Deciphering the key players within the bacterial microbiota associated with aerial crown gall tumors on rhododendron: Insights into the gallobiome

Tumorigenic agrobacteria are widespread plant pathogens causing crown gall and cane gall diseases on various agricultural crops. These pathogens genetically transform its host plant and thus form an ecological niche (galls), in which specific metabolites (i.e., opines) are produced. Opines provide the pathogen with multiple competitive advantages, but they can also be utilized by other bacteria colonizing galls. To gain a thorough understanding of disease processes and ecology, it is necessary to consider the pathogen in the context of its microbial environment within the diseased plant (i.e., the pathobiome). Therefore, in this study, we investigated the bacterial pathobiome associated with aerial crown gall tumors (gallobiome) on rhododendron. For this purpose, combination of cultivation-dependent and -independent approaches were applied, which also involved development of a novel amplicon sequencing approach targeting the recA housekeeping gene. The 16S rRNA and recA gene amplicon sequencing clearly indicated that Rhizobium rhododendri and the group of Agrobacterium spp., primarily belonging to the so-called “rubi” clade were the dominant members of bacterial microbiota in rhododendron galls. While the tumor-inducing (Ti) plasmid-harboring R. rhododendri strains are causative agents of crown gall disease, Agrobacterium spp. strains isolated in this study were nonpathogenic and carried genes for the catabolism of opines, enabling these bacteria to efficiently colonize tumor tissue. Taken together, our results clearly showed that the tumorigenic R. rhododendri and nonpathogenic opine-catabolizing Agrobacterium spp. were the key players within the bacterial microbiota associated with aerial crown gall tumors on rhododendron.