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Sample records for mannose-resistant proteus-like fimbriae

  1. MrpB Functions as the Terminator for Assembly of Proteus mirabilis Mannose-Resistant Proteus-Like Fimbriae

    PubMed Central

    Li, Xin; Mobley, Harry L. T.

    1998-01-01

    Insertional mutagenesis studies of mrpB, a putative pilin-encoding open reading frame of the mrp gene cluster, which encodes mannose-resistant Proteus-like (MR/P) fimbriae of Proteus mirabilis, indicate that MrpB functions as the terminator for fimbrial assembly. PMID:9529110

  2. Requirement of MrpH for Mannose-Resistant Proteus-Like Fimbria-Mediated Hemagglutination by Proteus mirabilis

    PubMed Central

    Li, Xin; Johnson, David E.; Mobley, Harry L. T.

    1999-01-01

    Two new genes, mrpH and mrpJ, were identified downstream of mrpG in the mrp gene cluster encoding mannose-resistant Proteus-like (MR/P) fimbriae of uropathogenic Proteus mirabilis. Since the predicted MrpH has 30% amino acid sequence identity to PapG, the Galα(1-4)Gal-binding adhesin of Escherichia coli P fimbriae, we hypothesized that mrpH encodes the functional MR/P hemagglutinin. MR/P fimbriae, expressed in E. coli DH5α, conferred on bacteria both the ability to cause mannose-resistant hemagglutination and the ability to aggregate to form pellicles on the broth surface. Both a ΔmrpH mutant expressed in E. coli DH5α and an isogenic mrpH::aphA mutant of P. mirabilis were unable to produce normal MR/P fimbriae efficiently, suggesting that MrpH was involved in fimbrial assembly. Amino acid residue substitution of the N-terminal cysteine residues (C66S and C128S) of MrpH abolished the receptor-binding activity (hemagglutinating ability) of MrpH but allowed normal fimbrial assembly, supporting the notion that MrpH was the functional MR/P hemagglutinin. Immunogold electron microscopy of P. mirabilis HI4320 revealed that MrpH was located at the tip of MR/P fimbriae, also consistent with its role in receptor binding. The isogenic mrpH::aphA mutant of HI4320 was less able to colonize the urine, bladder, and kidneys in a mouse model of ascending urinary tract infection (P < 0.01), and therefore MR/P fimbriae contribute significantly to bacterial colonization in mice. While there are similarities between P. mirabilis MR/P and E. coli P fimbriae, there are more notable differences: (i) synthesis of the MrpH adhesin is required to initiate fimbrial assembly, (ii) MR/P fimbriae confer an aggregation phenotype, (iii) site-directed mutation of specific residues can abolish receptor binding but allows fimbrial assembly, and (iv) mutation of the adhesin gene abolishes virulence in a mouse model of ascending urinary tract infection. PMID:10338487

  3. Requirement of MrpH for mannose-resistant Proteus-like fimbria-mediated hemagglutination by Proteus mirabilis.

    PubMed

    Li, X; Johnson, D E; Mobley, H L

    1999-06-01

    Two new genes, mrpH and mrpJ, were identified downstream of mrpG in the mrp gene cluster encoding mannose-resistant Proteus-like (MR/P) fimbriae of uropathogenic Proteus mirabilis. Since the predicted MrpH has 30% amino acid sequence identity to PapG, the Galalpha(1-4)Gal-binding adhesin of Escherichia coli P fimbriae, we hypothesized that mrpH encodes the functional MR/P hemagglutinin. MR/P fimbriae, expressed in E. coli DH5alpha, conferred on bacteria both the ability to cause mannose-resistant hemagglutination and the ability to aggregate to form pellicles on the broth surface. Both a DeltamrpH mutant expressed in E. coli DH5alpha and an isogenic mrpH::aphA mutant of P. mirabilis were unable to produce normal MR/P fimbriae efficiently, suggesting that MrpH was involved in fimbrial assembly. Amino acid residue substitution of the N-terminal cysteine residues (C66S and C128S) of MrpH abolished the receptor-binding activity (hemagglutinating ability) of MrpH but allowed normal fimbrial assembly, supporting the notion that MrpH was the functional MR/P hemagglutinin. Immunogold electron microscopy of P. mirabilis HI4320 revealed that MrpH was located at the tip of MR/P fimbriae, also consistent with its role in receptor binding. The isogenic mrpH::aphA mutant of HI4320 was less able to colonize the urine, bladder, and kidneys in a mouse model of ascending urinary tract infection (P < 0.01), and therefore MR/P fimbriae contribute significantly to bacterial colonization in mice. While there are similarities between P. mirabilis MR/P and E. coli P fimbriae, there are more notable differences: (i) synthesis of the MrpH adhesin is required to initiate fimbrial assembly, (ii) MR/P fimbriae confer an aggregation phenotype, (iii) site-directed mutation of specific residues can abolish receptor binding but allows fimbrial assembly, and (iv) mutation of the adhesin gene abolishes virulence in a mouse model of ascending urinary tract infection.

  4. Proteus mirabilis mannose-resistant, Proteus-like fimbriae: MrpG is located at the fimbrial tip and is required for fimbrial assembly.

    PubMed Central

    Li, X; Zhao, H; Geymonat, L; Bahrani, F; Johnson, D E; Mobley, H L

    1997-01-01

    The mannose-resistant, Proteus-like (MR/P) fimbria, responsible for mannose-resistant hemagglutination, is a virulence factor for uropathogenic Proteus mirabilis. Based on known fimbrial gene organization, we postulated that MrpG, a putative minor subunit of the MR/P fimbria, functions as an adhesin responsible for hemagglutination, while MrpA serves as the major structural subunit for the filamentous structure. To test this hypothesis, an mrpG mutant was constructed by allelic-exchange mutagenesis and verified by PCR and Southern blotting. The mrpG mutant was found to be negative for hemagglutination, while wild-type strain H14320 and the complemented mutant were positive. Western blots with antiserum raised against an overexpressed MrpG'-His6 fusion protein showed that MrpG was present in the fimbrial preparations of both the wild-type strain and the complemented mutant but absent in that of the mrpG mutant. The mrpG mutant was significantly less virulent in a CBA mouse model of ascending urinary tract infection. Western blots with antiserum to whole MR/P fimbriae showed that MrpA protein was also missing from the fimbrial preparation of the mrpG mutant. Using immunogold electron microscopy, we found that the normal MR/P-fimbrial structure was absent in the mrpG mutant, suggesting that MrpG is essential for initiation of normal fimbrial formation. In the wild-type strain, MrpG protein was localized to the tips of the fimbriae or at the surface of the cell when antiserum raised against overexpressed MrpG was used. Given the tip localization, MrpG may be required for initiation of assembly of MR/P fimbriae but does not appear to be the fimbrial adhesin. PMID:9119470

  5. Proteus mirabilis mannose-resistant, Proteus-like fimbriae: MrpG is located at the fimbrial tip and is required for fimbrial assembly.

    PubMed

    Li, X; Zhao, H; Geymonat, L; Bahrani, F; Johnson, D E; Mobley, H L

    1997-04-01

    The mannose-resistant, Proteus-like (MR/P) fimbria, responsible for mannose-resistant hemagglutination, is a virulence factor for uropathogenic Proteus mirabilis. Based on known fimbrial gene organization, we postulated that MrpG, a putative minor subunit of the MR/P fimbria, functions as an adhesin responsible for hemagglutination, while MrpA serves as the major structural subunit for the filamentous structure. To test this hypothesis, an mrpG mutant was constructed by allelic-exchange mutagenesis and verified by PCR and Southern blotting. The mrpG mutant was found to be negative for hemagglutination, while wild-type strain H14320 and the complemented mutant were positive. Western blots with antiserum raised against an overexpressed MrpG'-His6 fusion protein showed that MrpG was present in the fimbrial preparations of both the wild-type strain and the complemented mutant but absent in that of the mrpG mutant. The mrpG mutant was significantly less virulent in a CBA mouse model of ascending urinary tract infection. Western blots with antiserum to whole MR/P fimbriae showed that MrpA protein was also missing from the fimbrial preparation of the mrpG mutant. Using immunogold electron microscopy, we found that the normal MR/P-fimbrial structure was absent in the mrpG mutant, suggesting that MrpG is essential for initiation of normal fimbrial formation. In the wild-type strain, MrpG protein was localized to the tips of the fimbriae or at the surface of the cell when antiserum raised against overexpressed MrpG was used. Given the tip localization, MrpG may be required for initiation of assembly of MR/P fimbriae but does not appear to be the fimbrial adhesin.

  6. Mannose-Resistant Proteus-Like Fimbriae Are Produced by Most Proteus mirabilis Strains Infecting the Urinary Tract, Dictate the In Vivo Localization of Bacteria, and Contribute to Biofilm Formation

    PubMed Central

    Jansen, Angela M.; Lockatell, Virginia; Johnson, David E.; Mobley, Harry L. T.

    2004-01-01

    Proteus mirabilis, an etiologic agent of complicated urinary tract infections, expresses mannose-resistant Proteus-like (MR/P) fimbriae whose expression is phase variable. Here we examine the role of these fimbriae in biofilm formation and colonization of the urinary tract. The majority of wild-type P. mirabilis cells in transurethrally infected mice produced MR/P fimbriae. Mutants that were phase-locked for either constitutive expression (MR/P ON) or the inability to express MR/P fimbriae (MR/P OFF) were phenotypically distinct and swarmed at different rates. The number of P. mirabilis cells adhering to bladder tissue did not appear to be affected by MR/P fimbriation. However, the pattern of adherence to the bladder surface was strikingly different. MR/P OFF colonized the lamina propria underlying exfoliated uroepithelium, while MR/P ON colonized the luminal surfaces of bladder umbrella cells and not the exfoliated regions. Wild-type P. mirabilis was usually found colonizing intact uroepithelium, but it occasionally adhered to exfoliated areas. MR/P ON formed significantly more biofilm than either P. mirabilis HI4320 (P = 0.03) or MR/P OFF (P = 0.05). MR/P OFF was able to form a biofilm similar to that of the wild type. MR/P ON formed a three-dimensional biofilm structure as early as 18 h after the initiation of the biofilm, while MR/P OFF and the wild type did not. After 7 days, however, P. mirabilis HI4320 formed a 65-μm-thick biofilm, while the thickest MR/P ON and MR/P OFF biofilms were only 12 μm thick. We concluded that MR/P fimbriae are expressed by most P. mirabilis cells infecting the urinary tract, dictate the localization of bacteria in the bladder, and contribute to biofilm formation. PMID:15557655

  7. Characterization of type 1 and mannose-resistant fimbriae of Erwinia spp.

    PubMed Central

    Korhonen, T K; Kalkkinen, N; Haahtela, K; Old, D C

    1987-01-01

    Type 1 fimbriae from Erwinia carotovora subsp. carotovora and mannose-resistant fimbriae from Erwinia rhapontici were purified and characterized. The type 1 fimbrillin had an apparent molecular weight of 16,500; that of the mannose-resistant fimbrillin was 18,000. The amino-terminal amino acid sequences of the two fimbrillins were related, but tryptic peptide maps showed significant differences between the proteins. No serological cross-reaction was found between the two fimbrial filaments, nor did they cross-react with type 1 or type 3 fimbriae purified from other enterobacterial species. Immunofluorescent staining of bacterial populations revealed that they were heterogeneous with respect to fimbriation. Images PMID:2883172

  8. Proteus mirabilis fimbriae: identification, isolation, and characterization of a new ambient-temperature fimbria.

    PubMed Central

    Massad, G; Bahrani, F K; Mobley, H L

    1994-01-01

    Urinary tract infections involving Proteus mirabilis may lead to complications including bladder and kidney stones, acute pyelonephritis, and bacteremia. This bacterium produces a number of fimbriae, two of which, MR/P fimbria and P. mirabilis fimbria, have been shown to contribute to the ability of this pathogen to colonize the bladder and kidney. We have now purified and characterized a previously undescribed fimbria of P. mirabilis, named ambient-temperature fimbria (ATF). Electron microscopy of a pure preparation and immunogold labeling of cells demonstrated that ATF was fimbrial in nature. The major fimbrial subunit of ATF has an apparent molecular weight of 24,000. The N-terminal amino acid sequence, E-X-T-G-T-P-A-P-T-E-V-T-V-D-G-G-T-I-D-F, did not show significant similarity to that of any previously described fimbrial protein. ATF was expressed by all eight P. mirabilis strains examined. Culture conditions affected expression of ATF, with optimal expression observed in static broth cultures at 23 degrees C. This fimbria was not produced by cells grown at 42 degrees C or on solid medium. Expression of ATF did not correlate with mannose-resistant/Proteus-like (MR/P) or mannose-resistant/Klebsiella-like (MR/K) hemagglutination and represents a novel fimbria of P. mirabilis. Images PMID:7909538

  9. Proteus mirabilis fimbriae: identification, isolation, and characterization of a new ambient-temperature fimbria.

    PubMed

    Massad, G; Bahrani, F K; Mobley, H L

    1994-05-01

    Urinary tract infections involving Proteus mirabilis may lead to complications including bladder and kidney stones, acute pyelonephritis, and bacteremia. This bacterium produces a number of fimbriae, two of which, MR/P fimbria and P. mirabilis fimbria, have been shown to contribute to the ability of this pathogen to colonize the bladder and kidney. We have now purified and characterized a previously undescribed fimbria of P. mirabilis, named ambient-temperature fimbria (ATF). Electron microscopy of a pure preparation and immunogold labeling of cells demonstrated that ATF was fimbrial in nature. The major fimbrial subunit of ATF has an apparent molecular weight of 24,000. The N-terminal amino acid sequence, E-X-T-G-T-P-A-P-T-E-V-T-V-D-G-G-T-I-D-F, did not show significant similarity to that of any previously described fimbrial protein. ATF was expressed by all eight P. mirabilis strains examined. Culture conditions affected expression of ATF, with optimal expression observed in static broth cultures at 23 degrees C. This fimbria was not produced by cells grown at 42 degrees C or on solid medium. Expression of ATF did not correlate with mannose-resistant/Proteus-like (MR/P) or mannose-resistant/Klebsiella-like (MR/K) hemagglutination and represents a novel fimbria of P. mirabilis.

  10. Molecular organisation of the genes involved in the production of F7(2) fimbriae, causing mannose-resistant haemagglutination, of a uropathogenic Escherichia coli 06:K2:H1:F7 strain.

    PubMed

    van Die, I; van Megen, I; Hoekstra, W; Bergmans, H

    1984-01-01

    The genes responsible for the formation of the F7 (2) fimbriae of the uropathogenic E. coli strain AD110 (O6:K2:H1: F7 ) have been cloned on the recombinant plasmid pPIL110 -35 (Van Die et al. 1983). The F7 (2) fimbriae, like the F7 (1) fimbriae of AD110 , are responsible for mannose resistant haemagglutination ( MRHA ). The molecular organisation of the genes of pPIL110 -35 involved in the expression of MRHA was studied by: (a) analysis of transposon gamma delta and Tn5 insertion mutants. Mutations that cause an MRHA -deficient phenotype were located in discrete groups within an 11.5 kb restriction fragment of pPIL110 -35, separated by insertion mutations that do not inactivate MRHA . (b) complementation experiments. Restriction fragments of pPIL110 -35 subcloned in the vector pBR322 were tested for their ability to complement transposon insertion mutations in the corresponding regions of pPIL110 -35. Five complementation groups were distinguished. Five genes (designated A-E) involved in the expression of MRHA can be distinguished by these results. The products of these genes were analysed in minicells. The results indicate that gene B codes for a 75 K dalton protein, gene C for a 23 K dalton protein and gene E for a 36 K dalton protein. No product of gene D was observed. Gene A probably codes for the 17 K dalton subunit polypeptide of the F7 (2) fimbriae, as will be discussed.

  11. Proteus mirabilis fimbriae- and urease-dependent clusters assemble in an extracellular niche to initiate bladder stone formation

    PubMed Central

    Schaffer, Jessica N.; Norsworthy, Allison N.; Sun, Tung-Tien

    2016-01-01

    The catheter-associated uropathogen Proteus mirabilis frequently causes urinary stones, but little has been known about the initial stages of bladder colonization and stone formation. We found that P. mirabilis rapidly invades the bladder urothelium, but generally fails to establish an intracellular niche. Instead, it forms extracellular clusters in the bladder lumen, which form foci of mineral deposition consistent with development of urinary stones. These clusters elicit a robust neutrophil response, and we present evidence of neutrophil extracellular trap generation during experimental urinary tract infection. We identified two virulence factors required for cluster development: urease, which is required for urolithiasis, and mannose-resistant Proteus-like fimbriae. The extracellular cluster formation by P. mirabilis stands in direct contrast to uropathogenic Escherichia coli, which readily formed intracellular bacterial communities but not luminal clusters or urinary stones. We propose that extracellular clusters are a key mechanism of P. mirabilis survival and virulence in the bladder. PMID:27044107

  12. Proteus mirabilis fimbriae- and urease-dependent clusters assemble in an extracellular niche to initiate bladder stone formation.

    PubMed

    Schaffer, Jessica N; Norsworthy, Allison N; Sun, Tung-Tien; Pearson, Melanie M

    2016-04-19

    The catheter-associated uropathogenProteus mirabilisfrequently causes urinary stones, but little has been known about the initial stages of bladder colonization and stone formation. We found thatP. mirabilisrapidly invades the bladder urothelium, but generally fails to establish an intracellular niche. Instead, it forms extracellular clusters in the bladder lumen, which form foci of mineral deposition consistent with development of urinary stones. These clusters elicit a robust neutrophil response, and we present evidence of neutrophil extracellular trap generation during experimental urinary tract infection. We identified two virulence factors required for cluster development: urease, which is required for urolithiasis, and mannose-resistantProteus-like fimbriae. The extracellular cluster formation byP. mirabilisstands in direct contrast to uropathogenicEscherichia coli, which readily formed intracellular bacterial communities but not luminal clusters or urinary stones. We propose that extracellular clusters are a key mechanism ofP. mirabilissurvival and virulence in the bladder.

  13. Identification of MrpI as the sole recombinase that regulates the phase variation of MR/P fimbria, a bladder colonization factor of uropathogenic Proteus mirabilis.

    PubMed

    Li, Xin; Lockatell, C Virginia; Johnson, David E; Mobley, Harry L T

    2002-08-01

    Proteus mirabilis is a common cause of urinary tract infection (UTI) in individuals with structural abnormalities or long-term catheterization. The expression of mannose-resistant/Proteus-like (MR/P) fimbria is phase variable because of the inversion of a 251 bp DNA fragment that carries the promoter for the mrp operon. Previous studies have shown that mrpI, which is transcribed divergently from the mrp operon, encodes a recombinase capable of switching the orientation of this invertible element. In this study, we constructed isogenic mrpI null mutants from a clinical isolate of P. mirabilis, HI4320. A polymerase chain reaction (PCR)-based invertible element assay revealed that the isogenic mrpI null mutants were locked in one phase, either expressing (locked on) MR/P fimbriae or not (locked off), which indicated that MrpI was the sole recombinase that regulated the phase variation of MR/P fimbria. The locked-on and locked-off mutants were evaluated for virulence in the CBA mouse model of ascending UTI by co-challenges with each other and with the wild-type strain. Results from these experiments demonstrated conclusively that the MR/P fimbria was a critical bladder colonization factor of uropathogenic P. mirabilis and also suggested that the ability to switch off the expression of MR/P fimbria might be important for kidney colonization.

  14. Proteus mirabilis fimbriae: N-terminal amino acid sequence of a major fimbrial subunit and nucleotide sequences of the genes from two strains.

    PubMed

    Bahrani, F K; Cook, S; Hull, R A; Massad, G; Mobley, H L

    1993-03-01

    Proteus mirabilis, a common cause of urinary tract infection in hospitalized and catheterized patients, produces mannose-resistant/klebsiella-like (MR/K) and mannose-resistant/proteus-like (MR/P) hemagglutinins. The gene encoding the major structural subunit of a fimbria, possibly MR/K, was identified in two strains. A degenerate oligonucleotide probe based on the N terminus of the Proteus uroepithelial cell adhesin and antiserum raised against the denatured polypeptide were used to screen a cosmid gene bank of strain HU1069. A cosmid clone that reacted with the probe and antiserum was identified, and a fimbria-like open reading frame was determined by nucleotide sequencing. The predicted N-terminal amino acid sequence of the processed polypeptide, ENETPAPKVSSTKGEIQLKG (residues 23 to 42), did not match the uroepithelial cell adhesin N terminus but, rather, matched exactly the N-terminal amino acid sequence of a polypeptide with an apparent molecular size of 19.5 kDa isolated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of a fimbrial preparation from strain HI4320 expressing MR/K hemagglutinin. By using an oligonucleotide from the HU1069 open reading frame, the fimbrial gene was isolated and sequenced from a cosmid gene bank clone of strain HI4320. A 552-bp open reading frame predicts a 184-amino-acid polypeptide including a 22-amino-acid hydrophobic leader sequence. The unprocessed polypeptide is predicted to be 18,921 Da; the processed polypeptide is predicted to be 16,749 Da. The predicted amino acid sequence of the polypeptide encoded by the gene, designated pmfA, displayed 36% exact matches with the mannose-resistant fimbrial subunit encoded by smfA of Serratia marcescens but only 15% exact matches with the predicted sequence encoded by mrkA of Klebsiella pneumoniae.

  15. Distinct Residues Contribute to Motility Repression and Autoregulation in the Proteus mirabilis Fimbria-Associated Transcriptional Regulator AtfJ

    PubMed Central

    Bode, Nadine J.; Chan, Kun-Wei; Kong, Xiang-Peng

    2016-01-01

    ABSTRACT Proteus mirabilis contributes to a significant number of catheter-associated urinary tract infections, where coordinated regulation of adherence and motility is critical for ascending disease progression. Previously, the mannose-resistant Proteus-like (MR/P) fimbria-associated transcriptional regulator MrpJ has been shown to both repress motility and directly induce the transcription of its own operon; in addition, it affects the expression of a wide range of cellular processes. Interestingly, 14 additional mrpJ paralogs are included in the P. mirabilis genome. Looking at a selection of MrpJ paralogs, we discovered that these proteins, which consistently repress motility, also have nonidentical functions that include cross-regulation of fimbrial operons. A subset of paralogs, including AtfJ (encoded by the ambient temperature fimbrial operon), Fim8J, and MrpJ, are capable of autoinduction. We identified an element of the atf promoter extending from 487 to 655 nucleotides upstream of the transcriptional start site that is responsive to AtfJ, and we found that AtfJ directly binds this fragment. Mutational analysis of AtfJ revealed that its two identified functions, autoregulation and motility repression, are not invariably linked. Residues within the DNA-binding helix-turn-helix domain are required for motility repression but not necessarily autoregulation. Likewise, the C-terminal domain is dispensable for motility repression but is essential for autoregulation. Supported by a three-dimensional (3D) structural model, we hypothesize that the C-terminal domain confers unique regulatory capacities on the AtfJ family of regulators. IMPORTANCE Balancing adherence with motility is essential for uropathogens to successfully establish a foothold in their host. Proteus mirabilis uses a fimbria-associated transcriptional regulator to switch between these antagonistic processes by increasing fimbrial adherence while simultaneously downregulating flagella. The

  16. Distinct Residues Contribute to Motility Repression and Autoregulation in the Proteus mirabilis Fimbria-Associated Transcriptional Regulator AtfJ.

    PubMed

    Bode, Nadine J; Chan, Kun-Wei; Kong, Xiang-Peng; Pearson, Melanie M

    2016-08-01

    Proteus mirabilis contributes to a significant number of catheter-associated urinary tract infections, where coordinated regulation of adherence and motility is critical for ascending disease progression. Previously, the mannose-resistant Proteus-like (MR/P) fimbria-associated transcriptional regulator MrpJ has been shown to both repress motility and directly induce the transcription of its own operon; in addition, it affects the expression of a wide range of cellular processes. Interestingly, 14 additional mrpJ paralogs are included in the P. mirabilis genome. Looking at a selection of MrpJ paralogs, we discovered that these proteins, which consistently repress motility, also have nonidentical functions that include cross-regulation of fimbrial operons. A subset of paralogs, including AtfJ (encoded by the ambient temperature fimbrial operon), Fim8J, and MrpJ, are capable of autoinduction. We identified an element of the atf promoter extending from 487 to 655 nucleotides upstream of the transcriptional start site that is responsive to AtfJ, and we found that AtfJ directly binds this fragment. Mutational analysis of AtfJ revealed that its two identified functions, autoregulation and motility repression, are not invariably linked. Residues within the DNA-binding helix-turn-helix domain are required for motility repression but not necessarily autoregulation. Likewise, the C-terminal domain is dispensable for motility repression but is essential for autoregulation. Supported by a three-dimensional (3D) structural model, we hypothesize that the C-terminal domain confers unique regulatory capacities on the AtfJ family of regulators. Balancing adherence with motility is essential for uropathogens to successfully establish a foothold in their host. Proteus mirabilis uses a fimbria-associated transcriptional regulator to switch between these antagonistic processes by increasing fimbrial adherence while simultaneously downregulating flagella. The discovery of multiple

  17. Characterization of nonfimbrial mannose-resistant protein hemagglutinins of two Escherichia coli strains isolated from infants with enteritis.

    PubMed Central

    Williams, P H; Knutton, S; Brown, M G; Candy, D C; McNeish, A S

    1984-01-01

    Escherichia coli strains 444-3 and 469-3, isolated from patients with severe infantile enteritis, are able to adhere to and penetrate human epithelial cells in culture. In addition to type 1 fimbriae and glycocalyces , both strains elaborate mannose-resistant nonfimbrial protein hemagglutinins specific for human erythrocytes. Purified agglutinins are aggregates (greater than 4 X 10(6) daltons) of a single protein subunit of apparent Mr 14,000 (469-3) to 14,500 (444-3). The optimal temperature for expression of the agglutinins is 37 degrees C. Bacteria grown at 22 degrees C, which show 1% or less of maximal activity, and mutants deficient in the ability to agglutinate human erythrocytes do not synthesize detectable levels of these surface proteins and, moreover, do not adhere to cultured epithelial cells. Coupled with the observation that purified agglutinins competitively inhibit bacterial adherence to cultured cells, these data indicate that the nonfimbrial surface proteins expressed by strains 444-3 and 469-3 are essential for adherence both to erythrocytes and to cultured epithelial cells. Images PMID:6373609

  18. Haemagglutinins and fimbriae of soft rot Erwinias.

    PubMed

    Wallace, A; Pérombelon, M C

    1992-08-01

    Strains of phytopathogenic soft rot Erwinia spp. were examined for haemagglutinin (HA) production. Mannose-sensitive HA was found only in five of 15 strains of E. carotovora subsp. carotovora. Mannose-resistant HA (MRHA) was found in 12 of 15 strains of E.c. carotovora, ten of 13 strains of E.c. subsp. atroseptica and the single strain of E.c. subsp. betavasculorum, as well as all seven strains of E. chrysanthemi. MRHA, detectable only in a microtitre tray HA assay was of either broad- or narrow-spectrum activity when examined against blood of seven different animal species and could be inhibited by the beta-galactoside asialofetuin. Fimbriae of ca 10 nm diameter were found on MRHA(+) bacteria E.c. carotovora and E.c. atroseptica.

  19. Characterization of fimbriae produced by enteropathogenic Escherichia coli.

    PubMed Central

    Girón, J A; Ho, A S; Schoolnik, G K

    1993-01-01

    Enteropathogenic Escherichia coli (EPEC) express rope-like bundles of filaments, termed bundle-forming pili (BFP) (J. A. Girón, A. S. Y. Ho, and G. K. Schoolnik, Science 254:710-713, 1991). Expression of BFP is associated with localized adherence to HEp-2 cells and the presence of the EPEC adherence factor plasmid. In this study, we describe the identification of rod-like fimbriae and fibrillae expressed simultaneously on the bacterial surface of three prototype EPEC strains. Upon fimbrial extraction from EPEC B171 (O111:NM), three fimbrial subunits with masses of 16.5, 15.5, and 14.7 kDa were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Their N-terminal amino acid sequence showed homology with F9 and F7(2) fimbriae of uropathogenic E. coli and F1845 of diffuse-adhering E. coli, respectively. The mixture of fimbrial subunits (called FB171) exhibited mannose-resistant agglutination of human erythrocytes only, and this activity was not inhibited by alpha-D-Gal(1-4)-beta-Gal disaccharide or any other described receptor analogs for P, S, F, M, G, and Dr hemagglutinins of uropathogenic E. coli, which suggests a different receptor specificity. Hemagglutination was inhibited by extracellular matrix glycoproteins, i.e., collagen type IV, laminin, and fibronectin, and to a lesser extent by gangliosides, fetuin, and asialofetuin. Scanning electron microscopic studies performed on clusters of bacteria adhering to HEp-2 cells revealed the presence of structures resembling BFP and rod-like fimbriae linking bacteria to bacteria and bacteria to the eukaryotic cell membrane. We suggest a role of these surface appendages in the interaction of EPEC with eukaryotic cells as well as in the overall pathogenesis of intestinal disease caused by EPEC. Images PMID:7901197

  20. Biological relationship between F18ab and F18ac fimbriae of enterotoxigenic and verotoxigenic Escherichia coli from weaned pigs with oedema disease or diarrhoea.

    PubMed

    Nagy, B; Whipp, S C; Imberechts, H; Bertschinger, H U; Dean-Nystrom, E A; Casey, T A; Salajka, E

    1997-01-01

    Comparative fimbrial expression and adhesion studies were made on enterotoxigenic and verotoxigenic E. coli (ETEC and VTEC) strains isolated from cases of porcine postweaning diarrhoea or oedema disease. F107(F18ab) fimbriae--monitored by polyclonal and monoclonal antibodies and by electron microscopy--were poorly expressed on most VTEC strains. In contrast, 2134P(F18ac) fimbriae were more readily detected on most ETEC strains. The F18ac strains adhered in vivo to ligated intestinal loops in weaned pigs while the F18ab strains did not adhere or adhered weakly. Similarly, the F18ac strains adhered to isolated intestinal brush borders in weaned pigs but the F18ab strains (except for the F107 reference E. coli) did not adhere or adhered weakly in vitro. Neither the F18ab nor F18ac strains adhered to brush borders from newborn pigs. In vitro adhesion of F18ab and F18ac strains was mannose resistant and receptors for F18 seemed to differ from receptors for K88(F4). It is concluded that the antigenic variants of F18 fimbriae (F18ab and F18ac) are biologically distinct. F18ab fimbriae are expressed poorly both in vitro and in vivo and are frequently linked with the production of SLT-IIv and serogroup O139, while F18ac are more efficiently expressed in vitro and in vivo and most often are linked with enterotoxin (STa, STb) production, and serogroups O141, O157.

  1. Isolation and some properties of fimbriae of oral Streptococcus intermedius.

    PubMed

    Yamaguchi, Taihei; Matsunoshita, Nobuko

    2004-07-01

    Streptococcus intermedius 1208-1 carried linear fiber-like fimbriae that extended radially from the cell surface. The fimbriae were isolated by pipetting and sonication and were purified by ammonium sulfate precipitation followed by a column chromatography series. Heat treatment in the presence of sodium dodecyl sulfate resulted in the dissociation into smaller molecules. Rabbit antiserum raised against the purified protein reacted with fimbriae on the surface of bacteria under immunogold staining. Serotype g or g-related strains produced the fimbriae and aggregated in human saliva. The aggregation was inhibited by the anti-fimbriae immunoglobulin Fab fragment or the purified fimbriae.

  2. Type 1 fimbriae of Salmonella enteritidis.

    PubMed

    Müller, K H; Collinson, S K; Trust, T J; Kay, W W

    1991-08-01

    Salmonella enteritidis was previously shown to produce fimbriae composed of 14,000-molecular-weight (Mr) fimbrin monomers (J. Feutrier, W. W. Kay, and T. J. Trust, J. Bacteriol. 168:221-227, 1986). Another distinct fimbrial structure, comprising 21,000-Mr fimbrin monomers, has now been identified. These fimbriae are simply designated as SEF 14 and SEF 21, respectively (for S. enteritidis fimbriae and the Mr [in thousands] of the fimbrin monomer). A simple method for the purification of both structures was developed by using the different biochemical properties of these fimbriae. SEF 21 remained intact after being boiled in sodium dodecyl sulfate but readily dissociated into subunits of 21,000 Mr at pH 2.2. The overall amino acid composition and the N-terminal amino acid sequence of the SEF 21 fimbrin were distinct from those of SEF 14 but were virtually identical to the predicted sequence for type 1 fimbrin of Salmonella typhimurium. Immunoelectron microscopy of S. enteritidis clearly revealed fimbrial structures that reacted with immune serum specific to the 21,000-Mr fimbrin. Immune sera raised against this subunit were cross-reactive with type 1 fimbrins found in whole-cell lysates of S. typhimurium, Salmonella illinois, and Salmonella cubana. However, there was no cross-reaction with Escherichia coli type 1 fimbriae or with other fimbrins produced by S. enteritidis. Under certain growth conditions, S. enteritidis produced both SEF 14 and SEF 21. However, when S. enteritidis was grown at 30 degrees C or lower, only the 21,000-Mr SEF 21 fimbrin could be detected. There was a direct correlation between mannose-sensitive hemagglutination and the presence of SEF 21.

  3. Purification and characterization of serotype 6 fimbriae from Bordetella pertussis and comparison of their properties with serotype 2 fimbriae.

    PubMed Central

    Cowell, J L; Zhang, J M; Urisu, A; Suzuki, A; Steven, A C; Liu, T; Liu, T Y; Manclark, C R

    1987-01-01

    Fimbriae were removed from Bordetella pertussis (serotype 1.3.6) by mechanical shearing and purified by precipitation with ammonium sulfate, pH-dependent precipitation at pH 7.4, followed by two successive extractions of the precipitated fimbriae with 4 M urea. By electron microscopy, the precipitated fimbriae appeared as aggregated bundles of long, relatively straight filaments which were disaggregated to individual flexuous filaments at pH 10.5. These purified fimbriae were identified as serotype 6 agglutinogens, since antibody to the purified fimbriae agglutinated B. pertussis strains serotyped as 1.3.6, 1.2.3.6, or 1.2.3.4.6 but did not agglutinate strains of serotype 1.2.3.4, 1.2.3, or 1.3. In contrast, antibody to serotype 2 fimbriae only agglutinated B. pertussis strains containing serotype 2 agglutinogen. Purified type 6 and 2 fimbriae were found to be weakly cross-reactive by enzyme-linked immunosorbent assay, using polyclonal antibody to each type of fimbria. In an immunoblot assay, polyclonal antibodies to a 22,000-dalton subunit of fimbriae from B. bronchiseptica reacted strongly with the type 2 fimbrial subunit of B. pertussis, but only weakly with the type 6 subunit. When subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the protein subunit of the type 6 fimbriae migrated with a molecular weight of 21,500, whereas the type 2 fimbrial subunit had a molecular weight of 22,000. The two types of subunits had similar amino acid compositions and showed amino-terminal sequence homology in 15 of 21 amino acids. The amino-terminal amino acid sequences of the B. pertussis fimbriae were distinct from those reported for fimbriae from other gram-negative bacteria. Neither the type 6 nor the type 2 fimbriae caused hemagglutination when assayed with several types of erythrocytes. Images PMID:2881893

  4. Expression of a nonagglutinating fimbria by Proteus mirabilis.

    PubMed

    Tolson, D L; Barrigar, D L; McLean, R J; Altman, E

    1995-03-01

    We have clarified growth conditions and isolation strategies for the nonagglutinating fimbriae from Proteus mirabilis. Nonagglutinating fimbriae were expressed by all P. mirabilis strains we examined, and the major subunit proteins, which ranged from 23 to 29 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, had highly conserved N-terminal sequences.

  5. Expression of a nonagglutinating fimbria by Proteus mirabilis.

    PubMed Central

    Tolson, D L; Barrigar, D L; McLean, R J; Altman, E

    1995-01-01

    We have clarified growth conditions and isolation strategies for the nonagglutinating fimbriae from Proteus mirabilis. Nonagglutinating fimbriae were expressed by all P. mirabilis strains we examined, and the major subunit proteins, which ranged from 23 to 29 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, had highly conserved N-terminal sequences. PMID:7868237

  6. Isolation and characterization of a minor fimbria from Porphyromonas gingivalis.

    PubMed Central

    Hamada, N; Sojar, H T; Cho, M I; Genco, R J

    1996-01-01

    We have discovered two distinctly different fimbriae expressed by the same Porphyromonas gingivalis strain. The construction of a fimA mutant of P. gingivalis ATCC 33277 has previously been reported by N. Hamada et al. (Infect. Immun. 62:1696-1704, 1994). Expression of fimbriae on the surface of the fimA mutant and the wild-type strain, ATCC 33277, were investigated by electron microscopy. The wild-type strain produced long fimbrial structures extending from the cell surface, whereas those structures were not observed on the fimA mutant. However, short fimbrial structures were seen on the surface of the fimA mutant. The short fimbrial protein was purified from the fimA mutant by selective protein precipitation and chromatography on DEAE Sepharose CL-6B. We have found that the second fimbrial structure of P. gingivalis ATCC 33277 is distinct from the 41-kDa (43-kDa) major fimbrial protein (FimA). We provisionally call this protein minor fimbriae. The molecular mass of the minor fimbriae is 67 kDa as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions after boiling at 100 degrees C. The component shows a ladder-like pattern at 80 degrees C under nonreducing conditions, suggesting a tendency to aggregate or polymerize. In immunoblotting analysis, anti-minor fimbria serum reacted with both the 100 degrees C- and the 80 degrees C-treated minor fimbriae. The anti-minor fimbria serum also reacts with the same-molecular-size fimbrial preparation from the wild-type strain. Immunogold electron microscopy showed that the anti-minor fimbria serum bound to the minor fimbria on the cell surface of the wild-type strain. This is the first report on the identification of the minor fimbria produced by P. gingivalis. These results suggest that the minor fimbriae appearing on the fimA mutant strain are produced together with numerous long major fimbriae on the wild-type strain. Moreover, the minor fimbriae are different in size and

  7. Bordetella pertussis fimbriae (Fim): relevance for vaccines.

    PubMed

    Gorringe, Andrew R; Vaughan, Thomas E

    2014-10-01

    Bordetella pertussis produces two serologically distinct fimbriae, Fim2 and Fim3. Expression of these antigens is governed by the BvgA/S system and by the length of a poly(C) tract in the promoter of each gene. Fim2 and Fim3 are important antigens for whole cell pertussis vaccines as clinical trials have shown an association of anti-fimbriae antibody-mediated agglutination and protection. The current five component acellular pertussis vaccine contains co-purified Fim2/3 and provided good efficacy in clinical trials with the anti-Fim antibody response correlating with protection when pre and post exposure antibody levels were analysed. The predominant serotype of B. pertussis isolates has changed over time in most countries but it is not understood whether this is vaccine-driven or whether serotype is linked to the prevailing predominant genotype. Recent studies have shown that both Fim2 and Fim3 are expressed during infection and that Fim2 is more immunogenic than Fim3 in the acellular vaccine.

  8. Simple method for purification of enterotoxigenic E. coli fimbriae

    PubMed Central

    Curtis, Brittany; Grassel, Christen; Laufer, Rachel; Sears, Khandra; Pasetti, Marcela F.; Barry, Eileen M.; Simon, Raphael

    2016-01-01

    Enterotoxigenic E. coli (ETEC) are endemic pathogens in the developing world. They frequently cause illness in travelers, and are among the most prevalent causes of diarrheal disease in children. Pathogenic ETEC strains employ fimbriae as adhesion factors to bind the luminal surface of the intestinal epithelium and establish infection. Accordingly, there is marked interest in immunoprophylactic strategies targeting fimbriae to protect against ETEC infections. Multiple strategies have been reported for purification of ETEC fimbriae, however none is ideal. Purification has typically involved the use of highly virulent wild-type strains. We report here a simple and improved method to purify ETEC fimbriae, which was applied to obtain two different Class 5 fimbriae types of clinical relevance (CFA/I and CS4) expressed recombinantly in E. coli production strains. Following removal from cells by shearing, fimbriae proteins were purified by orthogonal purification steps employing ultracentrifugation, precipitation, and ion-exchange membrane chromatography. Purified fimbriae demonstrated the anticipated size and morphology by electron microscopy analysis, contained negligible levels of residual host cell proteins, nucleic acid, and endotoxin, and were recognized by convalescent human anti-sera. PMID:26581778

  9. Simple method for purification of enterotoxigenic Escherichia coli fimbriae.

    PubMed

    Curtis, Brittany; Grassel, Christen; Laufer, Rachel S; Sears, Khandra T; Pasetti, Marcela F; Barry, Eileen M; Simon, Raphael

    2016-03-01

    Enterotoxigenic Escherichia coli (ETEC) are endemic pathogens in the developing world. They frequently cause illness in travelers, and are among the most prevalent causes of diarrheal disease in children. Pathogenic ETEC strains employ fimbriae as adhesion factors to bind the luminal surface of the intestinal epithelium and establish infection. Accordingly, there is marked interest in immunoprophylactic strategies targeting fimbriae to protect against ETEC infections. Multiple strategies have been reported for purification of ETEC fimbriae, however none is ideal. Purification has typically involved the use of highly virulent wild-type strains. We report here a simple and improved method to purify ETEC fimbriae, which was applied to obtain two different Class 5 fimbriae types of clinical relevance (CFA/I and CS4) expressed recombinantly in E. coli production strains. Following removal from cells by shearing, fimbriae proteins were purified by orthogonal purification steps employing ultracentrifugation, precipitation, and ion-exchange membrane chromatography. Purified fimbriae demonstrated the anticipated size and morphology by electron microscopy analysis, contained negligible levels of residual host cell proteins, nucleic acid, and endotoxin, and were recognized by convalescent human anti-sera.

  10. P-antigen-recognizing fimbriae from human uropathogenic Escherichia coli strains.

    PubMed Central

    Korhonen, T K; Väisänen, V; Saxén, H; Hultberg, H; Svenson, S B

    1982-01-01

    P-antigen-recognizing fimbriae (P fimbriae) from four pyelonephritogenic Escherichia coli strains and type 1 fimbriae from an E. coli strain and a Salmonella typhimurium strain were purified. The P fimbriae were morphologically similar to type 1 fimbriae. The purified P fimbriae agglutinated neuraminidase-treated human P1 and P2k erythrocytes but not p erythrocytes, which lack all P-blood group-specific glycosphingolipids. However, coating of neuraminidase-treated p erythrocytes with globoside rendered such erythrocytes agglutinable by the P fimbriae. The hemagglutinations were in all instances fully inhibited by the synthetic alpha-D-Galp-(1-4)-beta-D-Galp-1-O-Me glycoside. The binding specificity of the P fimbriae could also be demonstrated by using fimbriae coated onto latex particles and nontreated erythrocytes. It was thus concluded that the P fimbriae recognize and bind to the alpha-D-Galp-(1-4)-beta-D-Galp carbohydrate sequence occurring in the series of P-blood group antigen-specific glycosphingolipids. In contrast to both type 1 fimbriae, all four P fimbriae preparations showed multiple bands in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Antisera were raised in rabbits against the various E. coli fimbriae. In enzyme-linked immunosorbent assays each one of the antisera to the P fimbriae reacted to titers of log 4 to 7 with both the homologous and the heterologous P fimbriae, but not with the type 1 fimbriae of E. coli. In a reciprocal fashion, the antiserum to the type 1 fimbriae of one E. coli strain reacted only with the homologous type 1 but not with any of the P fimbriae preparations. Images PMID:6125477

  11. Three-dimensional modeling of the human fallopian tube fimbriae

    PubMed Central

    Eddie, Sharon L.; Quartuccio, Suzanne M.; Zhu, Jie; Shepherd, Jessica A.; Kothari, Rajul; Kim, J. Julie; Woodruff, Teresa K.; Burdette, Joanna E.

    2015-01-01

    Objective Ovarian cancer is the most lethal gynecological malignancy that affects women. Recent data suggests the disease may originate in the fallopian fimbriae; however, the anatomical origin of ovarian carcinogenesis remains unclear. This is largely driven by our lack of knowledge regarding the structure and function of normal fimbriae and the relative paucity of models that accurately recapitulate the in vivo fallopian tube. Therefore, a human three-dimensional (3D) culture system was developed to examine the role of the fallopian fimbriae in serous tumorigenesis. Methods Alginate matrix was utilized to support human fallopian fimbriae ex vivo. Fimbriae were cultured with factors hypothesized to contribute to carcinogenesis, namely; H2O2 (1mM) a mimetic of oxidative stress, insulin (5 µg/ml) to stimulate glycolysis, and estradiol (E2, 10nM) which peaks before ovulation. Cultures were evaluated for changes in proliferation and p53 expression, criteria utilized to identify potential precursor lesions. Further, secretory factors were assessed after treatment with E2 to identify if steroid signaling induces a pro-tumorigenic microenvironment. Results 3D fimbriae cultures maintained normal tissue architecture up to 7 days, retaining both epithelial subtypes. Treatment of cultures with H2O2 or insulin significantly induced proliferation. However, p53 stabilization was unaffected by any particular treatment, although was induced by ex vivo culturing. Moreover, E2-alone treatment significantly induced its canonical target PR and expression of IL8, a factor linked to poor outcome. Conclusions 3D alginate cultures of human fallopian fimbriae provide an important microphysicological model, which can be further utilized to investigate serous tumorigenesis originating from the fallopian tube. PMID:25527363

  12. Antigenic characterisitcs of Moraxella nonliquefaciens fimbriae in double immunodiffusion.

    PubMed

    Frøholm, L O

    1978-08-01

    Rabbit antisera against purified fimbriae (pili) from Moraxella nonliquefaciens detected three fimbrial antigenic components, one or two of which appeared to be present in other fimbriated strains of M. nonliquefaciens and the closely related M. bovis. Maximal precipitation with the antisera required some denaturation of the antigen. Ultrasonication, repeated freeze-thawing, heating, and agents like KBr, NaSCN and urea were effective in liberating the antigen in diffusible forms. The morphology of the fimbriae was altered by heat treatment in 1 M KBr.

  13. Fimbriae have distinguishable roles in Proteus mirabilis biofilm formation.

    PubMed

    Scavone, Paola; Iribarnegaray, Victoria; Caetano, Ana Laura; Schlapp, Geraldine; Härtel, Steffen; Zunino, Pablo

    2016-07-01

    Proteus mirabilis is one of the most common etiological agents of complicated urinary tract infections, especially those associated with catheterization. This is related to the ability of P. mirabilis to form biofilms on different surfaces. This pathogen encodes 17 putative fimbrial operons, the highest number found in any sequenced bacterial species so far. The present study analyzed the role of four P. mirabilis fimbriae (MR/P, UCA, ATF and PMF) in biofilm formation using isogenic mutants. Experimental approaches included migration over catheter, swimming and swarming motility, the semiquantitative assay based on adhesion and crystal violet staining, and biofilm development by immunofluorescence and confocal microscopy. Different assays were performed using LB or artificial urine. Results indicated that the different fimbriae contribute to the formation of a stable and functional biofilm. Fimbriae revealed particular associated roles. First, all the mutants showed a significantly reduced ability to migrate across urinary catheter sections but neither swimming nor swarming motility were affected. However, some mutants formed smaller biofilms compared with the wild type (MRP and ATF) while others formed significantly larger biofilms (UCA and PMF) showing different bioarchitecture features. It can be concluded that P. mirabilis fimbriae have distinguishable roles in the generation of biofilms, particularly in association with catheters. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  14. Curli fimbria: an Escherichia coli adhesin associated with human cystitis.

    PubMed

    Cordeiro, Melina Aparecida; Werle, Catierine Hirsch; Milanez, Guilherme Paier; Yano, Tomomasa

    2016-01-01

    Escherichia coli is the major causative agent of human cystitis. In this study, a preliminary molecular analysis carried out by PCR (polymerase chain reaction) demonstrated that 100% of 31 E. coli strains isolated from patients with recurrent UTIs (urinary tract infections) showed the presence of the curli fimbria gene (csgA). Curli fimbria is known to be associated with bacterial biofilm formation but not with the adhesion of human cystitis-associated E. coli. Therefore, this work aimed to study how curli fimbria is associated with uropathogenic E. coli (UPEC) as an adhesion factor. For this purpose, the csgA gene was deleted from strain UPEC-4, which carries three adhesion factor genes (csgA, fimH and ompA). The wild-type UPEC-4 strain and its mutant (ΔcsgA) were analyzed for their adhesion ability over HTB-9 (human bladder carcinoma), Vero (kidney cells of African green monkey) and HUVEC (human umbilical vein) cells in the presence of α-d-mannose. All the wild-type UPEC strains tested (100%) were able to adhere to all three cell types, while the UPEC-4 ΔcsgA mutant lost its adherence to HTB-9 but continued to adhere to the HUVEC and Vero cells. The results suggest that curli fimbria has an important role in the adhesion processes associated with human UPEC-induced cystitis.

