Characterization of the fimA gene encoding bundle-forming fimbriae of the plant pathogen Xanthomonas campestris pv. vesicatoria

Abstract

The fimA gene of Xanthomonas campestris pv. vesicatoria was identified and characterized. A 20-mer degenerate oligonucleotide complementary to the N-terminal amino acid sequence of the purified 15.5-kDa fimbrillin was used to locate fimA on a 2.6-kb SalI fragment of the X. campestris pv. vesicatoria 3240 genome. The nucleotide sequence of a 1.4-kb fragment containing the fimA region revealed two open reading frames predicting highly homologous proteins FimA and FimB. FimA, which was composed of 136 amino acids and had a calculated molecular weight of 14,302, showed high sequence identity to the type IV fimbrillin precursors. fimB predicted a protein product of 135 amino acids and a molecular weight of 13,854. The open reading frame for fimB contained near the 5 end a palindromic sequence with a terminator loop potential, and the expression level of fimB in vitro and in Xanthomonas was considerably lower than that of fimA. We detected an efficiently transcribed fimA-specific mRNA of 600 bases as well as two weakly expressed, longer mRNA species that reacted with both fimA and fimB. A homolog of fimA but not of fimB was detected by Southern hybridization in strains of X. campestris pv. vesicatoria, campestris, begoniae, translucens, and graminis. A fimA::⍀ mutant of strain 3240 was not significantly reduced in virulence or adhesiveness to tomato leaves. However, the fimA mutant was dramatically reduced in cell aggregation in laboratory cultures and on infected tomato leaves. The fimA mutant strain also exhibited decreased tolerance to UV light.Xanthomonas campestris is a plant pathogen divided into more than 140 pathovars on the basis of the host plants of X. campestris isolates (48). Commonly, the pathovars exhibit a high degree of host specificity in causing the disease, which makes X. campestris infections an interesting target for studies on bacterium-plant interactions and pathogenetic mechanisms of the infections. The ability of X. campestris isolates to cause disease is controlled by hrp (hypersensitive reaction and pathogenicity) genes, whose pathogenetic functions have not yet been characterized. Some of the identified hrp gene products of X. campestris exhibit sequence homology to proteins functioning in secretion of virulence factors of bacteria causing infections in animals (reviewed in reference 5), suggesting that secreted proteins play a role in pathogenesis of X. campestris infections. Isolates of X. campestris secrete plant cell walldegrading enzymes whose function in the pathogenetic processes of X. campestris infections, however, has remained unclear (reviewed in reference 13).In contrast to bacterial infections in mammals, the importance of fimbriae and bacterial adhesion to plant tissue in the pathogenetic processes of plant pathogens has remained controversial (reviewed in references 4 and 34). Fimbriae have been indicated to mediate adhesion of Pseudomonas syringae to bean leaves and to affect the virulence of P. syringae in bean (35). Recently, van Doorn et al. (47) isolated fimbriae from ...

The Type III-Dependent Hrp Pilus Is Required for Productive Interaction of Xanthomonas campestris pv. vesicatoria with Pepper Host Plants

“…This was achieved in vitro by growth of the bacteria in hrp-inducing medium or by use of an hrpG* mutant which constitutively expresses the hrpG regulon. It is noteworthy to mention that no type IV fimbriae were observed under any of the growth conditions tested, although strain 85-10 was reported to carry a gene for type IV fimbriae (48). Altogether, these observations add another important plant pathogenic genus to the list of Hrp pilus-producing bacteria (32,49,59).…”

“…Pili were purified from strain 85E of X. campestris pv. vesicatoria by use of a deoxycholate-sucrose density gradient as described by Ojanen-Reuhs et al (48). Briefly, after mechanical detachment from the bacterial cells, cells were pelleted and the pili-containing supernatant was concentrated by precipitation with ammonium sulfate, dialyzed, and solubilized in buffer containing deoxycholate.…”

Twitching motility of Ralstonia solanacearum requires a type IV pilus system

“…vesicatoria, the pilin structural gene was found by reverse genetics and subsequently inactivated. The nonpiliated mutant retained full virulence on tomato plants when tested under controlled conditions, but because Tfp promoted aggregation on leaf surfaces and increased resistance to killing by UV light the authors speculated that piliation might increase survival on leaf surfaces (Ojanen-Reuhs et al, 1997). Similarly, the Tfp of Xanthomonas campestris pv.…”

“…Numerous Gram-negative bacteria, including some animal and plant pathogens, produce type IV pili (Tfp) (Ojanen-Reuhs et al, 1997 ;Roine et al, 1998 ;Strom & Lory, 1993 ;Su et al, 1999 ;van Doorn et al, 1994 ;. Depending on the organism, Tfp can be important for adhesion, aggregation, biofilm formation, horizontal gene transfer, multicellular development, pathogenesis and twitching motility (Fussenegger et al, 1997 ;Merz et al, 2000 ;Strom & Lory, 1993 ;.…”

Characterization of Xylella fastidiosa popP Gene Required for Pathogenicitya

“…Biofilms are complex structures involving X. fastidiosa cells and an extracellular matrix. X. fastidiosa produces abundant multiple putative afimbrial adhesins, such as exopolysaccharide (EPS), lipopolysaccharide (LPS), and surface proteins, which contribute to the virulence of X. fastidiosa by attaching to the xylem wall in the plant and enhancing biofilm formation [33,[40][41][42][43]. Previously study showed that X. fastidiosa EPS was coded by a cluster of nine gum genes closely related to the xanthan gum operon of Xanthomonas campestris pv.…”