Inter-species expression of an EF-HAND CALCIUM BINDING PROTEIN in Xanthomonas perforans leads to reduced virulence and decreased immune evasion in tomato plants.

ABSTRACT

Many phytopathogenic bacteria require a type three secretion system (TTSS) to activate effector triggered immunity (ETI). We identified a calcium binding protein, EfhXXfa, in the citrus pathogen, X. citri subsp. aurantifolii, that does not require a TTSS to activate reactive oxygen species (ROS) and elicit a hypersensitive reaction (HR) in tomato leaves following infection. Purified, recombinant EfhXXfa was shown to bind two moles of calcium per mole of protein, whereas mutation of the first of two EF-hands did not bind calcium . EfhXXfa expression was determined to be inducible in hrp-inducing medium. Additionally, growth of X. perforans transconjugants with and without the efhXXfa gene in hrp-inducing medium differed in intracellular calcium concentration; the transconjugant without efhXXfa yielded higher cell pellet masses and higher increased intracellular calcium concentrations relative to cells expressing EfhXXfa. An EfhXXfa homolog, EfhXXe, present in the pepper pathogen, X. euvesicatoria, when expressed in the tomato pathogen, X. perforans, triggered ROS production and an HR in tomato leaves and is a host-limiting factor. Interestingly, all tested X. perforans and X. euvesicatoria strains pathogenic on tomato contain a stop codon immediately upstream of the first EF-hand domain in the efhXXe gene, whereas most X. euvesicatoria strains pathogenic on pepper do not.