  15. Isolation and characterization of fimbriae from a sparsely fimbriated strain of Porphyromonas gingivalis.

    PubMed Central

    Sojar, H T; Hamada, N; Genco, R J

    1997-01-01

    Porphyromonas gingivalis W50 (ATCC 53978) possesses the gene for fimbriae; however, the surface-expressed fimbriae are sparse and have not been previously isolated and characterized. We purified fimbriae from strain W50 to homogeneity by ammonium sulfate precipitation and reverse-phase high-performance liquid chromatography [H. T. Sojar, N. Hamada, and R. J. Genco, Protein Expr. Purif. 9(1):49-52, 1997]. Negative staining of purified fimbriae viewed by electron microscopy revealed that the fimbriae were identical in diameter to fimbriae of other P. gingivalis strains, such as 2561, but were shorter in length. On sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis, the apparent molecular weight of isolated fimbrillin from strain W50 was found to be identical to that of the fimbrillin molecule of strain 2561. Unlike 2561 fimbriae, W50 fimbriae, under reducing condition, exhibited a monomeric structure on SDS-PAGE at room temperature. However, under nonreduced conditions, even at 100 degrees C, no monomer was observed. In immunoblot analysis as well as immunogold labeling of isolated fimbriae, polyclonal antibodies against 2561 fimbriae, as well as antibodies against peptide I (V-V-M-A-N-T-G-A-M-E-V-G-K-T-L-A-E-V-K-Cys) and peptide J (A-L-T-T-E-L-T-A-E-N-Q-E-A-A-G-L-I-M-T-A-E-P-Cys), reacted. However, antifimbrial antibodies against strain 2561 reacted very weakly compared to anti-peptide I and anti-peptide J. Negative staining of whole W50 cells, as well as immunogold electron microscopy with anti-peptide I and anti-peptide J, showed fimbriae shorter in length and very few in number compared to those of strain 2561. Purified fimbriae showed no hemagglutinating activity. Amino acid composition was very similar to that of previously reported fimbriae of the 2561 strain. PMID:9172351

  16. Confirmation that DNA encoding the major fimbrial subunit of Av24 fimbriae is homologous to DNA encoding the major fimbrial subunit of F107 fimbriae.

    PubMed

    Kennan, R M; Moncktor, R P; McDougall, B M; Conway, P L

    1995-01-01

    Plasmid DNA from porcine enterotoxigenic Escherichia coli strain Av24 (O141:K85ab) was cloned into recipient E. coli strain JM105 using the plasmid vector pUC18. Clones were obtained that produced fimbriae which reacted with antisera specific to the fimbriae produced by strain Av24. Restriction mapping of cloned DNA, PCR with fedA primers and DNA sequencing showed a portion of the cloned DNA to be homologous to that encoding the major fimbrial subunit of F107 fimbriae. This confirms that the fimbriae possessed by strains of E. coli causing both edema disease and post-weaning diarrhoea in piglets are variants of the same fimbriae.

  17. Type 1 Fimbria-Mediated Adhesion of Enteric Bacteria to Grass Roots

    PubMed Central

    Haahtela, Kielo; Tarkka, Eveliina; Korhonen, Timo K.

    1985-01-01

    Type 1 fimbriae of Klebsiella pneumoniae and Enterobacter agglomerans mediated bacterial adhesion to the roots of bluegrass, Poa pratensis. Purified, radiolabeled fimbriae bound to grass roots in vitro; binding was inhibited by α-methyl-d-mannoside or Fab fragments to the fimbriae. Anti-type 1 fimbriae Fab fragments and α-methyl-d-mannoside also inhibited adhesion of type 1-fimbriated bacteria to P. pratensis roots. It is proposed that associative nitrogen fixation by Klebsiella and Enterobacter strains also involves type 1 fimbriae, in addition to the type 3 fimbriae of Klebsiella spp. (T. K. Korhonen, E. Tarkka, H. Ranta, and K. Haahtela, J. Bacteriol. 155:860-865, 1983). Images PMID:16346792

  18. Structure, Function, and Assembly of Type 1 Fimbriae

    NASA Astrophysics Data System (ADS)

    Knight, Stefan D.; Bouckaert, Julie

    Bacterial infections constitute a major global health problem, acutely accentuated by the rapid spread of antibiotic resistant bacterial strains. The widespread need for bacteria to attach - adhere - to target cells before they can initiate an infection may be used to advantage by targeting the bacterial adhesion tools such as pili and fimbriae for development of novel anti-bacterial vaccines and drugs. Type 1 fimbriae are widely expressed by Escherichia coli. and are used by uropathogenic strains to mediate attachment to specific niches in the urinary tract. These fimbriae belong to a class of fibrillar adhesion organelles assembled through the chaperone/usher pathway, one of the terminal branches of the general secretion pathway in Gram-negative bacteria. Our understanding of the assembly, structure and function of these structures has evolved significantly over the last decade. Here, we summarize current understanding of the function and biogenesis of fibrillar adhesion organelles, and provide some examples of recent progress towards interfering with bacterial adhesion as a means to prevent infection.

  19. Sequences related to the major subunit gene fedA of F107 fimbriae in porcine Escherichia coli strains that express adhesive fimbriae.

    PubMed

    Imberechts, H; Van Pelt, N; De Greve, H; Lintermans, P

    1994-06-15

    Porcine Escherichia coli strains isolated from cases of postweaning diarrhea or edema disease were analysed for the presence of fedA, the major subunit gene of F107 fimbriae. The E. coli isolates were known to contain colonisation factor '8813', or to express F107, 2134P or other fimbriae, different from F4, F5, F6, and F41. PCR with fedA-specific primers, restriction enzyme digestion of the PCR product, and nucleotide sequence analysis demonstrated that 2134P pili, colonisation factor '8813' and fimbriae identified on Australian strains of the O141 serotype belong to one family of F107 fimbrial antigens.

  20. Biomechanical and Structural Features of CS2 Fimbriae of Enterotoxigenic Escherichia coli

    PubMed Central

    Mortezaei, Narges; Singh, Bhupender; Zakrisson, Johan; Bullitt, Esther; Andersson, Magnus

    2015-01-01

    Enterotoxigenic Escherichia coli (ETEC) are a major cause of diarrhea worldwide, and infection of children in under-developed countries often leads to high mortality rates. Isolated ETEC expresses a plethora of colonization factors (fimbriae/pili), of which CFA/I and CFA/II, which are assembled via the alternate chaperone pathway (ACP), are among the most common. Fimbriae are filamentous structures whose shafts are primarily composed of helically arranged single pilin-protein subunits, with a unique biomechanical ability to unwind and rewind. A sustained ETEC infection, under adverse conditions of dynamic shear forces, is primarily attributed to this biomechanical feature of ETEC fimbriae. Recent understanding about the role of fimbriae as virulence factors points to an evolutionary adaptation of their structural and biomechanical features. In this work, we investigated the biophysical properties of CS2 fimbriae from the CFA/II group. Homology modeling of its major structural subunit, CotA, reveals structural clues related to the niche in which they are expressed. Using optical-tweezers force spectroscopy, we found that CS2 fimbriae unwind at a constant force of 10 pN and have a corner velocity (i.e., the velocity at which the force required for unwinding rises exponentially with increased speed) of 1300 nm/s. The biophysical properties of CS2 fimbriae assessed in this work classify them into a low-force unwinding group of fimbriae together with the CFA/I and CS20 fimbriae expressed by ETEC strains. The three fimbriae are expressed by ETEC, colonize in similar gut environments, and exhibit similar biophysical features, but differ in their biogenesis. Our observation suggests that the environment has a strong impact on the biophysical characteristics of fimbriae expressed by ETEC. PMID:26153701

  1. Biomechanical and structural features of CS2 fimbriae of enterotoxigenic Escherichia coli.

    PubMed

    Mortezaei, Narges; Singh, Bhupender; Zakrisson, Johan; Bullitt, Esther; Andersson, Magnus

    2015-07-07

    Enterotoxigenic Escherichia coli (ETEC) are a major cause of diarrhea worldwide, and infection of children in under-developed countries often leads to high mortality rates. Isolated ETEC expresses a plethora of colonization factors (fimbriae/pili), of which CFA/I and CFA/II, which are assembled via the alternate chaperone pathway (ACP), are among the most common. Fimbriae are filamentous structures whose shafts are primarily composed of helically arranged single pilin-protein subunits, with a unique biomechanical ability to unwind and rewind. A sustained ETEC infection, under adverse conditions of dynamic shear forces, is primarily attributed to this biomechanical feature of ETEC fimbriae. Recent understanding about the role of fimbriae as virulence factors points to an evolutionary adaptation of their structural and biomechanical features. In this work, we investigated the biophysical properties of CS2 fimbriae from the CFA/II group. Homology modeling of its major structural subunit, CotA, reveals structural clues related to the niche in which they are expressed. Using optical-tweezers force spectroscopy, we found that CS2 fimbriae unwind at a constant force of 10 pN and have a corner velocity (i.e., the velocity at which the force required for unwinding rises exponentially with increased speed) of 1300 nm/s. The biophysical properties of CS2 fimbriae assessed in this work classify them into a low-force unwinding group of fimbriae together with the CFA/I and CS20 fimbriae expressed by ETEC strains. The three fimbriae are expressed by ETEC, colonize in similar gut environments, and exhibit similar biophysical features, but differ in their biogenesis. Our observation suggests that the environment has a strong impact on the biophysical characteristics of fimbriae expressed by ETEC. Copyright © 2015 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  2. Interactions of Streptococcus mutans Fimbria-Associated Surface Proteins with Salivary Components

    PubMed Central

    Ray, Chad A.; Gfell, Linda E.; Buller, Tiffany L.; Gregory, Richard L.

    1999-01-01

    Streptococcus mutans has been implicated as the major causative agent of human dental caries. S. mutans binds to saliva-coated tooth surfaces, and previous studies suggested that fimbriae may play a role in the initial bacterial adherence to salivary components. The objectives of this study were to establish the ability of an S. mutans fimbria preparation to bind to saliva-coated surfaces and determine the specific salivary components that facilitate binding with fimbriae. Enzyme-linked immunosorbent assay (ELISA) established that the S. mutans fimbria preparation bound to components of whole saliva. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot techniques were used to separate components of whole saliva and determine fimbria binding. SDS-PAGE separated 15 major protein bands from saliva samples, and Western blot analysis indicated significant binding of the S. mutans fimbria preparation to a 52-kDa salivary protein. The major fimbria-binding salivary protein was isolated by preparative electrophoresis. The ability of the S. mutans fimbria preparation to bind to the purified salivary protein was confirmed by Western blot analysis and ELISA. Incubation of the purified salivary protein with the S. mutans fimbria preparation significantly neutralized binding of the salivary protein-fimbria complex to saliva-coated surfaces. The salivary protein, whole saliva, and commercial amylase reacted similarly with antiamylase antibody in immunoblots. A purified 65-kDa fimbrial protein was demonstrated to bind to both saliva and amylase. These data indicated that the S. mutans fimbria preparation and a purified fimbrial protein bound to whole-saliva-coated surfaces and that amylase is the major salivary component involved in the binding. PMID:10225843

  3. Molecular Interactions of Porphyromonas gingivalis Fimbriae with Host Proteins: Kinetic Analyses Based on Surface Plasmon Resonance

    PubMed Central

    Amano, Atsuo; Nakamura, Takayuki; Kimura, Shigenobu; Morisaki, Ichijiro; Nakagawa, Ichiro; Kawabata, Shigetada; Hamada, Shigeyuki

    1999-01-01

    Fimbriae of Porphyromonas gingivalis are thought to play an important role in the colonization and invasion of periodontal tissues. In this study, we analyzed the interactions of P. gingivalis fimbriae with human hemoglobin, fibrinogen, and salivary components (i.e., proline-rich protein [PRP], proline-rich glycoprotein [PRG], and statherin) based on surface plasmon resonance (SPR) spectroscopy with a biomolecular interaction analyzing system (BIAcore). The real-time observation showed that the fimbriae interacted more quickly with hemoglobin and PRG than with other proteins and more intensely with fibrinogen. The significant association constant (ka) values obtained by BIAcore demonstrated that the interactions between fimbriae and these host proteins are specific. These estimated Ka values were not too different; however, the Ka values for hemoglobin (2.43 × 106) and fibrinogen (2.16 × 106) were statistically greater than those for the salivary proteins (1.48 × 106 to 1.63 × 106). The Ka value of anti-fimbriae immunoglobulin G for fimbriae was estimated to be 1.22 × 107, which was 6.55-fold higher than the mean Ka value of the host proteins. Peptide PRP-C, a potent inhibitor of PRP-fimbriae interaction, dramatically inhibited fimbrial association to PRP and PRG and was also inhibitory against other host proteins by BIAcore. The binding of fimbriae to these proteins was also evaluated by other methods with hydroxyapatite beads or polystyrene microtiter plates. The estimated binding abilities differed considerably, depending on the assay method that was used. It was noted that the binding capacity of PRP was strongly diminished by immobilization on a polystyrene surface. Taken together, these findings suggest that P. gingivalis fimbriae possess a strong ability to interact with the host proteins which promote bacterial adherence to the oral cavity and that SPR spectroscopy is a useful method for analyzing specific protein-fimbriae interactions. PMID:10225901

  4. Fragmentation of Escherichia coli type 1 fimbriae exposes cryptic D-mannose-binding sites.

    PubMed Central

    Ponniah, S; Endres, R O; Hasty, D L; Abraham, S N

    1991-01-01

    Cells of the gram-negative bacterium Escherichia coli are able to attach to various host cells by means of a mannose-specific adhesin associated with type 1 fimbriae. Here we show that fragmentation of type 1 fimbriae by freezing and thawing results in increased mannose-binding activity as demonstrated by increased hemagglutination, increased stimulation of human lymphocyte proliferation, and increased binding of the mannose-containing enzyme horseradish peroxidase. Increased activity in all three assays was mannose sensitive and was not exhibited by FimH- mutant type 1 fimbriae lacking the adhesin. Scatchard analysis of the data from peroxidase binding assays showed that unfrozen and frozen fimbriae contain binding sites displaying two classes of affinity. Frozen and thawed fimbriae expressed an increase in the number of high-affinity binding sites. These results show that fragmentation of the fimbrial structure exposes cryptic mannose-binding activity associated with type 1 fimbriae, presumably that of internally located adhesin molecules. Our data support earlier observations that adhesin moieties of type 1 fimbriae are located both at the tips and at intervals along the length of the fimbriae. In addition, our data suggest that only the adhesin moieties that are located at the fimbrial tips are functional in binding mannose. Adhesins located along the length of the fimbriae have their mannose-binding activity buried within the fimbrial structure and hence are not functional. We propose an updated model for the structure of type 1 fimbriae that is in agreement with the above observations. Images PMID:1676398

  5. Mat fimbriae promote biofilm formation by meningitis-associated Escherichia coli.

    PubMed

    Lehti, Timo A; Bauchart, Philippe; Heikkinen, Johanna; Hacker, Jörg; Korhonen, Timo K; Dobrindt, Ulrich; Westerlund-Wikström, Benita

    2010-08-01

    The mat (or ecp) fimbrial operon is ubiquitous and conserved in Escherichia coli, but its functions remain poorly described. In routine growth media newborn meningitis isolates of E. coli express the meningitis-associated and temperature-regulated (Mat) fimbria, also termed E. coli common pilus (ECP), at 20 degrees C, and here we show that the six-gene (matABCDEF)-encoded Mat fimbria is needed for temperature-dependent biofilm formation on abiotic surfaces. The matBCDEF deletion mutant of meningitis E. coli IHE 3034 was defective in an early stage of biofilm development and consequently unable to establish a detectable biofilm, contrasting with IHE 3034 derivatives deleted for flagella, type 1 fimbriae or S-fimbriae, which retained the wild-type biofilm phenotype. Furthermore, induced production of Mat fimbriae from expression plasmids enabled biofilm-deficient E. coli K-12 cells to form biofilm at 20 degrees C. No biofilm was detected with IHE 3034 or MG1655 strains grown at 37 degrees C. The surface expression of Mat fimbriae and the frequency of Mat-positive cells in the IHE 3034 population from 20 degrees C were high and remained unaltered during the transition from planktonic to biofilm growth and within the matured biofilm community. Considering the prevalence of the highly conserved mat locus in E. coli genomes, we hypothesize that Mat fimbria-mediated biofilm formation is an ancestral characteristic of E. coli.

  6. Bacterial fimbriae stimulate proinflammatory activation in the endothelium through distinct TLRs.

    PubMed

    Davey, Michael; Liu, Xinyan; Ukai, Takashi; Jain, Vishal; Gudino, Cynthia; Gibson, Frank C; Golenbock, Douglas; Visintin, Alberto; Genco, Caroline A

    2008-02-15

    The major and minor fimbriae proteins produced by the human pathogen Porphyromonas gingivalis are required for invasion of human aortic endothelial cells and for the stimulation of potent inflammatory responses. In this study, we report that native forms of both the major and minor fimbriae proteins bind to and signal through TLR2 for this response. Major and minor fimbriae bound to a human TLR2:Fc chimeric protein with an observed K(d) of 28.9 nM and 61.7 nM, respectively. Direct binding of the major and minor fimbriae to a human chimeric CD14-Fc protein also established specific binding of the major and minor fimbriae to CD14 with classic saturation kinetics. Using a P. gingivalis major and minor fimbriae mutant, we confirmed that TLR2 binding in whole cells is dependent on the expression of the major and minor fimbriae. Although we did not observe binding with the major or minor fimbriae to the TLR4-Fc chimeric protein, signaling through TLR4 for both proteins was demonstrated in human embryonic kidney 293 cells transfected with TLR4 and only in the presence MD-2. Transient transfection of dominant-negative forms of TLR2 or TLR4 reduced IL-8 production by human aortic endothelial cells following stimulation with major or minor fimbriae. The ability of two well-defined microbe-associated molecular patterns to select for innate immune recognition receptors based on accessory proteins may provide a novel way for a pathogen to sense and signal in appropriate host environments.

  7. Characterization of 17 chaperone-usher fimbriae encoded by Proteus mirabilis reveals strong conservation

    PubMed Central

    Kuan, Lisa; Schaffer, Jessica N.; Zouzias, Christos D.

    2014-01-01

    Proteus mirabilis is a Gram-negative enteric bacterium that causes complicated urinary tract infections, particularly in patients with indwelling catheters. Sequencing of clinical isolate P. mirabilis HI4320 revealed the presence of 17 predicted chaperone-usher fimbrial operons. We classified these fimbriae into three groups by their genetic relationship to other chaperone-usher fimbriae. Sixteen of these fimbriae are encoded by all seven currently sequenced P. mirabilis genomes. The predicted protein sequence of the major structural subunit for 14 of these fimbriae was highly conserved (≥95 % identity), whereas three other structural subunits (Fim3A, UcaA and Fim6A) were variable. Further examination of 58 clinical isolates showed that 14 of the 17 predicted major structural subunit genes of the fimbriae were present in most strains (>85 %). Transcription of the predicted major structural subunit genes for all 17 fimbriae was measured under different culture conditions designed to mimic conditions in the urinary tract. The majority of the fimbrial genes were induced during stationary phase, static culture or colony growth when compared to exponential-phase aerated culture. Major structural subunit proteins for six of these fimbriae were detected using MS of proteins sheared from the surface of broth-cultured P. mirabilis, demonstrating that this organism may produce multiple fimbriae within a single culture. The high degree of conservation of P. mirabilis fimbriae stands in contrast to uropathogenic Escherichia coli and Salmonella enterica, which exhibit greater variability in their fimbrial repertoires. These findings suggest there may be evolutionary pressure for P. mirabilis to maintain a large fimbrial arsenal. PMID:24809384

  8. Characterization of 17 chaperone-usher fimbriae encoded by Proteus mirabilis reveals strong conservation.

    PubMed

    Kuan, Lisa; Schaffer, Jessica N; Zouzias, Christos D; Pearson, Melanie M

    2014-07-01

    Proteus mirabilis is a Gram-negative enteric bacterium that causes complicated urinary tract infections, particularly in patients with indwelling catheters. Sequencing of clinical isolate P. mirabilis HI4320 revealed the presence of 17 predicted chaperone-usher fimbrial operons. We classified these fimbriae into three groups by their genetic relationship to other chaperone-usher fimbriae. Sixteen of these fimbriae are encoded by all seven currently sequenced P. mirabilis genomes. The predicted protein sequence of the major structural subunit for 14 of these fimbriae was highly conserved (≥ 95% identity), whereas three other structural subunits (Fim3A, UcaA and Fim6A) were variable. Further examination of 58 clinical isolates showed that 14 of the 17 predicted major structural subunit genes of the fimbriae were present in most strains (>85%). Transcription of the predicted major structural subunit genes for all 17 fimbriae was measured under different culture conditions designed to mimic conditions in the urinary tract. The majority of the fimbrial genes were induced during stationary phase, static culture or colony growth when compared to exponential-phase aerated culture. Major structural subunit proteins for six of these fimbriae were detected using MS of proteins sheared from the surface of broth-cultured P. mirabilis, demonstrating that this organism may produce multiple fimbriae within a single culture. The high degree of conservation of P. mirabilis fimbriae stands in contrast to uropathogenic Escherichia coli and Salmonella enterica, which exhibit greater variability in their fimbrial repertoires. These findings suggest there may be evolutionary pressure for P. mirabilis to maintain a large fimbrial arsenal. © 2014 The Authors.

  9. Antibodies Damage the Resilience of Fimbriae, Causing Them To Be Stiff and Tangled

    PubMed Central

    Singh, Bhupender; Mortezaei, Narges; Savarino, Stephen J.; Uhlin, Bernt Eric; Bullitt, Esther

    2016-01-01

    ABSTRACT As adhesion fimbriae are a major virulence factor for many pathogenic Gram-negative bacteria, they are also potential targets for antibodies. Fimbriae are commonly required for initiating the colonization that leads to disease, and their success as adhesion organelles lies in their ability to both initiate and sustain bacterial attachment to epithelial cells. The ability of fimbriae to unwind and rewind their helical filaments presumably reduces their detachment from tissue surfaces with the shear forces that accompany significant fluid flow. Therefore, the disruption of functional fimbriae by inhibiting this resilience should have high potential for use as a vaccine to prevent disease. In this study, we show that two characteristic biomechanical features of fimbrial resilience, namely, the extension force and the extension length, are significantly altered by the binding of antibodies to fimbriae. The fimbriae that were studied are normally expressed on enterotoxigenic Escherichia coli, which are a major cause of diarrheal disease. This alteration in biomechanical properties was observed with bivalent polyclonal antifimbrial antibodies that recognize major pilin subunits but not with the Fab fragments of these antibodies. Thus, we propose that the mechanism by which bound antibodies disrupt the uncoiling of natural fimbria under force is by clamping together layers of the helical filament, thereby increasing their stiffness and reducing their resilience during fluid flow. In addition, we propose that antibodies tangle fimbriae via bivalent binding, i.e., by binding to two individual fimbriae and linking them together. Use of antibodies to disrupt physical properties of fimbriae may be generally applicable to the large number of Gram-negative bacteria that rely on these surface-adhesion molecules as an essential virulence factor. IMPORTANCE Our study shows that the resiliency of colonization factor antigen I (CFA/I) and coli surface antigen 2 (CS2

  10. Antibodies Damage the Resilience of Fimbriae, Causing Them To Be Stiff and Tangled.

    PubMed

    Singh, Bhupender; Mortezaei, Narges; Savarino, Stephen J; Uhlin, Bernt Eric; Bullitt, Esther; Andersson, Magnus

    2017-01-01

    As adhesion fimbriae are a major virulence factor for many pathogenic Gram-negative bacteria, they are also potential targets for antibodies. Fimbriae are commonly required for initiating the colonization that leads to disease, and their success as adhesion organelles lies in their ability to both initiate and sustain bacterial attachment to epithelial cells. The ability of fimbriae to unwind and rewind their helical filaments presumably reduces their detachment from tissue surfaces with the shear forces that accompany significant fluid flow. Therefore, the disruption of functional fimbriae by inhibiting this resilience should have high potential for use as a vaccine to prevent disease. In this study, we show that two characteristic biomechanical features of fimbrial resilience, namely, the extension force and the extension length, are significantly altered by the binding of antibodies to fimbriae. The fimbriae that were studied are normally expressed on enterotoxigenic Escherichia coli, which are a major cause of diarrheal disease. This alteration in biomechanical properties was observed with bivalent polyclonal antifimbrial antibodies that recognize major pilin subunits but not with the Fab fragments of these antibodies. Thus, we propose that the mechanism by which bound antibodies disrupt the uncoiling of natural fimbria under force is by clamping together layers of the helical filament, thereby increasing their stiffness and reducing their resilience during fluid flow. In addition, we propose that antibodies tangle fimbriae via bivalent binding, i.e., by binding to two individual fimbriae and linking them together. Use of antibodies to disrupt physical properties of fimbriae may be generally applicable to the large number of Gram-negative bacteria that rely on these surface-adhesion molecules as an essential virulence factor. Our study shows that the resiliency of colonization factor antigen I (CFA/I) and coli surface antigen 2 (CS2) fimbriae, which are current

  11. Innate immune-stimulatory activity of Porphyromonas gingivalis fimbriae is eliminated by phase separation using Triton X-114.

    PubMed

    Nozoe, Kohji; Sanui, Terukazu; Takeshita, Masaaki; Fukuda, Takao; Haraguchi, Akira; Aida, Yoshitomi; Nishimura, Fusanori

    2017-02-01

    Fimbriae are virulence factors of Porphyromonas gingivalis (P. gingivalis). In this study, the action of fimbriae on neutrophil respiratory burst and cytokine production by mononuclear cells (MNC) were investigated. Native or denatured form of purified P. gingivalis fimbriae contained endotoxin at an equivalence of 1-3μglipopolysaccharides(LPS)/mg protein. The endotoxin could be reduced to the equivalent of 1ng-LPS/mg protein by phase separation using Triton X-114. Unfractionated fimbriae caused serum-dependent priming of neutrophils for enhanced respiratory burst, but both native and denatured forms of Triton X-114-fractionated fimbriae were not active at 100μg/mL. Unfractionated fimbriae induced serum-dependent production of IL-1β by MNC. Triton X-114-fractionated fimbriae (10μg/mL)-induced production of IL-1β, IL-8 or TNF-α was much lower than that induced by unfractionated fimbriae or 10ng/mL P. gingivalis-LPS preparation. Triton X-114-fractionated fimbriae immobilized on polystyrene tubes induced adhesion-stimulated superoxide release by LPS-primed neutrophils in a β2 integrin-dependent manner. P. gingivalis cells caused priming of neutrophils; however, Toll-like receptor (TLR) 4 antagonists did not affect this response. Thus, P. gingivalis fimbriae were ineffective in inducing innate immune response in leukocytes; however, they induced β2 integrin-mediated response by neutrophils. Immune-stimulatory components of P. gingivalis might be recognized by receptors other than TLR4.

  12. How type 1 fimbriae help Escherichia coli to evade extracellular antibiotics.

    PubMed

    Avalos Vizcarra, Ima; Hosseini, Vahid; Kollmannsberger, Philip; Meier, Stefanie; Weber, Stefan S; Arnoldini, Markus; Ackermann, Martin; Vogel, Viola

    2016-01-05

    To survive antibiotics, bacteria use two different strategies: counteracting antibiotic effects by expression of resistance genes or evading their effects e.g. by persisting inside host cells. Since bacterial adhesins provide access to the shielded, intracellular niche and the adhesin type 1 fimbriae increases bacterial survival chances inside macrophages, we asked if fimbriae also influenced survival by antibiotic evasion. Combined gentamicin survival assays, flow cytometry, single cell microscopy and kinetic modeling of dose response curves showed that type 1 fimbriae increased the adhesion and internalization by macrophages. This was caused by strongly decreased off-rates and affected the number of intracellular bacteria but not the macrophage viability and morphology. Fimbriae thus promote antibiotic evasion which is particularly relevant in the context of chronic infections.

  13. How type 1 fimbriae help Escherichia coli to evade extracellular antibiotics

    PubMed Central

    Avalos Vizcarra, Ima; Hosseini, Vahid; Kollmannsberger, Philip; Meier, Stefanie; Weber, Stefan S.; Arnoldini, Markus; Ackermann, Martin; Vogel, Viola

    2016-01-01

    To survive antibiotics, bacteria use two different strategies: counteracting antibiotic effects by expression of resistance genes or evading their effects e.g. by persisting inside host cells. Since bacterial adhesins provide access to the shielded, intracellular niche and the adhesin type 1 fimbriae increases bacterial survival chances inside macrophages, we asked if fimbriae also influenced survival by antibiotic evasion. Combined gentamicin survival assays, flow cytometry, single cell microscopy and kinetic modeling of dose response curves showed that type 1 fimbriae increased the adhesion and internalization by macrophages. This was caused by strongly decreased off-rates and affected the number of intracellular bacteria but not the macrophage viability and morphology. Fimbriae thus promote antibiotic evasion which is particularly relevant in the context of chronic infections. PMID:26728082

  14. Spatial and Temporal Deposition of Adhesive Extracellular Polysaccharide Capsule and Fimbriae by Hyphomonas Strain MHS-3

    PubMed Central

    Quintero, Ernesto J.; Busch, Kathryn; Weiner, Ronald M.

    1998-01-01

    Hyphomonas strain MHS-3, a member of a genus of primary colonizers of surfaces immersed in marine water, synthesizes two structures that mediate adhesion to solid substrata, namely, capsular exopolysaccharide and fimbriae. Specific stains, gold-labelled lectins, and monoclonal antibodies, along with transmission electron microscopy of synchronized populations, revealed that both structures are polarly and temporally expressed. The timed synthesis and placement of the fimbriae and capsule correlated with the timing and locus of MHS-3 adhesion. PMID:16349537

  15. Fimbriae and lipopolysaccharides are necessary for co-aggregation between Lactobacilli and Escherichia coli.

    PubMed

    Mizuno, Kouhei; Furukawa, Soichi; Usui, Yumi; Ishiba, Madoka; Ogihara, Hirokazu; Morinaga, Yasushi

    2014-01-01

    Cells of Lactobacilli co-aggregated with Escherichia coli K-12 cells to form co-aggregates under mixed-culture conditions at 37 °C for 24 h. Co-aggregation was inhibited by sodium dodecyl sulfate but not by protease. E. coli deletion mutants of fimbriae formation and lipopolysaccharide (LPS) formation did not co-aggregate with Lactobacilli. These results showed that fimbriae and LPS are necessary for co-aggregation between Lactobacilli and E. coli.

  16. Proteus mirabilis ambient-temperature fimbriae: cloning and nucleotide sequence of the aft gene cluster.

    PubMed Central

    Massad, G; Fulkerson, J F; Watson, D C; Mobley, H L

    1996-01-01

    Uropathogenic Proteus mirabilis produces at least four types of fimbriae. Amino acid sequences from two peptides, derived by tryptic digestion of the structural subunit of one type of these fimbriae, the ambient-temperature fimbriae, were determined: NVVPGQPSSTQ and LIEGENQLNYNA. PCR primers, based on these sequences and that of the N terminus, were used to amplify a 359-bp fragment. A cosmid clone, isolated from a P. mirabilis genomic library by hybridization with the 359-bp PCR product, was used to determine the nucleotide sequence of the atf gene cluster. A 3,903-bp region encodes three polypeptides: AtfA, the structural subunit; AtfB, the chaperone; and AtfC, the outer membrane molecular usher. No fimbria-related genes are evident either 5' or 3' to the three contiguous genes. AtfA demonstrates significant amino acid sequence identity with type 1 major fimbrial subunits of several enteric species. The 359-bp PCR product hybridized strongly with all Proteus isolates (n = 9) and 25% of 355 Escherichia coli isolates but failed to hybridize with any of 26 isolates among nine other uropathogenic species. Ambient-temperature fimbriae of P. mirabilis may represent a novel type of fimbriae of enteric species. PMID:8926119

  17. Proteus mirabilis ambient-temperature fimbriae: cloning and nucleotide sequence of the aft gene cluster.

    PubMed

    Massad, G; Fulkerson, J F; Watson, D C; Mobley, H L

    1996-10-01

    Uropathogenic Proteus mirabilis produces at least four types of fimbriae. Amino acid sequences from two peptides, derived by tryptic digestion of the structural subunit of one type of these fimbriae, the ambient-temperature fimbriae, were determined: NVVPGQPSSTQ and LIEGENQLNYNA. PCR primers, based on these sequences and that of the N terminus, were used to amplify a 359-bp fragment. A cosmid clone, isolated from a P. mirabilis genomic library by hybridization with the 359-bp PCR product, was used to determine the nucleotide sequence of the atf gene cluster. A 3,903-bp region encodes three polypeptides: AtfA, the structural subunit; AtfB, the chaperone; and AtfC, the outer membrane molecular usher. No fimbria-related genes are evident either 5' or 3' to the three contiguous genes. AtfA demonstrates significant amino acid sequence identity with type 1 major fimbrial subunits of several enteric species. The 359-bp PCR product hybridized strongly with all Proteus isolates (n = 9) and 25% of 355 Escherichia coli isolates but failed to hybridize with any of 26 isolates among nine other uropathogenic species. Ambient-temperature fimbriae of P. mirabilis may represent a novel type of fimbriae of enteric species.

  18. Salmonella enteritidis agfBAC operon encoding thin, aggregative fimbriae.

    PubMed

    Collinson, S K; Clouthier, S C; Doran, J L; Banser, P A; Kay, W W

    1996-02-01

    Salmonella enteritidis produces thin, aggregative fimbriae, named SEF17, which are composed of polymerized AgfA fimbrin proteins. DNA sequence analysis of a 2-kb region of S. enteritidis DNA revealed three contiguous genes, agfBAC. The 453-bp agfA gene encodes the AgfA fimbrin, which was predicted to be 74% identical and 86% similar in primary sequence to the Escherichia coli curli structural protein, CsgA. pHAG, a pUC18 derivative containing a 3.0-kb HindIII fragment encoding agfBAC, directed the in vitro expression of the major AgfA fimbrin, with an M(r) of 17,000, and a minor AgfB protein, with an M(r) of 16,000, encoded by the 453-bp agfB gene. AgfA was not expressed from pDAG, a pUC18 derivative containing a 3.1-kb DraI DNA fragment encoding agfA but not agfB. Primer extension analysis identified two adjacent transcription start sites located immediately upstream of agfB in positions analogous to those of the E. coli curlin csgBA operon. No transcription start sites were located immediately upstream of agfA or agfC. Northern (RNA) blot analysis confirmed that transcription of agfA was initiated from the agfB promoter region. Secondary-structure analysis of the putative mRNA transcript for agfBAC predicted the formation of a stem-loop structure (delta Gzero, -22 kcal/mol [-91 kJ/mol]) in the intercistronic region between agfA and agfC, which may be involved in stabilization of the agfBA portion of the agfBAC transcript. agfBAC and flanking regions had a high degree of sequence similarity with those counterparts of the E. coli curlin csgBA region for which sequence data are available. These data are demonstrative of the high degree of similarity between S. enteritidis SEF17 fimbriae and E. coli curli with respect to fimbrin amino acid sequence and genetic organization and, therefore, are indicative of a common and relatively recent ancestry.

  19. Evidence that Porphyromonas (Bacteroides) gingivalis fimbriae function in adhesion to Actinomyces viscosus.

    PubMed Central

    Goulbourne, P A; Ellen, R P

    1991-01-01

    Porphyromonas (Bacteroides) gingivalis adheres to gram-positive bacteria, such as Actinomyces viscosus, when colonizing the tooth surface. However, little is known of the adhesins responsible for this interaction. A series of experiments were performed to determine whether P. gingivalis fimbriae function in its coadhesion with A. viscosus. Fimbriae typical of P. gingivalis were isolated from strain 2561 (ATCC 33277) by the method of Yoshimura et al. (F. Yoshimura, K. Takahashi, Y. Nodasaka, and T. Suzuki, J. Bacteriol. 160:949-957, 1984) in fractions enriched with a 40-kDa subunit, the fimbrillin monomer, P. gingivalis-A. viscosus coaggregation was inhibited by purified rabbit antifimbrial immunoglobulin G (IgG) at dilutions eightfold higher than those of preimmune IgG, providing indirect evidence implicating P. gingivalis fimbriae in coadhesion. Three types of direct binding assays further supported this observation. (i) Mixtures of isolated P. gingivalis fimbriae and A. viscosus WVU627 cells were incubated for 1 h, washed vigorously with phosphate-buffered saline (pH 7.2), and subjected to electrophoresis. Transblots onto nitrocellulose were probed with antifimbrial antiserum. Fimbrillin labeled positively on these blots. No reaction occurred with the control protein, porcine serum albumin, when blots were exposed to anti-porcine serum albumin, (ii) A. viscosus cells incubated with P. gingivalis fimbriae were agglutinated only after the addition of antifimbrial antibodies. (iii) Binding curves generated from an enzyme immunoassay demonstrated concentration-dependent binding of P. gingivalis fimbriae to A. viscosus cells. From these lines of evidence, P. gingivalis fimbriae appear to be capable of binding to A. viscosus and mediating the coadhesion of these species. Images PMID:1679428

  20. Off-pathway assembly of fimbria subunits is prevented by chaperone CfaA of CFA/I fimbriae from enterotoxigenic E. coli.

    PubMed

    Bao, Rui; Liu, Yang; Savarino, Stephen J; Xia, Di

    2016-12-01

    The assembly of the class 5 colonization factor antigen I (CFA/I) fimbriae of enterotoxigenic E. coli was proposed to proceed via the alternate chaperone-usher pathway. Here, we show that in the absence of the chaperone CfaA, CfaB, the major pilin subunit of CFA/I fimbriae, is able to spontaneously refold and polymerize into cyclic trimers. CfaA kinetically traps CfaB to form a metastable complex that can be stabilized by mutations. Crystal structure of the stabilized complex reveals distinctive interactions provided by CfaA to trap CfaB in an assembly competent state through donor-strand complementation (DSC) and cleft-mediated anchorage. Mutagenesis indicated that DSC controls the stability of the chaperone-subunit complex and the cleft-mediated anchorage of the subunit C-terminus additionally assist in subunit refolding. Surprisingly, over-stabilization of the chaperone-subunit complex led to delayed fimbria assembly, whereas destabilizing the complex resulted in no fimbriation. Thus, CfaA acts predominantly as a kinetic trap by stabilizing subunit to avoid its off-pathway self-polymerization that results in energetically favorable trimers and could serve as a driving force for CFA/I pilus assembly, representing an energetic landscape unique to class 5 fimbria assembly. Published 2016. This article is a U.S. Government work and is in the public domain in the USA. Molecular Microbiology published by John Wiley & Sons Ltd.

  1. Role of overexpressed CFA/I fimbriae in bacterial swimming

    NASA Astrophysics Data System (ADS)

    Cao, Ling; Suo, Zhiyong; Lim, Timothy; Jun, SangMu; Deliorman, Muhammedin; Riccardi, Carol; Kellerman, Laura; Avci, Recep; Yang, Xinghong

    2012-06-01

    Enterotoxigenic Escherichia coli CFA/I is a protective antigen and has been overexpressed in bacterial vectors, such as Salmonella Typhimurium H683, to generate vaccines. Effects that overexpressed CFA/I may engender on the bacterial host remain largely unexplored. To investigate, we constructed a high CFA/I expression strain, H683-pC2, and compared it to a low CFA/I expression strain, H683-pC, and to a non-CFA/I expression strain, H683-pY. The results showed that H683-pC2 was less able to migrate into semisolid agar (0.35%) than either H683-pC or H683-pY. Bacteria that migrated showed motility halo sizes of H683-pC2 < H683-pC < H683-pY. In the liquid culture media, H683-pC2 cells precipitated to the bottom of the tube, while those of H683-pY did not. In situ imaging revealed that H683-pC2 bacilli tended to auto-agglutinate within the semisolid agar, while H683-pY bacilli did not. When the cfaBE fimbrial fiber encoding genes were deleted from pC2, the new plasmid, pC2(-), significantly recovered bacterial swimming capability. Our study highlights the negative impact of overexpressed CFA/I fimbriae on bacterial swimming motility.

  2. Dimethylsulfoniopropionate (DMSP) Increases Survival of Larval Sablefish, Anoplopoma fimbria.

    PubMed

    Lee, Jonathan S F; Poretsky, Rachel S; Cook, Matthew A; Reyes-Tomassini, Jose J; Berejikian, Barry A; Goetz, Frederick W

    2016-06-01

    High concentrations of dimethylsulfoniopropionate (DMSP), a chemical compound released by lysed phytoplankton, may indicate high rates of grazing by zooplankton and may thus be a foraging cue for planktivorous fishes. Previous studies have shown that some planktivorous fishes and birds aggregate or alter locomotory behavior in response to this chemical cue, which is likely adaptive because it helps them locate prey. These behavioral responses have been demonstrated in juveniles and adults, but no studies have tested for effects on larval fish. Larvae suffer from high mortality rates and are vulnerable to starvation. While larvae are generally thought to be visual predators, they actually have poor vision and cryptic prey. Thus, larval fish should benefit from a chemical cue that provides information on prey abundance. We reared larval sablefish, Anoplopoma fimbria, for one week and supplemented feedings with varying concentrations of DMSP to test the hypothesis that DMSP affects larval survival. Ecologically relevant DMSP concentrations increased larval survival by up to 70 %, which has implications for production in aquaculture and recruitment in nature. These results provide a new tool for increasing larval production in aquaculture and also suggest that larvae may use DMSP as an olfactory cue. The release of DMSP may be a previously unappreciated mechanism through which phytoplankton affect larval survival and recruitment.

  3. Role of overexpressed CFA/I fimbriae in bacterial swimming.

    PubMed

    Cao, Ling; Suo, Zhiyong; Lim, Timothy; Jun, Sangmu; Deliorman, Muhammedin; Riccardi, Carol; Kellerman, Laura; Avci, Recep; Yang, Xinghong

    2012-06-01

    Enterotoxigenic Escherichia coli CFA/I is a protective antigen and has been overexpressed in bacterial vectors, such as Salmonella Typhimurium H683, to generate vaccines. Effects that overexpressed CFA/I may engender on the bacterial host remain largely unexplored. To investigate, we constructed a high CFA/I expression strain, H683-pC2, and compared it to a low CFA/I expression strain, H683-pC, and to a non-CFA/I expression strain, H683-pY. The results showed that H683-pC2 was less able to migrate into semisolid agar (0.35%) than either H683-pC or H683-pY. Bacteria that migrated showed motility halo sizes of H683-pC2 < H683-pC < H683-pY. In the liquid culture media, H683-pC2 cells precipitated to the bottom of the tube, while those of H683-pY did not. In situ imaging revealed that H683-pC2 bacilli tended to auto-agglutinate within the semisolid agar, while H683-pY bacilli did not. When the cfaBE fimbrial fiber encoding genes were deleted from pC2, the new plasmid, pC2(-), significantly recovered bacterial swimming capability. Our study highlights the negative impact of overexpressed CFA/I fimbriae on bacterial swimming motility.

  4. Distinct Mutations Led to Inactivation of Type 1 Fimbriae Expression in Shigella spp.

    PubMed Central

    Bravo, Verónica; Puhar, Andrea; Sansonetti, Philippe; Parsot, Claude; Toro, Cecilia S.

    2015-01-01

    Shigella spp. are responsible for bacillary dysentery in humans. The acquisition or the modification of the virulence plasmid encoding factors promoting entry of bacteria into and dissemination within epithelial cells was a critical step in the evolution of these bacteria from their Escherichia coli ancestor(s). Incorporation of genomic islands (GI) and gene inactivation also shaped interactions between these pathogens and their human host. Sequence analysis of the GI inserted next to the leuX tRNA gene in S. boydii, S. dysenteriae, S. flexneri, S. sonnei and enteroinvasive E. coli (EIEC) suggests that this region initially carried the fec, yjhATS and fim gene clusters. The fim cluster encoding type I fimbriae is systematically inactivated in both reference strains and clinical isolates and distinct mutations are responsible for this inactivation in at least three phylogenetic groups. To investigate consequences of the presence of fimbriae on the outcome of the interaction of Shigella with host cells, we used a S. flexneri strain harboring a plasmid encoding the E. coli fim operon. Production of fimbriae by this recombinant strain increased the ability of bacteria to adhere to and enter into epithelial cells and had no effect on their ability to disseminate from cell to cell. The observations that production of type I fimbriae increases invasion of epithelial cells and that independent mutations abolish fimbriae production in Shigella suggest that these mutations correspond to pathoadaptive events. PMID:25811616

  5. Mucosal vaccination of mice with recombinant Proteus mirabilis structural fimbrial proteins.

    PubMed

    Scavone, Paola; Sosa, Vanessa; Pellegrino, Rafael; Galvalisi, Umberto; Zunino, Pablo

    2004-07-01

    Proteus mirabilis, a common cause of urinary tract infection (UTI), expresses several types of fimbria including mannose-resistant/Proteus-like fimbriae (MRP), uroepithelial cell adhesin (UCA), renamed non-agglutinating fimbriae (NAF) by some authors, and P. mirabilis fimbriae (PMF), which are potentially involved in adhesion to the uroepithelium. In this study, we immunised different groups of mice with recombinant structural subunits of these fimbriae (MrpA, UcaA and PmfA) using two mucosal routes (nasal and transurethral) and we transurethrally challenged the animals with a P. mirabilis uropathogenic isolate. Induction of specific serum and urine IgG and IgA was measured to assess the potential role of the humoral immune response in protection against experimental ascending P. mirabilis UTI. Intranasally MrpA- and UcaA-immunised mice were protected against P. mirabilis ascending UTI, since recovery of bacteria from kidneys and bladders was significantly lower than in PBS-treated mice, and both fimbrial subunits significantly induced specific serum and urine antibodies. Only MrpA and PmfA transurethrally immunised animals were protected only at the kidney level, and in this case only MrpA-immunised mice exhibited significant serum IgG induction. Correlation analysis did not show a significant relationship between serum and urine specific antibody response and protection observed against infection. Our results suggest that an immunisation strategy based on structural fimbrial proteins may be useful to prevent P. mirabilis UTI. Further studies are being carried out to characterise the immune and inflammatory response induced by P. mirabilis recombinant fimbrial subunits.

  6. Functional differences of Porphyromonas gingivalis Fimbriae in determining periodontal disease pathogenesis: a literature review.

    PubMed

    Moreno, Sandra; Contreras, Adolfo

    2013-01-01

    Porphyromonas gingivalis is implicated in chronic and aggressive periodontitis. This bacterium has numerous virulence factors and one is the Fimbriae, which is quite important for bacterial colonization. Fimbriae are appendices that anchor to the bacterial wall and are comprised of the protein FimBriline encoded by the FimA gene. Thus far, six genotypes have been identified, FimA I to V and Ib. Genotypes II and IV are associated with periodontal disease, while genotype I is related to gingival health. Genotype identification of P. gingivalis FimA in periodontitis would be important to confirm the pathogenic genotypes and to establish risk at population level. This review is about the P. gingivalis FimA genotype prevalence worldwide. A systematic search using Pubmed, Hinary, and Science Direct within the following descriptors: Porphyromonas gingivalis, bacterial adhesion, periodontitis, Fimbriae, FimA, genotipification was performed to April 2011.

  7. Expression of type 1 fimbriae and mannose-sensitive hemagglutinin by recombinant plasmids.

    PubMed Central

    Gerlach, G F; Clegg, S; Ness, N J; Swenson, D L; Allen, B L; Nichols, W A

    1989-01-01

    Deletions within the cloned genes (fimA) encoding the type 1 major fimbrial subunits of two isolates of Klebsiella pneumoniae resulted in a nonfimbriate but hemagglutinating phenotype after transformation of Escherichia coli HB101 or ORN103. Phenotypic expression of type 1 fimbriae could be restored by transformation with plasmids containing the fimA genes of the fimbrial gene clusters from different strains. The surface fimbriae expressed were serologically identical to those of the polymerized product of the introduced fimA gene. The fimA gene products of Salmonella typhimurium and Serratia marcescens could utilize the accessory fimbrial genes of K. pneumoniae to produce surface-associated, hemagglutinating fimbriae. The relatedness of the type 1 fimbrial gene clusters from multiple isolates of members of the family Enterobacteriaceae was examined by DNA hybridization techniques. These analyses demonstrated little nucleotide sequence agreement among distinct genera of the enteric bacteria. Images PMID:2563717

  8. Functional differences of Porphyromonas gingivalis Fimbriae in determining periodontal disease pathogenesis: a literature review

    PubMed Central

    Contreras, Adolfo

    2013-01-01

    Porphyromonas gingivalis is implicated in chronic and aggressive periodontitis. This bacterium has numerous virulence factors and one is the Fimbriae, which is quite important for bacterial colonization. Fimbriae are appendices that anchor to the bacterial wall and are comprised of the protein FimBriline encoded by the FimA gene. Thus far, six genotypes have been identified, FimA I to V and Ib. Genotypes II and IV are associated with periodontal disease, while genotype I is related to gingival health. Genotype identification of P. gingivalis FimA in periodontitis would be important to confirm the pathogenic genotypes and to establish risk at population level. This review is about the P. gingivalis FimA genotype prevalence worldwide. A systematic search using Pubmed, Hinary, and Science Direct within the following descriptors: Porphyromonas gingivalis, bacterial adhesion, periodontitis, Fimbriae, FimA, genotipification was performed to April 2011. PMID:24892323

  9. Monoclonal antibodies recognising fimbriae F107 (F18) of an oedema disease causing strain of Escherichia coli.

    PubMed

    Rosocha, J; Wray, C; Mikula, I

    1999-06-01

    Escherichia coli isolated from experimentally induced oedema disease in pigs was used for the isolation and purification of F107 fimbriae. The reference strain was probed using membrane DNA hybridisation for the presence of fed A gene. F107 fimbriae were purified on FPLC and purity was checked on HPLC and SDS PAGE. A protein with major subunit of 18.9 kDa was used for Mabs preparation. Mabs reacted with 18.9 kDa protein previously classified as a major fimbrial subunit and were able to detect F107 fimbriae in immunoelectron microscopy on the surface of the strains 107/86 and 8872. Other strains used in this study did not express any fimbriae. Western blot analysis and F107 ELISA confirmed, that Mabs react with 18.9 kDa subunit whereas strains passaged many times in laboratory did not express F107 fimbriae.

