Molecular analysis of H antigens reveals that human diarrheagenic Escherichia coli O26 strains that carry the eae gene belong to the H11 clonal complex.
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Fifty-seven Escherichia coli O26 strains isolated from patients in six countries were investigated by PCR restriction fragment length polymorphism (RFLP) analysis of the flagellin-encoding (fliC) gene (fliC RFLP analysis). The strains were determined by serotyping to belong to five different H types or were nonmotile. The fliC RFLP analysis revealed only two different patterns among the 57 strains. One fliC RFLP pattern was displayed by 54 strains and was identical to that of E. coli H11 reference strain Su4321-41. The other fliC RFLP pattern was observed for three strains and was identical to that for E. coli H32 reference strain K10. The 54 strains with the H11 fliC RFLP pattern included 22 strains of serotype O26:H11, 23 nonmotile strains, and 9 strains that were initially serotyped as H2, H8, H21, and H32 but that were confirmed to express H11 by repeat serotyping. All 54 strains with the H11 fliC RFLP pattern contained the attaching-and-effacing (eae) gene. The three strains with the H32 fliC RFLP pattern belonged to serotype O26:H32, and all were eae negative. The fliC genes of 14 selected E. coli O26:H11 strains isolated between 1964 and 1999 had identical nucleotide sequences. Our results demonstrate that E. coli O26 strains that carry the eae gene belong exclusively to the H11 clonal complex. Since there were no H11 fliC allelic variations among the O26 strains tested, E. coli O26:H11 may have emerged recently. The fliC PCR-RFLP test is a reliable, easy-to-perform, and rapid method for determination of the H types of E. coli O26 isolates. (+info)
Flagellin from an incompatible strain of Pseudomonas avenae induces a resistance response in cultured rice cells.
(66/1391)
The host range of Pseudomonas avenae is wide among monocotyledonous plants, but individual strains can infect only one or a few host species. The resistance response of rice cells to pathogens has been previously shown to be induced by a rice-incompatible strain, N1141, but not by a rice-compatible strain, H8301. To clarify the molecular mechanism of the host specificity in P. avenae, a strain-specific antibody that was raised against N1141 cells and then absorbed with H8301 cells was prepared. When a cell extract of strain N1141 was separated by SDS-polyacrylamide gel electrophoresis and immunostained with the N1141 strain-specific antibody, only a flagellin protein was detected. Purified N1141 flagellin induced the hypersensitive cell death in cultured rice cells within 6 h of treatment, whereas the H8301 flagellin did not. The hypersensitive cell death could be blocked by pretreatment with anti-N1141 flagellin antibody. Furthermore, a flagellin-deficient N1141 strain lost not only the induction ability of hypersensitive cell death but also the expression ability of the EL2 gene, which is thought to be one of the defense-related genes. These results demonstrated that the resistance response in cultured rice cells is induced by the flagellin existing in the incompatible strain of P. avenae but not in the flagellin of the compatible strain. (+info)
Elicitation of suspension-cultured tomato cells triggers the formation of phosphatidic acid and diacylglycerol pyrophosphate.
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Phosphatidic acid (PA) and its phosphorylated derivative diacylglycerol pyrophosphate (DGPP) are lipid molecules that have been implicated in plant cell signaling. In this study we report the rapid but transient accumulation of PA and DGPP in suspension-cultured tomato (Lycopersicon esculentum) cells treated with the general elicitors, N,N',N",N"'-tetraacetylchitotetraose, xylanase, and the flagellin-derived peptide flg22. To determine whether PA originated from the activation of phospholipase D or from the phosphorylation of diacylglycerol (DAG) by DAG kinase, a strategy involving differential radiolabeling with [(32)P]orthophosphate was used. DAG kinase was found to be the dominant producer of PA that was subsequently metabolized to DGPP. A minor but significant role for phospholipase D could only be detected when xylanase was used as elicitor. Since PA formation was correlated with the high turnover of polyphosphoinositides, we hypothesize that elicitor treatment activates phospholipase C to produce DAG, which in turn acts as substrate for DAG kinase. The potential roles of PA and DGPP in plant defense signaling are discussed. (+info)
Relative roles of the fla/che P(A), P(D-3), and P(sigD) promoters in regulating motility and sigD expression in Bacillus subtilis.
