Proteus mirabilis flagella and MR/P fimbriae: isolation, purification, N-terminal analysis, and serum antibody response following experimental urinary tract infection

Abstract

Urinary tract infection with Proteus mirabilis may lead to serious complications, including cystitis, acute pyelonephritis, fever, bacteremia, and death. In addition to the production of hemolysin and the enzyme urease, fimbriae and flagellum-mediated motility have been postulated as virulence factors for this species. We purified mannose-resistant/proteuslike (MR/P) fimbriae and flagella from strains CFT322 and HU2450, respectively. Electron microscopy revealed highly concentrated preparations of fimbriae and flagella. Fimbrial and flagellar structural subunits were estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis to be 18.5 and 41 kDa, respectively. N-terminal sequencing revealed that 10 of the first 20 amino acids of the major MR/P subunit matched the sequence of the P. mirabilis uroepithelial cell adhesin N terminus and 11 of 20 amino acids matched the predicted amino acid sequence of the Escherichia coli P fimbriae structural subunit, PapA. In addition, 90 and 80% homologies were found between the first 20 amino acids of P. mirabilis flagellin and those of Salmonella typhimurium phase-i flagellin and the E. coli hag gene product, respectively. An enzyme-linked immunosorbent assay using purified antigens showed a strong reaction between the MR/P fimbriae or flagella and sera of CBA mice challenged transurethrally with P. mirabilis. A possible role for MR/P fimbriae in the pathogenesis of urinary tract infection is supported by (i) a strong immune response to the antigen in experimentally infected animals, (ii) amino acid sequence similarity to other enteric surface structure, and (iii) our previously reported observation that MR/P fimbriae are expressed preferentially as the sole fimbrial type in human pyelonephritis isolates.

Expression of flagella and motility by Shigella

“…Sequence similarities between the N-termini of the flagellins of S. typhimurium, E. coil and P. mirabilis and the predicted N-terminal amino acid sequence derived from recombinant fliC of S. sonnei IID969 and S. flexneri IID642 were reported earlier (Bahrani et al, 1991;Tominaga et al, 1994). FlaA of Bordetella bronchiseptica was also reported to show similarities with FIiC of E. coliand S. typhimurium (Akerley and Miller, 1993).…”

“…To learn more about the biochemical properties of flash and its relatedness to other flagellins, flash was isolated from two well-characterized clinical shigellae strains, namely S. sonnei 53G and S. dysenteriae 1617 (Oaks et aL, 1986). Dissociation of flash from the bacteria was achieved by mechanical shearing and differential centrifugation (Bahrani et al, 1991). The partially purified preparation contained flagella as determined by electron microscopy, and the monomeric subunits were resolved by SDS-PAGE under denaturing conditions ( Fig.…”

Fimbriae of uropathogenicProteus mirabilis

“…The discrepancy in the results regarding the molecular mass of MrpA (21 and 18.5 kDa) indicates that although Sareneva et al (1990) and Bahrani et al (1991) have described functionally and morphologically similar fimbriae, they may be heterogeneous. The fact that the MR/P receptor has not been identified points to the possibility that the mannose-resistant hemagglutination observed may actually include several distinct fimbrial types (Bahrani et al, 1991).…”

“…The MR/P fimbriae were also characterized by Bahrani et al (1991Bahrani et al ( , 1993 using P. mirabilis strain HI4320. The MrpA protein, the major subunit of this fimbria type, has a molecular mass of 18.5 kDa and is encoded by the mrpA gene.…”

Repression of motility during fimbrial expression: identification of 14 mrpJ gene paralogues in Proteus mirabilis

“…Little is currently known about regulation of the 17 different potential fimbriae encoded by P. mirabilis. MR/P fimbriae are preferentially expressed in vitro during static culture at 37°C and are expressed during urinary tract infection in mice (Bahrani et al, 1991). Expression of MR/P fimbriae is controlled by an invertible element in the mrp promoter (Zhao et al, 1997) which is in turn regulated by the MrpI recombinase (Li et al, 2002).…”