  10. P-fimbriae in the presence of anti-PapA antibodies: new insight of antibodies action against pathogens

    NASA Astrophysics Data System (ADS)

    Mortezaei, Narges; Singh, Bhupender; Bullitt, Esther; Uhlin, Bernt Eric; Andersson, Magnus

    2013-12-01

    Uropathogenic strains of Escherichia coli establish urinary tract infections by attaching to host epithelial cells using adhesive organelles called fimbriae. Fimbriae are helix-like structures with a remarkable adaptability, offering safeguarding for bacteria exposed to changing fluid forces in the urinary tract. We challenged this property of P-fimbriae by cross-linking their subunits with shaft-specific antibodies and measuring the corresponding force response at a single organelle level. Our data show compromised extension and rewinding of P-fimbriae in the presence of antibodies and reduced fimbrial elasticity, which are important properties of fimbriae contributing to the ability of bacteria to cause urinary tract infections. The reduced elasticity found by cross-linking fimbrial subunits could thus be another assignment for antibodies; in addition to marking bacteria as foreign, antibodies physically compromise fimbrial function. We suggest that our assay and results will be a starting point for further investigations aimed at inhibiting sustained bacterial adhesion by antibodies.

  11. S-fimbriae from Escherichia coli bind to soluble glycoproteins from human milk.

    PubMed

    Schwertmann, A; Schroten, H; Hacker, J; Kunz, C

    1999-03-01

    Escherichia coli (E. coli) strains, expressing S-fimbriae, belong to the most common gram-negative pathogens that cause sepsis and meningitis in neonates. The attachment of S-fimbriae to the cell surface is mediated by membrane glycoconjugates, which often carry N-acetylneuraminic acid. Binding studies were performed with glycoproteins from the whey fraction of human milk to investigate whether they exert a potential inhibitory effect on bacterial adhesion. Whey glycoproteins were separated according to their molecular weight by fast protein liquid chromatography gel filtration. After sodium dodecyl sulfate-polyacrylamide gel electrophoresis, proteins were transferred to nitrocellulose membranes and incubated with isolated S-fimbriae from recombinant E. coli strain HB 101 (pANN 801-4). S-fimbriae recognized four whey proteins with a molecular mass of more than 200 kDa, 170 to 150 kDa, and 80 kDa. Their glycosylation pattern was investigated using the lectins Sambucus nigra, Maackia amurensis, Galanthus nivalis, and Arachis hypogaea. Thus the presence of N- and O-glycans in these proteins was confirmed. The preferential binding to N-acetylneuraminic acid containing glycoproteins was demonstrated by a complete abolishment of these reactions by incubation with acidic lactose-derived oligosaccharides. However, the cleavage of N-acetylneuraminic acid from glycoproteins by mild acid hydrolysis revealed a second binding site for S-fimbriae on milk proteins of a similar molecular weight range. Terminal galactose in human milk glycoconjugates were thought to react with S-fimbriae as well. These data further support the opinion that glycoproteins from human milk are potential receptor analogues for certain bacteria that may prevent microbial adhesion to the epithelial cell surface.

  12. Streptococcus salivarius Fimbriae Are Composed of a Glycoprotein Containing a Repeated Motif Assembled into a Filamentous Nondissociable Structure

    PubMed Central

    Lévesque, Céline; Vadeboncoeur, Christian; Chandad, Fatiha; Frenette, Michel

    2001-01-01

    Streptococcus salivarius, a gram-positive bacterium found in the human oral cavity, expresses flexible peritrichous fimbriae. In this paper, we report purification and partial characterization of S. salivarius fimbriae. Fimbriae were extracted by shearing the cell surface of hyperfimbriated mutant A37 (a spontaneous mutant of S. salivarius ATCC 25975) with glass beads. Preliminary experiments showed that S. salivarius fimbriae did not dissociate when they were incubated at 100°C in the presence of sodium dodecyl sulfate. This characteristic was used to separate them from other cell surface components by successive gel filtration chromatography procedures. Fimbriae with molecular masses ranging from 20 × 106 to 40 × 106 Da were purified. Examination of purified fimbriae by electron microscopy revealed the presence of filamentous structures up to 1 μm long and 3 to 4 nm in diameter. Biochemical studies of purified fimbriae and an amino acid sequence analysis of a fimbrial internal peptide revealed that S. salivarius fimbriae were composed of a glycoprotein assembled into a filamentous structure resistant to dissociation. The internal amino acid sequence was composed of a repeated motif of two amino acids alternating with two modified residues: A/X/T-E-Q-M/φ, where X represents a modified amino acid residue and φ represents a blank cycle. Immunolocalization experiments also revealed that the fimbriae were associated with a wheat germ agglutinin-reactive carbohydrate. Immunolabeling experiments with antifimbria polyclonal antibodies showed that antigenically related fimbria-like structures were expressed in two other human oral streptococcal species, Streptococcus mitis and Streptococcus constellatus. PMID:11292790

  13. Development of an Intranasal Vaccine To Prevent Urinary Tract Infection by Proteus mirabilis

    PubMed Central

    Li, Xin; Lockatell, C. Virginia; Johnson, David E.; Lane, M. Chelsea; Warren, John W.; Mobley, Harry L. T.

    2004-01-01

    Proteus mirabilis commonly infects the complicated urinary tract and is associated with urolithiasis. Stone formation is caused by bacterial urease, which hydrolyzes urea to ammonia, causing local pH to rise, and leads to the subsequent precipitation of magnesium ammonium phosphate (struvite) and calcium phosphate (apatite) crystals. To prevent these infections, we vaccinated CBA mice with formalin-killed bacteria or purified mannose-resistant, Proteus-like (MR/P) fimbriae, a surface antigen expressed by P. mirabilis during experimental urinary tract infection, via four routes of immunization: subcutaneous, intranasal, transurethral, and oral. We assessed the efficacy of vaccination using the CBA mouse model of ascending urinary tract infection. Subcutaneous or intranasal immunization with formalin-killed bacteria and intranasal or transurethral immunization with purified MR/P fimbriae significantly protected CBA mice from ascending urinary tract infection by P. mirabilis (P < 0.05). To investigate the potential of MrpH, the MR/P fimbrial tip adhesin, as a vaccine, the mature MrpH peptide (residues 23 to 275, excluding the signal peptide), and the N-terminal receptor-binding domain of MrpH (residues 23 to 157) were overexpressed as C-terminal fusions to maltose-binding protein (MBP) and purified on amylose resins. Intranasal immunization of CBA mice with MBP-MrpH (residues 23 to 157) conferred effective protection against urinary tract infection by P. mirabilis (P < 0.002). PMID:14688082

  14. Proteus mirabilis MR/P fimbrial operon: genetic organization, nucleotide sequence, and conditions for expression.

    PubMed

    Bahrani, F K; Mobley, H L

    1994-06-01

    Proteus mirabilis, an agent of urinary tract infection, expresses at least four fimbrial types. Among these are the MR/P (mannose-resistant/Proteus-like) fimbriae. MrpA, the structural subunit, is optimally expressed at 37 degrees C in Luria broth cultured statically for 48 h by each of seven strains examined. Genes encoding this fimbria were isolated, and the complete nucleotide sequence was determined. The mrp gene cluster encoded by 7,293 bp predicts eight polypeptides: MrpI (22,133 Da), MrpA (17,909 Da), MrpB (19,632 Da), MrpC (96,823 Da), MrpD (27,886 Da), MrpE (19,470 Da), MrpF (17,363 Da), and MrpG (13,169 Da). mrpI is upstream of the gene encoding the major structural subunit gene mrpA and is transcribed in the direction opposite to that of the rest of the operon. All predicted polypeptides share > or = 25% amino acid identity with at least one other enteric fimbrial gene product encoded by the pap, fim, smf, fan, or mrk gene clusters.

  15. Proteus mirabilis MR/P fimbrial operon: genetic organization, nucleotide sequence, and conditions for expression.

    PubMed Central

    Bahrani, F K; Mobley, H L

    1994-01-01

    Proteus mirabilis, an agent of urinary tract infection, expresses at least four fimbrial types. Among these are the MR/P (mannose-resistant/Proteus-like) fimbriae. MrpA, the structural subunit, is optimally expressed at 37 degrees C in Luria broth cultured statically for 48 h by each of seven strains examined. Genes encoding this fimbria were isolated, and the complete nucleotide sequence was determined. The mrp gene cluster encoded by 7,293 bp predicts eight polypeptides: MrpI (22,133 Da), MrpA (17,909 Da), MrpB (19,632 Da), MrpC (96,823 Da), MrpD (27,886 Da), MrpE (19,470 Da), MrpF (17,363 Da), and MrpG (13,169 Da). mrpI is upstream of the gene encoding the major structural subunit gene mrpA and is transcribed in the direction opposite to that of the rest of the operon. All predicted polypeptides share > or = 25% amino acid identity with at least one other enteric fimbrial gene product encoded by the pap, fim, smf, fan, or mrk gene clusters. Images PMID:7910820

  16. Genetic variability between complete mitochondrion genomes of the sablefish, Anoplopoma fimbria (Pallas, 1814).

    PubMed

    Galván-Tirado, Carolina; Del Río-Portilla, Miguel Angel; Delgado-Vega, Rigoberto; García-De León, Francisco J

    2016-07-01

    The complete mitochondrial genome of the sablefish, Anoplopoma fimbria (Genbank accession KP777542) is 16,507 bp in size and contains the typical 37 genes (13 protein-coding, 2 ribosomal RNA, and 22 transfer RNA) found in teleosts mitogenomes. The genome varies in 118 positions with respect to another mitogenome sablefish specimen.

  17. Long polar fimbriae contribute to colonization by Escherichia coli O157:H7 in vivo.

    PubMed

    Jordan, Dianna M; Cornick, Nancy; Torres, Alfredo G; Dean-Nystrom, Evelyn A; Kaper, James B; Moon, Harley W

    2004-10-01

    The contribution of long polar fimbriae to intestinal colonization by Escherichia coli O157:H7 was evaluated in sheep, conventional pigs, and gnotobiotic piglets. E. coli O157:H7 strains with lpfA1 and lpfA2 mutated were recovered in significantly lower numbers and caused fewer attachment and effacement lesions than the parent strain.

  18. Transcriptional profiling of human smooth muscle cells infected with gingipain and fimbriae mutants of Porphyromonas gingivalis

    PubMed Central

    Zhang, Boxi; Sirsjö, Allan; Khalaf, Hazem; Bengtsson, Torbjörn

    2016-01-01

    Porphyromonas gingivalis (P. gingivalis) is considered to be involved in the development of atherosclerosis. However, the role of different virulence factors produced by P. gingivalis in this process is still uncertain. The aim of this study was to investigate the transcriptional profiling of human aortic smooth muscle cells (AoSMCs) infected with wild type, gingipain mutants or fimbriae mutants of P. gingivalis. AoSMCs were exposed to wild type (W50 and 381), gingipain mutants (E8 and K1A), or fimbriae mutants (DPG-3 and KRX-178) of P. gingivalis. We observed that wild type P. gingivalis changes the expression of a considerable larger number of genes in AoSMCs compare to gingipain and fimbriae mutants, respectively. The results from pathway analysis revealed that the common differentially expressed genes for AoSMCs infected by 3 different wild type P. gingivalis strains were enriched in pathways of cancer, cytokine-cytokine receptor interaction, regulation of the actin cytoskeleton, focal adhesion, and MAPK signaling pathway. Disease ontology analysis showed that various strains of P. gingivalis were associated with different disease profilings. Our results suggest that gingipains and fimbriae, especially arginine-specific gingipain, produced by P. gingivalis play important roles in the association between periodontitis and other inflammatory diseases, including atherosclerosis. PMID:26907358

  19. Fur-dependent MrkHI regulation of type 3 fimbriae in Klebsiella pneumoniae CG43.

    PubMed

    Wu, Chien-Chen; Lin, Ching-Ting; Cheng, Wei-Yun; Huang, Ching-Jou; Wang, Zhe-Chong; Peng, Hwei-Ling

    2012-04-01

    Type 3 fimbriae play a crucial role in Klebsiella pneumoniae biofilm formation, but the mechanism of the regulation of the type 3 fimbrial operon is largely unknown. In K. pneumoniae CG43, three regulatory genes, mrkH, mrkI and mrkJ, are located downstream of the type 3 fimbrial genes mrkABCDF. The production of the major pilin MrkA is abolished by the deletion of mrkH or mrkI but slightly increased by the deletion of mrkJ. Additionally, quantitative RT-PCR and a promoter-reporter assay of mrkHI verified that the transcription of mrkHI was activated by MrkI, suggesting autoactivation of mrkHI transcription. In addition, sequence analysis of the mrkH promoter region revealed a putative ferric uptake regulator (Fur) box. Deletion of fur decreased the transcription of mrkH, mrkI and mrkA. The expression of type 3 fimbriae and bacterial biofilm formation were also reduced by the deletion of fur. Moreover, a recombinant Fur was found to be able to bind both promoters, with higher affinity for P(mrkH) than P(mrkA), implying that Fur controls type 3 fimbriae expression via MrkHI. We also proved that iron availability can influence type 3 fimbriae activity.

  20. Escherichia coli, fimbriae, bacterial persistence and host response induction in the human urinary tract.

    PubMed

    Bergsten, Göran; Wullt, Björn; Svanborg, Catharina

    2005-10-01

    Urinary tract infections (UTI) are among the most common bacterial infections in humans. Symptomatic UTIs may be acute, recurrent or chronic but the most frequent form of UTI is asymptomatic bacteruria (ABU). In ABU, the mucosa remains inert, despite the presence of large bacterial numbers in urine. The difference in disease severity reflects the virulence of the infecting strain and the propensity of the host to respond to infection. It is essential to understand the molecular basis of disease diversity and the molecular interactions between bacteria and host that determine asymptomatic carriage and the transition to disease. We discuss the initial interactions between bacteria and the mucosal surfaces in the human urinary tract, and the bacterial factors involved in the breach of mucosal inertia. Specifically, the contribution of P and type 1 fimbriae to bacterial establishment and host response induction are investigated. The results show that P fimbriae serve as independent virulence factors when expressed by an ABU strain, by promoting the establishment of bacteriuria and the innate host response, which is the cause of symptoms and tissue damage. P fimbriae thus fulfil the molecular Koch postulates as independent virulence factors in the human urinary tract. Type 1 fimbriae, in contrast, did not act as virulence factors in this model, and thus appear to serve a different function in man than in the murine model.

  1. Erythropoietin Promotes Neural Plasticity and Spatial Memory Recovery in Fimbria-Fornix-Lesioned Rats.

    PubMed

    Almaguer-Melian, William; Mercerón-Martínez, Daymara; Pavón-Fuentes, Nancy; Alberti-Amador, Esteban; Leon-Martinez, Rilda; Ledón, Nuris; Delgado Ocaña, Susana; Bergado Rosado, Jorge A

    2015-01-01

    Erythropoietin (EPO) upregulates the mitogen activated protein kinase (MAPK) cascade, a central signaling pathway in cellular plastic mechanisms, and is critical for normal brain development. We hypothesized that EPO could modulate the plasticity mechanisms supporting spatial memory recovery in fimbria-fornix-transected animals. Fimbria-fornix was transected in 3 groups of rats. Seven days later, EPO was injected daily for 4 consecutive days within 10 minutes after training on a water maze task. Our results show that EPO injections 10 minutes after training produced a substantial spatial memory recovery in fimbria-fornix-lesioned animals. In contrast, an EPO injection shortly after fimbria-fornix lesion surgery does not promote spatial-memory recovery. Neither does daily EPO injection 5 hours after the water maze performance. EPO, on the other hand, induced the expression of plasticity-related genes like arc and bdnf, but this effect was independent of training or lesion. This finding supports our working hypothesis that EPO can modulate transient neuroplastic mechanisms triggered by training in lesioned animals. Consequently, we propose that EPO administration can be a useful trophic factor to promote neural restoration when given in combination with training. © The Author(s) 2015.

  2. Receptor Structure for F1C Fimbriae of Uropathogenic Escherichia coli

    PubMed Central

    Khan, A. Salam; Kniep, Bernhard; Oelschlaeger, Tobias A.; Van Die, Irma; Korhonen, Timo; Hacker, Jörg

    2000-01-01

    F1C fimbriae are correlated with uropathogenic Escherichia coli strains. Although F1C fimbriae mediate binding to kidney tubular cells, their receptor is not known. In this paper, we demonstrate for the first time specific carbohydrate residues as receptor structure for F1C-fimbria-expressing E. coli. The binding of the F1C fimbriated recombinant E. coli strain HB101(pPIL110-54) and purified F1C fimbriae to reference glycolipids of different carbohydrate compositions was evaluated by using thin-layer chromatography (TLC) overlay and solid-phase binding assays. TLC fimbrial overlay analysis revealed the binding ability of purified F1C fimbriae only to glucosylceramide (GlcCer), β1-linked galactosylceramide 2 (GalCer2) with nonhydroxy fatty acids, lactosylceramide, globotriaosylceramide, paragloboside (nLc4Cer), lactotriaosylceramide, gangliotriaosylceramide (asialo-GM2 [GgO3Cer]) and gangliotetraosylceramide (asialo-GM1 [GgO4Cer]). The binding of purified F1C fimbriae as well as F1C fimbriated recombinant E. coli strain HB101(pPIL110-54) was optimal to microtiter plates coated with asialo-GM2 (GgO3Cer). The bacterial interaction with asialo-GM1 (GgO4Cer) and asialo-GM2 (GgO3Cer) was strongly inhibited only by disaccharide GalNAcβ1-4Galβ linked to bovine serum albumin. We observed no binding to globotetraosylceramide or Forssman antigen (Gb5Cer) glycosphingolipids or to sialic-acid-containing gangliosides. It was demonstrated that the presence of a GalCer or GlcCer residue alone is not sufficient for optimal binding, and additional carbohydrate residues are required for high-affinity adherence. Indeed, the binding efficiency of F1C fimbriated recombinant bacteria increased by 19-fold when disaccharide sequence GalNAcβ1-4Galβ is linked to glucosylceramide as in asialo-GM2 (GgO3Cer). Thus, it is suggested that the disaccharide sequence GalNAcβ1-4Galβ of asialo-GM2 (GgO3Cer) which is positioned internally in asialo-GM1 (GgO4Cer) is the high-affinity binding

  3. Salmonella enteritidis fimbriae displaying a heterologous epitope reveal a uniquely flexible structure and assembly mechanism.

    PubMed

    White, A P; Collinson, S K; Banser, P A; Dolhaine, D J; Kay, W W

    2000-02-18

    Two distinct Salmonella fimbrins, AgfA and SefA, comprising thin aggregative fimbriae SEF17 and SEF14, respectively, were each genetically engineered to carry PT3, an alpha-helical 16-amino acid Leishmania T-cell epitope derived from the metalloprotease gp63. To identify regions within AgfA and SefA fimbrins amenable to replacement with this epitope, PCR-generated chimeric fimbrin genes were constructed and used to replace the native chromosomal agfA and sefA genes in Salmonella enteritidis. Immunoblot analysis using anti-SEF17 and anti-PT3 sera demonstrated that all ten AgfA chimeric fimbrin proteins were expressed by S. enteritidis under normal growth conditions. Immunoelectron microscopy confirmed that eight of the AgfA::PT3 proteins were effectively assembled into cell surface-exposed fimbriae. The PT3 replacements in AgfA altered Congo red (CR) binding, cell-cell adhesion and cell surface properties of S. enteritidis to varying degrees. However, these chimeric fimbriae were still highly stable, being resistant to proteinase K digestion and requiring harsh formic acid treatment for depolymerization. In marked contrast to AgfA, none of the chimeric SefA proteins were expressed or assembled into fimbriae. Since each PT3 replacement constituted over 10% of the AgfA amino acid sequence and all ten replacements collectively represented greater than 75% of the entire AgfA primary sequence, the ability of AgfA to accept large sequence substitutions and still assemble into fibers is unique among fimbriae and other structural proteins. This structural flexibility may be related to the novel fivefold repeating sequence of AgfA and its recently proposed structure Proper formation of chimeric fimbrial fibers suggests an unusual assembly mechanism for thin aggregative fimbriae which tolerates aberrant structures. This study opens a range of possibilities for Salmonella thin aggregative fimbriae as a carrier of heterologous epitopes and as an experimental model for studies

  4. Gene cloning and characterization of Streptococcus intermedius fimbriae involved in saliva-mediated aggregation.

    PubMed

    Yamaguchi, Taihei; Matsumoto, Mitsuharu; Sugimoto, Yasushi; Soutome, Sakiko; Oho, Takahiko

    2009-12-01

    Streptococcus intermedius, an oral commensal and a cause of systemic pyogenic disease, expresses fimbriae. To identify the gene(s) encoding these fimbriae, we used a serum raised against purified fimbriae to screen libaries of recombinant lambda phages. The cloned gene cluster encoding S. intermedius fimbriae, (saliva-mediated aggregation and adherence-associated fimbriae), contained 4 ORFs, i.e. a putative ribonulease (Saf1), a putative adhesin (Saf2), the main pilus subunit (Saf3) and a sortase C (SrtC). Escherichia coli strains harboring recombinant phages and plasmids carrying the saf3 gene produced a 55kDa protein recognized by the antifimbriae serum. Saf3 contains an N-terminal signal sequence and a C-terminal cell-wall-anchoring motif LPXTG. Among strains of the Streptococcus anginosus group, only serotype g and untypable strains were found to contain the saf3 gene, to possess the fimbrial antigen and to exhibit saliva-mediated aggregation. Knockout mutants made by insertion of an erythromycin resistance gene into saf3 did not produce fimbrial structures or fimbrial antigens and did not participate in saliva-mediated aggregation. The adherent activity of mutants toward plastic wells coated with salivary agglutinin was about 65% that of the parental strain, and the reaction depended on calcium. There was no significant difference in adherence to hydroxyapatite beads pretreated with salivary agglutinin between the parental and mutant strains. These results demonstrated that Saf3 is associated with aggregation, and suggested that other molecule(s) are associated with adherence of S. intermedius.

  5. Porphyromonas gingivalis fimbriae dampen P2X7-dependent IL-1β secretion

    PubMed Central

    Morandini, Ana Carolina; Ramos-Junior, Erivan S.; Potempa, Jan; Nguyen, Ky-Anh; Oliveira, Ana Carolina; Bellio, Maria; Ojcius, David M.; Scharfstein, Julio; Coutinho-Silva, Robson

    2014-01-01

    Porphyromonas gingivalis is a major contributor to the pathogenesis of periodontitis, an infection-driven inflammatory disease that leads to bone destruction. This pathogen stimulates pro-IL-1β synthesis but not mature IL-1β secretion, unless the P2X7 receptor is activated by extracellular ATP. Here, we investigated the role of Pg fimbriae in eATP-induced IL-1β release. Bone marrow derived macrophages (BMDMs) from wild type (WT) or P2X7-deficient mice were infected with Pg (strain 381) or isogenic fimbriae deficient (strain DPG3) with or without subsequent eATP stimulation. DPG3 induced higher IL-1β secretion after eATP stimulation compared to 381 in WT BMDMs, but not in P2X7-deficient cells. This mechanism was dependent of K+ efflux and Ca2+-iPLA2 activity. Accordingly, non-fimbriated Pg failed to inhibit apoptosis via eATP/P2X7-pathway. Furthermore, Pg-driven stimulation of IL-1β was TLR2- and MyD88-dependent, and irrespective of fimbriae expression. Fimbriae-dependent down-modulation of IL-1β was selective, as levels of other cytokines remained unaffected by P2X7 deficiency. Confocal microscopy demonstrated the presence of discrete P2X7 expression in the absence of Pg stimulation which was enhanced by 381-stimulated cells. Notably, DPG3-infected macrophages revealed a distinct pattern of P2X7 receptor expression with a markedly foci formation. Collectively, these data demonstrate that eATP-induced IL-1β secretion is impaired by Pg fimbriae in a P2X7-dependent manner. PMID:24925032

  6. Erythrocyte and Porcine Intestinal Glycosphingolipids Recognized by F4 Fimbriae of Enterotoxigenic Escherichia coli

    PubMed Central

    Coddens, Annelies; Valis, Erik; Benktander, John; Ångström, Jonas; Breimer, Michael E.; Cox, Eric; Teneberg, Susann

    2011-01-01

    Enterotoxigenic F4-fimbriated Escherichia coli is associated with diarrheal disease in neonatal and postweaning pigs. The F4 fimbriae mediate attachment of the bacteria to the pig intestinal epithelium, enabling an efficient delivery of diarrhea-inducing enterotoxins to the target epithelial cells. There are three variants of F4 fimbriae designated F4ab, F4ac and F4ad, respectively, having different antigenic and adhesive properties. In the present study, the binding of isolated F4ab, F4ac and F4ad fimbriae, and F4ab/ac/ad-fimbriated E. coli, to glycosphingolipids from erythrocytes and from porcine small intestinal epithelium was examined, in order to get a comprehensive view of the F4-binding glycosphingolipids involved in F4-mediated hemagglutination and adhesion to the epithelial cells of porcine intestine. Specific interactions between the F4ab, F4ac and F4ad fimbriae and both acid and non-acid glycosphingolipids were obtained, and after isolation of binding-active glycosphingolipids and characterization by mass spectrometry and proton NMR, distinct carbohydrate binding patterns were defined for each fimbrial subtype. Two novel glycosphingolipids were isolated from chicken erythrocytes, and characterized as GalNAcα3GalNAcß3Galß4Glcß1Cer and GalNAcα3GalNAcß3Galß4GlcNAcß3Galß4Glcß1Cer. These two compounds, and lactosylceramide (Galß4Glcß1Cer) with phytosphingosine and hydroxy fatty acid, were recognized by all three variants of F4 fimbriae. No binding of the F4ad fimbriae or F4ad-fimbriated E. coli to the porcine intestinal glycosphingolipids occurred. However, for F4ab and F4ac two distinct binding patterns were observed. The F4ac fimbriae and the F4ac-expressing E. coli selectively bound to galactosylceramide (Galß1Cer) with sphingosine and hydroxy 24:0 fatty acid, while the porcine intestinal glycosphingolipids recognized by F4ab fimbriae and the F4ab-fimbriated bacteria were characterized as galactosylceramide, sulfatide (SO3-3Galß1Cer), sulf

  7. DNA-based diagnostic tests for Salmonella species targeting agfA, the structural gene for thin, aggregative fimbriae.

    PubMed Central

    Doran, J L; Collinson, S K; Burian, J; Sarlós, G; Todd, E C; Munro, C K; Kay, C M; Banser, P A; Peterkin, P I; Kay, W W

    1993-01-01

    Salmonella enteritidis 27655-3b and a few diarrheagenic Escherichia coli strains produce morphologically and antigenically related, thin, aggregative fimbriae, collectively named GVVPQ fimbriae (S. K. Collinson, L. Emödy, T. J. Trust, and W. W. Kay, J. Bacteriol. 174:4490-4495, 1992). To determine whether GVVPQ fimbriae are common to Salmonella spp. and other enteropathogenic members of the family Enterobacteriaceae, 113 isolates were phenotypically screened for Congo red binding and aggregative colony morphology. Presumptive positive and representative negative strains were examined by Western blotting (immunoblotting) by using antiserum to SEF 17, the native GVVPQ fimbria of S. enteritidis. Only four S. enteritidis strains and six E. coli isolates possessed substantial amounts of GVVPQ fimbriae after 24 h of incubation on T medium. Following 5 days of incubation, 56 of 93 Salmonella isolates (60%) and 1 of 7 additional E. coli clinical isolates possessed detectable levels of GVVPQ fimbriae. Since variable expression of GVVPQ fimbriae was observed among Salmonella isolates and some E. coli strains produced scant amounts, as revealed by immunoelectron microscopy, the ability to produce these fimbriae was evaluated by genotypic screening. The structural gene for the SEF 17 fimbrin, agfA, was amplified by the polymerase chain reaction, cloned, and sequenced to provide a characterized DNA probe. An agfA DNA fragment hybridized strongly to 603 of 604 (99.8%) Salmonella isolates but very weakly to 31 of 266 other members of the family Enterobacteriaceae including 26 of 137 E. coli strains, 3 of 14 Citrobacter spp., and single isolates of Shigella sonnei and Enterobacter cloacae. The agfA DNA probe proved to be a valuable diagnostic tool for Salmonella isolates arrayed on hydrophobic grid membrane filters. Unique agfA sequences were targeted in the development of a polymerase chain reaction assay specific for Salmonella spp. Images PMID:8104955

  8. Fimbria-dependent activation of pro-inflammatory molecules in Porphyromonas gingivalis infected human aortic endothelial cells.

    PubMed

    Takahashi, Yusuke; Davey, Michael; Yumoto, Hiromichi; Gibson, Frank C; Genco, Caroline Attardo

    2006-05-01

    Epidemiological studies support that chronic periodontal infections are associated with an increased risk of cardiovascular disease. Previously, we reported that the periodontal pathogen Porphyromonas gingivalis accelerated atherosclerotic plaque formation in hyperlipidemic apoE-/- mice, while an isogenic fimbria-deficient (FimA-) mutant did not. In this study, we utilized 41 kDa (major) and 67 kDa (minor) fimbria mutants to demonstrate that major fimbria are required for efficient P. gingivalis invasion of human aortic endothelial cells (HAEC). Enzyme-linked immunosorbent assay (ELISA) revealed that only invasive P. gingivalis strains induced HAEC production of pro-inflammatory molecules interleukin (IL)-1beta, IL-8, monocyte chemoattractant protein (MCP)-1, intracellular adhesion molecule (ICAM)-1, vascular cellular adhesion molecule (VCAM)-1 and E-selectin. The purified native forms of major and minor fimbria induced chemokine and adhesion molecule expression similar to invasive P. gingivalis, but failed to elicit IL-1beta production. In addition, the major and minor fimbria-mediated production of MCP-1 and IL-8 was inhibited in a dose-dependent manner by P. gingivalis lipopolysaccharide (LPS). Both P. gingivalis LPS and heat-killed organisms failed to stimulate HAEC. Treatment of endothelial cells with cytochalasin D abolished the observed pro-inflammatory MCP-1 and IL-8 response to invasive P. gingivalis and both purified fimbria, but did not affect P. gingivalis induction of IL-1beta. These results suggest that major and minor fimbria elicit chemokine production in HAEC through actin cytoskeletal rearrangements; however, induction of IL-1beta appears to occur via a separate mechanism. Collectively, these data support that invasive P. gingivalis and fimbria stimulate endothelial cell activation, a necessary initial event in the development of atherogenesis.

  9. Amended Description of the Genes for Synthesis of Actinomyces naeslundii T14V Type 1 Fimbriae and Associated Adhesin

    DTIC Science & Technology

    2007-05-07

    monoclonal antibody (MAb) (2). Eluted fimbriae were digested either with pepsin or by dilute-acid hydrolysis, and the resulting peptides were analyzed...Representative tandem mass spectrometry spectra of two such peptides, VVRNSDGTF derived by pepsin digestion and PSTPGAKPLTD from dilute-acid hydrolysis of...or -FimA). TABLE 1. FimQ peptides identified from type 1 fimbriae after pepsin and diluted-acid hydrolysis m/z Charge Mol wt Sequence (positions

  10. DNA-based diagnostic tests for Salmonella species targeting agfA, the structural gene for thin, aggregative fimbriae.

    PubMed

    Doran, J L; Collinson, S K; Burian, J; Sarlós, G; Todd, E C; Munro, C K; Kay, C M; Banser, P A; Peterkin, P I; Kay, W W

    1993-09-01

    Salmonella enteritidis 27655-3b and a few diarrheagenic Escherichia coli strains produce morphologically and antigenically related, thin, aggregative fimbriae, collectively named GVVPQ fimbriae (S. K. Collinson, L. Emödy, T. J. Trust, and W. W. Kay, J. Bacteriol. 174:4490-4495, 1992). To determine whether GVVPQ fimbriae are common to Salmonella spp. and other enteropathogenic members of the family Enterobacteriaceae, 113 isolates were phenotypically screened for Congo red binding and aggregative colony morphology. Presumptive positive and representative negative strains were examined by Western blotting (immunoblotting) by using antiserum to SEF 17, the native GVVPQ fimbria of S. enteritidis. Only four S. enteritidis strains and six E. coli isolates possessed substantial amounts of GVVPQ fimbriae after 24 h of incubation on T medium. Following 5 days of incubation, 56 of 93 Salmonella isolates (60%) and 1 of 7 additional E. coli clinical isolates possessed detectable levels of GVVPQ fimbriae. Since variable expression of GVVPQ fimbriae was observed among Salmonella isolates and some E. coli strains produced scant amounts, as revealed by immunoelectron microscopy, the ability to produce these fimbriae was evaluated by genotypic screening. The structural gene for the SEF 17 fimbrin, agfA, was amplified by the polymerase chain reaction, cloned, and sequenced to provide a characterized DNA probe. An agfA DNA fragment hybridized strongly to 603 of 604 (99.8%) Salmonella isolates but very weakly to 31 of 266 other members of the family Enterobacteriaceae including 26 of 137 E. coli strains, 3 of 14 Citrobacter spp., and single isolates of Shigella sonnei and Enterobacter cloacae. The agfA DNA probe proved to be a valuable diagnostic tool for Salmonella isolates arrayed on hydrophobic grid membrane filters. Unique agfA sequences were targeted in the development of a polymerase chain reaction assay specific for Salmonella spp.

  11. Construction and expression of recombinant plasmids encoding type 1 fimbriae of a urinary Klebsiella pneumoniae isolate.

    PubMed Central

    Purcell, B K; Clegg, S

    1983-01-01

    The type 1 fimbriae of Klebsiella pneumoniae have been implicated as important virulence factors in mediating Klebsiella urinary infections. The chromosomally encoded fimbrial genes were cloned by a cosmid cloning technique. Further subcloning was performed with the cloning vehicles pBR322 and pACYC184, and a recombinant plasmid containing the fimbrial genes was constructed. After transformation by this plasmid, both Escherichia coli and Salmonella typhimurium were shown to express fimbriae which reacted with Klebsiella fimbrial antiserum. The approximate location of the relevant genes on the chimeric plasmid was determined by insertion of the transposable element Tn5. Hemagglutination-negative phenotypes were used to estimate the minimum size of the DNA fragment necessary to encode fimbrial biosynthesis and expression. The size of the coding region of this fragment was found to be 5.5 kilobase pairs. PMID:6132874

  12. The Response Regulator RcsB Activates Expression of Mat Fimbriae in Meningitic Escherichia coli

    PubMed Central

    Lehti, Timo A.; Heikkinen, Johanna; Korhonen, Timo K.

    2012-01-01

    The common colonization factor of Escherichia coli, the Mat (also termed ECP) fimbria, functions to advance biofilm formation on inert surfaces as well as bacterial adherence to epithelial cells and subsequent colonization. We used global mini-Tn5 transposon mutagenesis to identify novel regulators of biofilm formation by the meningitic E. coli isolate IHE 3034. Of the 4,418 transformants, we found 17 that were impaired in biofilm formation. Most of these mutants were affected in lipopolysaccharide synthesis and were reduced in growth but not in Mat fimbria expression. In contrast, two mutants grew well but did not express Mat fimbria. The insertions in these two mutants were located at different sites of the rcsB gene, which encodes a DNA-binding response regulator of the Rcs response regulon. The mutations abrogated temperature-dependent biofilm formation by IHE 3034, and the phenotype correlated with loss of mat expression. The defect in biofilm formation in the rcsB mutant was reversed upon complementation with rcsB as well as by overexpression of structural mat genes but not by overexpression of the fimbria-specific activator gene matA. Monitoring of the mat operon promoter activity with chromosomal reporter fusions showed that the RcsB protein and an RcsAB box in the mat regulatory region, but not RcsC, RcsD, AckA, and Pta, are essential for initiation of mat transcription. Gel retardation assays showed that RcsB specifically binds to the mat promoter DNA, which enables its function in promoting biofilm formation by E. coli. PMID:22522901

  13. The response regulator RcsB activates expression of Mat fimbriae in meningitic Escherichia coli.

    PubMed

    Lehti, Timo A; Heikkinen, Johanna; Korhonen, Timo K; Westerlund-Wikström, Benita

    2012-07-01

    The common colonization factor of Escherichia coli, the Mat (also termed ECP) fimbria, functions to advance biofilm formation on inert surfaces as well as bacterial adherence to epithelial cells and subsequent colonization. We used global mini-Tn5 transposon mutagenesis to identify novel regulators of biofilm formation by the meningitic E. coli isolate IHE 3034. Of the 4,418 transformants, we found 17 that were impaired in biofilm formation. Most of these mutants were affected in lipopolysaccharide synthesis and were reduced in growth but not in Mat fimbria expression. In contrast, two mutants grew well but did not express Mat fimbria. The insertions in these two mutants were located at different sites of the rcsB gene, which encodes a DNA-binding response regulator of the Rcs response regulon. The mutations abrogated temperature-dependent biofilm formation by IHE 3034, and the phenotype correlated with loss of mat expression. The defect in biofilm formation in the rcsB mutant was reversed upon complementation with rcsB as well as by overexpression of structural mat genes but not by overexpression of the fimbria-specific activator gene matA. Monitoring of the mat operon promoter activity with chromosomal reporter fusions showed that the RcsB protein and an RcsAB box in the mat regulatory region, but not RcsC, RcsD, AckA, and Pta, are essential for initiation of mat transcription. Gel retardation assays showed that RcsB specifically binds to the mat promoter DNA, which enables its function in promoting biofilm formation by E. coli.

  14. Analysis of Spleen-Induced Fimbria Production in Recombinant Attenuated Salmonella enterica Serovar Typhimurium Vaccine Strains

    PubMed Central

    Łaniewski, Paweł; Baek, Chang-Ho; Roland, Kenneth L.

    2017-01-01

    ABSTRACT Salmonella enterica serovar Typhimurium genome encodes 13 fimbrial operons. Most of the fimbriae encoded by these operons are not produced under laboratory conditions but are likely to be synthesized in vivo. We used an in vivo expression technology (IVET) strategy to identify four fimbrial operons, agf, saf, sti, and stc that are expressed in the spleen. When any three of these operons were deleted, the strain retained wild-type virulence. However, when all four operons were deleted, the resulting strain was completely attenuated, indicating that these four fimbriae play functionally redundant roles critical for virulence. In mice, oral doses of as low as 1 × 105 CFU of the strain with four fimbrial operons deleted provided 100% protection against challenge with 1 × 109 CFU of wild-type S. Typhimurium. We also examined the possible effect of these fimbriae on the ability of a Salmonella vaccine strain to deliver a guest antigen. We modified one of our established attenuated vaccine strains, χ9088, to delete three fimbrial operons while the fourth operon was constitutively expressed. Each derivative was modified to express the Streptococcus pneumoniae antigen PspA. Strains that constitutively expressed saf or stc elicited a strong Th1 response with significantly greater levels of anti-PspA serum IgG and greater protective efficacy than strains carrying saf or stc deletions. The isogenic strain in which all four operons were deleted generated the lowest anti-PspA levels and did not protect against challenge with virulent S. pneumoniae. Our results indicate that these fimbriae play important roles, as yet not understood, in Salmonella virulence and immunogenicity. PMID:28830946

  15. Cyclic-AMP inhibition of fimbriae and prodigiosin production by Serratia marcescens is strain-dependent

    PubMed Central

    Stella, Nicholas A.; Shanks, Robert M. Q.

    2014-01-01

    The cyclic-nucleotide 3’,5’-cyclic AMP (cAMP) is an ancient and wide spread regulatory molecule. Previous studies have shown that fimbria production and secondary metabolite production are inhibited by cAMP in the prokaryote Serratia marcescens. This study used genetic manipulations to test the strain specificity of cAMP-CRP regulation of fimbria production and of the red pigment, prodigiosin. A surprising amount of variation was observed, as multicopy expression of the cAMP-phosphodiesterase gene, cpdS, conferred either an increase or decrease in fimbriae-dependent yeast agglutination and prodigiosin production depending upon the strain background. Mutation of crp, the gene coding for the cAMP-receptor protein similarly conferred strain-dependent phenotypes. This study shows that three distinct biological properties, modulated by a conserved genetic regulatory molecule, can vary significantly among strains. Such variation can complicate the functional analysis of bacterial phenotypic properties which are dependent upon global genetic regulators such as cAMP. PMID:24619531

  16. Streptococcus parasanguis Fimbria-Associated Adhesin Fap1 Is Required for Biofilm Formation

    PubMed Central

    Froeliger, Eunice H.; Fives-Taylor, Paula

    2001-01-01

    The sanguis streptococci are primary colonizers of the tooth surface and thus form the foundation for the complex multiple species biofilm known as dental plaque. In addition, these bacteria can colonize native and prosthetic heart valves and are a common cause of endocarditis. Little is known about the molecular mechanisms governing multiple or single species biofilm development within this group of organisms. Using an in vitro assay for biofilm formation, we determined that (i) Streptococcus parasanguis FW213 can form biofilms on inert surfaces such as polystyrene and (ii) environmental and nutritional factors, such as glucose, affect S. parasanguis biofilm formation. Several isogenic mutants of FW213 were tested in the biofilm assay. Strains containing mutations in fap1, a gene encoding a protein required for assembly of fimbriae, were deficient in biofilm formation. Mutants defective in recA, PepO endopeptidase activity, or the production of a fimbriae-associated protein, FimA, were still capable of biofilm formation. Phase-contrast microscopy was used to follow biofilm development by wild-type and fap1 mutant strains on plastic coverslips over time. Wild-type FW213 attached to the surface, formed aggregates of cells, and eventually formed a dense layer of cells that included microcolonies. In contrast, few fap1 mutant cells were observed attached to the surface, and no cell aggregates or microcolonies were formed. These results suggest that the long peritrichous fimbriae of FW213 are critical for the formation of biofilms on solid surfaces. PMID:11254614

  17. Influence of porcine intestinal pH and gastric digestion on antigenicity of F4 fimbriae for oral immunisation.

    PubMed

    Snoeck, Veerle; Cox, Eric; Verdonck, Frank; Joensuu, Jussi J; Goddeeris, Bruno M

    2004-01-14

    Newly weaned piglets can be orally immunised against F4+ enterotoxigenic Escherichia coli (ETEC) infection with F4 fimbriae. However, to efficiently develop a vaccine against ETEC induced postweaning diarrhoea, knowledge of the stability of the F4 fimbriae to different pH and gastric digestion is needed. The gastrointestinal pH in suckling and recently weaned piglets was measured and the stability of F4 fimbriae to different pH and to pepsin was assessed in vitro. In the stomach the lowest pH was found in the fundus gland region. Gastric pH values below 2.5 were not found in suckling piglets or at the day of weaning, in contrast to piglets 1 and 2 weeks postweaning. Along the first half of the small intestine and in the caecum, a negative correlation was found between pH and age. The F4 fimbriae were stable to pH 1.5 and 2 for 2 h, whereas longer incubation periods resulted in conversion of the multimeric forms into monomers. The F4 fimbriae were partially degraded by incubation for 15-30 min in simulated gastric fluid at pH 1.5 and 2, and completely digested from 3 h onwards. At pH 3, the fimbriae maintained their antigenicity for at least 4h. The results demonstrate that gastric digestion will only have a limited impact on oral immunisation since liquid passes through the stomach relatively quickly (50% within 2 h). However, we previously demonstrated that the transit times are prolonged shortly after weaning. Shortly after weaning it could be necessary to protect the F4 fimbriae against gastric digestion to obtain efficient oral immunisation of the piglets.

  18. Fimbria-Fornix Lesions Disrupt the Dead Reckoning (Homing) Component of Exploratory Behavior in Mice

    PubMed Central

    Gorny, Joanna H.; Gorny, Bogdan; Wallace, Douglas G.; Whishaw, Ian Q.

    2002-01-01

    Exploration is the primary way in which rodents gather information about their spatial surroundings. Thus, spatial theories propose that damage to the hippocampus, a structure thought to play a fundamental role in spatial behavior, should disrupt exploration. Exploration in rats is organized. The animals create home bases that are central to exploratory excursions and returns, and hippocampal formation damage alters the organization of exploration by disrupting returns. Mice do not appear to readily establish home bases in novel environments, thus, for this species, it is more difficult to establish the contribution of the hippocampus to exploration. The purpose of the present study was threefold: develop a task in which mice center their exploration from a home base, determine whether the exploratory behavior is organized, and evaluate the role of fimbria-fornix lesions on exploration. Mice were given a novel exploratory task in which their nesting material was placed on a large circular table. Video records of control and fimbria-fornix mice were made in both light and dark (infrared light) conditions. Exploration patterns (outward trips, stops, and homeward trips) were reconstructed from the video records. Control mice centered their activity on their bedding, from which they made circuitous outward trips marked by many stops, and periodic direct returns. The bedding-centered behavior and outward trips of the fimbria-fornix mice were similar to those of the control mice, but significantly fewer direct return trips occurred. The direct homeward trips observed under light and dark conditions were consistent with a dead-reckoning strategy, in which an animal computes its present position and homeward trajectory from self-movement cues generated on the outward trip. Because the fimbria-fornix lesions disrupted the homeward component of exploratory trips, we conclude that the fimbria-fornix may contribute to dead reckoning in mice. The results also show that the home

  19. Identification and Regulation of a Novel Citrobacter rodentium Gut Colonization Fimbria (Gcf)

    PubMed Central

    Caballero-Flores, Gustavo G.; Croxen, Matthew A.; Martínez-Santos, Verónica I.; Finlay, B. Brett

    2015-01-01

    ABSTRACT The Gram-negative enteric bacterium Citrobacter rodentium is a natural mouse pathogen that has been extensively used as a surrogate model for studying the human pathogens enteropathogenic and enterohemorrhagic Escherichia coli. All three pathogens produce similar attaching and effacing (A/E) lesions in the intestinal epithelium. During infection, these bacteria employ surface structures called fimbriae to adhere and colonize the host intestinal epithelium. For C. rodentium, the roles of only a small number of its genome-carried fimbrial operons have been evaluated. Here, we report the identification of a novel C. rodentium colonization factor, called gut colonization fimbria (Gcf), which is encoded by a chaperone-usher fimbrial operon. A gcfA mutant shows a severe colonization defect within the first 10 days of infection. The gcf promoter is not active in C. rodentium under several in vitro growth conditions; however, it is readily expressed in a C. rodentium Δhns1 mutant lacking the closest ortholog of the Escherichia coli histone-like nucleoid structuring protein (H-NS) but not in mutants with deletion of the other four genes encoding H-NS homologs. H-NS binds to the regulatory region of gcf, further supporting its direct role as a repressor of the gcf promoter that starts transcription 158 bp upstream of the start codon of its first open reading frame. The gcf operon possesses interesting novel traits that open future opportunities to expand our knowledge of the structure, regulation, and function during infection of these important bacterial structures. IMPORTANCE Fimbriae are surface bacterial structures implicated in a variety of biological processes. Some have been shown to play a critical role during host colonization and thus in disease. Pathogenic bacteria possess the genetic information for an assortment of fimbriae, but their function and regulation and the interplay between them have not been studied in detail. This work provides new insights

  20. Structure and characterization of AgfB from Salmonella enteritidis thin aggregative fimbriae.

    PubMed

    White, A P; Collinson, S K; Banser, P A; Gibson, D L; Paetzel, M; Strynadka, N C; Kay, W W

    2001-08-24

    The agfBAC operon of Salmonella enteritidis encodes thin aggregative fimbriae, fibrous, polymeric structures primarily composed of AgfA fimbrins. Although uncharacterized, AgfB shows a 51 % overall amino acid sequence similarity to AgfA. Using AgfB epitope-specific antiserum, AgfB was detected as a minor component of whole, purified fimbriae. Like AgfA, AgfB was released from purified fimbriae by >70 % formic acid, whereupon both AgfA-AgfA and AgfA-AgfB dimers as well as monomers were detected. This suggested that AgfB may form specific, highly stable, structural associations with AgfA in native fimbrial filaments, associations that were weakened in structurally unstable fibers derived from AgfA chimeric fimbrial mutants. Detailed sequence comparisons between AgfA and AgfB showed that AgfB harbored a similar fivefold repeated sequence pattern (x(6)QxGx(2)NxAx(3)Q), and contained structural motifs similar to the parallel beta helix model proposed for AgfA. Molecular modeling of AgfB revealed a 3D structure remarkably similar to that of AgfA, the structures differing principally in the surface disposition of non-conserved, basic, acidic and non-polar residues. Thus AgfB is a fimbrin-like structural homologue of AgfA and an integral, minor component of native thin aggregative fimbrial fibers. AgfB from an agfA deletion strain was detected as a non-fimbrial, SDS-insoluble form in the supernatant and was purified. AgfA from an agfB deletion strain was found in both SDS-soluble and insoluble, non-fimbrial forms. No AgfA-AgfA dimers were detected in the absence of AgfB. Fimbriae formation by intercellular complementation between agfB and agfA deletion strains could not be shown under a variety of conditions, indicating that AgfA and AgfB are not freely diffusible in S. enteritidis. This has important implications on the current assembly hypothesis for thin aggregative fimbriae.