(68/1391)
Three promoters have been identified as having potentially important regulatory roles in governing expression of the fla/che operon and of sigD, a gene that lies near the 3' end of the operon. Two of these promoters, fla/che P(A) and P(D-3), lie upstream of the >26-kb fla/che operon. The third promoter, P(sigD), lies within the operon, immediately upstream of sigD. fla/che P(A), transcribed by E sigma(A), lies >/=24 kb upstream of sigD and appears to be largely responsible for sigD expression. P(D-3), transcribed by E sigma(D), has been proposed to participate in an autoregulatory positive feedback loop. P(sigD), a minor sigma(A)-dependent promoter, has been implicated as essential for normal expression of the fla/che operon. We tested the proposed functions of these promoters in experiments that utilized strains that bear chromosomal deletions of fla/che P(A), P(D-3), or P(sigD). Our analysis of these strains indicates that fla/che P(A) is absolutely essential for motility, that P(D-3) does not function in positive feedback regulation of sigD expression, and that P(sigD) is not essential for normal fla/che expression. Further, our results suggest that an additional promoter(s) contributes to sigD expression. (+info)
A family of six flagellin genes contributes to the Caulobacter crescentus flagellar filament.
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The Caulobacter crescentus flagellar filament is assembled from multiple flagellin proteins that are encoded by six genes. The amino acid sequences of the FljJ and FljL flagellins are divergent from those of the other four flagellins. Since these flagellins are the first to be assembled in the flagellar filament, one or both might have specialized to facilitate the initiation of filament assembly. (+info)
The flagellar filament of Rhodobacter sphaeroides: pH-induced polymorphic transitions and analysis of the fliC gene.
(70/1391)
Flagellar motility in Rhodobacter sphaeroides is notably different from that in other bacteria. R. sphaeroides moves in a series of runs and stops produced by the intermittent rotation of the flagellar motor. R. sphaeroides has a single, plain filament whose conformation changes according to flagellar motor activity. Conformations adopted during swimming include coiled, helical, and apparently straight forms. This range of morphological transitions is larger than that in other bacteria, where filaments alternate between left- and right-handed helical forms. The polymorphic ability of isolated R. sphaeroides filaments was tested in vitro by varying pH and ionic strength. The isolated filaments could form open-coiled, straight, normal, or curly conformations. The range of transitions made by the R. sphaeroides filament differs from that reported for Salmonella enterica serovar Typhimurium. The sequence of the R. sphaeroides fliC gene, which encodes the flagellin protein, was determined. The gene appears to be controlled by a sigma(28)-dependent promoter. It encodes a predicted peptide of 493 amino acids. Serovar Typhimurium mutants with altered polymorphic ability usually have amino acid changes at the terminal portions of flagellin or a deletion in the central region. There are no obvious major differences in the central regions to explain the difference in polymorphic ability. In serovar Typhimurium filaments, the termini of flagellin monomers have a coiled-coil conformation. The termini of R. sphaeroides flagellin are predicted to have a lower probability of coiled coils than are those of serovar Typhimurium flagellin. This may be one reason for the differences in polymorphic ability between the two filaments. (+info)
Mutational analysis of genes encoding the early flagellar components of Helicobacter pylori: evidence for transcriptional regulation of flagellin A biosynthesis.
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We investigated the roles of fliF, fliS, flhB, fliQ, fliG, and fliI of Helicobacter pylori, predicted by homology to encode structural components of the flagellar basal body and export apparatus. Mutation of these genes resulted in nonmotile, nonflagellate strains. Western blot analysis showed that all the mutants had considerably reduced levels of both flagellin subunits and of FlgE, the flagellar hook protein. RNA slot blot hybridization showed reduced levels of flaA mRNA, indicating that transcription of the major flagellin gene is inhibited in the absence of the early components of the flagellar-assembly pathway. This is the first demonstration of a checkpoint in H. pylori flagellar assembly. (+info)
Construction of a multihybrid display system: flagellar filaments carrying two foreign adhesive peptides.
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A multivalent, bifunctional flagellum carrying two different adhesive peptides in separate flagellin subunits within a filament was constructed in Escherichia coli. The inserted peptides were the fibronectin-binding 115-mer D repeat region of Staphylococcus aureus and the 302-mer collagen-binding region of YadA of Yersinia enterocolitica. Western blotting, immunoelectron microscopy, and adhesion tests with hybrid flagella from an in trans-complemented DeltafliC E. coli strain showed that individual filaments consisted of both recombinant flagellins. (+info)