Genetic Determinants of Host Range Specificity of the Wellington Strain of Xanthomonas axonopodis pv. citri

Abstract

A new strain of the citrus canker bacterium (Xanthomonas axonopodis pv. citri) was detected in Florida. It was significant in that it was primarily pathogenic on Key lime trees, but not on grapefruit trees. This strain has been designated as Xac-Aw. This strain has caused concern among regulators with regard to how to treat this bacterium in the eradication program. Of major concern was the stability of the bacterium in regard to host specificity of the strain; for example, could the strain mutate to attack grapefruit and other citrus plants. We investigated the frequency of the development of mutants, the transfer of genes by conjugation, and the presence of an avirulence gene to grapefruit in the genome. We never were able to find a mutant, either natural or induced, that changed host range. In conjugation studies, transfer of marker genes on the chromosome to and from the Xac-Aw strain by conjugation was not successful. In an attempt to locate possible avirulence genes, a genetic library of Xac-Aw DNA was made using the pLAFR vector and transformed into Escherichia coli. This library was transferred to strain 91-118 of X. perforans, which is pathogenic to tomato, but causes a null reaction in grapefruit leaves. The transconjugants were screened for HR in grapefruit leaves. The inoculated leaves were observed for development of an HR in the infiltrated area. In the screening of the Xac-Aw genomic library we found an avirulence gene in the genome of Xac-Aw that interacts with grapefruit leaves to cause an HR. This is the first report of an avirulence gene in the genome of a citrus canker bacterium that interacts with a citrus species. This gene was designated as avrGf1, was sequenced, and then was characterized. The possibility of a second avr gene exists in the Xac-Aw strain, because the Xac-AwΩ lacking a functional avrGf1 did not cause exactly the same symptoms and develop the same population as the wild-type Xac-A strain when it was inoculated into grapefruit leaves.