  1. Hemagglutinin Typing as an Aid in Identification of Biochemically Atypical Escherichia coli Strains

    PubMed Central

    Crichton, Pamela B.; Ip, S. M.; Old, D. C.

    1981-01-01

    Tests for the presence of mannose-sensitive and mannose-resistant, eluting hemagglutinins and fimbriae were helpful in indicating whether biochemically atypical strains of the tribe Escherichieae might be escherichiae or shigellae. PMID:7334072

  2. The Shaft of the Type 1 Fimbriae Regulates an External Force to Match the FimH Catch Bond

    PubMed Central

    Zakrisson, Johan; Wiklund, Krister; Axner, Ove; Andersson, Magnus

    2013-01-01

    Type 1 fimbriae mediate adhesion of uropathogenic Escherichia coli to host cells. It has been hypothesized that due to their ability to uncoil under exposure to force, fimbriae can reduce fluid shear stress on the adhesin-receptor interaction by which the bacterium adheres to the surface. In this work, we develop a model that describes how the force on the adhesin-receptor interaction of a type 1 fimbria varies as a bacterium is affected by a time-dependent fluid flow mimicking in vivo conditions. The model combines in vivo hydrodynamic conditions with previously assessed biomechanical properties of the fimbriae. Numerical methods are used to solve for the motion and adhesion force under the presence of time-dependent fluid profiles. It is found that a bacterium tethered with a type 1 pilus will experience significantly reduced shear stress for moderate to high flow velocities and that the maximum stress the adhesin will experience is limited to ∼120 pN, which is sufficient to activate the conformational change of the FimH adhesin into its stronger state but also lower than the force required for breaking it under rapid loading. Our model thus supports the assumption that the type 1 fimbria shaft and the FimH adhesin-receptor interaction are optimized to each other, and that they give piliated bacteria significant advantages in rapidly changing fluidic environments. PMID:23708354

  3. P-fimbriae in the presence of anti-PapA antibodies: new insight of antibodies action against pathogens

    PubMed Central

    Mortezaei, Narges; Singh, Bhupender; Bullitt, Esther; Uhlin, Bernt Eric; Andersson, Magnus

    2013-01-01

    Uropathogenic strains of Escherichia coli establish urinary tract infections by attaching to host epithelial cells using adhesive organelles called fimbriae. Fimbriae are helix-like structures with a remarkable adaptability, offering safeguarding for bacteria exposed to changing fluid forces in the urinary tract. We challenged this property of P-fimbriae by cross-linking their subunits with shaft-specific antibodies and measuring the corresponding force response at a single organelle level. Our data show compromised extension and rewinding of P-fimbriae in the presence of antibodies and reduced fimbrial elasticity, which are important properties of fimbriae contributing to the ability of bacteria to cause urinary tract infections. The reduced elasticity found by cross-linking fimbrial subunits could thus be another assignment for antibodies; in addition to marking bacteria as foreign, antibodies physically compromise fimbrial function. We suggest that our assay and results will be a starting point for further investigations aimed at inhibiting sustained bacterial adhesion by antibodies. PMID:24292100

  4. Attachment of Streptomyces coelicolor is mediated by amyloidal fimbriae that are anchored to the cell surface via cellulose.

    PubMed

    de Jong, Wouter; Wösten, Han A B; Dijkhuizen, Lubbert; Claessen, Dennis

    2009-09-01

    The chaplin proteins ChpA-H enable the filamentous bacterium Streptomyces coelicolor to form reproductive aerial structures by assembling into surface-active amyloid-like fibrils. We here demonstrate that chaplins also mediate attachment of S. coelicolor to surfaces. Attachment coincides with the formation of fimbriae, which are connected to the cell surface via spike-shaped protrusions. Mass spectrometry, electron microscopy and Congo red treatment showed that these fimbriae are composed of bundled amyloid fibrils of chaplins. Attachment and fimbriae formation were abolished in a strain in which the chaplin genes chpA-H were inactivated. Instead, very thin fibrils emerged from the spike-shaped protrusions in this mutant. These fibrils were susceptible to cellulase treatment. This enzymatic treatment also released wild-type fimbriae from the cell surface, thereby abolishing attachment. The reduced attachment of a strain in which the gene of a predicted cellulose synthase was inactivated also indicates a role of cellulose in surface attachment. We propose that the mechanism of attachment via cellulose-anchored amyloidal fimbriae is widespread in bacteria and may function in initiation of infection and in formation of biofilms.

  5. Coaggregation of Streptococcus salivarius with periodontopathogens: evidence for involvement of fimbriae in the interaction with Prevotella intermedia.

    PubMed

    Lévesque, C; Lamothe, J; Frenette, M

    2003-10-01

    Streptococcus salivarius is divided into two serological subgroups that carry either fibrils or fimbriae. Although fimbriae have been observed on up to 50% of S. salivarius strains in the human oral cavity, no function has yet been assigned to them. To determine whether S. salivarius fimbriae have a role in adhesion, we examined the ability of S. salivarius to coaggregate with selected microorganisms involved in periodontal diseases. Our results show that S. salivarius coaggregated with Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia. However, only fimbriated S. salivarius cells were able to coaggregate with P. intermedia, suggesting a specific role for these structures in the interaction. Heat treatment, sensitivity to sugars, amino acids, and EDTA, as well as protease treatment were also used to further characterize coaggregation between S. salivarius and periodontopathogens.

  6. Proteus mirabilis uroepithelial cell adhesin (UCA) fimbria plays a role in the colonization of the urinary tract.

    PubMed

    Pellegrino, Rafael; Scavone, Paola; Umpiérrez, Ana; Maskell, Duncan J; Zunino, Pablo

    2013-03-01

    Urinary tract infections (UTIs) are among the most common bacterial infections in humans. Proteus mirabilis is an opportunistic pathogen, capable of causing severe UTIs, with serious kidney damage that may even lead to death. Several virulence factors are involved in the pathogenicity of this bacterium. Among these, adherence to the uroepithelium mediated by fimbriae appears to be a significant bacterial attribute related to urovirulence. Proteus mirabilis expresses several types of fimbriae that could be involved in the pathogenesis of UTI, including uroepithelial cell adhesin (UCA). In this report, we used an uropathogenic P. mirabilis wild-type strain and an isogenic ucaA mutant unable to express UCA to study the pathogenic role of this fimbria in UTI. Ability of the mutant to adhere to desquamated uroepithelial cells and to infect mice using different experimental UTI models was significantly impaired. These results allow us to conclude that P. mirabilis UCA plays an important role in the colonization of the urinary tract.

  7. sRNA-Mediated Regulation of P-Fimbriae Phase Variation in Uropathogenic Escherichia coli

    PubMed Central

    Khandige, Surabhi; Kronborg, Tina; Uhlin, Bernt Eric; Møller-Jensen, Jakob

    2015-01-01

    Uropathogenic Escherichia coli (UPEC) are capable of occupying physiologically distinct intracellular and extracellular niches within the urinary tract. This feat requires the timely regulation of gene expression and small RNAs (sRNAs) are known to mediate such rapid adjustments in response to changing environmental cues. This study aimed to uncover sRNA-mediated gene regulation in the UPEC strain UTI89, during infection of bladder epithelial cells. Hfq is an RNA chaperone known to facilitate and stabilize sRNA and target mRNA interactions with bacterial cells. The co-immunoprecipitation and high throughput RNA sequencing of Hfq bound sRNAs performed in this study, revealed distinct sRNA profiles in UPEC in the extracellular and intracellular environments. Our findings emphasize the importance of studying regulatory sRNAs in a biologically relevant niche. This strategy also led to the discovery of a novel virulence-associated trans-acting sRNA—PapR. Deletion of papR was found to enhance adhesion of UTI89 to both bladder and kidney cell lines in a manner independent of type-1 fimbriae. We demonstrate PapR mediated posttranscriptional repression of the P-fimbriae phase regulator gene papI and postulate a role for such regulation in fimbrial cross-talk at the population level in UPEC. Our results further implicate the Leucine responsive protein (LRP) as a transcriptional activator regulating PapR expression. Our study reports, for the first time, a role for sRNAs in regulation of P-fimbriae phase variation and emphasizes the importance of studying pathogenesis-specific sRNAs within a relevant biological niche. PMID:26291711

  8. Activation of dentate hilar neurons by stimulation of the fimbria in rat hippocampal slices

    PubMed Central

    Scharfman, Helen E.

    2012-01-01

    It is has been shown that the major afferent input to the dentate gyrus, the perforant path, excites dentate hilar neurons. However, little is known about the other inputs to hilar cells. Therefore, we examined the responses of hilar neurons to stimulation of the fimbria. We positioned our stimulating electrodes so that granule cells were not excited antidromically by fimbria stimulation, although action potentials were easily triggered in area CA3b and CA3c pyramidal cells by such stimulation. In these experiments, fimbria stimulation evoked responses from every hilar cell tested, including examples of both of the major cell types, the spiny hilar ‘mossy’ cells (n=15) and the relatively aspiny. ‘fast-spiking’ cells (putative interneurons, n=5). Hilar cell responses consisted primarily of EPSPs that could trigger action potentials, but small IPSPs were also evoked in some cases, particularly in the fast-spiking cells. Excitation was blocked by an antagonist of the AMPA/kainate receptor subtype of excitatory amino acid receptors, 6-cyano-7-nitroquinoxaline-2,3-dione(CNQX, 5μM, n=5), whereas the cholinergic antagonist atropine (10μM) had no effect (n=4). When sequential intracellular recordings were made from hilar cells and area CA3 pyramidal cells in the same slice, hilar cell EPSPs began after action potentials of CA3b pyramidal cells, and stimulus strengths required to evoke hilar cell EPSPs were above threshold for area CA3b pyramidal cells. Taken together with the evidence that area CA3 pyramidal cells use an excitatory amino acid as a neurotransmitter [7, 21], and the demonstrations of area CA3 axon collaterals in the hilus [11, 16], the results raise the possibility that some area CA3 pyramidal cells excite dentate hilar neurons. PMID:8105429

  9. Prevalence of F107 fimbriae on Escherichia coli isolated from pigs with oedema disease or postweaning diarrhoea.

    PubMed

    Imberechts, H; Bertschinger, H U; Stamm, M; Sydler, T; Pohl, P; De Greve, H; Hernalsteens, J P; Van Montagu, M; Lintermans, P

    1994-06-01

    The study comprises fifty 4 to 12 weeks old pigs that died from oedema disease or severe diarrhoea. Smears were prepared from the mucosa of duodenum, jejunum and ileum, and by immunofluorescence F107 fimbrial antigens were detected. E. coli strains were isolated from the intestines and were characterised by slide agglutination (serogroup and F107 fimbriae production), by their cytotoxicity for Vero cells, and by gene amplification (genes coding for the major F107 subunit FedA, the toxin causing oedema disease SLT-IIv, and enterotoxins LTI, STIa and STII). F107 fimbriae were demonstrated in association with E. coli of serogroups O139:K12 and O141:K85a,b but not of serogroup O149:K91:F4a,c. Expression in culture of F107 fimbriae by some isolates gave additional evidence for production of these fimbriae by ETEC strains. The genetic determinant of SLT-Ilv was found in association with F107, and could not be detected in serogroup O149:K91:F4a,c. Gene fedA was demonstrated in two isolates which were devoid of SLT-IIv. Most isolates from cases of oedema disease belonged to serogroup O139:K12 and did not contain enterotoxin genes. Isolates from pigs that suffered from diarrhoea were serotyped O141:K85a,b or O149:K91:F4a,c, and carried at least two enterotoxin genes in their genomes. In a small proportion of the cases F107 antigens were demonstrated in intestinal smears although gene fedA was not detected in the corresponding isolates. The results confirm the importance of F107 fimbriae as virulence factor in oedema disease E. coli strains, but also demonstrate that F107 fimbriae can be found in association with postweaning diarrhoea isolates. In these latter strains enterotoxins were always demonstrated, irrespective of the presence of toxin SLT-IIv.

  10. Identification of glyceraldehyde-3-phosphate dehydrogenase of epithelial cells as a second molecule that binds to Porphyromonas gingivalis fimbriae.

    PubMed

    Sojar, Hakimuddin T; Genco, Robert J

    2005-07-01

    Binding of Porphyromonas gingivalis to the host cells is an essential step in the pathogenesis of periodontal disease. P. gingivalis binds to and invades epithelial cells, and fimbriae are thought to be involved in this process. In our earlier studies, two major epithelial cell components of 40 and 50 kDa were identified as potential fimbrial receptors. Sequencing of a cyanogen bromide digestion fragment of the 50-kDa component resulted in an internal sequence identical to keratin I molecules, and hence this cytokeratin represents one of the epithelial cell receptors for P. gingivalis fimbriae. In this study, the 40-kDa component of KB cells was isolated and its amino-terminal sequence determined. The N-terminal amino sequence was found to be GKVKVGVNGF and showed perfect homology with human glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Furthermore, purified P. gingivalis fimbriae were found to bind to rabbit muscle GAPDH. Antibodies directed against internal peptide 49-68 and 69-90 of fimbrillin were shown to inhibit the binding of P. gingivalis and of fimbriae to epithelial cells. Antibodies against these peptides also inhibited the binding of fimbriae to GAPDH. Our results confirmed that the amino-terminal domain corresponding to amino residues 49-68 of the fimbrillin protein is the major GAPDH binding domain. These studies point to GAPDH as a major receptor for P. gingivalis major fimbriae and, as such, GAPDH likely plays a role in P. gingivalis adherence and colonization of the oral cavity, as well as triggering host cell processes involved in the pathogenesis of P. gingivalis infections.

  11. Oral Escherichia coli Colonization Factor Antigen I (CFA/I) Fimbriae Ameliorate Arthritis via IL-35, not IL-27

    PubMed Central

    Kochetkova, Irina; Thornburg, Theresa; Callis, Gayle; Holderness, Kathryn; Maddaloni, Massimo; Pascual, David W.

    2014-01-01

    A Salmonella therapeutic expressing enterotoxigenic E. coli colonization factor antigen I (CFA/I) fimbriae protects against collagen-induced arthritis (CIA) by eliciting two regulatory T cell (Treg) subsets: TGF-β-producing Foxp3−CD39+CD4+ and IL-10-producing Foxp3+CD39+CD4+ T cells. However, it is unclear if CFA/I fimbriae alone are protective, and if other regulatory cytokines are involved especially in the context for the EBI3-sharing cytokines, Treg-derived IL-35 and APC-derived IL-27, both capable of suppressing Th17 cells and regulating autoimmune diseases. Subsequent evaluation revealed that a single oral dose of purified, soluble CFA/I fimbriae protected against CIA as effectively as Salmonella-CFA/I, and found Foxp3+CD39+CD4+ T cells as the source of secreted IL-35, whereas IL-27 production by CD11c+ cells was inhibited. Inquiring into their relevance, CFA/I fimbriae-treated IL-27 receptor-deficient (WSX-1−/−) mice were equally protected against CIA as wild-type mice suggesting a limited role for IL-27. In contrast, CFA/I fimbriae-mediated protection was abated in EBI3−/− mice accompanied by the loss of TGF-β- and IL-10-producing Tregs. Adoptive transfer of B6 CD39+CD4+ T cells to EBI3−/− mice with concurrent CFA/I plus IL-35 treatment effectively stimulated Tregs suppressing proinflammatory CII-specific Th cells. Opposingly, recipients co-transferred with B6 and EBI3−/− CD39+CD4+ T cells and treated with CFA/I plus IL-35 failed in protecting mice implicating the importance for endogenous IL-35 to confer CFA/I-mediated protection. Thus, CFA/I fimbriae stimulate IL-35 required for the co-induction of TGF-β and IL-10. PMID:24337375

  12. Radial maze performance in three strains of mice - Role of the fimbria/fornix

    NASA Technical Reports Server (NTRS)

    Reinstein, D. K.; Deboissiere, T.; Robinson, N.; Wurtman, R. J.

    1983-01-01

    Three strains of mice were tested on an 8-arm radial maze, an index of hippocampus-dependent spatial memory. Levels of performance differed betweens strains with C57Br/cdj greater than Balb/cj greater than C57B1/6j. Lesions of the fimbria/fornix disrupted performance in the C57Br and Balb strains: the C57Bl mice never performed better than chance before or after surgery. Choline acetyltransferase activity in hippocampus was not correlated with radial maze performance. These findings suggest a possible genetic contribution towards radial maze behavior.

  13. Radial maze performance in three strains of mice: role of the fimbria/fornix.

    PubMed

    Reinstein, D K; DeBoissiere, T; Robinson, N; Wurtman, R J

    1983-03-14

    Three strains of mice were tested on an 8-arm radial maze, an index of hippocampus-dependent spatial memory. Levels of performance differed between strains with C57Br/cj greater than Balb/cj greater than C57Bl/6j. Lesions of the fimbria/fornix disrupted performance in the C57Br and Balb strains: the C57Bl mice never performed better than chance before or after surgery. Choline acetyltransferase activity in hippocampus was not correlated with radial maze performance. These findings suggest a possible genetic contribution towards radial maze behavior.

  14. Behavioral deficits after intrahippocampal fetal septal grafts in rats with selective fimbria-fornix lesions.

    PubMed

    Dalrymple-Alford, J C; Kelche, C; Cassel, J C; Toniolo, G; Pallage, V; Will, B E

    1988-01-01

    Fetal septal transplants have been shown to promote behavioral recovery in young adult rats with aspiration fimbria-fornix lesions, rats with septal lesions and in intact aged rats. The present study examined the behavioral impact of intrahippocampal septal cell suspension transplants (T) in young female rats that had received, 10 days earlier, either medial fimbria lesions (Group FI.T), dorsal (subcallosal) fornix lesions (Group FO.T) or these two lesions together (Group FIFO.T). Relative to rats with lesions only (groups FI, FO and FIFO), grafted rats, irrespective of lesion locus, displayed unexpected impairments in (i) a serial alternation learning task, 5 weeks and 6 months after transplantation, and (ii) in a radial maze, 7 months after transplantation. In the first alternation test, Group FIFO showed impaired performance relative to Groups FI, FO and the sham-operated controls (Group S). In the second alternation test, Groups FO.T and FO showed impaired performance relative to Groups FI.T and FI, and only the performance of Group FI did not differ from that of Group S. In the radial maze, Groups FI, FO and FIFO all showed impaired performance relative to Group S. By contrast, there were no deleterious effects of lesions or of grafts in the acquisition and retention of a step-through passive avoidance task, 10 weeks after transplantation. Our findings on the effects of selective fimbria-fornix lesions did not confirm the report that rats with FI lesions but not those with FO lesions are unable to learn a serial alternation task, nor the report that FO lesions impair passive avoidance retention. Acetylcholinesterase (AChE) histochemistry revealed that grafts were present but graft-derived innervation of the host hippocampus varied from extensive to almost non-existent in all transplant groups. AChE-positivity in the dorsal hippocampus (DH) was not related to behavioral performance. However, the grafts often grew to a considerable size within the host brain and

  15. Time-dependent effects of intrahippocampal grafts in rats with fimbria-fornix lesions.

    PubMed

    Cassel, J C; Kelche, C; Will, B

    1990-01-01

    Based on three experiments, this study examined whether behavioral and histological effects of fetal septal or hippocampal grafts placed in the denervated hippocampus depend on the duration of post-grafting delays. Each experiment included four groups of rats: sham-operated rats (Sham), rats with aspirative lesions of the fimbria-fornix (Fifo) and rats given both Fifo lesions and intrahippocampal fetal suspension grafts of either septal (Fifo.ST) or hippocampal (Fifo.HT) origin. All rats were tested (i) for home cage activity, (ii) for activity and reactivity in an open field and (iii) for learning ability in a 8-arm radial maze. Except for home cage activity which was also monitored preoperatively, behavioral tests were conducted between 1-2 months postgrafting in Experiment 1 (EXP1), 5-6 months post-grafting in Experiment 2 (EXP2) and 10-11 months post-grafting in Experiment 3 (EXP3). Each test period lasted 3 weeks. Histological controls consisted of acetylcholinesterase (AChE) and cresyl violet staining. Graft size was estimated by computerized image analysis. Normal rats performed well in each experiment. In all experiments, rats with fimbria-fornix lesions showed increased activity in both their familiar (home cage) and unfamiliar (open field) environments, and their performances in the radial maze task were impaired. In no experiment did grafts, whether hippocampal or septal, affect "noncognitive" behavioral variables. However, maze performance was improved by hippocampal grafts in EXP1 (short delay) and by septal grafts in EXP2 (intermediate delay). No graft-induced effect was found in EXP3 (long delay). Concerning AChE-positivity in the dorsal hippocampus, fimbria-fornix lesions reduced staining densities by at least 60%. Both types of grafts were undiscernably AChE-positive, but only septal grafts provided the denervated hippocampus with a significant AChE-positive fiber ingrowth. Differences among groups in density of hippocampal AChE staining were

  16. Radial maze performance in three strains of mice - Role of the fimbria/fornix

    NASA Technical Reports Server (NTRS)

    Reinstein, D. K.; Deboissiere, T.; Robinson, N.; Wurtman, R. J.

    1983-01-01

    Three strains of mice were tested on an 8-arm radial maze, an index of hippocampus-dependent spatial memory. Levels of performance differed betweens strains with C57Br/cdj greater than Balb/cj greater than C57B1/6j. Lesions of the fimbria/fornix disrupted performance in the C57Br and Balb strains: the C57Bl mice never performed better than chance before or after surgery. Choline acetyltransferase activity in hippocampus was not correlated with radial maze performance. These findings suggest a possible genetic contribution towards radial maze behavior.

  17. Transcriptome of Proteus mirabilis in the Murine Urinary Tract: Virulence and Nitrogen Assimilation Gene Expression▿†

    PubMed Central

    Pearson, Melanie M.; Yep, Alejandra; Smith, Sara N.; Mobley, Harry L. T.

    2011-01-01

    The enteric bacterium Proteus mirabilis is a common cause of complicated urinary tract infections. In this study, microarrays were used to analyze P. mirabilis gene expression in vivo from experimentally infected mice. Urine was collected at 1, 3, and 7 days postinfection, and RNA was isolated from bacteria in the urine for transcriptional analysis. Across nine microarrays, 471 genes were upregulated and 82 were downregulated in vivo compared to in vitro broth culture. Genes upregulated in vivo encoded mannose-resistant Proteus-like (MR/P) fimbriae, urease, iron uptake systems, amino acid and peptide transporters, pyruvate metabolism enzymes, and a portion of the tricarboxylic acid (TCA) cycle enzymes. Flagella were downregulated. Ammonia assimilation gene glnA (glutamine synthetase) was repressed in vivo, while gdhA (glutamate dehydrogenase) was upregulated in vivo. Contrary to our expectations, ammonia availability due to urease activity in P. mirabilis did not drive this gene expression. A gdhA mutant was growth deficient in minimal medium with citrate as the sole carbon source, and loss of gdhA resulted in a significant fitness defect in the mouse model of urinary tract infection. Unlike Escherichia coli, which represses gdhA and upregulates glnA in vivo and cannot utilize citrate, the data suggest that P. mirabilis uses glutamate dehydrogenase to monitor carbon-nitrogen balance, and this ability contributes to the pathogenic potential of P. mirabilis in the urinary tract. PMID:21505083

  18. Curli fimbriae are conditionally required in Escherichia coli O157:H7 for initial attachment and biofilm formation

    USDA-ARS?s Scientific Manuscript database

    Several species of enteric pathogens produce curli fimbriae, which may affect their interaction with surfaces and other microbes in nonhost environments. Here we used two E. coli O157:H7 outbreak strains with distinct genotypes to understand the role of curli in surface attachment and biofilm format...

  19. Production of Cellulose and Curli Fimbriae by Members of the Family Enterobacteriaceae Isolated from the Human Gastrointestinal Tract

    PubMed Central

    Zogaj, Xhavit; Bokranz, Werner; Nimtz, Manfred; Römling, Ute

    2003-01-01

    Citrobacter spp., Enterobacter spp., and Klebsiella spp. isolated from the human gut were investigated for the biosynthesis of cellulose and curli fimbriae (csg). While Citrobacter spp. produced curli fimbriae and cellulose and Enterobacter spp. produced cellulose with various temperature-regulatory programs, Klebsiella spp. did not show pronounced expression of those extracellular matrix components. Investigation of multicellular behavior in two Citrobacter species and Enterobacter sakazakii showed an extracellular matrix, cell clumping, pellicle formation, and biofilm formation associated with the expression of cellulose and curli fimbriae. In those three strains, the csgD-csgBA region and the cellulose synthase gene bcsA were conserved. PCR screening for the presence of csgD, csgA and bcsA revealed that besides Klebsiella pneumoniae and Klebsiella oxytoca, all species investigated harbored the genetic information for expression of curli fimbriae and cellulose. Since Citrobacter spp., Enterobacter spp., and Klebsiella spp. are frequently found to cause biofilm-related infections such as catheter-associated urinary tract infections, the human gut could serve as a reservoir for dissemination of biofilm-forming isolates. PMID:12819107

  20. CRP-Cyclic AMP Regulates the Expression of Type 3 Fimbriae via Cyclic di-GMP in Klebsiella pneumoniae

    PubMed Central

    Lin, Ching-Ting; Lin, Tien-Huang; Wu, Chien-Chen; Wan, Lei; Huang, Chun-Fa; Peng, Hwei-Ling

    2016-01-01

    Klebsiella pneumoniae is the predominant pathogen isolated from liver abscesses of diabetic patients in Asian countries. However, the effects of elevated blood glucose levels on the virulence of this pathogen remain largely unknown. Type 3 fimbriae, encoded by the mrkABCDF genes, are important virulence factors in K. pneumoniae pathogenesis. In this study, the effects of exogenous glucose and the intracellular cyclic AMP (cAMP) signaling pathway on type 3 fimbriae expression regulation were investigated. The production of MrkA, the major subunit of type 3 fimbriae, was increased in glucose-rich medium, whereas cAMP supplementation reversed the effect. MrkA production was markedly increased by cyaA or crp deletion, but slightly decreased by cpdA deletion. In addition, the mRNA levels of mrkABCDF genes and the activity of PmrkA were increased in Δcrp strain, as well as the mRNA levels of mrkHIJ genes that encode cyclic di-GMP (c-di-GMP)-related regulatory proteins that influence type 3 fimbriae expression. Moreover, the activities of PmrkHI and PmrkJ were decreased in ΔlacZΔcrp strain. These results indicate that CRP-cAMP down-regulates mrkABCDF and mrkHIJ at the transcriptional level. Further deletion of mrkH or mrkI in Δcrp strain diminished the production of MrkA, indicating that MrkH and MrkI are required for the CRP regulation of type 3 fimbriae expression. Furthermore, the high activity of PmrkHI in the ΔlacZΔcrp strain was diminished in ΔlacZΔcrpΔmrkHI, but increased in the ΔlacZΔcrpΔmrkJ strain. Deletion of crp increased the intracellular c-di-GMP concentration and reduced the phosphodiesterase activity. Moreover, we found that the mRNA levels of multiple genes related to c-di-GMP metabolism were altered in Δcrp strain. These indicate that CRP regulates type 3 fimbriae expression indirectly via the c-di-GMP signaling pathway. In conclusion, we found evidence of a coordinated regulation of type 3 fimbriae expression by the CRP-cAMP and c

  1. Bordetella filamentous hemagglutinin and fimbriae: critical adhesins with unrealized vaccine potential.

    PubMed

    Scheller, Erich V; Cotter, Peggy A

    2015-11-01

    Pertussis, or whooping cough, is a highly contagious respiratory disease that is caused by the Gram-negative bacterium Bordetella pertussis, which is transmitted exclusively from human to human. While vaccination against B. pertussis has been successful, replacement of the whole cell vaccine with an acellular component vaccine has correlated with reemergence of the disease, especially in adolescents and infants. Based on their presumed importance in mediating adherence to host tissues, filamentous hemagglutinin (FHA) and fimbria (FIM) were selected as components of most acellular pertussis vaccines. In this review, we describe the biogenesis of FHA and FIM, recent data that show that these factors do, in fact, play critical roles in adherence to respiratory epithelium, and evidence that they also contribute to persistence in the lower respiratory tract by modulating the host immune response. We also discuss shortcomings of whole cell and acellular pertussis vaccines and the possibility that FHA and FIM could serve as effective protective antigens in next-generation vaccines.

  2. FimH adhesin of Escherichia coli K1 type 1 fimbriae activates BV-2 microglia

    SciTech Connect

    Lee, Jongseok; Shin, Sooan; Teng, C.-H.; Hong, Suk Jin; Kim, Kwang Sik . E-mail: kwangkim@jhmi.edu

    2005-09-02

    The generation of intense inflammation in the subarachnoid space in response to meningitis-causing bacteria contributes to brain dysfunction and neuronal injury in bacterial meningitis. Microglia, the major immune effector cells in the central nervous system (CNS), become activated by bacterial components to produce proinflammatory immune mediators. In this study, we showed that FimH adhesin, a tip component of type 1 fimbriae of meningitis-causing Escherichia coli K1, activated the murine microglial cell line, BV-2, which resulted in the production of nitric oxide and the release of tumor necrosis factor-{alpha}. Mitogen-activated protein kinases, ERK and p-38, and nuclear factor-{kappa}B were involved in FimH adhesin-mediated microglial activation. These findings suggest that FimH adhesin contributes to the CNS inflammatory response by virtue of activating microglia in E. coli meningitis.

  3. Detection of antibodies against fimbria type 3 (Fim3) is useful diagnostic assay for pertussis.

    PubMed

    Oguchi, Kaoru; Miyata, Akiko; Kazuyama, Yukimasa; Noda, Atsuya; Suzuki, Eri; Watanabe, Mineo; Nakayama, Tetsuo

    2015-09-01

    Isolation of Bordetella pertussis and detection of the pertussis genome are not always successful because of low bacterial loads in adult patients with pertussis. Antibodies against pertussis toxin (PT) are measured but have low sensitivity in vaccinated subjects. There is no reliable diagnostic method at present. In this study, a fluorescent-EIA against several pertussis antigens and genome detection were investigated to establish clinical laboratory diagnostic methods for pertussis. The study was conducted in an outpatient clinic between September 2007 and 2013. Subjects consisted of 209 patients including adults suspected of pertussis and 35 staff members of the clinic. Loop-mediated isothermal amplification (LAMP) was performed to detect the pertussis genome in 5' UTR of the pertussis toxin (PT) gene. The catalytic region of the adenylate cyclase toxin (catACT), C-terminal of filamentous hemagglutinin (cFHA), and type 3 fimbria (Fim3) were selected, which are not pertussis vaccine component. Conventional PT and FHA antibodies were examined together with type 2 fimbria (Fim2) antibodies, and these are vaccine antigens. Pertussis DNA was detected in 23 (11%) out of 209. Detection sensitivity was high in young infants. Antibodies against Fim3 showed a higher positive rate in all age groups. Staff members at the pediatric outpatient clinic showed serological booster responses in Fim2 and Fim3 antibodies more sensitively than those in PT antibodies during outbreaks. LAMP was useful for detecting the pertussis genome in young infants, whereas a serological assay for fluorescent-EIA against Fim2 and Fim3 was preferable for adolescents and adults. Copyright © 2015. Published by Elsevier Ltd.

  4. Maternal immunity enhances systemic recall immune responses upon oral immunization of piglets with F4 fimbriae.

    PubMed

    Nguyen, Ut V; Melkebeek, Vesna; Devriendt, Bert; Goetstouwers, Tiphanie; Van Poucke, Mario; Peelman, Luc; Goddeeris, Bruno M; Cox, Eric

    2015-06-23

    F4 enterotoxigenic Escherichia coli (ETEC) cause diarrhoea and mortality in piglets leading to severe economic losses. Oral immunization of piglets with F4 fimbriae induces a protective intestinal immune response evidenced by an F4-specific serum and intestinal IgA response. However, successful oral immunization of pigs with F4 fimbriae in the presence of maternal immunity has not been demonstrated yet. In the present study we aimed to evaluate the effect of maternal immunity on the induction of a systemic immune response upon oral immunization of piglets. Whereas F4-specific IgG and IgA could be induced by oral immunization of pigs without maternal antibodies and by intramuscular immunization of pigs with maternal antibodies, no such response was seen in the orally immunized animals with maternal antibodies. Since maternal antibodies can mask an antibody response, we also looked by ELIspot assays for circulating F4-specific antibody secreting cells (ASCs). Enumerating the F4-specific ASCs within the circulating peripheral blood mononuclear cells, and the number of F4-specific IgA ASCs within the circulating IgA(+) B-cells revealed an F4-specific immune response in the orally immunized animals with maternal antibodies. Interestingly, results suggest a more robust IgA booster response by oral immunization of pigs with than without maternal antibodies. These results demonstrate that oral immunization of piglets with F4-specific maternal antibodies is feasible and that these maternal antibodies seem to enhance the secondary systemic immune response. Furthermore, our ELIspot assay on enriched IgA(+) B-cells could be used as a screening procedure to optimize mucosal immunization protocols in pigs with maternal immunity.

  5. Fimbriae-mediated outer membrane vesicle production and invasion of Porphyromonas gingivalis.

    PubMed

    Mantri, Chinmay K; Chen, Chin-Ho; Dong, Xinhong; Goodwin, Jeffery Shawn; Pratap, Siddharth; Paromov, Victor; Xie, Hua

    2015-02-01

    Porphyromonas gingivalis is a keystone periopathogen that plays an essential role in the progress of periodontitis. Like other gram-negative bacteria, the ability of P. gingivalis to produce outer membrane vesicles is a strategy used to interact with, and survive within its biological niches. Here we compared the protein components associated with vesicles derived from a fimbriated strain (33277) and an afimbriated strain (W83) of P. gingivalis using proteomic analyses. Some well-known virulence factors were identified in vesicles from both strains, such as gingipains and hemagglutinin. In contrast, FimC, FimD, and FimE, minor components of long fimbriae were found exclusively in 33277 vesicles, while proteins with a tetratricopeptide repeat (TPR) domain were unique to W83 vesicles. We found that significantly more 33277 than W83 vesicles were internalized into human oral keratinocytes and gingival fibroblasts. Interestingly, FimA, a well-known adhesin responsible for the attachment and invasion of P. gingivalis into host cells, was not essential for the invasive capabilities of P. gingivalis vesicles. Rather minor components of long fimbriae were required for an efficient invasive activity of vesicles. The most striking finding was that P. gingivalis strains lacking or having a reduced FimA expression showed a significant reduction in vesiculation. These results suggest that production and pathogenicity of P. gingivalis vesicles may largely depend on expression of the fim locus, and that the integration of vesicle production and pathogenicity with fimbrial expression may allow P. gingivalis to confer upon itself certain functional advantages. © 2014 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

  6. Cornel iridoid glycoside improves memory ability and promotes neuronal survival in fimbria-fornix transected rats.

    PubMed

    Zhao, Li-hong; Ding, Yue-xia; Zhang, Lan; Li, Lin

    2010-11-25

    Cornel iridoid glycoside (CIG) is a main component extracted from a traditional Chinese herb Cornus officinalis. Our previous study found that CIG improved neurological function in cerebral ischemic rats. The aim of this study was to investigate the therapeutic benefit of CIG in rats with fimbria-fornix transection (FFT) and explore the underlying molecular mechanisms. CIG (20, 60 and 180 mg/kg) or vehicle was intragastrically administered once daily to rats, starting immediately after the surgery and lasting for 4 weeks. Morris water maze and step-through tests showed that the memory deficits seen in FFT rats were significantly improved by CIG treatment. Immunohistochemical analysis showed that CIG treatment attenuated the loss of neurons in hippocampus. To elucidate the memory-improving mechanism of CIG, the neurotrophic factors, synaptic proteins and Bcl-2 family proteins in hippocampus were measured by Western blot analysis. FFT reduced hippocampal protein levels of nerve growth factor (NGF), tyrosine receptor kinase A (Trk A), brain-derived neurotrophic factor (BDNF), synaptophysin (SYP) and B-cell lymphoma-2 (Bcl-2), but not levels of tyrosine receptor kinase B (Trk B) and growth-associated protein 43 (GAP-43). FFT also elevated cytochorome C (Cyt c) and bcl-2-associated X protein (Bax). Administration of CIG to FFT rats significantly elevated the expression of NGF, TrkA, BDNF, SYP, GAP-43 and Bcl-2, and decreased the expression of Cyt c and Bax. These results indicated that CIG effectively counteracted cognitive impairments caused by fimbria-fornix lesions, and the mechanisms might be related to promoting neuronal survival and providing a beneficial environment for brain repair. Copyright © 2010 Elsevier B.V. All rights reserved.

  7. Fimbriae-mediated outer membrane vesicle production and invasion of Porphyromonas gingivalis

    PubMed Central

    Mantri, Chinmay K; Chen, Chin-Ho; Dong, Xinhong; Goodwin, Jeffery Shawn; Pratap, Siddharth; Paromov, Victor; Xie, Hua

    2015-01-01

    Porphyromonas gingivalis is a keystone periopathogen that plays an essential role in the progress of periodontitis. Like other gram-negative bacteria, the ability of P. gingivalis to produce outer membrane vesicles is a strategy used to interact with, and survive within its biological niches. Here we compared the protein components associated with vesicles derived from a fimbriated strain (33277) and an afimbriated strain (W83) of P. gingivalis using proteomic analyses. Some well-known virulence factors were identified in vesicles from both strains, such as gingipains and hemagglutinin. In contrast, FimC, FimD, and FimE, minor components of long fimbriae were found exclusively in 33277 vesicles, while proteins with a tetratricopeptide repeat (TPR) domain were unique to W83 vesicles. We found that significantly more 33277 than W83 vesicles were internalized into human oral keratinocytes and gingival fibroblasts. Interestingly, FimA, a well-known adhesin responsible for the attachment and invasion of P. gingivalis into host cells, was not essential for the invasive capabilities of P. gingivalis vesicles. Rather minor components of long fimbriae were required for an efficient invasive activity of vesicles. The most striking finding was that P. gingivalis strains lacking or having a reduced FimA expression showed a significant reduction in vesiculation. These results suggest that production and pathogenicity of P. gingivalis vesicles may largely depend on expression of the fim locus, and that the integration of vesicle production and pathogenicity with fimbrial expression may allow P. gingivalis to confer upon itself certain functional advantages. PMID:25524808

  8. Identification of Cell Surface-Exposed Proteins Involved in the Fimbria-Mediated Adherence of Enteroaggregative Escherichia coli to Intestinal Cells

    PubMed Central

    Izquierdo, Mariana; Navarro-Garcia, Fernando; Nava-Acosta, Raul; Nataro, James P.; Ruiz-Perez, Fernando

    2014-01-01

    Fimbria-mediated adherence to the intestinal epithelia is a key step in enteroaggregative Escherichia coli (EAEC) pathogenesis. To date, four fimbriae have been described for EAEC; aggregative adherence fimbria II (AAF/II) is the most important adherence factor for EAEC prototype strain 042. Previously, we described results showing that extracellular matrix (ECM) components might be involved in the recognition of AAF/II fimbriae by intestinal cells. In this study, we sought to identify novel potential receptors on intestinal epithelial cells recognized by the AAF/II fimbriae. Purified AafA-dsc protein, the major subunit of AAF/II fimbriae, was incubated with a monolayer of T84 cells, cross-linked to the surface-exposed T84 cell proteins, and immunoprecipitated by using anti-AafA antibodies. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis of cellular proteins bound to AafA-dsc protein identified laminin (previously recognized as a potential receptor for AAF/II) and cytokeratin 8 (CK8). Involvement of the major subunit of AAF/II fimbriae (AafA protein) in the binding to recombinant CK8 was confirmed by adherence assays with purified AAF/II fimbriae, AafA-dsc protein, and strain 042. Moreover, HEp-2 cells transfected with CK8 small interfering RNA (siRNA) showed reduced 042 adherence compared with cells transfected with scrambled siRNA as a control. Adherence of 042 to HEp-2 cells preincubated with antibodies against ECM proteins or CK8 was substantially reduced. Altogether, our results supported the idea of a role of CK8 as a potential receptor for EAEC. PMID:24516112

  9. The role of adhesive F107 fimbriae and of SLT-IIv toxin in the pathogenesis of edema disease in pigs.

    PubMed

    Imberechts, H; de Greve, H; Hernalsteens, J P; Schlicker, C; Bouchet, H; Pohl, P; Charlier, G; Bertschinger, H U; Wild, P; Vandekerckhove, J

    1993-04-01

    Colonization of the small intestine and the excretion of a toxin are important steps in the pathogenesis of edema disease in pigs. Although much is known about the chemical and biological characteristics of SLT-IIv toxin, its mode of action and its genetic determinant, F107 fimbriae were only recently described as colonization factors. Here we summarize our current knowledge about the virulence factors F107 fimbriae and SLT-IIv toxin.

  10. The stem-cell profile of ovarian surface epithelium is reproduced in the oviductal fimbriae, with increased stem-cell marker density in distal parts of the fimbriae.

    PubMed

    Auersperg, Nelly

    2013-09-01

    High-grade serous ovarian carcinomas are the most common and most lethal ovarian cancers, but their histologic origin is still controversial. Current evidence suggests that they may originate in the ovarian surface epithelium (OSE) and/or epithelium of oviductal fimbriae (FE). To further investigate this question we compared the stem-cell profiles of these epithelia. Formalin-fixed sections of normal FE (N=21) and ovaries (N=21) were stained immunohistochemically for the stem-cell markers NANOG, SFRP1, LHX9, ALDH1A1, and ALDH1A2. All markers were detected in both OSE and FE. A total of 75% to 100% of surface OSE expressed all markers except ALDH1A1, which occurred in about 25% of cells. Among epithelial inclusion cysts with flat-to-cuboidal epithelium, resembling OSE, ALDH1A1 was significantly increased, whereas SFRP1 was reduced compared with surface OSE, suggesting an increased trend towards malignant transformation. Similarly, among cysts lined by columnar cells resembling FE, SFRP1 expression was low, whereas ALDH1A1 approached 100% of the cysts. FE exhibited considerable variation between and within specimens. In about half of the samples, SFRP1 and NANOG were detected in ≤25% FE. The most widespread markers were ALDH1A1 and ALDH1A2. The highest proportion of all markers occurred in the distal parts of the FE, the site of the putative ovarian cancer precursors. Marker expression in tubal ampullae was low or absent except for ALDH1A1 and ALDH1A2. The results provide an explanation for the characteristic distal location of fimbrial high-grade serous ovarian carcinoma precursor lesions, and indicate that both OSE and FE have the capacity to undergo neoplastic transformation.

  11. F9 Fimbriae of Uropathogenic Escherichia coli Are Expressed at Low Temperature and Recognise Galβ1-3GlcNAc-Containing Glycans

    PubMed Central

    Wurpel, Daniël J.; Totsika, Makrina; Allsopp, Luke P.; Hartley-Tassell, Lauren E.; Day, Christopher J.; Peters, Kate M.; Sarkar, Sohinee; Ulett, Glen C.; Yang, Ji; Tiralongo, Joe; Strugnell, Richard A.; Jennings, Michael P.; Schembri, Mark A.

    2014-01-01

    Uropathogenic Escherichia coli (UPEC) is the leading causative agent of urinary tract infections (UTI) in the developed world. Among the major virulence factors of UPEC, surface expressed adhesins mediate attachment and tissue tropism. UPEC strains typically possess a range of adhesins, with type 1 fimbriae and P fimbriae of the chaperone-usher class the best characterised. We previously identified and characterised F9 as a new chaperone-usher fimbrial type that mediates biofilm formation. However, the regulation and specific role of F9 fimbriae remained to be determined in the context of wild-type clinical UPEC strains. In this study we have assessed the distribution and genetic context of the f9 operon among diverse E. coli lineages and pathotypes and demonstrated that f9 genes are significantly more conserved in a UPEC strain collection in comparison to the well-defined E. coli reference (ECOR) collection. In the prototypic UPEC strain CFT073, the global regulator protein H-NS was identified as a transcriptional repressor of f9 gene expression at 37°C through its ability to bind directly to the f9 promoter region. F9 fimbriae expression was demonstrated at 20°C, representing the first evidence of functional F9 fimbriae expression by wild-type E. coli. Finally, glycan array analysis demonstrated that F9 fimbriae recognise and bind to terminal Galβ1-3GlcNAc structures. PMID:24671091

  12. Type 3 Fimbriae Encoded on Plasmids Are Expressed from a Unique Promoter without Affecting Host Motility, Facilitating an Exceptional Phenotype That Enhances Conjugal Plasmid Transfer

    PubMed Central

    Madsen, Jonas Stenløkke; Riber, Leise; Kot, Witold; Basfeld, Alrun; Burmølle, Mette; Hansen, Lars Hestbjerg; Sørensen, Søren Johannes

    2016-01-01

    Horizontal gene transfer (HGT), the transmission of genetic material to a recipient that is not the progeny of the donor, is fundamental in bacterial evolution. HGT is often mediated by mobile genetic elements such as conjugative plasmids, which may be in conflict with the chromosomal elements of the genome because they are independent replicons that may petition their own evolutionary strategy. Here we study differences between type 3 fimbriae encoded on wild type plasmids and in chromosomes. Using known and newly characterized plasmids we show that the expression of type 3 fimbriae encoded on plasmids is systematically different, as MrkH, a c-di-GMP dependent transcriptional activator is not needed for strong expression of the fimbriae. MrkH is required for expression of type 3 fimbriae of the Klebsiella pneumoniae chromosome, wherefrom the fimbriae operon (mrkABCDF) of plasmids is believed to have originated. We find that mrkABCDFs of plasmids are highly expressed via a unique promoter that differs from the original Klebsiella promoter resulting in fundamental behavioral consequences. Plasmid associated mrkABCDFs did not influence the swimming behavior of the host, that hereby acquired an exceptional phenotype being able to both actively swim (planktonic behavior) and express biofilm associated fimbriae (sessile behavior). We show that this exceptional phenotype enhances the conjugal transfer of the plasmid. PMID:27627107

  13. F9 fimbriae of uropathogenic Escherichia coli are expressed at low temperature and recognise Galβ1-3GlcNAc-containing glycans.

    PubMed

    Wurpel, Daniël J; Totsika, Makrina; Allsopp, Luke P; Hartley-Tassell, Lauren E; Day, Christopher J; Peters, Kate M; Sarkar, Sohinee; Ulett, Glen C; Yang, Ji; Tiralongo, Joe; Strugnell, Richard A; Jennings, Michael P; Schembri, Mark A

    2014-01-01

    Uropathogenic Escherichia coli (UPEC) is the leading causative agent of urinary tract infections (UTI) in the developed world. Among the major virulence factors of UPEC, surface expressed adhesins mediate attachment and tissue tropism. UPEC strains typically possess a range of adhesins, with type 1 fimbriae and P fimbriae of the chaperone-usher class the best characterised. We previously identified and characterised F9 as a new chaperone-usher fimbrial type that mediates biofilm formation. However, the regulation and specific role of F9 fimbriae remained to be determined in the context of wild-type clinical UPEC strains. In this study we have assessed the distribution and genetic context of the f9 operon among diverse E. coli lineages and pathotypes and demonstrated that f9 genes are significantly more conserved in a UPEC strain collection in comparison to the well-defined E. coli reference (ECOR) collection. In the prototypic UPEC strain CFT073, the global regulator protein H-NS was identified as a transcriptional repressor of f9 gene expression at 37°C through its ability to bind directly to the f9 promoter region. F9 fimbriae expression was demonstrated at 20°C, representing the first evidence of functional F9 fimbriae expression by wild-type E. coli. Finally, glycan array analysis demonstrated that F9 fimbriae recognise and bind to terminal Galβ1-3GlcNAc structures.

  14. Involvement of polyphosphate kinase in virulence and stress tolerance of uropathogenic Proteus mirabilis.

    PubMed

    Peng, Liang; Jiang, Qiao; Pan, Jia-Yun; Deng, Cong; Yu, Jing-Yi; Wu, Xiao-Man; Huang, Sheng-He; Deng, Xiao-Yan

    2016-04-01

    Proteus mirabilis (P. mirabilis), a gram-negative enteric bacterium, frequently causes urinary tract infections. Many virulence factors of uropathogenic P. mirabilis have been identified, including urease, flagella, hemolysin and fimbriae. However, the functions of polyphosphate kinase (PPK), which are related to the pathogenicity of many bacteria, remain entirely unknown in P. mirabilis. In this study, a ppk gene encoding the PPK insertional mutant in P. mirabilis strain HI4320 was constructed, and its biological functions were examined. The results of survival studies demonstrated that the ppk mutant was deficient in resistance to oxidative, hyperosmotic and heat stress. The swarming and biofilm formation abilities of P. mirabilis were also attenuated after the ppk interruption. In vitro and in vivo experiments suggested that ppk was required for P. mirabilis to invade the bladder. The negative phenotypes of the ppk mutant could be restored by ppk gene complementation. Furthermore, two-dimensional gel electrophoresis and liquid chromatography-mass spectrometry were used to analyze the proteomes of the wild-type strain and the ppk mutant. Compared with the wild-type strain, seven proteins including TonB-dependent receptor, universal stress protein G, major mannose-resistant/Proteus-like fimbrial protein (MR/P fimbriae), heat shock protein, flagellar capping protein, putative membrane protein and multidrug efflux protein were down-regulated, and four proteins including exported peptidase, repressor protein for FtsI, FKBP-type peptidyl-prolyl cis-trans isomerase and phosphotransferase were up-regulated in the ppk mutant. As a whole, these results indicate that PPK is an important regulator and plays a crucial role in stress tolerance and virulence in uropathogenic P. mirabilis.

