Abstract:
Ralstonia solanacearum, the causal agent of bacterial wilt disease, has been found to affect
numerous economically important plants. Understanding the molecular basis of resistance,
tolerance and susceptibility of plants to pathogens such as R. solanacearum is a major goal
of molecular plant pathologists. Prior to this study it was thought that Arabidopsis accession
Kil-0 shows gene-for-gene “resistance” to an African Eucalyptus isolate of R. solanacearum,
BCCF402. However, a subsequent preliminary study indicated that Kil-0 may exhibit
“tolerance” which is defined as the plant’s ability to support high pathogen numbers without
displaying disease symptoms or a reduction in host fitness. The aim of this study was to
determine if Kil-0 was tolerant to R. solanacearum BCCF402. The bacterial load of
R. solanacearum was quantified in accessions Kil-0 and Be-0 using dilution plating and
quantitative PCR methods. The cytC gene region was used to quantify R. solanacearum in
Arabidopsis plants and the amount of bacterial DNA was normalized to “alien” DNA that was
spiked into each sample. High bacterial concentrations of BCCF402 were found in Kil-0 but
plants exhibited no wilting symptoms. Additionally, Kil-0 plants inoculated with BCCF402
showed no significant reduction in fitness compared to control Kil-0 plants. In contrast, high
bacterial numbers and severe disease symptoms were observed in the susceptible Be-0
plants, whereas Nd1 plants contained a low number of bacteria and no disease symptoms
indicative of a resistance response. These results illustrated that Kil-0 is tolerant to
R. solanacearum isolate BCCF402. A tool for the visualization of R. solanacearum in
Arabidopsis plants was designed. R. solanacearum isolate BCCF402 was tagged with two
mCherry-containing plasmids under the constitutive expression of the tac promoter. The
expression levels of mCherry were suitable for successful visualization in planta. BCCF402
cells transformed with the mCherry-containing plasmids were not affected in terms of
virulence or disease progression compared to wildtype BCCF402 cells. A plasmid loss of
30-35% was observed in mCherry-tagged BCCF402 cells at later stages of Arabidopsis
infection. mCherry-tagged BCCF402 was successfully visualized in Kil-0 leaves at early
infection stages.
