Generation and characterization of a T3SS mutant of A.
citrulli. Deletion of the AAC00-1 hrcC gene was confirmed by
PCR amplification (Fig. 1). Using primers for the hrcC gene, a
1.08-kb DNA fragment was amplified from wild-type AAC00-1
and the complemented strain, AAC00-1ΔhrcCcomp. However, the
1.08-kb fragment was absent from AAC00-1ΔhrcC and a smaller
0.4-kb fragment was amplified, indicating deletion of the hrcC
gene. As expected, both amplicons were produced with DNA from
AAC00-1ΔhrcCcomp (Fig. 1). hrcC deletion resulted in the loss of
T3S in AAC00-1, which was confirmed by lack of an HR when
AAC00-1ΔhrcC was infiltrated into tobacco leaves (Fig. 2A). In
contrast, AAC00-1 (wild-type) and AAC00-1ΔhrcCcomp induced a
typical HR on tobacco leaves (Fig. 2A). As expected, leaf infiltrations
with PBS did not yield an HR on tobacco. When infiltrated
into watermelon seedlings, AAC00-1 and AAC00-1ΔhrcCcomp
produced typical water-soaking and necrosis by 2 days post inoculation
(dpi) (Fig. 2B). In contrast, seedlings infiltrated with
AAC00-1ΔhrcC did not develop BFB symptoms and yielded results
that were comparable with seedlings inoculated with PBS
(Fig. 2B).