  15. Purification and partial immunochemical characterization of proteins of fimbriae F107 from Escherichia coli isolated from edema disease of pigs.

    PubMed

    Rosocha, J; Mikula, I; Kalinácová, V; Kollárová, Z

    1995-01-01

    The paper describes the isolation, purification and characterization of F107-fimbrial proteins, obtained by thermoelution from Escherichia coli 107/86. Isolation of the pure F107 protein was done by FPLC chromatography, employing Superose 12, Mono Q, and Phenyl-Superose columns. The highest purity of the F107 protein was achieved with Superose 12 HR 10/30. Purity checking by a HPLC system Waters 625 LC (Millipore) proved the absence of protein admixtures in a fraction from Superose 12. Analysis of the molar mass of F107 proteins by SDS PAGE revealed that F107 fimbriae consist of two proteins, one of M = 43 kDa (minor), and other of M = 18.9 kDa (major). Western blot analysis with rabbit polyclonal antiserum confirmed that the 18.9 kDa protein was the major characteristic unit of F107 fimbriae.

  16. Glutathione Deficit Affects the Integrity and Function of the Fimbria/Fornix and Anterior Commissure in Mice: Relevance for Schizophrenia

    PubMed Central

    Corcoba, Alberto; Steullet, Pascal; Duarte, João M. N.; Van de Looij, Yohan; Monin, Aline; Cuenod, Michel; Gruetter, Rolf

    2016-01-01

    Background: Structural anomalies of white matter are found in various brain regions of patients with schizophrenia and bipolar and other psychiatric disorders, but the causes at the cellular and molecular levels remain unclear. Oxidative stress and redox dysregulation have been proposed to play a role in the pathophysiology of several psychiatric conditions, but their anatomical and functional consequences are poorly understood. The aim of this study was to investigate white matter throughout the brain in a preclinical model of redox dysregulation. Methods: In a mouse model with impaired glutathione synthesis (Gclm KO), a state-of-the-art multimodal magnetic resonance protocol at high field (14.1 T) was used to assess longitudinally the white matter structure, prefrontal neurochemical profile, and ventricular volume. Electrophysiological recordings in the abnormal white matter tracts identified by diffusion tensor imaging were performed to characterize the functional consequences of fractional anisotropy alterations. Results: Structural alterations observed at peri-pubertal age and adulthood in Gclm KO mice were restricted to the anterior commissure and fornix-fimbria. Reduced fractional anisotropy in the anterior commissure (-7.5%±1.9, P<.01) and fornix-fimbria (-4.5%±1.3, P<.05) were accompanied by reduced conduction velocity in fast-conducting fibers of the posterior limb of the anterior commissure (-14.3%±5.1, P<.05) and slow-conducting fibers of the fornix-fimbria (-8.6%±2.6, P<.05). Ventricular enlargement was found at peri-puberty (+25%±8 P<.05) but not in adult Gclm KO mice. Conclusions: Glutathione deficit in Gclm KO mice affects ventricular size and the integrity of the fornix-fimbria and anterior commissure. This suggests that redox dysregulation could contribute during neurodevelopment to the impaired white matter and ventricle enlargement observed in schizophrenia and other psychiatric disorders. PMID:26433393

  17. Role of Fimbriae, Flagella and Cellulose on the Attachment of Salmonella Typhimurium ATCC 14028 to Plant Cell Wall Models.

    PubMed

    Tan, Michelle S F; White, Aaron P; Rahman, Sadequr; Dykes, Gary A

    2016-01-01

    Cases of foodborne disease caused by Salmonella are frequently associated with the consumption of minimally processed produce. Bacterial cell surface components are known to be important for the attachment of bacterial pathogens to fresh produce. The role of these extracellular structures in Salmonella attachment to plant cell walls has not been investigated in detail. We investigated the role of flagella, fimbriae and cellulose on the attachment of Salmonella Typhimurium ATCC 14028 and a range of isogenic deletion mutants (ΔfliC fljB, ΔbcsA, ΔcsgA, ΔcsgA bcsA and ΔcsgD) to bacterial cellulose (BC)-based plant cell wall models [BC-Pectin (BCP), BC-Xyloglucan (BCX) and BC-Pectin-Xyloglucan (BCPX)] after growth at different temperatures (28°C and 37°C). We found that all three cell surface components were produced at 28°C but only the flagella was produced at 37°C. Flagella appeared to be most important for attachment (reduction of up to 1.5 log CFU/cm2) although both cellulose and fimbriae also aided in attachment. The csgD deletion mutant, which lacks both cellulose and fimbriae, showed significantly higher attachment as compared to wild type cells at 37°C. This may be due to the increased expression of flagella-related genes which are also indirectly regulated by the csgD gene. Our study suggests that bacterial attachment to plant cell walls is a complex process involving many factors. Although flagella, cellulose and fimbriae all aid in attachment, these structures are not the only mechanism as no strain was completely defective in its attachment.

  18. Role of Fimbriae, Flagella and Cellulose on the Attachment of Salmonella Typhimurium ATCC 14028 to Plant Cell Wall Models

    PubMed Central

    Tan, Michelle S. F.; White, Aaron P.; Rahman, Sadequr

    2016-01-01

    Cases of foodborne disease caused by Salmonella are frequently associated with the consumption of minimally processed produce. Bacterial cell surface components are known to be important for the attachment of bacterial pathogens to fresh produce. The role of these extracellular structures in Salmonella attachment to plant cell walls has not been investigated in detail. We investigated the role of flagella, fimbriae and cellulose on the attachment of Salmonella Typhimurium ATCC 14028 and a range of isogenic deletion mutants (ΔfliC fljB, ΔbcsA, ΔcsgA, ΔcsgA bcsA and ΔcsgD) to bacterial cellulose (BC)-based plant cell wall models [BC-Pectin (BCP), BC-Xyloglucan (BCX) and BC-Pectin-Xyloglucan (BCPX)] after growth at different temperatures (28°C and 37°C). We found that all three cell surface components were produced at 28°C but only the flagella was produced at 37°C. Flagella appeared to be most important for attachment (reduction of up to 1.5 log CFU/cm2) although both cellulose and fimbriae also aided in attachment. The csgD deletion mutant, which lacks both cellulose and fimbriae, showed significantly higher attachment as compared to wild type cells at 37°C. This may be due to the increased expression of flagella-related genes which are also indirectly regulated by the csgD gene. Our study suggests that bacterial attachment to plant cell walls is a complex process involving many factors. Although flagella, cellulose and fimbriae all aid in attachment, these structures are not the only mechanism as no strain was completely defective in its attachment. PMID:27355584

  19. Structural and Functional Insight into the Carbohydrate Receptor Binding of F4 Fimbriae-producing Enterotoxigenic Escherichia coli *

    PubMed Central

    Moonens, Kristof; Van den Broeck, Imke; De Kerpel, Maia; Deboeck, Francine; Raymaekers, Hanne; Remaut, Han; De Greve, Henri

    2015-01-01

    Enterotoxigenic Escherichia coli (ETEC) strains are important causes of intestinal disease in humans and lead to severe production losses in animal farming. A range of fimbrial adhesins in ETEC strains determines host and tissue tropism. ETEC strains expressing F4 fimbriae are associated with neonatal and post-weaning diarrhea in piglets. Three naturally occurring variants of F4 fimbriae (F4ab, F4ac, and F4ad) exist that differ in the primary sequence of their major adhesive subunit FaeG, and each features a related yet distinct receptor binding profile. Here the x-ray structure of FaeGad bound to lactose provides the first structural insight into the receptor specificity and mode of binding by the poly-adhesive F4 fimbriae. A small D′-D″-α1-α2 subdomain grafted on the immunoglobulin-like core of FaeG hosts the carbohydrate binding site. Two short amino acid stretches Phe150–Glu152 and Val166–Glu170 of FaeGad bind the terminal galactose in the lactosyl unit and provide affinity and specificity to the interaction. A hemagglutination-based assay with E. coli expressing mutant F4ad fimbriae confirmed the elucidated co-complex structure. Interestingly, the crucial D′-α1 loop that borders the FaeGad binding site adopts a different conformation in the two other FaeG variants and hints at a heterogeneous binding pocket among the FaeG serotypes. PMID:25631050

  20. cAMP receptor protein regulates mouse colonization, motility, fimbria-mediated adhesion, and stress tolerance in uropathogenic Proteus mirabilis.

    PubMed

    Tsai, Yi-Lin; Chien, Hsiung-Fei; Huang, Kuo-Tong; Lin, Wen-Yuan; Liaw, Shwu-Jen

    2017-08-04

    Cyclic AMP receptor protein (Crp) is a major transcriptional regulator in bacteria. This study demonstrated that Crp affects numerous virulence-related phenotypes, including colonization of mice, motility, fimbria-mediated adhesion, and glucose stress tolerance in uropathogenic Proteus mirabilis. Diabetic mice were more susceptible to kidney colonization by wild-type strain than nondiabetic mice, in which the crp mutant exhibited increased kidney colonization. Loss of crp or addition of 10% glucose increased the P. mirabilis adhesion to kidney cells. Direct negative regulation of pmpA (which encodes the major subunit of P-like fimbriae) expression by Crp was demonstrated using a reporter assay and DNase I footprinting. Moreover, the pmpA/crp double mutant exhibited reduced kidney adhesion comparable to that of the pmpA mutant, and mouse kidney colonization by the pmpA mutant was significantly attenuated. Hence, the upregulation of P-like fimbriae in the crp mutant substantially enhanced kidney colonization. Moreover, increased survival in macrophages, increased stress tolerance, RpoS upregulation, and flagellum deficiency leading to immune evasion may promote kidney colonization by the crp mutant. This is the first study to elucidate the role of Crp in the virulence of uropathogenic P. mirabilis, underlying mechanisms, and related therapeutic potential.

  1. Shigella flexneri transformants expressing type 1 (mannose-specific) fimbriae bind to, activate, and are killed by phagocytic cells.

    PubMed Central

    Gbarah, A; Mirelman, D; Sansonetti, P J; Verdon, R; Bernhard, W; Sharon, N

    1993-01-01

    Shigella flexneri M90T (invasive) and BS176 (noninvasive) are typical nonfimbriated organisms that do not bind to or activate phagocytic cells. We demonstrate that S. flexneri M90Tp and BS176p, obtained by transformation of the strains named above with the cluster of genes encoding type 1 (mannose-specific) fimbriae of Escherichia coli, express the functional fimbriae, as shown by electron microscopy, by binding of antifimbria antibodies and by yeast cell aggregation. The transformants, but not the parental strains, bound to human granulocytes and mouse peritoneal macrophages. This binding was inhibited by methyl alpha-D-mannoside but not by methyl alpha-D-galactoside. The bound bacteria induced oxidative burst activation and degranulation of the granulocytes in vitro. With mouse peritoneal macrophages, the binding of the fimbriated bacteria induced degranulation in vitro. Injection of the bacteria into mouse peritoneum also induced degranulation of the macrophages in vivo; no such effect was observed with the nonfimbriated strains. The bound fimbriated transformants were effectively killed by the human granulocytes in vitro in the absence of opsonins or after opsonization with human anti-S. flexneri antiserum. The nonfimbriated strains were killed only after opsonization. These results provide further evidence for the role of type 1 fimbriae in lectin-mediated nonopsonic phagocytosis. Images PMID:8097492

  2. CfaE tip mutations in enterotoxigenic Escherichia coli CFA/I fimbriae define critical human intestinal binding sites.

    PubMed

    Baker, K K; Levine, M M; Morison, J; Phillips, A; Barry, E M

    2009-05-01

    Enterotoxigenic Escherichia coli (ETEC) use colonization factors to attach to the human intestinal mucosa, followed by enterotoxin expression that induces net secretion and diarrhoeal illness. ETEC strain H10407 expresses CFA/I fimbriae, which are composed of multiple CfaB structural subunits and a CfaE tip subunit. Currently, the contribution of these individual fimbrial subunits in intestinal binding remains incompletely defined. To identify the role of CfaE in attachment in the native ETEC background, an R181A single-amino-acid substitution was introduced by recombination into the H10407 genome. The substitution of R181A eliminated haemagglutination and binding of intestinal mucosa biopsies in in vitro organ culture assays, without loss of CFA/I fimbriae expression. Wild-type in trans plasmid-expressed cfaE restored the binding phenotype. In contrast, in trans expression of cfaE containing amino acid 181 substitutions with similar amino acids, lysine, methionine and glutamine did not restore the binding phenotype, indicating that the loss of the binding phenotype was due to localized areas of epitope disruption. R181 appears to have an irreplaceable role in the formation of a receptor-binding feature on CFA/I fimbriae. The results specifically indicate that the CfaE tip protein is a required binding factor in CFA/I-mediated ETEC colonization, making it a potentially important vaccine antigen. © 2009 Blackwell Publishing Ltd.

  3. Role of fimbriae F18 for actively acquired immunity against porcine enterotoxigenic Escherichia coli.

    PubMed

    Sarrazin, E; Bertschinger, H U

    1997-02-01

    Enterotoxigenic (ETEC) and enterotoxaemic (ETEEC) Escherichia (E.) coli that express F18 (F107) fimbriate colonize the small intestine and cause diarrhoea and/or oedema disease in weaned pigs. So far, two antigenic variants of F18 can be distinguished with a common antigenic factor designated 'a' and two specific factors called 'b' and 'c'. In this study the existence of crosswise anti-colonization immunity between E. coli strains that express F18ab or F18ac fimbrial variants, respectively, was demonstrated. Weaned pigs of susceptible genotype with respect to susceptibility to adhesion of E. coli with fimbriae F18 were inoculated with E. coli strains 3064STM (0157:K-:H-:F18ab; resistant to streptomycin) and 8199RIF (0141ab:K-:H4:F18ac; resistant to rifampicin). The faecal shedding was compared subsequent to immunization and homologous or heterologous challenge. An enzyme-linked immunosorbent assay (ELISA) was applied to measure IgA, IgM and IgG antibodies against the F18ab and F18ac antigens in saliva, faeces, serum and intestinal wash samples. About 8 log CFU/g of the inoculated strains were found in faeces of all pigs following immunization as well as in non-immunized controls after challenge. Bacterial counts of the inoculated strains after challenge were between 2 and 5 log lower, without any difference between homologous and heterologous challenge. Intestinal colonization with fimbriated E. coli resulted in production of significantly increased levels of anti-fimbrial antibodies, especially IgA, in serum and intestinal wash samples. There were higher levels of homologous than of heterologous anti-fimbrial antibodies. Production of antibodies against F18a or against another common fimbrial antigen is probably responsible for crosswise anti-colonization immunity between E. coli strains with F18ab and F18ac fimbrial variants. Serum F18-specific IgA may be a useful indicator of a mucosal immune response directed against F18 fimbriae.

  4. Structural predictions of AgfA, the insoluble fimbrial subunit of Salmonella thin aggregative fimbriae.

    PubMed

    Collinson, S K; Parker, J M; Hodges, R S; Kay, W W

    1999-07-16

    The unusually stable and multifunctional, thin aggregative fimbriae common to all Salmonella spp. are principally polymers of the fimbrin subunit, AgfA. AgfA of Salmonella enteritidis consists of two domains: a protease-sensitive, 22 amino acid residue N-terminal region and a protease-resistant, 109 residue C-terminal core. The unusual amino acid sequence of the AgfA core region comprises two-, five- and tenfold internal sequence homology patterns reflected in five conserved, 18-residue tandem repeats. These repeats have the consensus sequence, Sx5QxGx2NxAx3Q and are linked together by four or five residues, (x)xAx2. The predicted secondary structure for this unusual arrangement of tandem repeats in AgfA indicates mainly extended conformation with the beta strands linked by four to six residues. Candidate proteins of known structure with motifs of alternating beta strands and short loops were selected from folds described in SCOP as a source of coordinates for AgfA model construction. Three all-beta class motifs selected from the Serratia marcescens metalloprotease, myelin P2 protein or vitelline membrane outer protein I were used for initial AgfA homology build-up procedures ultimately resulting in three structural models; beta barrel, beta prism and parallel beta helix. The beta barrel model is a compact, albeit irregular structure, with the beta strands arranged in two antiparallel beta sheet faces. The beta prism model does not reflect the 5 or 10-fold symmetry of the AgfA primary sequence. However, the favored, parallel beta helix model is a compact coil of ten helically arranged beta strands forming two parallel beta sheet faces. This arrangement predicts a regular, potentially stable, C-terminal core region consistent with the observed tandem repeat sequences, protease-resistance and strong tendency of this fimbrin to oligomerize and aggregate. Positional conservation of amino acid residues in AgfA and the Escherichia coli AgfA homologue, CsgA, provides

  5. Crystallization of the FaeE chaperone of Escherichia coli F4 fimbriae

    PubMed Central

    Van Molle, Inge; Buts, Lieven; Coppens, Fanny; Qiang, Liu; Wyns, Lode; Loris, Remy; Bouckaert, Julie; De Greve, Henri

    2005-01-01

    F4 (formerly K88) fimbriae from enterotoxigenic Escherichia coli are assembled via the FaeE/FaeD chaperone/usher pathway. The chaperone FaeE crystallizes in three crystal forms, all belonging to space group C2. Crystals of form 1 diffract to 2.3 Å and have unit-cell parameters a = 195.7, b = 78.5, c = 184.6 Å, β = 102.2°. X-ray data for crystal form 2 were collected to 2.7 Å using an SeMet variant of FaeE. The crystals have unit-cell parameters a = 136.4, b = 75.7, c = 69.4 Å, β = 92.8°. Crystals of form 3 were formed in a solution containing the FaeE–FaeG complex and diffract to 2.8 Å. Unit-cell parameters are a = 109.7, b = 78.6, c = 87.8 Å, β = 96.4°. PMID:16511060

  6. Enterohemorrhagic Escherichia coli pathogenesis: role of Long polar fimbriae in Peyer’s patches interactions

    PubMed Central

    Cordonnier, Charlotte; Etienne-Mesmin, Lucie; Thévenot, Jonathan; Rougeron, Amandine; Rénier, Sandra; Chassaing, Benoit; Darfeuille-Michaud, Arlette; Barnich, Nicolas; Blanquet-Diot, Stéphanie; Livrelli, Valérie

    2017-01-01

    Enterohemorrhagic Escherichia coli (EHEC) are major food-borne pathogens whose survival and virulence in the human digestive tract remain unclear owing to paucity of relevant models. EHEC interact with the follicle-associated epithelium of Peyer’s patches of the distal ileum and translocate across the intestinal epithelium via M-cells, but the underlying molecular mechanisms are still unknown. Here, we investigated the involvement of Long polar fimbriae (Lpf) in EHEC pathogenesis. Of the 236 strains tested, a significant association was observed between the presence of lpf operons and pathogenicity. In sophisticated in vitro models of the human gastro-intestinal tract, lpf expression was induced during transit through the simulated stomach and small intestine, but not in the colonic compartment. To investigate the involvement of Lpf in EHEC pathogenesis, lpf isogenic mutants and their relative trans-complemented strains were generated. Translocation across M-cells, interactions with murine ileal biopsies containing Peyer’s patches and the number of hemorrhagic lesions were significantly reduced with the lpf mutants compared to the wild-type strain. Complementation of lpf mutants fully restored the wild-type phenotypes. Our results indicate that (i) EHEC might colonize the terminal ileum at the early stages of infection, (ii) Lpf are an important player in the interactions with Peyer’s patches and M-cells, and could contribute to intestinal colonization. PMID:28317910

  7. The Role of Thin Aggregative Fimbriae on Pathogenic Bacterial Transport Through Porous Media

    NASA Astrophysics Data System (ADS)

    Salvucci, A. E.; Fuka, D. R.; Marjerison, R. D.; Hay, A. G.; Zhang, W.; Caballero, L. A.; Zevi, Y.; Richards, B. K.; Steenhuis, T. S.

    2008-05-01

    Pathogenic bacteria, such as Escherichia coli and Salmonella sp., are responsible for many deaths worldwide every year. Their survival in the natural environment is enhanced by the production of biofilms, which provide a resistance to environmental stresses. However, it remains unclear how these biofilms affect the bacterias' ability to move through the soil matrix and potentially contaminate groundwater or water from drainage systems. In this presentation, we discuss the role of thin aggregative fimbriae (curli), a key biofilm component, on transport through porous media. An experiment was performed consisting of 96 sand columns created using a deep-well microtiter plate. We used well-characterized strains of E. coli, one with the ability to form curli and one without. Pulsing the E. coli strains through the sand column, mimicking natural leaching processes, showed less transport, by greater retention, in the strains that produce curli versus those strains that do not. In addition, when cultured in conditions unfavorable to curli production, transport between strains did not differ significantly. These preliminary results indicate that curli, and to a larger extent biofilms, could be an important component influencing the transport of bacterial strains through the soil matrix. This determination of pathogens' ability to move through the environment, as related to how well they form biofilms, will facilitate a better understanding of the fate of pathogenic bacteria in the environment.

  8. Role of pili (fimbriae) in attachment of Bradyrhizobium japonicum to soybean roots

    SciTech Connect

    Vesper, S.J.; Bauer, W.D.

    1986-07-01

    Pili (fimbriae) were observed on cells of each of the five strains of Bradyrhizobium japonicum and the one strain of Rhizobium trifolii examined. Pili on B. japonicum were about 4 nm in diameter and polarly expressed. Piliated cells were estimated by transmission electron microscopy and hydrophobic attachment to polystyrene to constitute only a small percentage of the total population. The proportion of piliated cells in these populations was dependent on culture age in some strains. Piliated B. japonicum cells were selectively and quantitatively removed from suspension when cultures were incubated with either soybean roots or hydrophobic plastic surfaces, indicating that pili were involved in the attachment of the bacteria to these surfaces. Pili from B. japonicum 110 ARS were purified and found to have a subunit molecular weight of approximately 21,000. Treatment of B. japonicum suspensions with antiserum against the isolated pili reduced attachment to soybean roots by about 90% and nodulation by about 80%. Pili appear to be important mediators of attachment of B. japonicum to soybean roots under the conditions examined.

  9. AgfC and AgfE facilitate extracellular thin aggregative fimbriae synthesis in Salmonella enteritidis.

    PubMed

    Gibson, D L; White, A P; Rajotte, C M; Kay, W W

    2007-04-01

    Salmonella thin aggregative fimbriae (Tafi; curli) are important in pathogenesis and biofilm formation; however, less is known of their structure and morphogenesis. In the Salmonella agfBAC Tafi operon, the transcription and role of agfC have been elusive. In this study, agfBAC transcripts were detected using a sensitive reverse transcriptase technique. Native AgfC was not detected using polyclonal antibodies generated against purified hexahistidine-tagged AgfC; however, in trans expression revealed that AgfC was localized to the periplasm as a mature form. An isogenic DeltaagfC mutant displayed an abundance of 20 nm fibres, in addition to native Tafi (5-7 nm), and had an increase in cell surface hydrophobicity. Purified 20 nm fibres were depolymerized under exceptionally stringent conditions to release what proved to be AgfA subunits. This revealed that the 20 nm fibres represented a different form of Tafi. The role of AgfC in Tafi assembly was investigated further using an antibody-capture assay of isogenic Deltaagf mutants. A soluble antibody-accessible form of AgfA was captured in wild-type (wt), DeltaagfB and DeltaagfF strains, in support of the extracellular nucleation-precipitation pathway of Tafi assembly, but not in DeltaagfC or DeltaagfE mutants. This indicates that AgfC and AgfE are important for AgfA extracellular assembly, facilitating the synthesis of Tafi.

  10. Promoting crystallisation of the Salmonella enteritidis fimbriae 14 pilin SefD using deuterium oxide

    SciTech Connect

    Liu, Bing; Garnett, James A.; Lee, Wei-chao; Lin, Jing; Salgado, Paula; Taylor, Jonathan; Xu, Yingqi; Lambert, Sebastian; Cota, Ernesto; Matthews, Steve

    2012-05-04

    Highlights: Black-Right-Pointing-Pointer The benefits of D{sub 2}O in screening for crystallisation was explored. Black-Right-Pointing-Pointer The crystal structures of the SefD pilin in both H{sub 2}O and D{sub 2}O reveal differences. Black-Right-Pointing-Pointer Crystallisation improvements are explained by altered interactions in D{sub 2}O crystals. Black-Right-Pointing-Pointer D{sub 2}O is useful additive in sparse-matrix screening for crystallisation. -- Abstract: The use of heavy water (D{sub 2}O) as a solvent is commonplace in many spectroscopic techniques for the study of biological macromolecules. A significant deuterium isotope effect exists where hydrogen-bonding is important, such as in protein stability, dynamics and assembly. Here we illustrate the use of D{sub 2}O in additive screening for the production of reproducible diffraction-quality crystals for the Salmonella enteritidis fimbriae 14 (SEF14) putative tip adhesin, SefD.

  11. Bordetella filamentous hemagglutinin and fimbriae: critical adhesins with unrealized vaccine potential

    PubMed Central

    Scheller, Erich V.; Cotter, Peggy A.

    2015-01-01

    Pertussis, or whooping cough, is a highly contagious respiratory disease that is caused by the Gram-negative bacterium Bordetella pertussis, which is transmitted exclusively from human to human. While vaccination against B. pertussis has been successful, replacement of the whole cell vaccine with an acellular component vaccine has correlated with reemergence of the disease, especially in adolescents and infants. Based on their presumed importance in mediating adherence to host tissues, filamentous hemagglutinin (FHA) and fimbria (FIM) were selected as components of most acellular pertussis vaccines. In this review, we describe the biogenesis of FHA and FIM, recent data that show that these factors do, in fact, play critical roles in adherence to respiratory epithelium, and evidence that they also contribute to persistence in the lower respiratory tract by modulating the host immune response. We also discuss shortcomings of whole cell and acellular pertussis vaccines and the possibility that FHA and FIM could serve as effective protective antigens in next-generation vaccines. PMID:26416077

  12. Structural insight into host recognition by aggregative adherence fimbriae of enteroaggregative Escherichia coli.

    PubMed

    Berry, Andrea A; Yang, Yi; Pakharukova, Natalia; Garnett, James A; Lee, Wei-chao; Cota, Ernesto; Marchant, Jan; Roy, Saumendra; Tuittila, Minna; Liu, Bing; Inman, Keith G; Ruiz-Perez, Fernando; Mandomando, Inacio; Nataro, James P; Zavialov, Anton V; Matthews, Steve

    2014-09-01

    Enteroaggregative Escherichia coli (EAEC) is a leading cause of acute and persistent diarrhea worldwide. A recently emerged Shiga-toxin-producing strain of EAEC resulted in significant mortality and morbidity due to progressive development of hemolytic-uremic syndrome. The attachment of EAEC to the human intestinal mucosa is mediated by aggregative adherence fimbria (AAF). Using X-ray crystallography and NMR structures, we present new atomic resolution insight into the structure of AAF variant I from the strain that caused the deadly outbreak in Germany in 2011, and AAF variant II from archetype strain 042, and propose a mechanism for AAF-mediated adhesion and biofilm formation. Our work shows that major subunits of AAF assemble into linear polymers by donor strand complementation where a single minor subunit is inserted at the tip of the polymer by accepting the donor strand from the terminal major subunit. Whereas the minor subunits of AAF have a distinct conserved structure, AAF major subunits display large structural differences, affecting the overall pilus architecture. These structures suggest a mechanism for AAF-mediated adhesion and biofilm formation. Binding experiments using wild type and mutant subunits (NMR and SPR) and bacteria (ELISA) revealed that despite the structural differences AAF recognize a common receptor, fibronectin, by employing clusters of basic residues at the junction between subunits in the pilus. We show that AAF-fibronectin attachment is based primarily on electrostatic interactions, a mechanism not reported previously for bacterial adhesion to biotic surfaces.

  13. [Comparative study of bilateral lesions in the entorhinal cortex and in the fimbria fornix].

    PubMed

    Almaguer-Melian, W; Vallejo, A; Ramírez, M; Capdevila, V; Rosillo-Martí, J C; Bergado-Rosado, J A

    Numerous reports show that lesions to hippocampus afferents, such as the entorhinal cortex (EC) and the fimbria fornix (FF), exert an effect on memory in rodents. There are, however, no long term comparative studies that show which of these lesions could be most useful as a model for studies into neuroplasticity. Young male Sprague Dawley rats were used. Bilateral electrolytic lesion was caused to the EC or the FF was damaged by transection. One, four or 12 weeks later the animals were evaluated in a Morris water maze, first with an invisible platform and then with the platform within view. The results from the two groups were compared to each other and to those obtained from healthy controls and subjects with false lesions by means of a variance analysis. In the test with an invisible platform, both types of lesion gave rise to serious, irreparable involvement of the spatial memory of the animals, at least up to 12 weeks after the lesion. The test with the visible platform revealed significant differences between animals with lesion to the EC evaluated at 12 weeks, which suggests the development of some visual or motor deterioration in these animals. Although both lesions gave rise to behavioural deterioration that was irreversible in the long term in rodents, the lesion to the FF seems to be a better model for evaluating specific effects on learning and memory, since the lesion to the EC apparently triggers additional sensory and motor involvement.

  14. Presence of F107, 2134P and Av24 fimbriae on strains of Escherichia coli isolated from Swedish piglets with diarrhoea.

    PubMed

    Kennan, R; Söderlind, O; Conway, P

    1995-02-01

    A total of 109 Escherichia coli isolates from piglets with diarrhoea, that had previously been shown to be enterotoxin producers, but negative for the adhesive fimbriae K88, K99, 987P and F41 were tested for the presence of more recently characterised fimbriae. Testing was done by immunodot assay with absorbed polyclonal antisera against Av24 and F107 fimbriae, and unabsorbed polyclonal antiserum and monoclonal antiserum against 2134P fimbriae. Strains were also tested by polymerase chain reaction for the presence of genes encoding the major subunit of F107 fimbriae. After elimination of possible non-specific reactions, antisera testing produced 10 strains positive with all 4 antisera, 1 strain that reacted with all antisera except F107, 2 strains that reacted with all antisera except the 2134P monoclonal, 3 strains that reacted with 2134P polyclonal and F107 and 2 that reacted with F107 only. The PCR testing confirmed the results of the antisera, but also produced an additional 14 positive strains, giving a total of 30% of the strains tested reacting positively by PCR. Furthermore, all 33 isolates positive by PCR came from pigs that were older than 1 week, which is 45% of the 72 isolates tested which came from older pigs.

  15. Aggregative adherence fimbriae I (AAF/I) mediate colonization of fresh produce and abiotic surface by Shiga toxigenic enteroaggregative Escherichia coli O104:H4.

    PubMed

    Nagy, Attila; Xu, Yunfeng; Bauchan, Gary R; Shelton, Daniel R; Nou, Xiangwu

    2016-07-16

    The Shiga toxigenic Escherichia coli O104:H4 isolated during the 2011 European outbreak expresses Shiga toxin 2a and possess virulence genes associated with the enteroaggregative E. coli (EAEC) pathotype. It produces plasmid encoded aggregative adherence fimbriae I (AAF/I) which mediate cell aggregation and biofilm formation in human intestine and promote Shiga-toxin adsorption, but it is not clear whether the AAF/I fimbriae are involved in the colonization and biofilm formation on food and environmental matrices such as the surface of fresh produce. We deleted the gene encoding for the AAF/I fimbriae main subunit (AggA) from an outbreak associated E. coli O104:H4 strain, and evaluated the role of AAF/I fimbriae in the adherence and colonization of E. coli O104:H4 to spinach and abiotic surfaces. The deletion of aggA did not affect the adherence of E. coli O104:H4 to these surfaces. However, it severely diminished the colonization and biofilm formation of E. coli O104:H4 on these surfaces. Strong aggregation and biofilm formation on spinach and abiotic surfaces were observed with the wild type strain but not the isogenic aggA deletion mutant, suggesting that AAF/I fimbriae play a crucial role in persistence of O104:H4 cells outside of the intestines of host species, such as on the surface of fresh produce.

  16. More than one way to control hair growth: regulatory mechanisms in enterobacteria that affect fimbriae assembled by the chaperone/usher pathway.

    PubMed

    Clegg, Steven; Wilson, Janet; Johnson, Jeremiah

    2011-05-01

    Many gram-negative enterobacteria produce surface-associated fimbriae that facilitate attachment and adherence to eucaryotic cells and tissues. These organelles are believed to play an important role during infection by enabling bacteria to colonize specific niches within their hosts. One class of these fimbriae is assembled using a periplasmic chaperone and membrane-associated scaffolding protein that has been referred to as an usher because of its function in fimbrial biogenesis. The presence of multiple types of fimbriae assembled by the chaperone/usher pathway can be found both within a single bacterial species and also among different genera. One way of controlling fimbrial assembly in these bacteria is at the genetic level by positively or negatively regulating fimbrial gene expression. This minireview considers the mechanisms that have been described to control fimbrial gene expression and uses specific examples to demonstrate both unique and shared properties of such regulatory mechanisms.

  17. More than One Way To Control Hair Growth: Regulatory Mechanisms in Enterobacteria That Affect Fimbriae Assembled by the Chaperone/Usher Pathway▿

    PubMed Central

    Clegg, Steven; Wilson, Janet; Johnson, Jeremiah

    2011-01-01

    Many Gram-negative enterobacteria produce surface-associated fimbriae that facilitate attachment and adherence to eucaryotic cells and tissues. These organelles are believed to play an important role during infection by enabling bacteria to colonize specific niches within their hosts. One class of these fimbriae is assembled using a periplasmic chaperone and membrane-associated scaffolding protein that has been referred to as an usher because of its function in fimbrial biogenesis. The presence of multiple types of fimbriae assembled by the chaperone/usher pathway can be found both within a single bacterial species and also among different genera. One way of controlling fimbrial assembly in these bacteria is at the genetic level by positively or negatively regulating fimbrial gene expression. This minireview considers the mechanisms that have been described to control fimbrial gene expression and uses specific examples to demonstrate both unique and shared properties of such regulatory mechanisms. PMID:21398554

  18. Inner-membrane protein MorC is involved in fimbriae production and biofilm formation in Aggregatibacter actinomycetemcomitans.

    PubMed

    Smith, Kenneth P; Ruiz, Teresa; Mintz, Keith P

    2016-03-01

    Fimbrial subunit synthesis, secretion and assembly on the surface of the periodontal pathogen Aggregatibacter actinomycetemcomitans are essential for biofilm formation. A recent quantitative proteomics study employing an afimbriated strain and a developed mutant isogenic for the inner-membrane protein morphogenesis protein C (MorC) revealed that the abundance of the proteins of the fimbrial secretion apparatus in the membrane is dependent on MorC. To investigate further the relationship between MorC and fimbriation, we identified and complemented the defect in fimbriae production in the afimbriated laboratory strain. The transformed strain expressing a plasmid containing genes encoding the WT fimbrial subunit and the prepilin peptidase displayed all of the hallmarks of a fimbriated bacterium including the distinct star-like colony morphology, robust biofilm formation, biofilm architecture composed of discrete microcolonies and the presence of fimbriae. When the identical plasmid was transformed into a morC mutant strain, the bacterium did not display any of the phenotypes of fimbriated strains. Extension of these studies to a naturally fimbriated clinical strain showed that the resulting morC mutant maintained the characteristic colony morphology of fimbriated strains. There was, however, a reduction in the secretion of fimbrial subunits, and fewer fimbriae were observed on the surface of the mutant strain. Furthermore, the morC mutant of the fimbriated strain displayed a significantly altered biofilm microcolony architecture, while maintaining a similar biofilm mass to the parent strain. These results suggest that MorC influences fimbrial secretion and microcolony formation in A. actinomycetemcomitans.

  19. Diversification of the Salmonella Fimbriae: A Model of Macro- and Microevolution

    PubMed Central

    Yue, Min; Rankin, Shelley C.; Blanchet, Ryan T.; Nulton, James D.; Edwards, Robert A.; Schifferli, Dieter M.

    2012-01-01

    Bacteria of the genus Salmonella comprise a large and evolutionary related population of zoonotic pathogens that can infect mammals, including humans and domestic animals, birds, reptiles and amphibians. Salmonella carries a plethora of virulence genes, including fimbrial adhesins, some of them known to participate in mammalian or avian host colonization. Each type of fimbria has its structural subunit and biogenesis genes encoded by one fimbrial gene cluster (FGC). The accumulation of new genomic information offered a timely opportunity to better evaluate the number and types of FGCs in the Salmonella pangenome, to test the use of current classifications based on phylogeny, and to infer potential correlations between FGC evolution in various Salmonella serovars and host niches. This study focused on the FGCs of the currently deciphered 90 genomes and 60 plasmids of Salmonella. The analysis highlighted a fimbriome consisting of 35 different FGCs, of which 16 were new, each strain carrying between 5 and 14 FGCs. The Salmonella fimbriome was extremely diverse with FGC representatives in 8 out of 9 previously categorized fimbrial clades and subclades. Phylogenetic analysis of Salmonella suggested macroevolutionary shifts detectable by extensive FGC deletion and acquisition. In addition, microevolutionary drifts were best depicted by the high level of allelic variation in predicted or known adhesins, such as the type 1 fimbrial adhesin FimH for which 67 different natural alleles were identified in S. enterica subsp. I. Together with strain-specific collections of FGCs, allelic variation among adhesins attested to the pathoadaptive evolution of Salmonella towards specific hosts and tissues, potentially modulating host range, strain virulence, disease progression, and transmission efficiency. Further understanding of how each Salmonella strain utilizes its panel of FGCs and specific adhesin alleles for survival and infection will support the development of new approaches

  20. Porphyromonas gingivalis fimbria-dependent activation of inflammatory genes in human aortic endothelial cells.

    PubMed

    Chou, Hsin-Hua; Yumoto, Hiromichi; Davey, Michael; Takahashi, Yusuke; Miyamoto, Takanari; Gibson, Frank C; Genco, Caroline A

    2005-09-01

    Epidemiological and pathological studies have suggested that infection with the oral pathogen Porphyromonas gingivalis can potentiate atherosclerosis and human coronary heart disease. Furthermore, infection with invasive, but not noninvasive P. gingivalis has been demonstrated to accelerate atherosclerosis in apolipoprotein E-deficient (ApoE(-/-)) mice and to accelerate local inflammatory responses in aortic tissue. In the present study, using high-density oligonucleotide microarrays, we have defined the gene expression profile of human aortic endothelial cells (HAEC) after infection with invasive and noninvasive P. gingivalis. After infection of HAEC with invasive P. gingivalis strain 381, we observed the upregulation of 68 genes. Genes coding for the cytokines Gro2 and Gro3; the adhesion molecules intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule (VCAM)-1, and ELAM-1 (E-selectin); the chemokine interleukin-8 (IL-8); and the proinflammatory molecules IL-6 and cyclooxygenase-2 were among the most highly upregulated genes in P. gingivalis 381-infected HAEC compared to uninfected HAEC control. Increased mRNA levels for signaling molecules, transcriptional regulators, and cell surface receptors were also observed. Of note, only 4 of these 68 genes were also upregulated in HAEC infected with the noninvasive P. gingivalis fimA mutant. Reverse transcription-PCR, enzyme-linked immunosorbent assay, and fluorescence-activated cell sorting analysis confirmed the expression of ICAM-1, VCAM-1, E-/P-selectins, IL-6, and IL-8 in HAEC infected with invasive P. gingivalis. We also demonstrated that increased expression of ICAM-1 and VCAM-1 in aortic tissue of ApoE(-/-) mice orally challenged with invasive P. gingivalis but not with the noninvasive P. gingivalis fimA mutant by immunohistochemical analysis. Taken together, these results demonstrate that P. gingivalis fimbria-mediated invasion upregulates inflammatory gene expression in HAEC and in aortic

  1. Gonadal sex differentiation and effects of dietary methyltestosterone treatment in sablefish (Anoplopoma fimbria).

    PubMed

    Luckenbach, J Adam; Fairgrieve, William T

    2016-02-01

    Methods for sex control are needed to establish monosex aquaculture of sablefish (Anoplopoma fimbria). Here we conducted the first characterization of sex differentiation by histology and hormonal sex reversal experiment in sablefish. Ovarian differentiation was first discernible at ~80 mm fork length (FL) and characterized by development of lamellar structures and onset of meiosis. Testes exhibited a dual-lobe appearance over much of their length and remained non-meiotic until males were ≥520 mm FL (2 years post-fertilization). Juveniles with undifferentiated gonads were provided diets containing 0 (control), 5 or 50 mg 17α-methyltestosterone (MT)/kg for 2 months. Following treatment, controls possessed either ovaries or non-meiotic testes, whereas MT-treated fish exhibited meiotic testes (60% of the fish), intersex gonads (~30%), or gonads that appeared sterile (~10%). A genetic sex marker revealed that all intersex fish were genetic females, although other females appeared to be completely sex reversed (i.e., neomales). One year after treatment, MT-treated fish possessed non-meiotic testes similar to control males or intersex gonads with reduced ovarian features, presumably due to atresia following MT withdrawal. Milt collected from neomales and genetic males 3 years post-treatment permitted sperm motility analyses; however, neomale sperm were virtually immotile. These results demonstrated that sablefish are differentiated gonochorists and that MT treatment from 76 to 196 mm FL induced permanent masculinization of a portion of the genetic females, but acquisition of sperm motility was impaired. Earlier administration of MT may be necessary to sex reverse a higher proportion of genetic females and reduce negative effects on fertility.

  2. Development of approaches to induce puberty in cultured female sablefish (Anoplopoma fimbria).

    PubMed

    Guzmán, José M; Luckenbach, J Adam; da Silva, Denis A M; Ylitalo, Gina M; Swanson, Penny

    2015-09-15

    Efforts to establish sustainable and efficient aquaculture production of sablefish (Anoplopoma fimbria) have been constrained by delayed puberty in cultured females. This study integrates a series of experiments aimed at gaining an understanding of the reproductive physiology of puberty in female sablefish. We detected transcripts for the dopamine D2 receptor (drd2) in brain, pituitary and ovary of sablefish, and prepubertal females exhibited significantly elevated brain and pituitary drd2 expression relative to wild maturing females. Treatments with sustained-release cholesterol pellets containing testosterone (T) and the dopamine D2 receptor antagonist, metoclopramide (Met), stimulated expression of pituitary luteinizing hormone beta subunit (lhb) and follicle-stimulating hormone beta subunit (fshb), respectively, in prepubertal females, whereas a combination of T and gonadotropin-releasing hormone agonist (GnRHa) had a strong synergistic effect on lhb expression (2000-fold higher than control). Although T induced a significant increase in the maximum ovarian follicle volume, none of the treatments tested stimulated onset of vitellogenesis. Using liquid chromatography/tandem mass spectrometry, we demonstrated that Met stimulated production of T by previtellogenic ovarian follicles in vitro, whereas gonadotropin preparations enhanced 17α-hydroxyprogesterone, androstenedione (A4), T and 17β-estradiol (E2) production. Treatment with T increased production of A4, 11β-hydroxyandrostenedione, 11β-hydroxytestosterone, E2, 11-ketotestosterone, and 5α-dihydrotestosterone (DHT). Interestingly, in the presence of high doses of T the previtellogenic ovary preferentially produced A4 and DHT over any other metabolite. Our data suggest the existence of dopamine inhibition of the reproductive axis in female sablefish. Treatments with Met and T elevated gonadotropin mRNAs in prepubertal females but failed to stimulate the transition into vitellogenic growth, suggesting a

  3. Extracellular polysaccharides associated with thin aggregative fimbriae of Salmonella enterica serovar enteritidis.

    PubMed

    White, A P; Gibson, D L; Collinson, S K; Banser, P A; Kay, W W

    2003-09-01

    Lipopolysaccharide (LPS) O polysaccharide was identified as the principle factor impeding intercellular formation of intact thin aggregative fimbriae (Tafi) in Salmonella enterica serovar Enteritidis. The extracellular nucleation-precipitation assembly pathway for these organelles was investigated by quantifying fimbrial formation between deltaagfA (AgfA recipient) and deltaagfB (AgfA donor) cells harboring mutations in LPS (galE::Tn10) and/or cellulose (deltabcsA) synthesis. Intercellular complementation could be detected between deltaagfA and deltaagfB strains only when both possessed the galE mutation. LPS O polysaccharide appears to be an impenetrable barrier to AgfA assembly between cells but not within individual cells. The presence of cellulose did not restrict Tafi formation between cells. Transmission electron microscopy of w+ S. enterica serovar Enteritidis 3b cells revealed diffuse Tafi networks without discernible fine structure. In the absence of cellulose, however, individual Tafi fibers were clearly visible, appeared to be occasionally branched, and showed the generally distinctive appearance described for Escherichia coli K-12 curli. A third extracellular matrix component closely associated with cellulose and Tafi was detected on Western blots by using immune serum raised to whole, purified Tafi aggregates. Cellulose was required to tightly link this material to cells. Antigenically similar material was also detected in S. enterica serovar Typhimurium and one diarrheagenic E. coli isolate. Preliminary analysis indicated that this material represented an anionic, extracellular polysaccharide that was distinct from colanic acid. Therefore, Tafi in their native state appear to exist as a complex with cellulose and at least one other component.

  4. Diversification of the Salmonella fimbriae: a model of macro- and microevolution.

    PubMed

    Yue, Min; Rankin, Shelley C; Blanchet, Ryan T; Nulton, James D; Edwards, Robert A; Schifferli, Dieter M

    2012-01-01

    Bacteria of the genus Salmonella comprise a large and evolutionary related population of zoonotic pathogens that can infect mammals, including humans and domestic animals, birds, reptiles and amphibians. Salmonella carries a plethora of virulence genes, including fimbrial adhesins, some of them known to participate in mammalian or avian host colonization. Each type of fimbria has its structural subunit and biogenesis genes encoded by one fimbrial gene cluster (FGC). The accumulation of new genomic information offered a timely opportunity to better evaluate the number and types of FGCs in the Salmonella pangenome, to test the use of current classifications based on phylogeny, and to infer potential correlations between FGC evolution in various Salmonella serovars and host niches. This study focused on the FGCs of the currently deciphered 90 genomes and 60 plasmids of Salmonella. The analysis highlighted a fimbriome consisting of 35 different FGCs, of which 16 were new, each strain carrying between 5 and 14 FGCs. The Salmonella fimbriome was extremely diverse with FGC representatives in 8 out of 9 previously categorized fimbrial clades and subclades. Phylogenetic analysis of Salmonella suggested macroevolutionary shifts detectable by extensive FGC deletion and acquisition. In addition, microevolutionary drifts were best depicted by the high level of allelic variation in predicted or known adhesins, such as the type 1 fimbrial adhesin FimH for which 67 different natural alleles were identified in S. enterica subsp. I. Together with strain-specific collections of FGCs, allelic variation among adhesins attested to the pathoadaptive evolution of Salmonella towards specific hosts and tissues, potentially modulating host range, strain virulence, disease progression, and transmission efficiency. Further understanding of how each Salmonella strain utilizes its panel of FGCs and specific adhesin alleles for survival and infection will support the development of new approaches

  5. Purification and characterization of the fimbria F18ac (2134P) isolated from enterotoxigenic Escherichia coli (ETEC).

    PubMed

    Amorim, C R; Matsuura, M S; Rosa, J C; Greene, L J; Leite, D S; Yano, T

    2000-09-15

    The adhesin F18ac purified on Sepharose CL 4B column chromatography and SDS-PAGE stained with Coomassie Blue and Western blotting using specific anti-F18ac serum presented one band of approximately 17kDa. Gold immunolabeling revealed that the adhesin F18ac has a fimbrial structure on the bacterial surface. The first 27 amino acid residues of the N-terminal portion of the adhesin F18ac, showed 92.5% homology (25 amino acids) with the F107 (F18ab) fimbriae.

