Affiliation:
1. Department of Plant Pathology, University of Wisconsin—Madison, Madison, Wisconsin 53706
Abstract
ABSTRACT
Ralstonia solanacearum
, a widely distributed and economically important plant pathogen, invades the roots of diverse plant hosts from the soil and aggressively colonizes the xylem vessels, causing a lethal wilting known as bacterial wilt disease. By examining bacteria from the xylem vessels of infected plants, we found that
R. solanacearum
is essentially nonmotile in planta, although it can be highly motile in culture. To determine the role of pathogen motility in this disease, we cloned, characterized, and mutated two genes in the
R. solanacearum
flagellar biosynthetic pathway. The genes for flagellin, the subunit of the flagellar filament (
fliC
), and for the flagellar motor switch protein (
fliM
) were isolated based on their resemblance to these proteins in other bacteria. As is typical for flagellins, the predicted FliC protein had well-conserved N- and C-terminal regions, separated by a divergent central domain. The predicted
R. solanacearum
FliM closely resembled motor switch proteins from other proteobacteria. Chromosomal mutants lacking
fliC
or
fliM
were created by replacing the genes with marked interrupted constructs. Since
fliM
is embedded in the
fliLMNOPQR
operon, the
aphA
cassette was used to make a nonpolar
fliM
mutation. Both mutants were completely nonmotile on soft agar plates, in minimal broth, and in tomato plants. The
fliC
mutant lacked flagella altogether; moreover, sheared-cell protein preparations from the
fliC
mutant lacked a 30-kDa band corresponding to flagellin. The
fliM
mutant was usually aflagellate, but about 10% of cells had abnormal truncated flagella. In a biologically representative soil-soak inoculation virulence assay, both nonmotile mutants were significantly reduced in the ability to cause disease on tomato plants. However, the
fliC
mutant had wild-type virulence when it was inoculated directly onto cut tomato petioles, an inoculation method that did not require bacteria to enter the intact host from the soil. These results suggest that swimming motility makes its most important contribution to bacterial wilt virulence in the early stages of host plant invasion and colonization.
Publisher
American Society for Microbiology
Subject
Molecular Biology,Microbiology
Cited by
266 articles.
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