P-223 - Unveiling the mechanisms determining tissue specificity in the plant pathogen Clavibacter michiganensis using experimental evolution

The plant pathogenic bacterium Clavibacter michiganensis (Cm) is a systemic vascular pathogen that thrives in host xylem vessels and intracellular apoplast tissues. To identify traits linked to adaptation in these environments, we performed tissue-specific experimental evolution, adapting twenty independent clone lineages to vascular or apoplastic lifestyles through repeated passaging in tomato stems or leaves. Evolved Cm populations were analyzed for virulence-associated traits. Results showed clear associations of these traits with the adapted tissue. Most vascular-adapted clones exhibited enhanced biofilm formation, reduced cellulase activity, decreased exopolysaccharide (EPS) production, and attenuated virulence compared to the parent clone. Conversely, apoplast-adapted clones displayed reduced biofilm formation and increased EPS production while preserving their aggression. To pinpoint loci related to tissue specificity, we sequenced the entire genome of all evolved clones and compared genomes to identify alterations. Notably, six of the ten vascular-adapted clones carried two independent mutations in a putative Cro/CI-type transcriptional regulator, indicating its potential role in vascular adaptation. These experiments enhance our understanding of the tissue adaptation of plant pathogenic bacteria and suggest that cellulase activity and biofilm formation contribute to tissue specificity.