  6. Type 1 Fimbriae, a Colonization Factor of Uropathogenic Escherichia coli, Are Controlled by the Metabolic Sensor CRP-cAMP

    PubMed Central

    Müller, Claudia M.; Åberg, Anna; Straseviçiene, Jurate; Emődy, Levente; Uhlin, Bernt Eric; Balsalobre, Carlos

    2009-01-01

    Type 1 fimbriae are a crucial factor for the virulence of uropathogenic Escherichia coli during the first steps of infection by mediating adhesion to epithelial cells. They are also required for the consequent colonization of the tissues and for invasion of the uroepithelium. Here, we studied the role of the specialized signal transduction system CRP-cAMP in the regulation of type 1 fimbriation. Although initially discovered by regulating carbohydrate metabolism, the CRP-cAMP complex controls a major regulatory network in Gram-negative bacteria, including a broad subset of genes spread into different functional categories of the cell. Our results indicate that CRP-cAMP plays a dual role in type 1 fimbriation, affecting both the phase variation process and fimA promoter activity, with an overall repressive outcome on fimbriation. The dissection of the regulatory pathway let us conclude that CRP-cAMP negatively affects FimB-mediated recombination by an indirect mechanism that requires DNA gyrase activity. Moreover, the underlying studies revealed that CRP-cAMP controls the expression of another global regulator in Gram-negative bacteria, the leucine-responsive protein Lrp. CRP-cAMP-mediated repression is limiting the switch from the non-fimbriated to the fimbriated state. Consistently, a drop in the intracellular concentration of cAMP due to altered physiological conditions (e.g. growth in presence of glucose) increases the percentage of fimbriated cells in the bacterial population. We also provide evidence that the repression of type 1 fimbriae by CRP-cAMP occurs during fast growth conditions (logarithmic phase) and is alleviated during slow growth (stationary phase), which is consistent with an involvement of type 1 fimbriae in the adaptation to stress conditions by promoting biofilm growth or entry into host cells. Our work suggests that the metabolic sensor CRP-cAMP plays a role in coupling the expression of type 1 fimbriae to environmental conditions, thereby

  7. Structural insight in the inhibition of adherence of F4 fimbriae producing enterotoxigenic Escherichia coli by llama single domain antibodies.

    PubMed

    Moonens, Kristof; Van den Broeck, Imke; Okello, Emmanuel; Pardon, Els; De Kerpel, Maia; Remaut, Han; De Greve, Henri

    2015-02-24

    Enterotoxigenic Escherichia coli that cause neonatal and post-weaning diarrhea in piglets express F4 fimbriae to mediate attachment towards host receptors. Recently we described how llama single domain antibodies (VHHs) fused to IgA, produced in Arabidopsis thaliana seeds and fed to piglets resulted in a progressive decline in shedding of F4 positive ETEC bacteria. Here we present the structures of these inhibiting VHHs in complex with the major adhesive subunit FaeG. A conserved surface, distant from the lactose binding pocket, is targeted by these VHHs, highlighting the possibility of targeting epitopes on single-domain adhesins that are non-involved in receptor binding.

  8. Structural basis for Myf and Psa fimbriae-mediated tropism of pathogenic strains of Yersinia for host tissues.

    PubMed

    Pakharukova, Natalia; Roy, Saumendra; Tuittila, Minna; Rahman, Mohammad M; Paavilainen, Sari; Ingars, Anna-Karin; Skaldin, Maksym; Lamminmäki, Urpo; Härd, Torleif; Teneberg, Susann; Zavialov, Anton V

    2016-11-01

    Three pathogenic species of the genus Yersinia assemble adhesive fimbriae via the FGL-chaperone/usher pathway. Closely related Y. pestis and Y. pseudotuberculosis elaborate the pH6 antigen (Psa), which mediates bacterial attachment to alveolar cells of the lung. Y. enterocolitica, instead, assembles the homologous fimbriae Myf of unknown function. Here, we discovered that Myf, like Psa, specifically recognizes β1-3- or β1-4-linked galactose in glycosphingolipids, but completely lacks affinity for phosphatidylcholine, the main receptor for Psa in alveolar cells. The crystal structure of a subunit of Psa (PsaA) complexed with choline together with mutagenesis experiments revealed that PsaA has four phosphatidylcholine binding pockets that enable super-high-avidity binding of Psa-fibres to cell membranes. The pockets are arranged as six tyrosine residues, which are all missing in the MyfA subunit of Myf. Conversely, the crystal structure of the MyfA-galactose complex revealed that the galactose-binding site is more extended in MyfA, enabling tighter binding to lactosyl moieties. Our results suggest that during evolution, Psa has acquired a tyrosine-rich surface that enables it to bind to phosphatidylcholine and mediate adhesion of Y. pestis/pseudotuberculosis to alveolar cells, whereas Myf has specialized as a carbohydrate-binding adhesin, facilitating the attachment of Y. enterocolitica to intestinal cells. © 2016 John Wiley & Sons Ltd.

  9. Organization of Biogenesis Genes for Aggregative Adherence Fimbria II Defines a Virulence Gene Cluster in Enteroaggregative Escherichia coli

    PubMed Central

    Elias, Waldir P.; Czeczulin, John R.; Henderson, Ian R.; Trabulsi, Luiz R.; Nataro, James P.

    1999-01-01

    Several virulence-related genes have been described for prototype enteroaggregative Escherichia coli (EAEC) strain 042, which has been shown to cause diarrhea in human volunteers. Among these factors are the enterotoxins Pet and EAST and the fimbrial antigen aggregative adherence fimbria II (AAF/II), all of which are encoded on the 65-MDa virulence plasmid pAA2. Using nucleotide sequence analysis and insertional mutagenesis, we have found that the genes required for the expression of each of these factors, as well as the transcriptional activator of fimbrial expression AggR, map to a distinct cluster on the pAA2 plasmid map. The cluster is 23 kb in length and includes two regions required for expression of the AAF/II fimbria. These fimbrial biogenesis genes feature a unique organization in which the chaperone, subunit, and transcriptional activator lie in one cluster, whereas the second, unlinked cluster comprises a silent chaperone gene, usher, and invasin reminiscent of Dr family fimbrial clusters. This plasmid-borne virulence locus may represent an important set of virulence determinants in EAEC strains. PMID:10074069

  10. Environmental regulation of the long polar fimbriae 2 of enterohemorrhagic Escherichia coli O157:H7

    PubMed Central

    Arenas-Hernández, Margarita M.P.; Rojas-López, Maricarmen; Medrano-López, Abraham; Nuñez-Reza, Karen J.; Puente, José Luis; Martínez-Laguna, Ygnacio; Torres, Alfredo G.

    2014-01-01

    The molecular mechanisms controlling expression of the Long Polar Fimbriae 2 (Lpf2) of enterohemorrhagic E. coli (EHEC) O157:H7 were evaluated. Primer extension was used to locate the lpfA2 transcriptional start site in EHEC strain EDL933 at 171 bp upstream of the lpfA2 start codon. Semi-quantitative RT-PCR demonstrated that the highest lpfA2 expression occurs between an OD600 of 1.0 and 1.2 in DMEM at pH 6.5 and 37°C. The level of lpfA2 transcription at OD600 1.2 and pH 6.5 was 4-times greater than that at pH 7.2. Although lpfA2 expression was decreased under iron-depleted conditions, its expression was increased in a Ferric-uptake-regulator (Fur) mutant strain. The lpfA2 transcript was 0.7 and 2-times more abundant in wt EHEC grown in DMEM pH 6.5 plus iron and MacConkey broth at 25°C, respectively, than in DMEM at pH 6.5. The lpf2 expression in DMEM pH 6.5 plus iron and bile salts was 2.7-times more abundant and similar to MacConkey. Further, transcription in the EDL933Δfur was 0.6 and 0.8-times higher as compared to the wt strain grown in DMEM pH 6.5 plus iron and MacConkey broth, respectively. Electrophoretic mobility shift assays (EMSA) showed that purified Fur interacts with the lpf2 regulatory region, indicating that Fur-repression is exerted by direct binding to the promoter region. In summary, we demonstrated that the EHEC lpf2 operon is regulated in response to temperature, pH, bile salts and iron, during exponential phase of growth, and controlled by Fur. PMID:24966050

  11. Environmental regulation of the long polar fimbriae 2 of enterohemorrhagic Escherichia coli O157:H7.

    PubMed

    Arenas-Hernández, Margarita M; Rojas-López, Maricarmen; Medrano-López, Abraham; Nuñez-Reza, Karen J; Puente, José Luis; Martínez-Laguna, Ygnacio; Torres, Alfredo G

    2014-08-01

    The molecular mechanisms controlling expression of the long polar fimbriae 2 (Lpf2) of enterohemorrhagic Escherichia coli (EHEC) O157:H7 were evaluated. Primer extension was used to locate the lpfA2 transcriptional start site in EHEC strain EDL933 at 171 bp upstream of the lpfA2 start codon. Semi-quantitative RT-PCR demonstrated that the highest lpfA2 expression occurs between an OD600 of 1.0 and 1.2 in DMEM at pH 6.5 and 37 °C. The level of lpfA2 transcription at OD600 1.2 and pH 6.5 was four times greater than that at pH 7.2. Although lpfA2 expression was decreased under iron-depleted conditions, its expression was increased in a ferric-uptake-regulator (Fur) mutant strain. The lpfA2 transcript was 0.7 and 2 times more abundant in wt EHEC grown in DMEM pH 6.5 plus iron and MacConkey broth at 25 °C, respectively, than in DMEM at pH 6.5. The lpf2 expression in DMEM pH 6.5 plus iron and bile salts was 2.7 times more abundant than baseline conditions. Further, transcription in the EDL933∆fur was 0.6 and 0.8 times higher as compared with the wt strain grown in DMEM pH 6.5 plus iron and MacConkey broth, respectively. Electrophoretic mobility shift assays showed that purified Fur interacts with the lpf2 regulatory region, indicating that Fur repression is exerted by direct binding to the promoter region. In summary, we demonstrated that the EHEC lpf2 operon is regulated in response to temperature, pH, bile salts and iron, during the exponential phase of growth, and is controlled by Fur.

  12. The prevalence of F107 fimbriae and their association with Shiga-like toxin II in Escherichia coli strains from weaned Australian pigs.

    PubMed

    Hide, E J; Connaughton, I D; Driesen, S J; Hasse, D; Monckton, R P; Sammons, N G

    1995-12-01

    A total of 480 haemolytic Escherichia coli (HEC) strains from weaned pigs were tested using an oligonucleotide probe to determine the prevalence of F107 fimbriae in Australia. Of these, 62% were positive. F107 was detected in serogroups O141, O138, O8, O45, O139, O157 and O98 but not in O149 nor O147. 81% of E. coli strains not producing other fimbriae (K88, 987P, K99 or F41) were positive for F107; 5% of strains with K88 fimbriae also had F107. Serological investigation of the expression of F107 fimbriae indicated that serogroups O141, O138, O8, O45 and O157 produced variant F107ac. Variant F107ab was found on O139 strains only. The F107 fimbrial subunits of both variants had a molecular weight of approximately 16 kDa. A total of 350 of the HEC strains were tested to determine the prevalence of the Shiga-like toxin II (SLT II) gene. 29.0% of these strains were positive. SLT II was detected in serogroups O141, O138, O149, O98, O45, O8 and O157 but not in O139. 25% of these strains were positive for both F107 and SLT II.

  13. The Bordetella avium BAV1965-1962 fimbrial locus is regulated by temperature and produces fimbriae involved in adherence to turkey tracheal tissue.

    PubMed

    Loker, Stewart B; Temple, Louise M; Preston, Andrew

    2011-06-01

    Bordetella pertussis, Bordetella parapertussis, and Bordetella bronchiseptica cause respiratory tract disease in mammals, whereas Bordetella avium causes respiratory tract disease in avian hosts. While there are striking similarities between the diseases caused by the mammalian- and avian-adapted bordetellae, differences at the genetic level may account for their different host tropisms. Bacterial pathogens utilize the chaperone-usher pathway to assemble extracellular multisubunit structures (fimbriae) that play a role in virulence. Fimbriae of the mammalian bordetellae mediate attachment to the host respiratory epithelium. They are assembled by a single chaperone/usher system encoded by the fimbrial biogenesis operon fimA-D. B. avium contains a homologous fimbrial operon (BAV1965-1962), and we report here the functionality of this locus. Reverse transcription (RT)-PCR and quantitative PCR analyses demonstrated that transcription of the locus is regulated by temperature. By immuno-transmission electron microscopy (TEM), BAV1965-containing fimbriae were observed on bacteria grown at 37°C but not those grown at 22°C. A mutant in which BAV1965-1962 was deleted displayed significantly lower levels of adherence to turkey tracheal rings than the wild type. Thus, the BAV1965-1962 fimbrial locus is functional, its expression is regulated in response to temperature, and it produces fimbriae involved in adherence to host respiratory tract tissue.

  14. Aggregative adherence fimbriae I (AAF/I) mediate colonization of fresh produce and abiotic surface by Shiga toxigenic enteroaggregative Escherichia coli O104:H4

    USDA-ARS?s Scientific Manuscript database

    The Shiga toxigenic Escherichia coli O104:H4 bares the characteristics of both enterohemorrhagic (EHEC) and enteroaggregative (EAEC) E. coli. It produces plasmid encoded aggregative adherence fimbriae I (AAF/I) which mediate cell aggregation and biofilm formation in human intestine and promote Shiga...

  15. Development and Efficacy Assessment of an Enteric Coated Porous Tablet Loaded With F4 Fimbriae for Oral Vaccination of Piglets against F4+ Escherichia coli Infections.

    PubMed

    Srivastava, Atul; Gowda, D V; Madhunapantula, SubbaRao V; Siddaramaiah

    2016-01-01

    Enterotoxigenic Escherichia coli (ETEC) infection is one of the major causes contributing to the development of diarrhoea and mortality in new born, suckling and newly weaned piglets. To date, no preventive/treatment strategy showed promising results, which could be due to the lack of potent vaccines, and/or due to the development of resistance of ETEC to antibiotics. Therefore, in the present investigation, a novel porous sodium alginate (SA) tablet formulation loaded with F4 fimbriae antigen was developed and tested for efficacy against ETEC infections in piglet models. Precompression parameters of the powder mixes and post compression parameters of tablets have been evaluated and results were found to be satisfactory. Loading of F4 fimbrial antigens into the tablets was achieved by inducing pores in the tablets via the sublimation of camphor followed by incubation with purified F4 fimbriae. The loaded tablets have been coated with Eudragit L100 to protect the F4 fimbriae from (a) highly acidic gastric environment; (b) proteolytic cleavage by pepsin; and (c) to promote subsequent release in the intestine. Evaluation of developed F4 fimbrial tablets in a Pig model demonstrated induction of mucosal immunity, and a significant reduction of F4+ E. coli in faeces. Therefore, F4 fimbriae loaded porous tablets could be a novel oral vaccination candidate to induce mucosal and systemic immunity against ETEC infections.

  16. Both flagella and F4 fimbriae from F4ac+ enterotoxigenic Escherichia coli contribute to attachment to IPEC-J2 cells in vitro.

    PubMed

    Zhou, Mingxu; Duan, Qiangde; Zhu, Xiaofang; Guo, Zhiyan; Li, Yinchau; Hardwidge, Philip R; Zhu, Guoqiang

    2013-05-13

    The role of flagella in the pathogenesis of F4ac+ Enterotoxigenic Escherichia coli (ETEC) mediated neonatal and post-weaning diarrhea (PWD) is not currently understood. We targeted the reference C83902 ETEC strain (O8:H19:F4ac+ LT+ STa+ STb+), to construct isogenic mutants in the fliC (encoding the major flagellin protein), motA (encoding the flagella motor), and faeG (encoding the major subunit of F4 fimbriae) genes. Both the ΔfliC and ΔfaeG mutants had a reduced ability to adhere to porcine intestinal epithelial IPEC-J2 cells. F4 fimbriae expression was significantly down-regulated after deleting fliC, which revealed that co-regulation exists between flagella and F4 fimbriae. However, there was no difference in adhesion between the ΔmotA mutant and its parent strain. These data demonstrate that both flagella and F4 fimbriae are required for efficient F4ac+ ETEC adhesion in vitro.

  17. Live attenuated Salmonella displaying HIV-1 10E8 epitope on fimbriae: systemic and mucosal immune responses in BALB/c mice by mucosal administration

    PubMed Central

    Li, Qing-Hai; Jin, Gang; Wang, Jia-Ye; Li, Hai-Ning; Liu, Huidi; Chang, Xiao-Yun; Wang, Fu-Xiang; Liu, Shu-Lin

    2016-01-01

    The HIV-1 membrane proximal external region (MPER) that is targeted by several broadly neutralizing antibodies (BNAbs) has been considered a potential immunogen for vaccine development. However, to date the immunogenicity of these BNAb epitopes has not been made sufficiently adequate. In the present work, we used live attenuated Salmonella as a platform to present the HIV-1 MPER 10E8 epitope in the fimbriae. The insertion of the 10E8 epitope into the fimbriae had no significant influence on the expression and the absorption capacity of bacterial fimbriae, nor on the virulence and invasiveness of the attenuated Salmonella. After oral administration of the vaccine construct to mice followed by 10E8 epitope peptide boost, specific antibody responses in serum and mucosa as well as memory lymphocytes in spleen and plasma cells in bone marrow were induced. We also found that the live attenuated Salmonella vector directed the immunity toward Th1 bias, induced Th1 and Th2 cytokine responses and stimulated significant B cell differentiation into GC B, memory B and plasma cells. Therefore, we propose that the live attenuated Salmonella constitutively expressing HIV-1 BNAb epitopes on the fimbriae will be an effective approach to improving immune microenvironment and enhancing the immunogenicity of HIV-1 epitope vaccines. PMID:27411313

  18. Preliminary X-ray diffraction analysis of CfaA, a molecular chaperone essential for the assembly of CFA/I fimbriae of human enterotoxigenic Escherichia coli

    SciTech Connect

    Bao, Rui; Esser, Lothar; Poole, Steven; McVeigh, Annette; Chen, Yu-xing; Savarino, Stephen J.; Xia, Di

    2014-01-21

    The molecular chaperone CfaA plays a critical role in the bioassembly of the surface-adhesive CFA/I fimbriae of enterotoxigenic E. coli. Purified CfaA was crystallized and the phase solution was determined by the multiple isomorphous replacement coupled with anomalous scattering method.

  19. Altered regulation of the Diguanylate Cyclase YaiC reduces production of Type 1 Fimbriae in a Pst Mutant of Uropathogenic E. coli CFT073.

    PubMed

    Crépin, Sébastien; Porcheron, Gaëlle; Houle, Sébastien; Harel, Josée; Dozois, Charles M

    2017-09-18

    The pst gene cluster encodes the phosphate specific transport system (Pst). Inactivation of the Pst system constitutively activates the two-component regulatory system PhoBR and attenuates virulence of pathogenic bacteria. In uropathogenic E. coli strain CFT073, attenuation by inactivation of pst is predominantly attributed to the decreased expression of type 1 fimbriae. However, the molecular mechanisms connecting the Pst system and type 1 fimbriae are unknown. To address this, a transposon library was constructed in the pst mutant, and clones were tested for a regain in type 1 fimbriae production. Among them, the diguanylate cyclase encoded by yaiC (adrA in Salmonella) was identified to connect the Pst system and type 1 fimbrial expression. In the pst mutant, the decreased expression of type 1 fimbriae is connected by the induction of yaiC This is predominantly due to altered expression of the FimBE-like recombinases ipuA and ipbA, affecting at the same time, the inversion of the fim promoter switch (fimS). In the pst mutant, inactivation of yaiC restored fim-dependent adhesion to bladder cells and virulence. Interestingly, expression of yaiC was activated by PhoB, since transcription of yaiC was linked to the PhoB-dependent phoA-psiF operon. As YaiC is involved in c-di-GMP biosynthesis, an increased accumulation of c-di-GMP was observed in the pst mutant. Hence, results suggest that one mechanism by which deletion of the Pst system reduces expression of type 1 fimbriae is through PhoBR-mediated activation of yaiC, which in turn increases accumulation of c-di-GMP, represses the fim operon and consequently, attenuates virulence in the mouse urinary tract infection model.IMPORTANCE Urinary tract infections (UTIs) are common bacterial infections in humans. They are mainly caused by Uropathogenic Escherichia coli (UPEC). We previously showed that interfering with phosphate homeostasis decreases expression of type 1 fimbriae and attenuates UPEC virulence. Herein, we

  20. Micro-evolutionary processes in sablefish Anoplopoma fimbria, based on polymorphism of the two sites of mitochondrial DNA.

    PubMed

    Orlova, S Y; Orlov, A M; Volkov, A A; Novikov, R N

    2014-01-01

    Sablefish Anoplopoma fimbria is a deep-sea fish, endemic to the North Pacific Ocean, with continuous range from southern California to the central part of Honshu Island, including the Bering and Okhotsk Seas. It is an important commercial species and a promising object for aquaculture. Compared to the eastern part of the range the population structure of sablefish in Asian waters is poorly studied. It is believed that sablefish goes to the Bering Sea and Pacific waters of Kamchatka and the Kuril Islands from the northeastern Pacific, and Asian waters are its eviction zone. Other authors suggest that replenishment of sablefish off the eastern Kamchatka and the Kuril Islands is not only due to migration of the adult fish from the Bering Sea along the continental slope, but also due to the drift of yearlings by Aleutian current over the American coast.

  1. Characterization of Nonenterotoxigenic Escherichia coli Strains Producing F17 Fimbriae Isolated from Diarrheic Lambs and Goat Kids

    PubMed Central

    Cid, D.; Sanz, R.; Marín, I.; de Greve, H.; Ruiz-Santa-Quiteria, J. A.; Amils, R.; de la Fuente, R.

    1999-01-01

    Forty-five ovine and caprine nonenterotoxigenic Escherichia coli strains producing F17-related fimbriae were characterized with respect to the fimbrial structural subunit and adhesin subtypes produced. In addition, several characteristics related to the virulence of strains producing F17 fimbriae were studied. Most of the strains (73%) possessed the f17cA structural subunit gene, whereas the f17aA and f17dA genes were detected only on three (6%) and two (4%) strains, respectively. The f17bA gene was not detected. All but one of these strains possessed the f17G genes of the adhesin subfamily II. The only strain having the f17G gene of subfamily I possessed the structural subunit gene f17dA. Sequencing of the f17A and f17G genes of four selected strains confirmed the association of f17cA and f17dA structural subunit genes with the f17G genes of the adhesin subfamily II. These results indicated that adhesins of the subfamily II are prominent among ovine and caprine isolates and that they are indistinctly associated with the F17 structural subunit subtypes on these field strains. CS31A- and CNF2-related genes were not detected. Most of the strains adhered in vitro to ovine intestinal brush borders (36 of 45) and agglutinated the erythrocytes of different species in the presence of d-mannose (39 of 45). F17-positive strains produced colicin V (57%) and were resistant to the bactericidal effect of serum (91%) in significantly higher percentages than F17-negative strains (34% produced colicin V, and 66% were serum resistant). Thus, most of the studied ovine and caprine strains showed phenotypic characteristics of septicemic strains. PMID:10203489

  2. SEF14 fimbriae from Salmonella enteritidis play a role in pathogenitic to cell model in vitro and host in vivo.

    PubMed

    Zhu, Chunhong; Meng, Xia; Duan, Xiaoli; Tao, Zhiyun; Gong, Jiansen; Hou, Huayan; Zhu, Guoqiang

    2013-11-01

    The role of SEF14 fimbriae in virulence remains to be elucidated and in this study, we showed that sefA mutant constructed in the wild-type (WT) Salmonella enteritidis strain 50336 displayed increased invasion to IPEC-J2 cell lines and survival in mouse peritoneal macrophages, and the lethal dose 50% (LD50) in 6-week-old Balb/c mice intra-peritoneally injected with WT S. enteritidis strain decreased significantly upon deletion of sefA indicating their role in virulence. Overall, these results demonstrated that expression of sefA of SEF14 fimbriae enhances S. enteritidis adhesion to epithelial cells and survival in macrophages and contributes to S. enteritidis virulence in mice.

  3. The plasmid-encoded Ipf and Klf fimbriae display different expression and varying roles in the virulence of Salmonella enterica serovar Infantis in mouse vs. avian hosts

    PubMed Central

    Mikhlin, Svetlana; Cohen, Helit; Vitman Zilber, Shaul; Grassl, Guntram A.

    2017-01-01

    Salmonella enterica serovar Infantis is one of the prevalent Salmonella serovars worldwide. Different emergent clones of S. Infantis were shown to acquire the pESI virulence-resistance megaplasmid affecting its ecology and pathogenicity. Here, we studied two previously uncharacterized pESI-encoded chaperone-usher fimbriae, named Ipf and Klf. While Ipf homologs are rare and were found only in S. enterica subspecies diarizonae and subspecies VII, Klf is related to the known K88-Fae fimbria and klf clusters were identified in seven S. enterica subspecies I serovars, harboring interchanging alleles of the fimbria major subunit, KlfG. Regulation studies showed that the klf genes expression is negatively and positively controlled by the pESI-encoded regulators KlfL and KlfB, respectively, and are activated by the ancestral leucine-responsive regulator (Lrp). ipf genes are negatively regulated by Fur and activated by OmpR. Furthermore, induced expression of both klf and ipf clusters occurs under microaerobic conditions and at 41°C compared to 37°C, in-vitro. Consistent with these results, we demonstrate higher expression of ipf and klf in chicks compared to mice, characterized by physiological temperature of 41.2°C and 37°C, respectively. Interestingly, while Klf was dispensable for S. Infantis colonization in the mouse, Ipf was required for maximal colonization in the murine ileum. In contrast to these phenotypes in mice, both Klf and Ipf contributed to a restrained infection in chicks, where the absence of these fimbriae has led to moderately higher bacterial burden in the avian host. Taken together, these data suggest that physiological differences between host species, such as the body temperature, can confer differences in fimbriome expression, affecting Salmonella colonization and other host-pathogen interplays. PMID:28817673

  4. Construction of non-toxic Escherichia coli and Vibrio cholerae strains expressing high and immunogenic levels of enterotoxigenic E. coli colonization factor I fimbriae.

    PubMed

    Tobias, Joshua; Lebens, Michael; Bölin, Ingrid; Wiklund, Gudrun; Svennerholm, Ann-Mari

    2008-02-06

    To express high quantities of colonization factor antigen I (CFA/I) derived from enterotoxigenic Escherichia coli (ETEC) for use in ETEC vaccines, the entire CFA/I operon consisting of four genes (cfa-A, -B, -C, -E) was cloned into plasmid expression vectors that could be maintained either with or without antibiotic selection. Expression from the powerful tac promoter was under the control of the lacIq repressor present on the plasmids. Fimbriae were expressed on the surface of both a non-toxigenic E. coli K12 strain and a non-toxigenic strain of Vibrio cholerae following induction with isopropyl-beta-D-thiogalactopyranoside (IPTG). It was found that the recombinant E. coli strains expressed up to 16-fold higher levels of CFA/I fimbriae compared to a reference strain which had previously been shown to be among the highest natural producers of the CFA/I fimbriae among tested wild type ETEC strains. Oral immunization with formalin-killed recombinant E. coli bacteria over-expressing CFA/I induced significantly higher serum IgA and IgG+M antibodies responses compared to the reference strain. Oral immunization with formalin-killed recombinant V. cholerae bacteria also induce strong CFA/I-specific serum IgA and IgG+M responses. We conclude that our constructs may be useful as candidate strains in an oral killed CF-ETEC vaccine.

  5. Curli fimbriae are conditionally required in Escherichia coli O157:H7 for initial attachment and biofilm formation.

    PubMed

    Carter, Michelle Qiu; Louie, Jacqueline W; Feng, Doris; Zhong, Wayne; Brandl, Maria T

    2016-08-01

    Several species of enteric pathogens produce curli fimbriae, which may affect their interaction with surfaces and other microbes in nonhost environments. Here we used two Escherichia coli O157:H7 outbreak strains with distinct genotypes to understand the role of curli in surface attachment and biofilm formation in several systems relevant to fresh produce production and processing. Curli significantly enhanced the initial attachment of E. coli O157:H7 to spinach leaves and stainless steel surfaces by 5-fold. Curli was also required for E. coli O157:H7 biofilm formation on stainless steel and enhanced biofilm production on glass by 19-27 fold in LB no-salt broth. However, this contribution was not observed when cells were grown in sterile spinach lysates. Furthermore, both strains of E. coli O157:H7 produced minimal biofilms on polypropylene in LB no-salt broth but considerable amounts in spinach lysates. Under the latter conditions, curli appeared to slightly increase biofilm production. Importantly, curli played an essential role in the formation of mixed biofilm by E. coli O157:H7 and plant-associated microorganisms in spinach leaf washes, as revealed by confocal microscopy. Little or no E. coli O157:H7 biofilms were detected at 4 °C, supporting the importance of temperature control in postharvest and produce processing environments.

  6. Environmental covariates of sablefish (Anoplopoma fimbria) and Pacific ocean perch (Sebastes alutus) recruitment in the Gulf of Alaska

    NASA Astrophysics Data System (ADS)

    Coffin, Brendan; Mueter, Franz

    2016-10-01

    The sablefish (Anoplopoma fimbria) and Pacific ocean perch (POP; Sebastes alutus) fisheries in the Gulf of Alaska (GOA) are both highly lucrative and variable. Determining environmental factors that drive variability in their recruitment may improve our understanding of forces affecting their early life survival, which may be helpful when evaluating management strategies. Here we examine relationships between sablefish and POP recruitment and multiple environmental indices associated with circulation in the GOA. We used hierarchical cluster analysis to determine spatially and seasonally relevant scales for analyzing these relationships. We then used structural equation modeling to examine sequential relationships between large-scale climate variables, regional (eastern and western GOA) environmental variables, and recruitment using both hypothesis-testing and exploratory approaches. Exploratory analyses revealed that sablefish recruitment was positively related to July upwelling-favorable winds and negatively related to late winter freshwater discharge in the eastern GOA during age 1. POP recruitment was negatively related to June upwelling-favorable winds in both regions during ages 0 and 1 and positively related to late spring freshwater discharge throughout the GOA during age 1. These results suggest that upwelling-favorable winds and freshwater discharge may affect recruitment of both species through productivity-related mechanisms, and may additionally affect POP recruitment through advection-related mechanisms. Targeted studies at the appropriate scales are needed to provide greater certainty in the potential mechanisms behind these relationships.

  7. Crohn disease--associated adherent-invasive E. coli bacteria target mouse and human Peyer's patches via long polar fimbriae.

    PubMed

    Chassaing, Benoit; Rolhion, Nathalie; de Vallée, Amélie; Salim, Sa'ad Y; Prorok-Hamon, Maelle; Neut, Christel; Campbell, Barry J; Söderholm, Johan D; Hugot, Jean-Pierre; Colombel, Jean-Frédéric; Darfeuille-Michaud, Arlette

    2011-03-01

    Crohn disease (CD) is a multifactorial disease in which an abnormal immune response in the gastrointestinal (GI) tract leads to chronic inflammation. The small intestine, particularly the ileum, of patients with CD is colonized by adherent-invasive E. coli (AIEC)--a pathogenic group of E. coli able to adhere to and invade intestinal epithelial cells. As the earliest inflammatory lesions are microscopic erosions of the epithelium lining the Peyer's patches (PPs), we investigated the ability of AIEC bacteria to interact with PPs and the virulence factors involved. We found that AIEC bacteria could interact with mouse and human PPs via long polar fimbriae (LPF). An LPF-negative AIEC mutant was highly impaired in its ability to interact with mouse and human PPs and to translocate across monolayers of M cells, specialized epithelial cells at the surface of PPs. The prevalence of AIEC strains harboring the lpf operon was markedly higher in CD patients compared with controls. In addition, increased numbers of AIEC, but not LPF-deficient AIEC, bacteria were found interacting with PPs from Nod2(-/-) mice compared with WT mice. In conclusion, we have identified LPF as a key factor for AIEC to target PPs. This could be the missing link between AIEC colonization and the presence of early lesions in the PPs of CD patients.

  8. A Novel pAA Virulence Plasmid Encoding Toxins and Two Distinct Variants of the Fimbriae of Enteroaggregative Escherichia coli

    PubMed Central

    Jønsson, Rie; Struve, Carsten; Boll, Erik J.; Boisen, Nadia; Joensen, Katrine G.; Sørensen, Camilla A.; Jensen, Betina H.; Scheutz, Flemming; Jenssen, Håvard; Krogfelt, Karen A.

    2017-01-01

    Enteroaggregative Escherichia coli (EAEC) is an increasingly recognized pathogen associated with acute and persistent diarrhea worldwide. While EAEC strains are considered highly heterogeneous, aggregative adherence fimbriae (AAFs) are thought to play a pivotal role in pathogenicity by facilitating adherence to the intestinal mucosa. In this study, we optimized an existing multiplex PCR to target all known AAF variants, which are distinguished by differences in their pilin subunits. We applied the assay on a collection of 162 clinical Danish EAEC strains and interestingly found six, by SNP analysis phylogenetically distinct, strains harboring the major pilin subunits from both AAF/III and AAF/V. Whole-genome and plasmid sequencing revealed that in these six strains the agg3A and agg5A genes were located on a novel pAA plasmid variant. Moreover, the plasmid also encoded several other virulence genes including some not previously found on pAA plasmids. Thus, this plasmid endows the host strains with a remarkably high number of EAEC associated virulence genes hereby likely promoting strain pathogenicity. PMID:28275371

  9. Pathogenic Role of SEF14, SEF17, and SEF21 Fimbriae in Salmonella enterica Serovar Enteritidis Infection of Chickens

    PubMed Central

    Rajashekara, Gireesh; Munir, Shirin; Alexeyev, Mikhail F.; Halvorson, David A.; Wells, Carol L.; Nagaraja, Kakambi V.

    2000-01-01

    Very little is known about the contribution of surface appendages of Salmonella enterica serovar Enteritidis to pathogenesis in chickens. This study was designed to clarify the role of SEF14, SEF17, and SEF21 fimbriae in serovar Enteritidis pathogenesis. Stable, single, defined sefA (SEF14), agfA (SEF17), and fimA (SEF21) insertionally inactivated fimbrial gene mutants of serovar Enteritidis were constructed. All mutant strains invaded Caco-2 and HT-29 enterocytes at levels similar to that of the wild type. Both mutant and wild-type strains were ingested equally well by chicken macrophage cell lines HD11 and MQ-NCSU. There were no significant differences in the abilities of these strains to colonize chicken ceca. The SEF14− strain was isolated in lower numbers from the livers of infected chickens and was cleared from the spleens faster than other strains. No significant differences in fecal shedding of these strains were observed. PMID:10742278

  10. Type 3 Fimbriae and Biofilm Formation Are Regulated by the Transcriptional Regulators MrkHI in Klebsiella pneumoniae▿

    PubMed Central

    Johnson, Jeremiah G.; Murphy, Caitlin N.; Sippy, Jean; Johnson, Tylor J.; Clegg, Steven

    2011-01-01

    Klebsiella pneumoniae is an opportunistic pathogen which frequently causes hospital-acquired urinary and respiratory tract infections. K. pneumoniae may establish these infections in vivo following adherence, using the type 3 fimbriae, to indwelling devices coated with extracellular matrix components. Using a colony immunoblot screen, we identified transposon insertion mutants which were deficient for type 3 fimbrial surface production. One of these mutants possessed a transposon insertion within a gene, designated mrkI, encoding a putative transcriptional regulator. A site-directed mutant of this gene was constructed and shown to be deficient for fimbrial surface expression under aerobic conditions. MrkI mutants have a significantly decreased ability to form biofilms on both abiotic and extracellular matrix-coated surfaces. This gene was found to be cotranscribed with a gene predicted to encode a PilZ domain-containing protein, designated MrkH. This protein was found to bind cyclic-di-GMP (c-di-GMP) and regulate type 3 fimbrial expression. PMID:21571997

  11. Fimbria-fornix and entorhinal cortex differential contribution to contextual and cued fear conditioning consolidation in rats.

    PubMed

    Baldi, Elisabetta; Liuzzo, Antonino; Bucherelli, Corrado

    2013-04-10

    The Fimbria-Fornix (FF) and Entorhinal Cortex (EC) are the primary interfaces between the hippocampus and, respectively, subcortical structures and cortical areas. Their mnemonic role has been repeatedly proposed. In order to investigate their role in fear conditioning, FF and EC were subjected to bilateral fully reversible tetrodotoxin (TTX) inactivation during consolidation in adult male Wistar rats that had undergone training for fear conditioning to an acoustic stimulus (CS) and context. TTX was stereotaxically injected into animals of different groups at increasing post-acquisition delays. Memory was assessed as conditioned freezing duration measured during retention testing, performed 72 and 96 h after TTX administration in a counterbalanced manner. The results showed that FF inactivation, performed immediately after conditioning, did not disrupt consolidation of either contextual or auditory fear memory. On the contrary, EC inactivation performed at the same time was followed by both contextual and CS fear response retention impairment. EC inactivation performed 1.5h post-acquisition impaired only contextual fear response retention. EC inactivation performed 24h after acquisition training had no effect on the consolidation process. The present findings show a clearly different role of FF and EC in fear conditioning consolidation in the rat. The results are discussed in relation to their known connections with the hippocampus.

  12. Comparative proteomics of uropathogenic Escherichia coli during growth in human urine identify UCA-like (UCL) fimbriae as an adherence factor involved in biofilm formation and binding to uroepithelial cells.

    PubMed

    Wurpel, Daniël J; Totsika, Makrina; Allsopp, Luke P; Webb, Richard I; Moriel, Danilo G; Schembri, Mark A

    2016-01-10

    Uropathogenic Escherichia coli (UPEC) are the primary cause of urinary tract infection (UTI) in humans. For the successful colonisation of the human urinary tract, UPEC employ a diverse collection of secreted or surface-exposed virulence factors including toxins, iron acquisition systems and adhesins. In this study, a comparative proteomic approach was utilised to define the UPEC pan and core surface proteome following growth in pooled human urine. Identified proteins were investigated for subcellular origin, prevalence and homology to characterised virulence factors. Fourteen core surface proteins were identified, as well as eleven iron uptake receptor proteins and four distinct fimbrial types, including type 1, P, F1C/S and a previously uncharacterised fimbrial type, designated UCA-like (UCL) fimbriae in this study. These pathogenicity island (PAI)-associated fimbriae are related to UCA fimbriae of Proteus mirabilis, associated with UPEC and exclusively found in members of the E. coli B2 and D phylogroup. We further demonstrated that UCL fimbriae promote significant biofilm formation on abiotic surfaces and mediate specific attachment to exfoliated human uroepithelial cells. Combined, this study has defined the surface proteomic profiles and core surface proteome of UPEC during growth in human urine and identified a new type of fimbriae that may contribute to UTI. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

  13. Immunohistochemical analysis of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) toxicity on the developmental dentate gyrus and hippocampal fimbria in fetal mice

    PubMed Central

    KOBAYASHI, Yoshihiro; HIRANO, Tetsushi; OMOTEHARA, Takuya; HASHIMOTO, Rie; UMEMURA, Yuria; YUASA, Hideto; MASUDA, Natsumi; KUBOTA, Naoto; MINAMI, Kiichi; YANAI, Shogo; ISHIHARA-SUGANO, Mitsuko; MANTANI, Youhei; YOKOYAMA, Toshifumi; KITAGAWA, Hiroshi; HOSHI, Nobuhiko

    2015-01-01

    Dioxins are widespread persistent environmental contaminants with adverse impacts on humans and experimental animals. Behavioral and cognitive functions are impaired by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure. TCDD exerts its toxicity via the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor. The hippocampus, which plays important roles in episodic memory and spatial function, is considered vulnerable to TCDD-induced neurotoxicity, because it contains the AhR. We herein investigated the effects of TCDD toxicity on hippocampal development in embryonic mice. TCDD was administered to dams at 8.5 days postcoitum with a single dose of 20, 200, 2,000 and 5,000 ng/kg body weight (groups T20, T200, T2000 and T5000, respectively), and the brains were dissected from their pups at embryonic day 18.5. Immunohistochemical analysis demonstrated that the Glial Fibrillary Acidic Protein (GFAP) immunoreactivities in the dentate gyrus (DG) were reduced in the T5000 group. Granular GFAP immunoreactivity was observed in the hippocampal fimbria, and the number of immunoreactive fimbria was significantly decreased in the T5000 group. The number of Proliferating Cell Nuclear Antigen (PCNA)-positive cells was decreased in all TCDD-exposed groups and significantly reduced in the T20, T200 and T5000 groups. Together, these results demonstrate that maternal TCDD exposure has adverse impacts on neural stem cells (NSCs), neural precursor cells (NPCs) and granular cells in the DG and disrupts the NSC maintenance and timing of differentiation in the hippocampal fimbria, which in turn interrupt neuronal development in future generations of mice. PMID:26096965

  14. Preliminary X-ray diffraction analysis of CfaA, a molecular chaperone essential for the assembly of CFA/I fimbriae of human enterotoxigenic Escherichia coli.

    PubMed

    Bao, Rui; Esser, Lothar; Poole, Steven; McVeigh, Annette; Chen, Yu Xing; Savarino, Stephen J; Xia, Di

    2014-02-01

    Understanding of pilus bioassembly in Gram-negative bacteria stems mainly from studies of P pili and type 1 fimbriae of uropathogenic Escherichia coli, which are mediated by the classic chaperone-usher pathway (CUP). However, CFA/I fimbriae, a class 5 fimbria and intestinal colonization factor for enterotoxigenic E. coli (ETEC), are proposed to assemble via the alternate chaperone pathway (ACP). Both CUP and ACP fimbrial bioassembly pathways require the function of a periplasmic chaperone, but their corresponding proteins share very low similarity in primary sequence. Here, the crystallization of the CFA/I periplasmic chaperone CfaA by the hanging-drop vapor-diffusion method is reported. X-ray diffraction data sets were collected from a native CfaA crystal to 2 Å resolution and to 1.8 and 2.8 Å resolution, respectively, from a lead and a platinum derivative. These crystals displayed the symmetry of space group C2, with unit-cell parameters a = 103.6, b = 28.68, c = 90.60 Å, β = 119.7°. Initial phases were derived from multiple isomorphous replacement with anomalous scattering experiments using the data from the platinum and lead derivatives. This resulted in an interpretable electron-density map showing one CfaA molecule in an asymmetric unit. Sequence assignments were aided by anomalous signals from the heavy-atom derivatives. Refinement of the atomic model of CfaA is ongoing, which is expected to further understanding of the essential aspects and allowable variations in tertiary structure of the greater family of chaperones involved in chaperone-usher mediated bioassembly.

  15. Immunization with the recombinant Cholera toxin B fused to Fimbria 2 protein protects against Bordetella pertussis infection.

    PubMed

    Olivera, Noelia; Castuma, Celina E; Hozbor, Daniela; Gaillard, María E; Rumbo, Martín; Gómez, Ricardo M

    2014-01-01

    This study examined the immunogenic properties of the fusion protein fimbria 2 of Bordetella pertussis (Fim2)-cholera toxin B subunit (CTB) in the intranasal murine model of infection. To this end B. pertussis Fim2 coding sequence was cloned downstream of the cholera toxin B subunit coding sequence. The expression and assembly of the fusion protein into pentameric structures (CTB-Fim2) were evaluated by SDS-PAGE and monosialotetrahexosylgaglioside (GM1-ganglioside) enzyme-linked immunosorbent assay (ELISA). To evaluate the protective capacity of CTB-Fim2, an intraperitoneal or intranasal mouse immunization schedule was performed with 50 μg of CTB-Fim2. Recombinant (rFim2) or purified (BpFim2) Fim2, CTB, and phosphate-buffered saline (PBS) were used as controls. The results showed that mice immunized with BpFim2 or CTB-Fim2 intraperitoneally or intranasally presented a significant reduction in bacterial lung counts compared to control groups (P < 0.01 or P < 0.001 , resp.). Moreover, intranasal immunization with CTB-Fim2 induced significant levels of Fim2-specific IgG in serum and bronchoalveolar lavage (BAL) and Fim2-specific IgA in BAL. Analysis of IgG isotypes and cytokines mRNA levels showed that CTB-Fim2 results in a mixed Th1/Th2 (T-helper) response. The data presented here provide support for CTB-Fim2 as a promising recombinant antigen against Bordetella pertussis infection.

  16. Molecular characterization of the gonadal kisspeptin system: Cloning, tissue distribution, gene expression analysis and localization in sablefish (Anoplopoma fimbria).

    PubMed

    Fairgrieve, Marian R; Shibata, Yasushi; Smith, Elizabeth K; Hayman, Edward S; Luckenbach, J Adam

    2016-01-01

    The kisspeptin system plays pivotal roles in the regulation of vertebrate reproduction. Classically, kisspeptin produced in the brain stimulates brain gonadotropin-releasing hormone signaling, which in turn activates the pituitary-gonad axis. Expression of the kisspeptin system has also been documented in peripheral tissues, including gonads of mammals and fishes. However, the fish gonadal kisspeptin system remained uncharacterized. Herein we report identification and characterization of four kisspeptin system mRNAs (kisspeptin 1 (kiss1), kiss2, and G protein-coupled receptor 54-1 (gpr54-1) and gpr54-2) in sablefish, Anoplopoma fimbria. Sablefish predicted protein sequences were highly similar to those of other marine teleosts, but less so to freshwater teleosts. Tissue distribution analyses revealed that all four kisspeptin-system transcripts were expressed in both brain and gonad. However, kiss2 was the predominant transcript in the gonads and the only transcript detected in ovulated eggs. Ontogenetic analysis of kiss2 expression in juvenile sablefish gonads demonstrated that levels were low during sex differentiation but increased with fish size and gonadal development. Dramatic increases in kiss2 mRNA occurred during primary oocyte growth, while levels remained relatively low in testes. In situ hybridization revealed that kiss2 mRNA was localized to cytoplasm of perinucleolus stage oocytes, suggesting it could play a local role in oogenesis or could be synthesized and stored within oocytes as maternal mRNA. This represents the first study to focus on the gonadal kisspeptin system in fishes and provides important tools for further investigation of both the gonadal and brain kisspeptin systems in sablefish.

  17. Post-training stimulation of the basolateral amygdala improves spatial learning in rats with lesion of the fimbria-fornix.

    PubMed

    Almaguer, William; Capdevila, Vladimir; Ramírez, Magaly; Vallejo, Araceli; Rosillo, Juan C; Bergado, Jorge A

    2005-01-01

    To evaluate the capacity of amygdala stimulation to improve neural plasticity in animals bearing lesions of the fimbria-fornix (FF) system. The animals were lesioned under narcosis (chloral hydrate, 420 mg/kg ip.) using a bilateral transection of the FF procedure. During the same surgery some animals were implanted with an electrode in the right basolateral amygdala (BLA) to allow the electrical stimulation of this structure. Training was carried out one week after surgery using a Morris water maze. Animals were trained in four consecutive days (8 trials/day) in the non-visible platform condition except in the fourth day in which only 4 trials were performed followed by a probe trial in which the escape platform was removed. On day 5 of training 8 trials with visible platform were performed. After each of the first 3 training days one group of animals received trains of electrical stimulation to the BLA, while control groups were not stimulated. A group of non-lesioned animals served as control. The location of the electrode was confirmed histologically after the end of the experiments. The learning capacity of the lesioned animals was improved by the electrical stimulation of the amygdala. The latency to find the submerged platform within this group approaches that of the non lesioned animals in the course of training (2-way ANOVA with repeated measures), while other lesioned animals continued to show severely impaired learning abilities. This is the first evidence that stimulating the BLA can positively influence the learning abilities of lesioned animals. Further experiments should contribute to improve the stimulation paradigms to make it more effective, if possible.

  18. Induction of Th1 polarized immune responses by thiolated Eudragit-coated F4 and F18 fimbriae of enterotoxigenic Escherichia coli.

    PubMed

    Lee, Won-Jung; Cha, Seungbin; Shin, Minkyoung; Islam, Mohammad Ariful; Cho, Chong-su; Yoo, Han Sang

    2011-10-01

    Diarrhea in newborn and weaned piglets is mainly induced by enterotoxigenic Escherichia coli (ETEC) with fimbriae F4 (K88) and F18 (F107). In this study, we evaluated F4 and F18 coated with thiolated Eudragit microspheres (TEMS) as a candidate for an oral vaccine. The average particle sizes of TEMS, F4-loaded TEMS, and F18-loaded TEMS were measured as 4.2±0.75 μm, 4.7±0.50 μm, and 4.5±0.37 μm, respectively. F4 is more efficiently encapsulated than F18 in the loading with TEMS. In the release test, F4 and F18 fimbriae were protected in acidic circumstances, whereas most were released at pH 7.4 of intestine circumstances. Production of TNF-α and NO from RAW 264.7 cells was increased in a time-dependent manner after exposure to all groups, whereas only F4- or F18-loaded TEMS-stimulated IL-6 secretion. The levels of IFN-γ from mouse splenocytes after exposure to F4 or F18 were increased while IL-4 was not detectable. These results suggest that F4- and F18-loaded TEMS may effectively induce immune response with the efficient release of antigens to appropriate target sites. Copyright © 2011 Elsevier B.V. All rights reserved.

  19. Role of Proteus mirabilis MR/P fimbriae and flagella in adhesion, cytotoxicity and genotoxicity induction in T24 and Vero cells.

    PubMed

    Scavone, Paola; Villar, Silvia; Umpiérrez, Ana; Zunino, Pablo

    2015-06-01

    Proteus mirabilis is frequently associated with complicated urinary tract infections (UTI). It is proposed that several virulence factors are associated with P. mirabilis uropathogenicity. The aim of this work was to elucidate genotoxic and cytotoxic effects mediated by MR/P fimbriae and flagella in eukaryotic cells in vitro. Two cell lines (kidney- and bladder-derived) were infected with a clinical wild-type P. mirabilis strain and an MR/P and a flagellar mutant. We evaluated adhesion, genotoxicity and cytotoxicity by microscopy, comet assay and triple staining technique, respectively. Mutant strains displayed lower adhesion rates than the P. mirabilis wild-type strain and were significantly less effective to induce genotoxic and cytotoxic effects compared to the wild type. We report for the first time that P. mirabilis MR/P fimbriae and flagella mediate genotoxic and cytotoxic effects on eukaryotic cells, at least in in vitro conditions. These results could contribute to design new strategies for the control of UTI.

