relevant biological features (growth rate, biofilm formation, cell-cell aggregation, and twitching motility) with those of the type strain Temecula1. The experiments clearly showed that the strain De Donno did not show fringe on the agar plates, produced larger amounts of biofilm and had a more aggregative behavior than the strain Temecula1. Repeated attempts to transform, by natural competence, the strain De Donno failed to produce a GFP-expressing and a knockout mutant for the gene. Computational prediction allowed us to identify potentially deleterious sequence variations most likely affecting the natural competence and the lack of fringe formation. GFP and - mutants were successfully obtained by co-electroporation in the presence of an inhibitor of the type I restriction-modification system. The availability of De Donno mutant strains will open for new explorations of its interactions with hosts and insect vectors.