  20. Salicylate increases the expression of marA and reduces in vitro biofilm formation in uropathogenic Escherichia coli by decreasing type 1 fimbriae expression.

    PubMed

    Vila, Jordi; Soto, Sara M

    2012-05-01

    Escherichia coli is one of the most frequent bacteria implicated in biofilm formation, which is a dynamic process whose first step consists in bacteria adhesion to surfaces through type 1 fimbriae. Salicylate induces a number of morphological and physiological alterations in bacteria including the activation of the transcriptional regulator MarA. In this report the effects of salicylate on biofilm formation and their relationship with MarA were studied. An inverse relationship was observed between in vitro biofilm formation and salicylate concentration added to the culture medium. Salicylate increases the expression of marA and decreases the expression of fimA and fimB genes in the wild-type strain. In addition, the fimA and fimB expression was decreased in a MarR mutant in which marA was also overexpressed. In conclusion, the expression of type 1 fimbriae in presence of salicylate may be regulated by the level of marA expression through fimB regulator, albeit through neither the ompX nor the tolC genes.

  1. Salicylate increases the expression of marA and reduces in vitro biofilm formation in uropathogenic Escherichia coli by decreasing type 1 fimbriae expression

    PubMed Central

    Vila, Jordi; Soto, Sara M.

    2012-01-01

    Escherichia coli is one of the most frequent bacteria implicated in biofilm formation, which is a dynamic process whose first step consists in bacteria adhesion to surfaces through type 1 fimbriae. Salicylate induces a number of morphological and physiological alterations in bacteria including the activation of the transcriptional regulator MarA. In this report the effects of salicylate on biofilm formation and their relationship with MarA were studied. An inverse relationship was observed between in vitro biofilm formation and salicylate concentration added to the culture medium. Salicylate increases the expression of marA and decreases the expression of fimA and fimB genes in the wild-type strain. In addition, the fimA and fimB expression was decreased in a MarR mutant in which marA was also overexpressed. In conclusion, the expression of type 1 fimbriae in presence of salicylate may be regulated by the level of marA expression through fimB regulator, albeit through neither the ompX nor the tolC genes. PMID:22546909

  2. Path integration absent in scent-tracking fimbria-fornix rats: evidence for hippocampal involvement in "sense of direction" and "sense of distance" using self-movement cues.

    PubMed

    Whishaw, I Q; Gorny, B

    1999-06-01

    Allothetic and idiothetic navigation strategies use very different cue constellations and computational processes. Allothetic navigation requires the use of the relationships between relatively stable external (visual, olfactory, auditory) cues, whereas idiothetic navigation requires the integration of cues generated by self-movement and/or efferent copy of movement commands. The flexibility with which animals can switch between these strategies and the neural structures that support these strategies are not well understood. By capitalizing on the proclivity of foraging rats to carry large food pellets back to a refuge for eating, the present study examined the contribution of the hippocampus to the use of allothetic versus idiothetic navigation strategies. Control rats and fimbria-fornix-ablated rats were trained to follow linear, polygonal, and octagonal scent trails that led to a piece of food. The ability of the rats to return to the refuge with the food via the shortest route using allothetic cues (visual cues and/or the odor trail available) or using ideothetic cues (the odor trail removed and the rats blindfolded or tested in infrared light) was examined. Control rats "closed the polygon" by returning directly home in all cue conditions. Fimbria-fornix rats successfully used allothetic cues (closed the polygon using visual cues or tracked back on the string) but were insensitive to the direction and distance of the refuge and were lost when restricted to idiothetic cues. The results support the hypothesis that the hippocampal formation is necessary for navigation requiring the integration of idiothetic cues.

  3. Increased Expression of Type-1 Fimbriae by Nonpathogenic Escherichia coli 83972 Results in an Increased Capacity for Catheter Adherence and Bacterial Interference

    PubMed Central

    Trautner, Barbara W.; Cevallos, Manuel E.; Li, Huaiguang; Riosa, Sarah; Hull, Richard A.; Hull, Sheila I.; Tweardy, David J.; Darouiche, Rabih O.

    2010-01-01

    Background In vitro, urinary catheter colonization by avirulent Escherichia coli 83972 impedes subsequent catheter colonization by a variety of uropathogenic organisms. However, E. coli 83972 shows a low efficacy of adherence to silicone urinary catheter material, possibly because the fim operon encoding adhesive type 1 fimbriae is incomplete. We hypothesized that improving the catheter adherence of E. coli 83972 would improve its bacterial interference properties. Methods We created adhesive mutants by transforming wild-type E. coli 83972 with fim+ plasmids. Adherence to urinary catheters and ability to prevent uropathogenic E. coli from colonizing urinary catheters were studied by use of a sonication assay. Results The addition of a single-copy fim+ plasmid increased adherence to urinary catheters 10-fold, and addition of an 18-copy fim+ plasmid increased adherence 100-fold. The more adherent 18-copy fim+ plasmid strain was more effective at blocking catheter colonization by pathogenic E. coli than was the wild-type parental strain. Neither Δfim nor fim+ E. coli 83972 adhered to shed urinary epithelial cells. Conclusions Our results indicate that improving urinary catheter adherence augments the bacterial interference capabilities of benign E. coli 83972. Increased expression of type-1 fimbriae may enhance bacterial interference without conferring virulence on E. coli 83972. PMID:18643750

  4. Crystallization and preliminary X-ray diffraction analysis of CfaE, the adhesive subunit of the CFA/I fimbriae from human enterotoxigenic Escherichia coli

    SciTech Connect

    Li, Yong-Fu; Poole, Steven; Rasulova, Fatima; Esser, Lothar; Savarino, Stephen J.; Xia, Di

    2006-02-01

    The adhesin CfaE of the CFA/I fimbriae from human enterotoxigenic E. coli has been crystallized. CfaE crystals diffracted X-rays to better than 2.4 Å and phasing was solved by the SIRAS method. Enterotoxigenic Escherichia coli (ETEC) represents a formidable food and waterborne diarrheal disease threat of global importance. The first step in ETEC pathogenesis is bacterial attachment to small-intestine epithelial cells via adhesive fimbriae, many of which are genetically related to the prototype colonization factor antigen I (CFA/I). The minor fimbrial subunit CfaE is required for initiation of CFA/I fimbrial assembly and mediates bacterial attachment to host cell-surface receptors. A donor-strand complemented variant of CfaE (dscCfaE) was expressed with a hexahistidine tag, purified to homogeneity and crystallized using the hanging-drop vapor-diffusion method. X-ray diffraction data sets were collected to 2.4 Å resolution for both native and derivatized crystals and showed the symmetry of space group P6{sub 2}22, with unit-cell parameters a = b = 142.9, c = 231.9 Å. Initial phases were derived from the SIRAS approach and electron density showed two molecules in the crystallographic asymmetric unit. Sequence assignments were aided by anomalous signals from the selenium of an SeMet-derivatized crystal and from S atoms of a native crystal.

  5. Construction and expression of immunogenic hybrid enterotoxigenic Escherichia coli CFA/I and CS2 colonization fimbriae for use in vaccines.

    PubMed

    Tobias, Joshua; Svennerholm, Ann-Mari; Holmgren, Jan; Lebens, Michael

    2010-07-01

    Enterotoxigenic Escherichia coli (ETEC) are an important cause of diarrheal morbidity in developing countries, especially in children and also of traveler's diarrhea. Colonization factors (CFs) of ETEC, like CFA/I and CS2 which are genetically and structurally related, play a substantial role in pathogenicity, and since intestinal-mucosal immune responses against CFs appear to be protective, much effort has focused on the development of a CF-based ETEC vaccine. We have constructed hybrid operons in which the major CS2 subunit-encoding cotA gene was inserted into the CFA/I operon, either replacing (hybrid I) or being added to the major CFA/I subunit-encoding cfaB gene (hybrid II). Using specific monoclonal antibodies against the major subunits of CFA/I and CS2, high levels of surface expression of both fimbrial subunits were shown in E. coli carrying the hybrid II operon. Oral immunization of mice with formalin-killed bacteria expressing hybrid II fimbriae induced strong CFA/I- and CS2-specific serum IgG + IgM and fecal IgA antibody responses, which were higher than those achieved by similar immunization with the reference strains. Bacteria expressing hybrid fimbriae are potential candidate strains in an oral-killed CF-ETEC vaccine, and the approach represents an attractive and novel means of producing a broad-spectrum ETEC vaccine.

  6. Amyloid-β induces synaptic dysfunction through G protein-gated inwardly rectifying potassium channels in the fimbria-CA3 hippocampal synapse.

    PubMed

    Nava-Mesa, Mauricio O; Jiménez-Díaz, Lydia; Yajeya, Javier; Navarro-Lopez, Juan D

    2013-01-01

    Last evidences suggest that, in Alzheimer's disease (AD) early stage, Amyloid-β (Aβ) peptide induces an imbalance between excitatory and inhibitory neurotransmission systems resulting in the functional impairment of neural networks. Such alterations are particularly important in the septohippocampal system where learning and memory processes take place depending on accurate oscillatory activity tuned at fimbria-CA3 synapse. Here, the acute effects of Aβ on CA3 pyramidal neurons and their synaptic activation from septal part of the fimbria were studied in rats. A triphasic postsynaptic response defined by an excitatory potential (EPSP) followed by both early and late inhibitory potentials (IPSP) was evoked. The EPSP was glutamatergic acting on ionotropic receptors. The early IPSP was blocked by GABAA antagonists whereas the late IPSP was removed by GABAB antagonists. Aβ perfusion induced recorded cells to depolarize, increase their input resistance and decrease the late IPSP. Aβ action mechanism was localized at postsynaptic level and most likely linked to GABAB-related ion channels conductance decrease. In addition, it was found that the specific pharmacological modulation of the GABAB receptor effector, G-protein-coupled inward rectifier potassium (GirK) channels, mimicked all Aβ effects previously described. Thus, our findings suggest that Aβ altering GirK channels conductance in CA3 pyramidal neurons might have a key role in the septohippocampal activity dysfunction observed in AD.

  7. Amyloid-β induces synaptic dysfunction through G protein-gated inwardly rectifying potassium channels in the fimbria-CA3 hippocampal synapse

    PubMed Central

    Nava-Mesa, Mauricio O.; Jiménez-Díaz, Lydia; Yajeya, Javier; Navarro-Lopez, Juan D.

    2013-01-01

    Last evidences suggest that, in Alzheimer's disease (AD) early stage, Amyloid-β (Aβ) peptide induces an imbalance between excitatory and inhibitory neurotransmission systems resulting in the functional impairment of neural networks. Such alterations are particularly important in the septohippocampal system where learning and memory processes take place depending on accurate oscillatory activity tuned at fimbria-CA3 synapse. Here, the acute effects of Aβ on CA3 pyramidal neurons and their synaptic activation from septal part of the fimbria were studied in rats. A triphasic postsynaptic response defined by an excitatory potential (EPSP) followed by both early and late inhibitory potentials (IPSP) was evoked. The EPSP was glutamatergic acting on ionotropic receptors. The early IPSP was blocked by GABAA antagonists whereas the late IPSP was removed by GABAB antagonists. Aβ perfusion induced recorded cells to depolarize, increase their input resistance and decrease the late IPSP. Aβ action mechanism was localized at postsynaptic level and most likely linked to GABAB-related ion channels conductance decrease. In addition, it was found that the specific pharmacological modulation of the GABAB receptor effector, G-protein-coupled inward rectifier potassium (GirK) channels, mimicked all Aβ effects previously described. Thus, our findings suggest that Aβ altering GirK channels conductance in CA3 pyramidal neurons might have a key role in the septohippocampal activity dysfunction observed in AD. PMID:23898239

  8. In vivo emergence of enterotoxigenic Escherichia coli variants lacking genes for K99 fimbriae and heat-stable enterotoxin.

    PubMed Central

    Mainil, J G; Sadowski, P L; Tarsio, M; Moon, H W

    1987-01-01

    Neonatal pigs were inoculated with porcine enterotoxigenic Escherichia coli 431, which carries genes for K99 fimbriae and STaP enterotoxin. Colonies of strain 431 were recovered from feces of pigs for up to 17 days after inoculation and tested for hybridization with gene probes for K99 and STaP. Variants of strain 431 that did not hybridize with the probes were considered to have lost the genes. Variants were recovered from 10 of 13 suckling pigs that survived the infection. Only 0.4% of the isolates recovered during the first 2 days after inoculation were variants. Of the isolates recovered 3 to 5 days after inoculation, 20 to 36% were variants. Variant colonies were detected more frequently among pigs in some litters than in others. The litter with the highest number of variant-shedding pigs had the dam with the highest titer of K99 antibody in her colostrum. Variants also occurred in colostrum-deprived, artificially reared pigs. However, the number of variants detected was lower and they occurred later in the course of the infection in colostrum-deprived pigs than in suckling pigs. More variants were detected and they were detected earlier in colostrum-deprived pigs fed anti-K99 monoclonal antibody than in controls fed anti-K88 monoclonal antibody. Loss of STaP appeared to be secondary to loss of K99 in that some variants lacked only K99 (K99- STaP+) and some lacked both genes (K99- STaP-), but none was of the K99+ STaP- type. Our results confirmed reports of gene loss from enterotoxigenic E. coli during infection. They are consistent with the hypothesis that variants emerge under in vivo selection pressure of K99 antibody and with the speculation that gene loss may be an important component of protection in vaccinated populations. However, the emergence of variants did not appear to play a major role in the recovery of individual pigs from clinical disease. PMID:2890584

  9. Retrograde cell changes in medial septum and diagonal band following fimbria-fornix transection: quantitative temporal analysis.

    PubMed

    Gage, F H; Wictorin, K; Fischer, W; Williams, L R; Varon, S; Bjorklund, A

    1986-09-01

    Complete unilateral fimbria-fornix transections, including the overlying cingulate cortex, were administered to female rats. At time points from 1 day to 6 weeks, the septal-diagonal band region was examined using acetylcholinesterase histochemistry, Cresyl Violet cell staining, and choline acetyltransferase biochemistry. As early as 1 day following the transection a decrease in acetylcholinesterase positive cell body staining was observed in the medial septum; however, no loss of Nissl-stained neurons was measured in Cresyl Violet stained sections until 1 week after the lesion. Maximal loss of acetylcholinesterase-positive cells, as visualized after irreversible acetylcholinesterase inhibition, was measured at 1 week, and no further change was observed at time points up to 6 weeks after operation. The loss of acetyltransferase-positive cells was greatest in the medial septal area (-65%) and the vertical limb of the diagonal band (-55%). Little cell loss was measured in the horizontal limb of the diagonal band. This is consistent with the known projections of these cell bodies. Remaining acetylcholinesterase-positive cell bodies in the medial septum had shrunk by about 20% (measured as the diameter along the major axis). A marked neuronal cell loss (about 50%) was demonstrable in the medial septum and vertical limb of the diagonal band in the Cresyl Violet-stained sections, too. A pile-up of acetylcholinesterase-stained material was observed in the dorsal-lateral quadrant of the septal area just proximal to the lesion at 1 day following transection. This pile-up occurred in the medial septum and diagonal band area up to 1 week following the transection, and had nearly disappeared by 2 weeks post-transection. Choline acetyltransferase biochemical activity, measured in samples of whole septum, decreased significantly at 1 day but subsequently returned to control levels. By 2 weeks following transection, an increase in acetylcholinesterase-positive stained fibers was

  10. MrkH, a Novel c-di-GMP-Dependent Transcriptional Activator, Controls Klebsiella pneumoniae Biofilm Formation by Regulating Type 3 Fimbriae Expression

    PubMed Central

    Wilksch, Jonathan J.; Yang, Ji; Clements, Abigail; Gabbe, Jacinta L.; Short, Kirsty R.; Cao, Hanwei; Cavaliere, Rosalia; James, Catherine E.; Whitchurch, Cynthia B.; Schembri, Mark A.; Chuah, Mary L. C.; Liang, Zhao-Xun; Wijburg, Odilia L.; Jenney, Adam W.; Lithgow, Trevor; Strugnell, Richard A.

    2011-01-01

    Klebsiella pneumoniae causes significant morbidity and mortality worldwide, particularly amongst hospitalized individuals. The principle mechanism for pathogenesis in hospital environments involves the formation of biofilms, primarily on implanted medical devices. In this study, we constructed a transposon mutant library in a clinical isolate, K. pneumoniae AJ218, to identify the genes and pathways implicated in biofilm formation. Three mutants severely defective in biofilm formation contained insertions within the mrkABCDF genes encoding the main structural subunit and assembly machinery for type 3 fimbriae. Two other mutants carried insertions within the yfiN and mrkJ genes, which encode GGDEF domain- and EAL domain-containing c-di-GMP turnover enzymes, respectively. The remaining two isolates contained insertions that inactivated the mrkH and mrkI genes, which encode for novel proteins with a c-di-GMP-binding PilZ domain and a LuxR-type transcriptional regulator, respectively. Biochemical and functional assays indicated that the effects of these factors on biofilm formation accompany concomitant changes in type 3 fimbriae expression. We mapped the transcriptional start site of mrkA, demonstrated that MrkH directly activates transcription of the mrkA promoter and showed that MrkH binds strongly to the mrkA regulatory region only in the presence of c-di-GMP. Furthermore, a point mutation in the putative c-di-GMP-binding domain of MrkH completely abolished its function as a transcriptional activator. In vivo analysis of the yfiN and mrkJ genes strongly indicated their c-di-GMP-specific function as diguanylate cyclase and phosphodiesterase, respectively. In addition, in vitro assays showed that purified MrkJ protein has strong c-di-GMP phosphodiesterase activity. These results demonstrate for the first time that c-di-GMP can function as an effector to stimulate the activity of a transcriptional activator, and explain how type 3 fimbriae expression is coordinated

  11. MrkH, a novel c-di-GMP-dependent transcriptional activator, controls Klebsiella pneumoniae biofilm formation by regulating type 3 fimbriae expression.

    PubMed

    Wilksch, Jonathan J; Yang, Ji; Clements, Abigail; Gabbe, Jacinta L; Short, Kirsty R; Cao, Hanwei; Cavaliere, Rosalia; James, Catherine E; Whitchurch, Cynthia B; Schembri, Mark A; Chuah, Mary L C; Liang, Zhao-Xun; Wijburg, Odilia L; Jenney, Adam W; Lithgow, Trevor; Strugnell, Richard A

    2011-08-01

    Klebsiella pneumoniae causes significant morbidity and mortality worldwide, particularly amongst hospitalized individuals. The principle mechanism for pathogenesis in hospital environments involves the formation of biofilms, primarily on implanted medical devices. In this study, we constructed a transposon mutant library in a clinical isolate, K. pneumoniae AJ218, to identify the genes and pathways implicated in biofilm formation. Three mutants severely defective in biofilm formation contained insertions within the mrkABCDF genes encoding the main structural subunit and assembly machinery for type 3 fimbriae. Two other mutants carried insertions within the yfiN and mrkJ genes, which encode GGDEF domain- and EAL domain-containing c-di-GMP turnover enzymes, respectively. The remaining two isolates contained insertions that inactivated the mrkH and mrkI genes, which encode for novel proteins with a c-di-GMP-binding PilZ domain and a LuxR-type transcriptional regulator, respectively. Biochemical and functional assays indicated that the effects of these factors on biofilm formation accompany concomitant changes in type 3 fimbriae expression. We mapped the transcriptional start site of mrkA, demonstrated that MrkH directly activates transcription of the mrkA promoter and showed that MrkH binds strongly to the mrkA regulatory region only in the presence of c-di-GMP. Furthermore, a point mutation in the putative c-di-GMP-binding domain of MrkH completely abolished its function as a transcriptional activator. In vivo analysis of the yfiN and mrkJ genes strongly indicated their c-di-GMP-specific function as diguanylate cyclase and phosphodiesterase, respectively. In addition, in vitro assays showed that purified MrkJ protein has strong c-di-GMP phosphodiesterase activity. These results demonstrate for the first time that c-di-GMP can function as an effector to stimulate the activity of a transcriptional activator, and explain how type 3 fimbriae expression is coordinated

  12. Effects of dorsal-striatum lesions and fimbria-fornix lesions on the problem-solving strategies of rats in a shallow water maze.

    PubMed

    Okaichi, H

    2001-09-01

    In solving a spatial problem, animals can use a place, cue, or response strategy. The present research was designed to evaluate the role of dorsal striatum (DS) in spatial problem solving and to compare it with that of fimbria fornix (FF). Rats were trained with a place + cue task in a shallow pool, then were divided into three groups (DS, FF, control), and lesions were made in the corresponding areas. After retraining, four probe tests were given: Test 1 (start position moved), Test 2 (goal and start positions moved), Test 3 (invisible goal), and Test 4 (curtain test). The test results suggest that the DS and Control groups performed the original task by using the place strategy, whereas the FF group used the cue strategy, which strongly implies that the DS group was impaired in the use of the cue strategy. This research also provides evidence supporting the usefulness of a shallow pool in evaluating animal behavior.

  13. Inhibition of type 1 fimbriae-mediated Escherichia coli adhesion and biofilm formation by trimeric cluster thiomannosides conjugated to diamond nanoparticles

    NASA Astrophysics Data System (ADS)

    Khanal, Manakamana; Larsonneur, Fanny; Raks, Victoriia; Barras, Alexandre; Baumann, Jean-Sébastien; Martin, Fernando Ariel; Boukherroub, Rabah; Ghigo, Jean-Marc; Ortiz Mellet, Carmen; Zaitsev, Vladimir; Garcia Fernandez, Jose M.; Beloin, Christophe; Siriwardena, Aloysius; Szunerits, Sabine

    2015-01-01

    Recent advances in nanotechnology have seen the development of a number of microbiocidal and/or anti-adhesive nanoparticles displaying activity against biofilms. In this work, trimeric thiomannoside clusters conjugated to nanodiamond particles (ND) were targeted for investigation. NDs have attracted attention as a biocompatible nanomaterial and we were curious to see whether the high mannose glycotope density obtained upon grouping monosaccharide units in triads might lead to the corresponding ND-conjugates behaving as effective inhibitors of E. coli type 1 fimbriae-mediated adhesion as well as of biofilm formation. The required trimeric thiosugar clusters were obtained through a convenient thiol-ene ``click'' strategy and were subsequently conjugated to alkynyl-functionalized NDs using a Cu(i)-catalysed ``click'' reaction. We demonstrated that the tri-thiomannoside cluster-conjugated NDs (ND-Man3) show potent inhibition of type 1 fimbriae-mediated E. coli adhesion to yeast and T24 bladder cells as well as of biofilm formation. The biofilm disrupting effects demonstrated here have only rarely been reported in the past for analogues featuring such simple glycosidic motifs. Moreover, the finding that the tri-thiomannoside cluster (Man3N3) is itself a relatively efficient inhibitor, even when not conjugated to any ND edifice, suggests that alternative mono- or multivalent sugar-derived analogues might also be usefully explored for E. coli-mediated biofilm disrupting properties.Recent advances in nanotechnology have seen the development of a number of microbiocidal and/or anti-adhesive nanoparticles displaying activity against biofilms. In this work, trimeric thiomannoside clusters conjugated to nanodiamond particles (ND) were targeted for investigation. NDs have attracted attention as a biocompatible nanomaterial and we were curious to see whether the high mannose glycotope density obtained upon grouping monosaccharide units in triads might lead to the corresponding

  14. F1C Fimbriae Play an Important Role in Biofilm Formation and Intestinal Colonization by the Escherichia coli Commensal Strain Nissle 1917▿

    PubMed Central

    Lasaro, Melissa A.; Salinger, Nina; Zhang, Jing; Wang, Yantao; Zhong, Zhengtao; Goulian, Mark; Zhu, Jun

    2009-01-01

    Bacterial biofilm formation is thought to enhance survival in natural environments and during interaction with hosts. A robust colonizer of the human gastrointestinal tract, Escherichia coli Nissle 1917, is widely employed in probiotic therapy. In this study, we performed a genetic screen to identify genes that are involved in Nissle biofilm formation. We found that F1C fimbriae are required for biofilm formation on an inert surface. In addition, these structures are also important for adherence to epithelial cells and persistence in infant mouse colonization. The data suggest a possible connection between Nissle biofilm formation and the survival of this commensal within the host. Further study of the requirements for robust biofilm formation may improve the therapeutic efficacy of Nissle 1917. PMID:18997018

  15. F1C fimbriae play an important role in biofilm formation and intestinal colonization by the Escherichia coli commensal strain Nissle 1917.

    PubMed

    Lasaro, Melissa A; Salinger, Nina; Zhang, Jing; Wang, Yantao; Zhong, Zhengtao; Goulian, Mark; Zhu, Jun

    2009-01-01

    Bacterial biofilm formation is thought to enhance survival in natural environments and during interaction with hosts. A robust colonizer of the human gastrointestinal tract, Escherichia coli Nissle 1917, is widely employed in probiotic therapy. In this study, we performed a genetic screen to identify genes that are involved in Nissle biofilm formation. We found that F1C fimbriae are required for biofilm formation on an inert surface. In addition, these structures are also important for adherence to epithelial cells and persistence in infant mouse colonization. The data suggest a possible connection between Nissle biofilm formation and the survival of this commensal within the host. Further study of the requirements for robust biofilm formation may improve the therapeutic efficacy of Nissle 1917.

  16. Sensitization of human aortic endothelial cells to lipopolysaccharide via regulation of Toll-like receptor 4 by bacterial fimbria-dependent invasion.

    PubMed

    Yumoto, Hiromichi; Chou, Hsin-Hua; Takahashi, Yusuke; Davey, Michael; Gibson, Frank C; Genco, Caroline A

    2005-12-01

    Toll-like receptors (TLRs) are differentially up-regulated in response to microbial infection and chronic inflammatory diseases such as atherosclerosis. Epidemiological data support the idea that periodontal disease may be a risk factor for acceleration of atherosclerosis. Porphyromonas gingivalis, the etiological agent of periodontal disease, invades endothelium, has been detected in human atheromatous tissue, and accelerates atheroma formation in apolipoprotein E-/- mice with concurrent induction of TLRs in the aorta. As endothelial cells can present antigen via TLRs and play an important role in the development of atherosclerosis, we examined TLR expression in human aortic endothelial cells (HAEC) cultured with wild-type P. gingivalis, a fimbria-deficient mutant, and purified antigens. We observed increased TLR expression in HAEC infected with wild-type P. gingivalis by fluorescence-activated cell sorter, but not with noninvasive, fimbria-deficient mutant or purified P. gingivalis antigens. Following a wild-type P. gingivalis challenge, functional TLR2 and TLR4 activation was assessed by subsequent stimulation with TLR agonists Staphylococcus aureus lipoteichoic acid (SLTA; TLR2 ligand) and Escherichia coli lipopolysaccharide (LPS; TLR4 ligand). Unchallenged HAEC failed to elicit monocyte chemoattractant protein 1 (MCP-1) in response to LPS or SLTA but did so when cultured with wild-type P. gingivalis. P. gingivalis-induced TLR2 and -4 expression on HAEC functionally reacted to SLTA and E. coli LPS as measured by a further increase in MCP-1 production. Furthermore, MCP-1 expression elicited by E. coli LPS was inhibitable with TLR4-specific antibody and polymyxin B. These results indicate that invasive P. gingivalis stimulates TLR expression on the surface of endothelium and these primed cells respond to defined TLR-specific ligands.

  17. Role of fimbriae expressed by nontypeable Haemophilus influenzae in pathogenesis of and protection against otitis media and relatedness of the fimbrin subunit to outer membrane protein A.

    PubMed Central

    Sirakova, T; Kolattukudy, P E; Murwin, D; Billy, J; Leake, E; Lim, D; DeMaria, T; Bakaletz, L

    1994-01-01

    Nontypeable Haemophilus influenzae is a primary pathogen in both acute otitis media (OM) and chronic OM, yet the pathogenesis of this disease is not fully understood. Although fimbriae have been observed on all clinical OM isolates examined to date, their role in pathogenesis remains unclear. Therefore, the gene which codes for the fimbrial subunit protein (fimbrin) in nontypeable H. influenzae 1128 was isolated, cloned, and sequenced. The nucleotide sequence of the fimbrin gene was found to contain an open reading frame of 1,077 bp which would encode a mature fimbrin protein consisting of 338 amino acid with a calculated molecular mass of 36.4 kDa. The translated amino acid sequence was found to be homologous with various OmpA proteins of other gram-negative bacteria, and algorithmic analysis predicted that this protein is organized as a coiled coil. To directly test whether fimbriae are involved in pathogenesis, the fimbrin gene was disrupted, and the biological consequences of disruption were absence of both expression of the fimbrial appendage and the specific immunogold labeling thereof with antisera directed against isolated fimbrial protein, reduced adherence to human oropharyngeal cells in vitro, augmented clearance from the tympanum post-transbullar inoculation, and significantly reduced induction of OM post-intranasal inoculation in a chinchilla model compared with the fimbriated parent strain. We additionally find that either passive immunization or active immunization against isolated fimbrial protein confers partial protection against transbullar challenge. A Western blot (immunoblot) indicated a degree of serological relatedness among fimbrin proteins of 15 nontypeable and type b isolates. These data suggest that fimbrin could be useful as a component of a vaccine to protect against OM. Images PMID:7909539

  18. Implementation of a Functional Observation Battery for the Assessment of Postoperative Well-being in Rats Subjected to Fimbria-Fornix Transection.

    PubMed

    Marschner, Linda; Wogensen, Elise; Mogensen, Jesper; Abelson, Klas

    2016-01-01

    The postoperative well-being of Wistar rats subjected to fimbria-fornix transections was assessed using a functional observational battery (FOB), including observations of relative body weight change, general condition, fur quality, body posture and movement, appetite, and pica behavior. Fimbria-fornix transected animals (FF), sham-operated animals (Sham), and two non-operated control groups with and without administration of buprenorphine (+BUP and -BUP, respectively) were observed twice daily for seven days after surgery. Buprenorphine (0.4 mg/kg) mixed in a nut paste for voluntary ingestion was supplied twice daily for 84 h to all groups except the -BUP control group starting on the day of surgery. Body weight was slightly decreased postoperatively in both surgical groups (FF and Sham) compared to control groups. The +BUP control group lost weight starting at day four after discontinuation of buprenorphine. Furthermore, the FF group exhibited significantly reduced general condition one day after surgery, with significantly affected body posture and movement for two days after surgery. In addition, mild pica behavior was observed in the FF group during the first postsurgical day. In conclusion, the FOB implemented in the present study appears to be a sensitive and accurate protocol for assessing animal well-being in the experimental setup applied. It is apparent that the FF transection is an invasive procedure that causes mildly adverse postoperative effects on the rats' well-being. We therefore recommend that this FOB is applied as a routine welfare monitoring protocol in experiments using mechanical central nervous system injury models, such as FF transection.

  19. Development and application of pathovar-specific monoclonal antibodies that recognize the lipopolysaccharide O antigen and the type IV fimbriae of Xanthomonas hyacinthi

    SciTech Connect

    Doorn, J. van; Ojanen-Reuhs, T.; Hollinger, T.C.; Reuhs, B.L.; Schots, A.; Boonekamp, P.M.; Oudega, B.

    1999-09-01

    The objective of this study was to develop a specific immunological diagnostic assay for yellow disease in hyacinths, using monoclonal antibodies (MAbs). Mice were immunized with a crude cell wall preparation (shear fraction) from Xanthomonas hyacinthi and with purified type IV fimbriae. Hybridomas were screened for a positive reaction with X. hyacinthi cells or fimbriae and for a negative reaction with X. translucens pv. graminis or Erwinia carotovora subsp. carotovora. Nine MAbs recognized fimbrial epitopes, as shown by immunoblotting, immunofluorescence, enzyme-linked immunosorbent assay (ELISA), and immunoelectron microscopy; however, three of these MAbs had weak cross-reactions with two X. translucens pathovars in immunoblotting experiments. Seven MAbs reacted with lipopolysaccharides and yielded a low-mobility ladder pattern on immunoblots. Subsequent analysis of MAb 2E5 showed that it specifically recognized an epitope on the O antigen, which was found to consist of rhamnose and fucose in a 2:1 molar ratio. The cross-reaction of MAb 2E5 with all X. hyacinthi strains tested showed that this O antigen is highly conserved within this species. MAb 1B10 also reacted with lipopolysaccharides. MAbs 2E5 and 1B10 were further tested in ELISA and immunoblotting experiments with cells and extracts from other pathogens. No cross-reaction was found with 27 other Xanthomonas pathovars tested or with 14 other bacterial species from other genera, such as Erwinia and Pseudomonas, indicating the high specificity of these antibodies. MAbs 2E5 and 1B10 were shown to be useful in ELISA for the detection of X. hyacinthi in infected hyacinths.

  20. Development and Application of Pathovar-Specific Monoclonal Antibodies That Recognize the Lipopolysaccharide O Antigen and the Type IV Fimbriae of Xanthomonas hyacinthi

    PubMed Central

    van Doorn, J.; Ojanen-Reuhs, T.; Hollinger, T. C.; Reuhs, B. L.; Schots, A.; Boonekamp, P. M.; Oudega, B.

    1999-01-01

    The objective of this study was to develop a specific immunological diagnostic assay for yellow disease in hyacinths, using monoclonal antibodies (MAbs). Mice were immunized with a crude cell wall preparation (shear fraction) from Xanthomonas hyacinthi and with purified type IV fimbriae. Hybridomas were screened for a positive reaction with X. hyacinthi cells or fimbriae and for a negative reaction with X. translucens pv. graminis or Erwinia carotovora subsp. carotovora. Nine MAbs recognized fimbrial epitopes, as shown by immunoblotting, immunofluorescence, enzyme-linked immunosorbent assay (ELISA), and immunoelectron microscopy; however, three of these MAbs had weak cross-reactions with two X. translucens pathovars in immunoblotting experiments. Seven MAbs reacted with lipopolysaccharides and yielded a low-mobility ladder pattern on immunoblots. Subsequent analysis of MAb 2E5 showed that it specifically recognized an epitope on the O antigen, which was found to consist of rhamnose and fucose in a 2:1 molar ratio. The cross-reaction of MAb 2E5 with all X. hyacinthi strains tested showed that this O antigen is highly conserved within this species. MAb 1B10 also reacted with lipopolysaccharides. MAbs 2E5 and 1B10 were further tested in ELISA and immunoblotting experiments with cells and extracts from other pathogens. No cross-reaction was found with 27 other Xanthomonas pathovars tested or with 14 other bacterial species from other genera, such as Erwinia and Pseudomonas, indicating the high specificity of these antibodies. MAbs 2E5 and 1B10 were shown to be useful in ELISA for the detection of X. hyacinthi in infected hyacinths. PMID:10473431

  1. The Radical Scavenging Activity and Cytotoxicity of Resveratrol, Orcinol and 4-Allylphenol and their Inhibitory Effects on Cox-2 Gene Expression and Nf-κb Activation in RAW264.7 Cells Stimulated with Porphyromonas gingivalis-fimbriae.

    PubMed

    Murakami, Yukio; Kawata, Akifumi; Ito, Shigeru; Katayama, Tadashi; Fujisawa, Seiichiro

    2015-01-01

    Resveratrol is a polyphenol with efficient anti-oxidative and anti-inflammatory activity. To clarify the molecular mechanism responsible for its anti-inflammatory action, we investigated the radical scavenging activity, cytotoxicity and anti-inflammatory activity of resveratrol and its related compounds, orcinol and 4-allylphenol. The radical scavenging activities of these compounds were determined by the DPPH (2,2'-diphenyl-1-picrylhydrazyl) assay and their cytotoxicities against RAW264.7 cells were determined using a cell-counting kit (CCK-8). The inhibitory effects of these compounds on cyclooxygenase-2 (Cox2) expression in RAW264.7 cells stimulated with Porphyromonas gingivalis (Pg) fimbriae were also determined using real-time polymerase chain reaction and western blot analysis, while inhibition of the fimbria-stimulated activation of nuclear factor-kappa B (Nf-κb) was evaluated using western blot analysis and enzyme-linked immunosorbent assay-like microwell colorimetric transcription factor activity assay, respectively. The quantum chemical parameters were calculated on the basis of the density function theory (DFT) BLYP/6-31G*. DPPH radical scavenging activity declined in the order resveratrol > orcinol > 4-allylphenol. The cytotoxicity of the compounds was in the order 4-allylphenol > resveratrol > orcinol. The inhibitory effect on Pg fimbria-stimulated Cox2 expression and Nf-κb activation was enhanced by resveratrol-alone. Resveratrol showed high electronegativity (χ) and softness (σ) values, as determined by quantum chemical calculations. Resveratrol exerts potent anti-inflammatory activity in RAW264.7 cells stimulated with Pg-fimbriae and may be applicable as a therapeutic agent for inflammatory periodontal disease as a manifestation of systemic disease. Copyright © 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  2. Characterization of F107 fimbriae of Escherichia coli 107/86, which causes edema disease in pigs, and nucleotide sequence of the F107 major fimbrial subunit gene, fedA.

    PubMed

    Imberechts, H; De Greve, H; Schlicker, C; Bouchet, H; Pohl, P; Charlier, G; Bertschinger, H; Wild, P; Vandekerckhove, J; Van Damme, J

    1992-05-01

    F107 fimbriae were isolated and purified from edema disease strain 107/86 of Escherichia coli. Plasmid pIH120 was constructed, which contains the gene cluster that codes for adhesive F107 fimbriae. The major fimbrial subunit gene, fedA, was sequenced. An open reading frame that codes for a protein with 170 amino acids, including a 21-amino-acid signal peptide, was found. The protein without the signal sequence has a calculated molecular mass of 15,099 Da. Construction of a nonsense mutation in the open reading frame of fedA abolished both fimbrial expression and the capacity to adhere to isolated porcine intestinal villi. In a screening of 28 reference edema disease strains and isolates from clinically ill piglets, fedA was detected in 24 cases (85.7%). In 20 (83.3%) of these 24 strains, fedA was found in association with Shiga-like toxin II variant genes, coding for the toxin that is characteristic for edema disease strains of E. coli. The fimbrial subunit gene was not detected in enterotoxigenic E. coli strains. Because of the capacity of E. coli HB101(pIH120) transformants to adhere to isolated porcine intestinal villi, the high prevalence of fedA in edema disease strains, and the high correlation with the Shiga-like toxin II variant toxin-encoding genes, we suggest that F107 fimbriae are an important virulence factor in edema disease strains of E. coli.

  3. The PprA-PprB two-component system activates CupE, the first non-archetypal Pseudomonas aeruginosa chaperone-usher pathway system assembling fimbriae.

    PubMed

    Giraud, Caroline; Bernard, Christophe S; Calderon, Virginie; Yang, Liang; Filloux, Alain; Molin, Søren; Fichant, Gwennaele; Bordi, Christophe; de Bentzmann, Sophie

    2011-03-01

    The opportunistic pathogen Pseudomonas aeruginosa has redundant molecular systems that contribute to its pathogenicity. Those assembling fimbrial structures promote complex organized community lifestyle. We characterized a new 5.8 kb genetic locus, cupE, that includes the conserved usher- and chaperone-encoding genes. This locus, widely conserved in different bacterial species, contains four additional genes encoding non-archetypal fimbrial subunits. We first evidenced that the cupE gene cluster was specifically expressed in biofilm conditions and was responsible for fibre assembly containing at least CupE1 protein, at the bacterial cell surface. These fimbriae not only played a significant role in the early stages (microcolony and macrocolony formation) but also in shaping 3D mushrooms during P. aeruginosa biofilm development. Using wide-genome transposon mutagenesis, we identified the PprAB two-component system (TCS) as a regulator of cupE expression, and further demonstrated the involvement of the PprAB TCS in direct CupE fimbrial assembly activation. Thus, this TCS represents a new regulatory element controlling the transition between planktonic and community lifestyles in P. aeruginosa.

  4. The Klebsiella pneumoniae YfgL (BamB) lipoprotein contributes to outer membrane protein biogenesis, type-1 fimbriae expression, anti-phagocytosis, and in vivo virulence.

    PubMed

    Hsieh, Pei-Fang; Hsu, Chun-Ru; Chen, Chun-Tang; Lin, Tzu-Lung; Wang, Jin-Town

    2016-07-03

    Klebsiella pneumoniae is an opportunistic pathogen that causes several kinds of infections, including pneumonia, bacteremia, urinary tract infection and community-acquired pyogenic liver abscess (PLA). Adhesion is the critical first step in the infection process. Our previous work demonstrated that the transcellular translocation is exploited by K. pneumoniae strains to migrate from the gut flora into other tissues, resulting in systemic infections. However, the initial stages of K. pneumoniae infection remain unclear. In this study, we demonstrated that a K. pneumoniae strain deleted for yfgL (bamB) exhibited reduced adherence to and invasion of host cells; changed biogenesis of major β-barrel outer membrane proteins; decreased transcriptional expression of type-1 fimbriae; and increased susceptibility to vancomycin and erythromycin. The yfgL deletion mutant also had reduced ability to against neutrophil phagocytosis; exhibited decreased induction of host IL-6 production; and was profoundly attenuated for virulence in a K. pneumoniae model of bacteremia. Thus, the K. pneumoniae YfgL lipoprotein mediates in outer membrane proteins biogenesis and is crucial for anti-phagocytosis and survival in vivo. These data provide a new insight for K. pneumoniae attachment and such knowledge could facilitate preventive therapies or alternative therapies against K. pneumoniae.

  5. A double, long polar fimbria mutant of Escherichia coli O157:H7 expresses Curli and exhibits reduced in vivo colonization.

    PubMed

    Lloyd, Sonja J; Ritchie, Jennifer M; Rojas-Lopez, Maricarmen; Blumentritt, Carla A; Popov, Vsevolod L; Greenwich, Jennifer L; Waldor, Matthew K; Torres, Alfredo G

    2012-03-01

    Escherichia coli O157:H7 causes food and waterborne enteric infections that can result in hemorrhagic colitis and life-threatening hemolytic uremic syndrome. Intimate adherence of the bacteria to intestinal epithelial cells is mediated by intimin, but E. coli O157:H7 also possess several other putative adhesins, including curli and two operons that encode long polar fimbriae (Lpf). To assess the importance of Lpf for intestinal colonization, we performed competition experiments between E. coli O157:H7 and an isogenic ΔlpfA1 ΔlpfA2 double mutant in the infant rabbit model. The mutant was outcompeted in the ileum, cecum, and midcolon, suggesting that Lpf contributes to intestinal colonization. In contrast, the ΔlpfA1 ΔlpfA2 mutant showed increased adherence to colonic epithelial cells in vitro. Transmission electron microscopy revealed curli-like structures on the surface of the ΔlpfA1 ΔlpfA2 mutant, and the presence of curli was confirmed by Congo red binding, immunogold-labeling electron microscopy, immunoblotting, and quantitative real-time reverse transcription-PCR (qRT-PCR) measuring csgA expression. However, deletion of csgA, which encodes the major curli subunit, does not appear to affect intestinal colonization. In addition to suggesting that Lpf can contribute to EHEC intestinal colonization, our observations indicate that the regulatory pathways governing the expression of Lpf and curli are interdependent.

  6. Crohn disease–associated adherent-invasive E. coli bacteria target mouse and human Peyer’s patches via long polar fimbriae

    PubMed Central

    Chassaing, Benoit; Rolhion, Nathalie; de Vallée, Amélie; Salim, Sa’ad Y.; Prorok-Hamon, Maelle; Neut, Christel; Campbell, Barry J.; Söderholm, Johan D.; Hugot, Jean-Pierre; Colombel, Jean-Frédéric; Darfeuille-Michaud, Arlette

    2011-01-01

    Crohn disease (CD) is a multifactorial disease in which an abnormal immune response in the gastrointestinal (GI) tract leads to chronic inflammation. The small intestine, particularly the ileum, of patients with CD is colonized by adherent-invasive E. coli (AIEC) — a pathogenic group of E. coli able to adhere to and invade intestinal epithelial cells. As the earliest inflammatory lesions are microscopic erosions of the epithelium lining the Peyer’s patches (PPs), we investigated the ability of AIEC bacteria to interact with PPs and the virulence factors involved. We found that AIEC bacteria could interact with mouse and human PPs via long polar fimbriae (LPF). An LPF-negative AIEC mutant was highly impaired in its ability to interact with mouse and human PPs and to translocate across monolayers of M cells, specialized epithelial cells at the surface of PPs. The prevalence of AIEC strains harboring the lpf operon was markedly higher in CD patients compared with controls. In addition, increased numbers of AIEC, but not LPF-deficient AIEC, bacteria were found interacting with PPs from Nod2–/– mice compared with WT mice. In conclusion, we have identified LPF as a key factor for AIEC to target PPs. This could be the missing link between AIEC colonization and the presence of early lesions in the PPs of CD patients. PMID:21339647

  7. Characterization of F107 fimbriae of Escherichia coli 107/86, which causes edema disease in pigs, and nucleotide sequence of the F107 major fimbrial subunit gene, fedA.

    PubMed Central

    Imberechts, H; De Greve, H; Schlicker, C; Bouchet, H; Pohl, P; Charlier, G; Bertschinger, H; Wild, P; Vandekerckhove, J; Van Damme, J

    1992-01-01

    F107 fimbriae were isolated and purified from edema disease strain 107/86 of Escherichia coli. Plasmid pIH120 was constructed, which contains the gene cluster that codes for adhesive F107 fimbriae. The major fimbrial subunit gene, fedA, was sequenced. An open reading frame that codes for a protein with 170 amino acids, including a 21-amino-acid signal peptide, was found. The protein without the signal sequence has a calculated molecular mass of 15,099 Da. Construction of a nonsense mutation in the open reading frame of fedA abolished both fimbrial expression and the capacity to adhere to isolated porcine intestinal villi. In a screening of 28 reference edema disease strains and isolates from clinically ill piglets, fedA was detected in 24 cases (85.7%). In 20 (83.3%) of these 24 strains, fedA was found in association with Shiga-like toxin II variant genes, coding for the toxin that is characteristic for edema disease strains of E. coli. The fimbrial subunit gene was not detected in enterotoxigenic E. coli strains. Because of the capacity of E. coli HB101(pIH120) transformants to adhere to isolated porcine intestinal villi, the high prevalence of fedA in edema disease strains, and the high correlation with the Shiga-like toxin II variant toxin-encoding genes, we suggest that F107 fimbriae are an important virulence factor in edema disease strains of E. coli. Images PMID:1348723

  8. Attenuated Shigella flexneri 2a ΔguaBA Strain CVD 1204 Expressing Enterotoxigenic Escherichia coli (ETEC) CS2 and CS3 Fimbriae as a Live Mucosal Vaccine against Shigella and ETEC Infection

    PubMed Central

    Altboum, Zeev; Barry, Eileen M.; Losonsky, Genevieve; Galen, James E.; Levine, Myron M.

    2001-01-01

    To construct a prototype hybrid vaccine against Shigella and enterotoxigenic Escherichia coli (ETEC), the genes encoding the production of ETEC CS2 and CS3 fimbriae were isolated and expressed in attenuated Shigella flexneri 2a guaBA strain CVD 1204. The CS2 cotA to -D genes, isolated from ETEC strain C91F, and the CS3 cstA to -H genes, subcloned from plasmid pCS100, were cloned into ∼15-copy-number-stabilized pGA1 behind the osmotically regulated ompC promoter, resulting in high expression of both fimbriae. Under nonselective in vitro growth conditions, pGA1-CS2 and pGA1-CS3 were stable in CVD 1204, exhibiting a plasmid loss of only approximately 1% per duplication. Expression of CS2 and CS3 reduced the invasiveness of Shigella for HeLa cells and slowed the intracellular growth rate. Guinea pigs immunized intranasally with CVD 1204(pGA1-CS2) or CVD 1204(pGA1-CS3), or with a mixture of these strains, developed secretory immunoglobulin A (IgA) in tears and serum IgG antibodies against Shigella lipopolysaccharide, CS2, and CS3 antigens. Moreover, the animals were protected against keratoconjunctivitis following conjunctival challenge with virulent S. flexneri 2a strain 2457T. Animals immunized with Shigella expressing CS2 or CS3 developed serum antibodies that agglutinated Shigella as well as an ETEC strain bearing the homologous fimbriae, whereas animals immunized with combined CVD 1204(pGA1-CS2) and CVD 1204(pGA1-CS3) developed antibodies that agglutinated all three test strains. These observations support the feasibility of a multivalent vaccine against shigellosis and ETEC diarrhea consisting of multiple Shigella live vectors expressing relevant ETEC antigens. PMID:11292735

  9. Diel behavioral rhythms in sablefish (Anoplopoma fimbria) and other benthic species, as recorded by the Deep-sea cabled observatories in Barkley canyon (NEPTUNE-Canada)

    NASA Astrophysics Data System (ADS)

    Doya, C.; Aguzzi, J.; Pardo, M.; Matabos, M.; Company, J. B.; Costa, C.; Mihaly, S.; Canals, M.

    2014-02-01

    Recent advances in cabled observatory video-imaging now enable faunal monitoring over extended periods of time. These platforms can be used to avoid biases in population and biodiversity assessments due to behavioral rhythms (i.e. massive population displacements). In this study we used video monitoring to examine the interplay between day-night and internal tidal cycles in regulating the behavior of sablefish (also referred to as black cod; Anoplopoma fimbria), hagfish (Eptatretus spp.) and crabs. We counted the number of animals in 50 s video-recordings taken at 30 min intervals with 3 NEPTUNE-Canada cameras located in Barkley canyon at approximately 1000 m depth (one in the axis and two on the wall of the canyon). Current data just above the seafloor was recorded as an indicator of the local internal tidal regime. Chi-Square periodogram analysis did not show significant (p < 0.05) day-night or tidal-based rhythms for the three species. The same analysis conducted for the sablefish (i.e. the most abundant) at each camera separately revealed different and significant (p < 0.05) 12- and 24-h based periods. Waveform analysis for these time series showed a temporal phase shift among cameras, suggesting diel displacements within the canyon axis. Our results highlight how some Deep-sea fish may present diel rhythmic displacements along canyons according to the day-night and internal tidal temporization. In this context, bathymetric networks of cabled video-stations can be an effective sampling tool to monitor this kind of behavior.

  10. The study of adhesive forces between the type-3 fimbriae of Klebsiella pneumoniae and collagen-coated surfaces by using optical tweezers

    NASA Astrophysics Data System (ADS)

    Chan, Chiahan; Fan, Chia-chieh; Huang, Ying-Jung; Peng, Hwei-Ling; Long, Hsu

    2004-10-01

    Adherence to host cells by a bacterial pathogen is a critical step for establishment of infection. It will contribute greatly to the understanding of bacterial pathogenesis by studying the biological force between a single pair of pathogen and host cell. In our experiment, we use a calibrated optical tweezers system to detach a single Klebsiella pneumoniae, the pathogen, from collagen, the host. By gradually increasing the laser power of the optical tweezers until the Klebsiella pneumoniae is detached from the collagen, we obtain the magnitude of the adhesive force between them. This happens when the adhesive force is barely equal to the trapping force provided by the optical tweezers at that specific laser power. This study is important because Klebsiella pneumoniae is an opportunistic pathogen which causes suppurative lesions, urinary and respiratory tract infections. It has been proved that type 3 fimbrial adhesin (mrkD) is strongly associated with the adherence of Klebsiella pneumoniae. Besides, four polymorphic mrkD alleles: namely, mrkDv1, v2, v3, and v4, are typed by using RFLP. In order to investigate the relationship between the structure and the function for each of these variants, DNA fragments encoding the major fimbrial proteins mrkA, mrkB, mrkC are expressed together with any of the four mrkD adhesins in E. coli JM109. Our study shows that the E. coli strain carrying the mrkDv3 fimbriae has the strongest binding activity. This suggests that mrkDv3 is a key factor that enhances the adherence of Klebsiella Pneumoniae to human body.

  11. Real-time quantitative reverse transcription-PCR analysis of expression stability of Aggregatibacter actinomycetemcomitans fimbria-associated gene in response to photodynamic therapy.

    PubMed

    Pourhajibagher, Maryam; Monzavi, Abbas; Chiniforush, Nasim; Monzavi, Mohammad Moein; Sobhani, Shaghayegh; Shahabi, Sima; Bahador, Abbas

    2017-06-01

    Aggregatibacter actinomycetemcomitans is an etiological agent of both chronic and aggressive periodontitis. Dissemination of A. actinomycetemcomitans from the oral cavity and initiation of systemic infections has led to new approaches for treatment being needed. In this study, a series of experiments presented investigated the effect of methylene blue (MB)-mediated antimicrobial photodynamic therapy (aPDT) on cell viability and expression of fimbria-associated gene (rcpA) in A. actinomycetemcomitans. To determine the dose-depended effects of aPDT, A. actinomycetemcomitans ATCC 33384 strain photosensitized with MB was irradiated with diode laser following bacterial viability measurements. Cell-surviving assay and expression ratio of rcpA were assessed by colony forming unit and real-time quantitative reverse transcription-PCR (qRT-PCR) assays, respectively. In the current study, MB-mediated aPDT using 100μg/mL showed significant reduction in A. actinomycetemcomitans growth when compared to the control (P<0.05). Sub-lethal dose of aPDT against A. actinomycetemcomitans was 25μg/mL MB at fluency of 93.75J/cm(2). Sub-lethal dose of aPDT could lead to about four-fold suppression of expression of rcpA. High doses of MB-mediated aPDT could potentially exhibit antimicrobial activity, and the expression of rcpA as an important virulence factor of this strain is reduced in cells surviving aPDT with MB. So, aPDT can be a valuable tool for the treatment of A. actinomycetemcomitans infections. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. The lpf Gene Cluster for Long Polar Fimbriae Is Not Involved in Adherence of Enteropathogenic Escherichia coli or Virulence of Citrobacter rodentium

    PubMed Central

    Tatsuno, Ichiro; Mundy, Rosanna; Frankel, Gad; Chong, Yuwen; Phillips, Alan D.; Torres, Alfredo G.; Kaper, James B.

    2006-01-01

    Using the enteropathogenic Escherichia coli (EPEC) genome sequence, we found that EPEC E2348/69 has an lpfABCDE gene cluster homologous (about 60% identical at the protein level) to the Salmonella long polar fimbria (LPF) operon. To determine whether this operon is essential for adherence, the lpfABCDE23 genes were deleted from EPEC strain E2348/69 by allelic exchange. Analysis of the resulting EPECΔlpfABCDE23 strain showed no change in adherence to HeLa cells or to human intestinal biopsy cells in the in vitro organ culture (IVOC) system compared to the wild type. Sera from volunteers experimentally infected with E2348/69 showed no antibody response to the major subunit protein, LpfA. These results suggested that the lpfE23 gene cluster is not necessary for EPEC adherence and attaching/effacing (A/E) lesion formation on human biopsy samples and is not expressed during human infection. We also identified an lpf gene cluster in Citrobacter rodentium strain ICC168 (lpfcr). A ΔlpfAcr mutant of ICC168 retained wild-type adherence and A/E lesion-forming activity on HeLa cells. C3H/HeJ mice were infected with a wild-type C. rodentium strain and its lpfAcr isogenic mutant. Both strains were recovered at high levels in stools, and there were no significant differences between the groups both in terms of the number of CFU/organ (colon and cecum) and in terms of the amount of hyperplasia, as measured by weight. Similar results were observed in a second mouse strain, C57BL/6. These data suggest that in addition to playing no apparent role in EPEC pathogenesis, lpfcr is not required for C. rodentium virulence in either the C3H/HeJ or C57BL/6 mouse model. PMID:16368980

  13. Effects of the dimeric PSD-95 inhibitor UCCB01-144 on functional recovery after fimbria-fornix transection in rats.

    PubMed

    Sommer, Jens Bak; Bach, Anders; Malá, Hana; Strømgaard, Kristian; Mogensen, Jesper; Pickering, Darryl S

    2017-10-01

    Pharmacological inhibition of PSD-95 is a promising therapeutic strategy in the treatment of stroke, and positive effects of monomeric and dimeric PSD-95 inhibitors have been reported in numerous studies. However, whether therapeutic effects will generalize to other types of acute brain injury such as traumatic brain injury (TBI), which has pathophysiological mechanisms in common with stroke, is currently uncertain. We have previously found a lack of neuroprotective effects of dimeric PSD-95 inhibitors in the controlled cortical impact model of TBI in rats. However, as no single animal model is currently able to mimic the complex and heterogeneous pathophysiology of TBI, it is necessary to assess treatment effects across a range of models. In this preliminary study we investigated the neuroprotective abilities of the dimeric PSD-95 inhibitor UCCB01-144 after fimbria-fornix (FF) transection in rats. UCCB01-144 or saline was injected into the lateral tail vein of rats immediately after sham surgery or FF-transection, and effects on spatial delayed alternation in a T-maze were assessed over a 28-day period. Task acquisition was significantly impaired in FF-transected animals, but there were no significant effects of UCCB01-144 on spatial delayed alternation after FF-transection or sham surgery, although decelerated learning curves were seen after treatment with UCCB01-144 in FF-transected animals. The results of the present study are consistent with previous research showing a lack of neuroprotective effects of PSD-95 inhibition in experimental models of TBI. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Contributions of EspA Filaments and Curli Fimbriae in Cellular Adherence and Biofilm Formation of Enterohemorrhagic Escherichia coli O157:H7

    PubMed Central

    Sharma, Vijay K.; Kudva, Indira T.; Bearson, Bradley L.; Stasko, Judith A.

    2016-01-01

    In Escherichia coli O157:H7 (O157), the filamentous structure of the type III secretion system is produced from the polymerization of the EspA protein. EspA filaments are essential for O157 adherence to epithelial cells. In previous studies, we demonstrated that O157 hha deletion mutants showed increased adherence to HEp-2 cells and produced abundant biofilms. Transcriptional analysis revealed increased expression of espA as well as the csgA gene, which encodes curli fimbriae that are essential for biofilm formation. In the present study, we constructed hha espA, hha csgA, and hha csgA espA deletion mutants to determine the relative importance of EspA and CsgA in O157 adherence to HEp-2 cells and biofilm formation. In vitro adherence assays, conducted at 37°C in a tissue culture medium containing 0.1% glucose, showed that HEp-2 cell adherence required EspA because hha espA and hha csgA espA mutants adhered to HEp-2 cells at higher levels only when complemented with an espA-expressing plasmid. Biofilm assays performed at 28°C in a medium lacking glucose showed dependency of biofilm formation on CsgA; however EspA was not produced under these conditions. Despite production of detectable levels of EspA at 37°C in media supplemented with 0.1% glucose, the biofilm formation occurred independent of EspA. These results indicate dependency of O157 adherence to epithelial cells on EspA filaments, while CsgA promoted biofilm formation under conditions mimicking those found in the environment (low temperature with nutrient limitations) and in the digestive tract of an host animal (higher temperature and low levels of glucose). PMID:26900701

  15. Role of F1C fimbriae, flagella, and secreted bacterial components in the inhibitory effect of probiotic Escherichia coli Nissle 1917 on atypical enteropathogenic E. coli infection.

    PubMed

    Kleta, Sylvia; Nordhoff, Marcel; Tedin, Karsten; Wieler, Lothar H; Kolenda, Rafal; Oswald, Sibylle; Oelschlaeger, Tobias A; Bleiss, Wilfried; Schierack, Peter

    2014-05-01

    Enteropathogenic Escherichia coli (EPEC) is recognized as an important intestinal pathogen that frequently causes acute and persistent diarrhea in humans and animals. The use of probiotic bacteria to prevent diarrhea is gaining increasing interest. The probiotic E. coli strain Nissle 1917 (EcN) is known to be effective in the treatment of several gastrointestinal disorders. While both in vitro and in vivo studies have described strong inhibitory effects of EcN on enteropathogenic bacteria, including pathogenic E. coli, the underlying molecular mechanisms remain largely unknown. In this study, we examined the inhibitory effect of EcN on infections of porcine intestinal epithelial cells with atypical enteropathogenic E. coli (aEPEC) with respect to single infection steps, including adhesion, microcolony formation, and the attaching and effacing phenotype. We show that EcN drastically reduced the infection efficiencies of aEPEC by inhibiting bacterial adhesion and growth of microcolonies, but not the attaching and effacing of adherent bacteria. The inhibitory effect correlated with EcN adhesion capacities and was predominantly mediated by F1C fimbriae, but also by H1 flagella, which served as bridges between EcN cells. Furthermore, EcN seemed to interfere with the initial adhesion of aEPEC to host cells by secretion of inhibitory components. These components do not appear to be specific to EcN, but we propose that the strong adhesion capacities enable EcN to secrete sufficient local concentrations of the inhibitory factors. The results of this study are consistent with a mode of action whereby EcN inhibits secretion of virulence-associated proteins of EPEC, but not their expression.

  16. The YfcO fimbriae gene enhances adherence and colonization abilities of avian pathogenic Escherichia coli in vivo and in vitro.

    PubMed

    Li, Yaxin; Wang, Haojin; Ren, Jianluan; Chen, Ling; Zhuge, Xiangkai; Hu, Lin; Li, Dezhi; Tang, Fang; Dai, Jianjun

    2016-11-01

    Chaperone-usher (CU) fimbriae, which are adhesive surface organelles found in many Gram-negative bacteria, mediate tissue tropism through the interaction of fimbrial adhesins with specific receptors expressed on the host cell surface. A CU fimbrial gene yfcO, was identified in avian pathogenic E. coli (APEC) strain DE205B via gene functional analysis. In this study, yfcO was found in 13.41% (11/82) of E. coli strains, including phylogenetic groups A, B1, B2 and D, with the highest percentage in group B2. The expression of yfcO in biofilm forming bacteria was significantly higher (P < 0.05) than that in the planktonic bacteria. A yfcO deletion mutant was constructed, and adherence to DF-1 chicken embryo fibroblast cells was analyzed in vitro. Compared to the wild-type (WT), adherence of the mutant to DF-1 cells was significantly decreased (P < 0.01). The mutant bacterial loads in the heart, brain and liver were significantly lower (P < 0.05) than those of the WT strain. Resistance of the mutant to acidic (acetic, pH 4.0, 20 min) and high osmolarity (2.5 M NaCl, 1 h) stress conditions decreased by 51.28% (P < 0.001) and 80.34% (P < 0.01), respectively. These results suggest that yfcO contributes to APEC virulence through bacterial adherence to host tissues.

  17. Comparative inhibitory effects of magnolol, honokiol, eugenol and bis-eugenol on cyclooxygenase-2 expression and nuclear factor-kappa B activation in RAW264.7 macrophage-like cells stimulated with fimbriae of Porphyromonas gingivalis.

    PubMed

    Murakami, Yukio; Kawata, Akifumi; Seki, Yuya; Koh, Teho; Yuhara, Kenji; Maruyama, Takehisa; Machino, Mamoru; Ito, Shigeru; Kadoma, Yoshinori; Fujisawa, Seiichiro

    2012-01-01

    The anti-inflammatory activity of magnolol and related compounds is currently a focus of interest. In the present study, the inhibitory effects of these compounds on cyclooxygenase (COX-2) expression and nuclear factor-kappa B (NF-κB) activation were investigated in RAW264.7 macrophage-like cells stimulated with the fimbriae of Porphyromonas gingivalis, an oral anaerobe. The cytotoxicity of magnolol, honokiol, eugenol and bis-eugenol against RAW264.7 cells was determined using a cell counting kit (CCK-8). The regulatory effect of these compounds on the expression of COX-2 mRNA, stimulated by exposure to the fimbriae was investigated by real-time polymerase chain reaction (PCR). NF-κB activation was evaluated by enzyme-linked immunosorbent assay (ELISA)-like microwell colorimetric transcription factor activity assay (Trans-AM) and western blot analysis. The radical-scavenging activity was determined using the induction period method in the methyl methacrylate-azobisisobutyronitrile (AIBN) polymerization system under nearly anaerobic conditions. The phenolic bond dissociation enthalpy (BDE) and orbital energy were calculated at the density functional theory (DFT) B3LYP/6-31G* level. The cytotoxicity against RAW264.7 cells declined in the order bis-eugenol>eugenol> honokiol>magnolol, whereas the radical-scavenging activity declined in the order honokiol, bis-eugenol>magnolol> eugenol. Magnolol and honokiol significantly inhibited the fimbria-induced expression of COX-2 at non-cytotoxic concentrations. Both the fimbria-stimulated binding of NF-κB to its consensus sequence and phosphorylation-dependent proteolysis of inhibitor κB-α were markedly inhibited by magnilol and honokiol, whereas eugenol and bis-eugenol did not inhibit COX-2 expression and NF-κB activation. Magnolol and honokiol possessed a high electronegativity (χ) value. Magnolol and honokiol exhibit antioxidative activity, low cytotoxicity, and anti-inflammatory activity. These compounds may be

  18. Manual segmentation of the fornix, fimbria, and alveus on high-resolution 3T MRI: Application via fully-automated mapping of the human memory circuit white and grey matter in healthy and pathological aging.

    PubMed

    Amaral, Robert S C; Park, Min Tae M; Devenyi, Gabriel A; Lynn, Vivian; Pipitone, Jon; Winterburn, Julie; Chavez, Sofia; Schira, Mark; Lobaugh, Nancy J; Voineskos, Aristotle N; Pruessner, Jens C; Chakravarty, M Mallar

    2016-10-18

    Recently, much attention has been focused on the definition and structure of the hippocampus and its subfields, while the projections from the hippocampus have been relatively understudied. Here, we derive a reliable protocol for manual segmentation of hippocampal white matter regions (alveus, fimbria, and fornix) using high-resolution magnetic resonance images that are complementary to our previous definitions of the hippocampal subfields, both of which are freely available at https://github.com/cobralab/atlases. Our segmentation methods demonstrated high inter- and intra-rater reliability, were validated as inputs in automated segmentation, and were used to analyze the trajectory of these regions in both healthy aging (OASIS), and Alzheimer's disease (AD) and mild cognitive impairment (MCI; using ADNI). We observed significant bilateral decreases in the fornix in healthy aging while the alveus and cornu ammonis (CA) 1 were well preserved (all p's<0.006). MCI and AD demonstrated significant decreases in fimbriae and fornices. Many hippocampal subfields exhibited decreased volume in both MCI and AD, yet no significant differences were found between MCI and AD cohorts themselves. Our results suggest a neuroprotective or compensatory role for the alveus and CA1 in healthy aging and suggest that an improved understanding of the volumetric trajectories of these structures is required.

  19. Inhibitory effects of tocopherols on expression of the cyclooxygenase-2 gene in RAW264.7 cells stimulated by lipopolysaccharide, tumor necrosis factor-α or Porphyromonas gingivalis fimbriae.

    PubMed

    Murakami, Yukio; Kawata, Akifumi; Koh, Teho; Seki, Yuya; Tamura, Seiko; Katayama, Tadashi; Fujisawa, Seiichiro

    2013-01-01

    Tocopherols, which include α-, β-, γ-, and δ-tocopherol, protect cells against harmful free radicals and play an important role in preventing many human diseases such as cancer, inflammatory disorders, and ageing itself. However, the causal relationships between periodontal or oral chronic diseases and tocopherols have not been sufficiently studied. The present study investigated the inhibitory effects of these compounds on the expression of cyclooxygenase-2 (COX2) mRNA in RAW264.7 cells stimulated with lipopolysaccharide (LPS), tumor necrosis factor-α (TNFα) or fimbriae of Poryphyromonas gingivalis (Pg), an oral anaerobe. The cytotoxicity (EC₅₀) of tocopherols toward RAW cells was determined using a cell counting kit (CCK-8). The regulatory effect of these compounds on the expression of COX2 mRNA stimulated with LPS, TNFα or Pg fimbriae was investigated using real-time polymerase chain reaction (PCR). Each tocopherol had similarly low cytotoxicity. COX2 gene expression in RAW cells after exposure to the three different macrophage activators was inhibited by the tocopherols (p<0.01). Compared to α-tocopherol, β-, γ- and δ-tocopherol exhibited greater inhibitory effects (p<0.05). Tocopherols exhibit anti-inflammatory activity, and β-, γ- and δ-tocopherol have particularly more potent anti-inflammatory activity than α-tocopherol. Tocopherols may have potential utility for prevention of periodontal and chronic oral diseases.

  20. Bone mesenchymal stem cells for gene transfer of NGF to the adult rat brain: rescue the NGFR p75 positive neurons from fimbria-fornix lesion-induced degeneration.

    PubMed

    Leng, Shuilong; He, Junfeng; Fan, Wenguo; Cheng, Shaowu; Long, Dahong; He, Hongwen

    2008-12-31

    The study was to evaluate the therapeutic benefit of transplanted bone mesenchymal stem cells (BMSCs) transfected never growth factor (NGF) gene and GFP gene (as a reporter gene), in treating the rat with fimbria-fornix lesion. After transduction of NGF gene via recombinant retroviral vectors into the rat BMSCs, BMSCs were therefore transformed into the GFP-NGF positive BMSCs, nearly 100% of BMSCs expressed NGF, and then transplanted into basal forebrain of rat with fimbria-fornix lesion. After 2 weeks post-transplantation, the GFP-NGF positive BMSCs survive and fuse in vivo with astroglia or NGFR p75 positive neurons in the basal forebrain, no evidence of transdifferentiation was observed in this study. The number of NGFR p75 positive neurons in basal forebrain of NGF group was significantly higher than those of the void plasmid group (p < 0.05) or the PBS group (p < 0.01). These results indicate that the GFP-NGF positive BMSCs provide, by way of paracrine, NGF that effectively perform the functions of neuroprotection, which cell fusion may be also contribute to.

  1. The long polar fimbriae genes identified in Shiga toxin-producing Escherichia coli are present in other diarrheagenic E. coli and in the standard E. coli collection of reference (ECOR) strains.

    PubMed

    Toma, Claudia; Higa, Naomi; Iyoda, Sunao; Rivas, Marta; Iwanaga, Masaaki

    2006-03-01

    Long polar fimbriae (LPF) are related to type I fimbriae in genetic organization and were first identified in Salmonella enterica serovar Typhimurium. Four lpfA genetic variants designated lpfA(O157/OI-141), lpfA(O157/OI-154), lpfA(O26) and lpfA(O113) have been identified in Shiga toxin-producing Escherichia coli (STEC). In this study, PCR was employed to determine the distribution of STEC-lpfAs in enteropathogenic, enteroaggregative, enterotoxigenic and enteroinvasive E. coli (EPEC, EAEC, ETEC and EIEC) and in the standard E. coli collection of reference (ECOR). Among the 97 diarrheagenic strains from our collection, only 2 EPEC strains of serotypes O55:H7 and O119:NM were positive for both lpfA(O157/OI-141) and lpfA(O157/OI-154). lpfA(O157/OI-141) was also positive in 1 of 25 ETEC strains. lpfA(O113) was present in 51 of 97 strains and lpfA(O26) in 13 of 97 strains of diverse diarrheagenic categories. STEC-lpfAs were also present in non-pathogenic ECOR strains of all phylogenetic groups. This study showed that the lpfA genes identified in the genome of STEC strains are not specific to this category. Our results suggest that there is a relationship between the lpfA variant and the phylogenetic group.

  2. A video demonstration of preserved piloting by scent tracking but impaired dead reckoning after fimbria-fornix lesions in the rat.

    PubMed

    Whishaw, Ian Q; Gorny, Boguslaw P

    2009-04-24

    ; Martin et al., 1997; Maaswinkel and Whishaw, 1999). The objective of the present video demonstrations was to solve the problem of cue specification in order to examine the relative contribution of the hippocampus in the use of these strategies. The rats were trained in a new task in which they followed linear or polygon scented trails to obtain a large food pellet hidden on an open field. Because rats have a proclivity to carry the food back to the refuge, accuracy and the cues used to return to the home base were dependent variables (Whishaw and Tomie, 1997). To force an animal to use a a dead reckoning strategy to reach its refuge with the food, the rats were tested when blindfolded or under infrared light, a spectral wavelength in which they cannot see, and in some experiments the scent trail was additionally removed once an animal reached the food. To examine the relative contribution of the hippocampus, fimbria-fornix (FF) lesions, which disrupt information flow in the hippocampal formation (Bland, 1986), impair memory (Gaffan and Gaffan, 1991), and produce spatial deficits (Whishaw and Jarrard, 1995), were used.

  3. Afa/Dr-expressing, diffusely adhering Escherichia coli strain C1845 triggers F1845 fimbria-dependent phosphatidylserine externalization on neutrophil-like differentiated PLB-985 cells through an apoptosis-independent mechanism.

    PubMed

    Sémiramoth, Nicolas; Gleizes, Aude; Turbica, Isabelle; Sandré, Catherine; Marin-Esteban, Viviana; Gorges, Roseline; Servin, Alain; Chollet-Martin, Sylvie

    2010-07-01

    The enterovirulent Escherichia coli strains potentially involved in inflammatory bowel diseases include diffusely adherent strains expressing Afa/Dr fimbriae (Afa/Dr DAEC). We have previously observed type 1 pilus-mediated interleukin-8 (IL-8) hyperproduction in infected neutrophils. As pathogen induction of host cell death programs and clearance of apoptotic infected cells are crucial for innate immune system homeostasis and host integrity, we examined modulation of neutrophil cell death by Afa/Dr DAEC. Using the human PLB-985 cell line differentiated into fully mature neutrophils, we found that the wild-type enterovirulent E. coli strain C1845 and the recombinant strain DH5alpha/pF1845 (expressing the fimbrial adhesin F1845) similarly induced time-dependent phosphatidylserine (PS) externalization, suggesting a major specific role of this virulence factor. Using small interfering RNA (siRNA) decay-accelerating factor (DAF)-transfected PLB-985 cells, we then showed that this PS externalization was triggered in part by glycosylphosphatidylinositol (GPI)-anchored DAF receptor engagement (leading to tyrosine kinase and protein kinase C activation) and that it required cytoskeleton and lipid raft architectural integrity. PS externalization under these conditions was not dependent on caspases, mitochondria, lysosomes, or reactive oxygen or nitrogen species. F1845-mediated PS externalization was sufficient to enable macrophage engulfment of infected differentiated PLB-985 cells. These findings provide new insights into the neutrophil response to Afa/Dr DAEC infection and highlight a new role for F1845 fimbriae. Interestingly, although apoptosis pathways were not engaged, C1845-infected PLB-985 cells displayed enhanced removal by macrophages, a process that may participate in the resolution of Afa/Dr DAEC infection and related inflammation.

  4. Radical-scavenging and Anti-inflammatory Activity of Quercetin and Related Compounds and Their Combinations Against RAW264.7 Cells Stimulated with Porphyromonas gingivalis Fimbriae. Relationships between Anti-inflammatory Activity and Quantum Chemical Parameters.

    PubMed

    Murakami, Yukio; Kawata, Akifumi; Ito, Shigeru; Katayama, Tadashi; Fujisawa, Seiichiro

    2015-01-01

    The flavonoid quercetin exerts significant anti-inflammatory activity against chronic infections, including periodontal disease. However, it is unclear whether combination of quercetin with other flavonoids enhances antioxidant and anti-inflammatory activity. To clarify the molecular mechanism responsible for the anti-inflammatory activity of quercetin, we investigated the antioxidant, cytotoxicity and anti-inflammatory activity of quercetin and its related compounds, catechin and epicatechin, and their combinations. Radical-scavenging activities were determined by the 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay, and cytotoxicity against RAW264.7 cells was determined using a cell counting kit (CCK-8). The inhibitory effects of these compounds on the mRNA expression of cyclooxygenase-2 (Cox2), tumor necrosis factor-alpha (Tnfα) and nitric oxide synthase 2 (Nos2), in RAW264.7 cells stimulated with Porphyromonas gingivalis (Pg) fimbriae, was also determined using real-time polymerase chain reaction analysis. The phenolic O-H bond dissociation enthalpy (BDE) and quantum chemical parameters were calculated on the basis of density function theory (DFT) BLYP/6-31G*. The DPPH(•) radical-scavenging activity (EC50) of quercetin, catechin and epicatechin was 5.5, 7.7 and 6.2 μM, respectively, whereas the cytotoxicity (LC50) was 4.45, 4.80 and 4.95 mM, respectively. Quercetin had slightly higher cytotoxicity and anti-DPPH(•) activity than catechin and epicatechin. The BDE for the three flavonoids at the 4'-OH in the B ring, which is the initial active site, was about 75 kcal/mol. Furthermore, various combinations of quercetin with catechin or epicatechin exerted an antagonistic effect on anti-DPPH(•) activity. Gene expression of Cox2, Tnfα and Nos2 stimulated by exposure to Pg-fimbriae was markedly suppressed by quercetin, but was not modulated by its combination with epicatechin. The 50% inhibitory concentration of quercetin for Cox2 expression was approximately 10

  5. Localization of a domain in the FimH adhesin of Escherichia coli type 1 fimbriae capable of receptor recognition and use of a domain-specific antibody to confer protection against experimental urinary tract infection.

    PubMed Central

    Thankavel, K; Madison, B; Ikeda, T; Malaviya, R; Shah, A H; Arumugam, P M; Abraham, S N

    1997-01-01

    The FimH subunit of type 1-fimbriated Escherichia coli has been implicated as an important determinant of bacterial adherence and colonization of the urinary tract. Here, we sought to localize the functionally important domain(s) within the FimH molecule and to determine if antibodies against this domain would block adherence of type 1-fimbriated E. coli to the bladder mucosa in situ and in vivo in an established mouse model of cystitis. We generated translational fusion proteins of disparate regions of the FimH molecule with an affinity tag MalE, and tested each of the fusion products in vitro for functional activity. The minimum region responsible for binding mouse bladder epithelial cells and a soluble mannoprotein, horseradish peroxidase, was contained within residues 1-100 of the FimH molecule. We validated and extended these findings by demonstrating that antibodies directed at the putative binding region of FimH or at synthetic peptides corresponding to epitopes within the binding domain could specifically block type 1 fimbriae-mediated bacterial adherence to bladder epithelial cells in situ and yeast cells in vitro. Next, we compared the ability of mice passively immunized intraperitoneally with antisera raised against residues 1-25 and 253-264 of FimH or 1-13 of FimA to resist bladder colonization in vivo after intravesicular challenge with type 1-fimbriated E. coli. Only the antibody directed at the putative binding region of FimH (anti- s-FimH1-25) significantly reduced E. coli bladder infections in the experimental mouse model of urinary tract infections. Similar results were obtained when the mice were actively immunized with synthetic peptides corresponding to residues 1-25 and 253-264 of FimH or 1-13 of FimA. The mechanism of protection was attributed, at least in part, to inhibition of bacterial adherence to the bladder surface by s-FimH1-25-specific antibody molecules that had filtered through the kidneys into the urine. The level of Fim

  6. “Kinin danger signals proteolytically released by gingipain induce fimbriae-specific IFN-γ and IL-17-producing T cells in mice infected intramucosally with Porphyromonas gingivalis”

    PubMed Central

    Monteiro, Ana Carolina; Scovino, Aline; Raposo, Susane; Gaze, Vinicius Mussa; Cruz, Catia; Svensjö, Erik; Narciso, Marcelo Sampaio; Colombo, Ana Paula; Pesquero, João B.; Feres-Filho, Eduardo; Nguyen, Ky-Anh; Sroka, Aneta; Potempa, Jan; Scharfstein, Julio

    2009-01-01

    Porphyromonas gingivalis, a gram-negative bacterium that causes periodontitis, activates the kinin system via the cysteine protease R-gingipain. Using a model of buccal infection based on P. gingivalis inoculation in the anterior mandibular vestibule, here we studied whether kinins released by gingipain may link mucosal inflammation to T cell-dependent immunity through the activation of bradykinin B2 receptors (B2R). Our data show that P. gingivalis W83 (WT), but not gingipain deficient mutant or WT bacteria pretreated with gingipain inhibitors, elicited buccal edema and gingivitis in Balb/C or C57BL/6 mice. Studies in TLR2−/−, B2R−/− and neutrophil-depleted C57Bl/6 mice revealed that P. gingivalis induced edema through the sequential activation of TLR2/neutrophils, with the initial plasma leakage being amplified by gingipain-dependent release of vasoactive kinins from plasma-borne kininogens. We then used fimbriae (Fim) Ag as a read-out to verify if activation of the TLR2>PMN>B2R axis at early-stages of mucosal infection had impact on adaptive immunity. Analyzes of T cell recall responses indicated that gingipain drives B2R-dependent generation of IFN-γ-producing Fim T cells in submandibular draining LNs of Balb/C and C57BL/6 mice while IL-17-producing Fim T cells were generated only in Balb/C mice. In summary, our studies suggest that two virulence factors, LPS (an atypical TLR2 ligand) and gingipain, forges a trans-cellular cross-talk between TLR2/B2R, thus forming an innate axis that guides the development of Fim-specific T cells in mice challenged intrabuccally by P. gingivalis. Ongoing research may clarify if kinin-driven modulation of T cell responses may also influence the severity of chronic periodontitis. PMID:19687097

  7. Colistin inhibition of mannose-resistant haemagglutination by K88-positive and K99-positive escherichia coli strains. A preliminary report.

    PubMed

    Søgaard, H; Larsen, J L; Christensen, S

    1983-03-01

    Two enteropathogenic E. coli strains isolated from a calf and a piglet succumbed to diarrhoea were studied. The bovine strain carried K99-antigen and the porcine strain was K88-positive. Both strains agglutinated pig erythrocytes in the presence of D-mannose. In the test a bacterial cell density of 3 x 10(9) per ml and doubling dilutions hereof were used. The haemagglutination titres were 16 and 128, respectively. When the bacteria were exposed to colistin before mixing with the red cells, haemagglutination was inhibited completely with 1.0 and 0.5 microgram/mg of colistin. At a colistin concentration of 0.25 microgram/ml (1/4-1/2 of the MIC's) the titres were lowered by a factor of 16-32.

  8. A Tripartite Fusion, FaeG-FedF-LT192A2:B, of Enterotoxigenic Escherichia coli (ETEC) Elicits Antibodies That Neutralize Cholera Toxin, Inhibit Adherence of K88 (F4) and F18 Fimbriae, and Protect Pigs against K88ac/Heat-Labile Toxin Infection ▿

    PubMed Central

    Ruan, Xiaosai; Liu, Mei; Casey, Thomas A.; Zhang, Weiping

    2011-01-01

    Enterotoxigenic Escherichia coli (ETEC) strains expressing K88 (F4) or F18 fimbriae and heat-labile (LT) and/or heat-stable (ST) toxins are the major cause of diarrhea in young pigs. Effective vaccines inducing antiadhesin (anti-K88 and anti-F18) and antitoxin (anti-LT and anti-ST) immunity would provide broad protection to young pigs against ETEC. In this study, we genetically fused nucleotides coding for peptides from K88ac major subunit FaeG, F18 minor subunit FedF, and LT toxoid (LT192) A2 and B subunits for a tripartite adhesin-adhesin-toxoid fusion (FaeG-FedF-LT192A2:B). This fusion was used for immunizations in mice and pigs to assess the induction of antiadhesin and antitoxin antibodies. In addition, protection by the elicited antiadhesin and antitoxin antibodies against a porcine ETEC strain was evaluated in a gnotobiotic piglet challenge model. The data showed that this FaeG-FedF-LT192A2:B fusion elicited anti-K88, anti-F18, and anti-LT antibodies in immunized mice and pigs. In addition, the anti-porcine antibodies elicited neutralized cholera toxin and inhibited adherence against both K88 and F18 fimbriae. Moreover, immunized piglets were protected when challenged with ETEC strain 30302 (K88ac/LT/STb) and did not develop clinical disease. In contrast, all control nonvaccinated piglets developed severe diarrhea and dehydration after being challenged with the same ETEC strain. This study clearly demonstrated that this FaeG-FedF-LT192A2:B fusion antigen elicited antibodies that neutralized LT toxin and inhibited the adherence of K88 and F18 fimbrial E. coli strains and that this fusion could serve as an antigen for vaccines against porcine ETEC diarrhea. In addition, the adhesin-toxoid fusion approach used in this study may provide important information for developing effective vaccines against human ETEC diarrhea. PMID:21813665

  9. Hemolysin and K antigens in relation to serotype and hemagglutination type of Escherichia coli isolated from extraintestinal infections.

    PubMed Central

    Evans, D J; Evans, D G; Höhne, C; Noble, M A; Haldane, E V; Lior, H; Young, L S

    1981-01-01

    Escherichia coli isolated from cases of bacteremia and from a variety of urinary tract infections were characterized according to serotype (O:H antigenicity), K type (possession of K1, K2, K3, K12, or K13), hemagglutination (HA) type, and production of beta-hemolysin. Results obtained with the bacteremia and urinary tract infection isolates were similar except for more hemolytic isolated from urine than from blood (42 versus 29%) and more K1+ isolates from blood than from urine (50 versus 29%). A close correlation was found between Ha type VI (production of fimbriae which mediate mannose-resistant HA of human and African green monkey erythrocytes) and the production of hemolysin or K1 capsular antigen or both. Most (95 of 98, or 95%) of the HA type VI+ blood isolates and most (146 of 164, or 89%) of the HA type VI+ urine isolates produced hemolysin or K1 or both, in contrast to 22 and 26%, respectively, of those belonging to HA types other than HA type VI. Also, 76% of all hemolytic and 70% of all K1+ isolates belonged to HA type VI. Remarkably few of the HA type VI+ isolates (13%) and even fewer of the HA type VI- isolates (3%) produced both K1 and hemolysin; these belonged mainly to serotypes O16:H6, O18:H7 and O2:H4. Other major serogroups were usually K1+/hemolysin- (O1, O7) or K1-/hemolysin+ (O2, O4, O6). At least 74% (262 of 351) and possibly as many as 83% (293 of 351) of those isolates which produced mannose-resistant HA of human erythrocytes were classified as HA type VI+; 31 isolates produced mannose-resistant HA with all erythrocytes tested. Taking serogroup and serotype into consideration, we conclude that the E. coli fimbrial hemagglutinin(s) responsible for the HA type VI phenotype will prove to be the same as the virulence-associated mannose-resistant adhesins of uropathogenic E. coli which other investigators have characterized as unique fimbrial antigens detectable by mannose-resistant HA of human erythrocytes. PMID:7007421

  10. Increased biofilm formation ability in Klebsiella pneumoniae after short-term exposure to a simulated microgravity environment.

    PubMed

    Wang, Haili; Yan, Yanfeng; Rong, Dan; Wang, Jing; Wang, Hongduo; Liu, Zizhong; Wang, Jiaping; Yang, Ruifu; Han, Yanping

    2016-10-01

    Biofilm formation is closely related to the pathogenetic processes of Klebsiella pneumoniae, which frequently causes infections in immunocompromised individuals. The immune system of astronauts is compromised in spaceflight. Accordingly, K. pneumoniae, which used to be isolated from orbiting spacecraft and astronauts, poses potential threats to the health of astronauts and mission security. Microgravity is a key environmental cue during spaceflight. Therefore, determining its effects on bacterial biofilm formation is necessary. In this study, K. pneumoniae ATCC BAA-1705 was exposed to a simulated microgravity (SMG) environment. K. pneumoniae grown under SMG formed thicker biofilms compared with those under normal gravity (NG) control after 2 weeks of subculture. Two indicative dyes (i.e., Congo red and calcofluor) specifically binding to cellulose fibers and/or fimbriae were utilized to reconfirm the enhanced biofilm formation ability of K. pneumoniae grown under SMG. Further analysis showed that the biofilms formed by SMG-treated K. pneumoniae were susceptible to cellulase digestion. Yeast cells mannose-resistant agglutination by K. pneumoniae type 3 fimbriae was more obvious in the SMG group, which suggests that cellulose production and type 3 fimbriae expression in K. pneumoniae were both enhanced under the SMG condition. Transcriptomic analysis showed that 171 genes belonging to 15 functional categories were dysregulated in this organism exposed to the SMG conditions compared with those in the NG group, where the genes responsible for the type 3 fimbriae (mrkABCDF) and its regulator (mrkH) were upregulated.

  11. Pathogenicity of Vietnamese enterotoxigenic Escherichia coli strains in colostrum-deprived one-day-old piglets.

    PubMed

    Do, T N; Wilkie, I; Driesen, S J; Fahy, V A; Trott, D J

    2006-03-01

    Preweaning colibacillosis is a major cause of economic loss to the swine industry in Vietnam. The aim of this study was to examine the enteropathogenicity of representative enterotoxigenic Escherichia coli (ETEC) strains obtained during an earlier epidemiologic survey conducted in five provinces in North Vietnam. This included isolates belonging to serotype O8 that produced heat-stable and heat-labile enterotoxins but did not produce any of the recognized fimbriae (F4, F5, F6, F41, F18). In vitro hemagglutination (unique mannose-resistant hemagglutination activity with guinea pig, sheep, human, and chicken red blood cells at 37 degrees C, but not at 18 degrees C) and enterocyte brush border attachment assays suggested that the F- ETEC strains produced an unidentified colonization factor that promoted adherence to the intestinal epithelium. Colostrum-deprived 1-day-old piglets challenged with an F- strain (1-2 x 10(9) bacteria) developed acute watery diarrhea within 4 hours of inoculation and suffered up to 20% weight loss, with comparable severity to piglets challenged with conventional F4 and F5 strains. At necropsy, viable counts and histopathologic examination of intestinal sections demonstrated colonization of the duodenum, jejunum, and ileum by F4-positive strains. In comparison, the F- and F5-positive strains attached exclusively to the ileum. Transmission electron micrographs of negatively stained F- cells grown at 37 degrees C demonstrated the presence of fimbriae. These results confirm the presence of a potentially new pathogenic ETEC fimbrial type in piggeries in Vietnam, with a unique hemagglutination property and attachment characteristics similar to ETEC bearing F5 fimbriae.

  12. Isolation and characterization of Escherichia coli pili from diverse clinical sources.

    PubMed Central

    Salit, I E; Vavougios, J; Hofmann, T

    1983-01-01

    Bacteria which attach to different mucous membranes should have differing specificities of adherence in vitro. Human Escherichia coli isolates from blood and urine (pathogens) and from stool and throat (commensals) were characterized as to the patterns of hemagglutination (HA), as well as the structure and function of their pili. Bacterial HA was done in microtiter plates and on slides after bacterial growth in broth or agar. Human erythrocytes were agglutinated by 95% of the pathogens and 65 to 70% of the commensals grown in broth or agar. Mannose-resistant HA was characteristically caused by pathogens, and commensals characteristically caused mannose-sensitive HA of guinea pig cells. Strains often had both mannose-resistant and mannose-sensitive reactions, or even a mannose-paradoxical reaction. Pathogens more often caused HA, but titers were lower than those for commensals. Slide HA was less sensitive than the microtiter method. All isolates were piliated. Commensals also had more pili than pathogens when grown in broth (117.8 versus 38.3 pili per bacterium), but pathogens had more pili after growth on agar (32.1 versus 8.1 pili per bacterium). Isolates causing high-titer HA had large numbers of pili (greater than 85 pili per bacterium), but some well-piliated strains were non-hemagglutinating. Pili were purified from seven E. coli strains from different sites of isolation and with different erythrocyte-binding specificity. Pili usually migrated as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. However, more than one type of pilus could be copurified from some strains since there were two or more bands after separation in octyl-glucoside and two different amino terminal sequences. Protein sequencing was done on five different pili: four resembled type 1 pili and one was a P fimbria. The type 1-like pili (strains 2239 and 9353) had an initial variable sequence of 1 to 5 residues, followed by a common region of 21 residues. The P

  13. Virulence factors in Escherichia coli urinary tract infection.

    PubMed Central

    Johnson, J R

    1991-01-01

    Uropathogenic strains of Escherichia coli are characterized by the expression of distinctive bacterial properties, products, or structures referred to as virulence factors because they help the organism overcome host defenses and colonize or invade the urinary tract. Virulence factors of recognized importance in the pathogenesis of urinary tract infection (UTI) include adhesins (P fimbriae, certain other mannose-resistant adhesins, and type 1 fimbriae), the aerobactin system, hemolysin, K capsule, and resistance to serum killing. This review summarizes the virtual explosion of information regarding the epidemiology, biochemistry, mechanisms of action, and genetic basis of these urovirulence factors that has occurred in the past decade and identifies areas in need of further study. Virulence factor expression is more common among certain genetically related groups of E. coli which constitute virulent clones within the larger E. coli population. In general, the more virulence factors a strain expresses, the more severe an infection it is able to cause. Certain virulence factors specifically favor the development of pyelonephritis, others favor cystitis, and others favor asymptomatic bacteriuria. The currently defined virulence factors clearly contribute to the virulence of wild-type strains but are usually insufficient in themselves to transform an avirulent organism into a pathogen, demonstrating that other as-yet-undefined virulence properties await discovery. Virulence factor testing is a useful epidemiological and research tool but as yet has no defined clinical role. Immunological and biochemical anti-virulence factor interventions are effective in animal models of UTI and hold promise for the prevention of UTI in humans. Images PMID:1672263

  14. Production and regulation of functional amyloid curli fimbriae by Shiga toxin-producing Escherichia coli

    USDA-ARS?s Scientific Manuscript database

    Functional amyloid, in the form of adhesive fimbrial proteins termed curli, was first described in Salmonella and Escherichia coli. Curli fibers adhere to various host cells and structural proteins, interact with components of the host immune system, and participate in biofilm formation. Shiga toxin...

  15. Epigenetic reprogramming of fallopian tube fimbriae in BRCA mutation carriers defines early ovarian cancer evolution

    PubMed Central

    Bartlett, Thomas E.; Chindera, Kantaraja; McDermott, Jacqueline; Breeze, Charles E.; Cooke, William R.; Jones, Allison; Reisel, Daniel; Karegodar, Smita T.; Arora, Rupali; Beck, Stephan; Menon, Usha; Dubeau, Louis; Widschwendter, Martin

    2016-01-01

    The exact timing and contribution of epigenetic reprogramming to carcinogenesis are unclear. Women harbouring BRCA1/2 mutations demonstrate a 30–40-fold increased risk of high-grade serous extra-uterine Müllerian cancers (HGSEMC), otherwise referred to as ‘ovarian carcinomas', which frequently develop from fimbrial cells but not from the proximal portion of the fallopian tube. Here we compare the DNA methylome of the fimbrial and proximal ends of the fallopian tube in BRCA1/2 mutation carriers and non-carriers. We show that the number of CpGs displaying significant differences in methylation levels between fimbrial and proximal fallopian tube segments are threefold higher in BRCA mutation carriers than in controls, correlating with overexpression of activation-induced deaminase in their fimbrial epithelium. The differentially methylated CpGs accurately discriminate HGSEMCs from non-serous subtypes. Epigenetic reprogramming is an early pre-malignant event integral to BRCA1/2 mutation-driven carcinogenesis. Our findings may provide a basis for cancer-preventative strategies. PMID:27216078

  16. Host Cell Contact-Induced Transcription of the Type IV Fimbria Gene Cluster of Actinobacillus pleuropneumoniae

    PubMed Central

    Boekema, Bouke K. H. L.; Van Putten, Jos P. M.; Stockhofe-Zurwieden, Norbert; Smith, Hilde E.

    2004-01-01

    Type IV pili (Tfp) of gram-negative species share many characteristics, including a common architecture and conserved biogenesis pathway. Much less is known about the regulation of Tfp expression in response to changing environmental conditions. We investigated the diversity of Tfp regulatory systems by searching for the molecular basis of the reported variable expression of the Tfp gene cluster of the pathogen Actinobacillus pleuropneumoniae. Despite the presence of an intact Tfp gene cluster consisting of four genes, apfABCD, no Tfp were formed under standard growth conditions. Sequence analysis of the predicted major subunit protein ApfA showed an atypical alanine residue at position −1 from the prepilin peptidase cleavage site in 42 strains. This alanine deviates from the consensus glycine at this position in Tfp from other species. Yet, cloning of the apfABCD genes under a constitutive promoter in A. pleuropneumoniae resulted in pilin and Tfp assembly. Tfp promoter-luxAB reporter gene fusions demonstrated that the Tfp promoter was intact but tightly regulated. Promoter activity varied with bacterial growth phase and was detected only when bacteria were grown in chemically defined medium. Infection experiments with cultured epithelial cells demonstrated that Tfp promoter activity was upregulated upon adherence of the pathogen to primary cultures of lung epithelial cells. Nonadherent bacteria in the culture supernatant exhibited virtually no promoter activity. A similar upregulation of Tfp promoter activity was observed in vivo during experimental infection of pigs. The host cell contact-induced and in vivo-upregulated Tfp promoter activity in A. pleuropneumoniae adds a new dimension to the diversity of Tfp regulation. PMID:14742510

  17. Genomic Subtractive Hybridization and Selective Capture of Transcribed Sequences Identify a Novel Salmonella typhimurium Fimbrial Operon and Putative Transcriptional Regulator That Are Absent from the Salmonella typhi Genome

    PubMed Central

    Morrow, Brian J.; Graham, James E.; Curtiss, Roy

    1999-01-01

    Salmonella typhi, the etiologic agent of typhoid fever, is adapted to the human host and unable to infect nonprimate species. The genetic basis for host specificity in S. typhi is unknown. The avirulence of S. typhi in animal hosts may result from a lack of genes present in the broad-host-range pathogen Salmonella typhimurium. Genomic subtractive hybridization was successfully employed to isolate S. typhimurium genomic sequences which are absent from the S. typhi genome. These genomic subtracted sequences mapped to 17 regions distributed throughout the S. typhimurium chromosome. A positive cDNA selection method was then used to identify subtracted sequences which were transcribed by S. typhimurium following macrophage phagocytosis. A novel putative transcriptional regulator of the LysR family was identified as transcribed by intramacrophage S. typhimurium. This putative transcriptional regulator was absent from the genomes of the human-adapted serovars S. typhi and Salmonella paratyphi A. Mutations within this gene did not alter the level of S. typhimurium survival within macrophages or virulence within mice. A subtracted genomic fragment derived from the ferrichrome operon also hybridized to the intramacrophage cDNA. Nucleotide sequence analysis of S. typhimurium and S. typhi chromosomal sequences flanking the ferrichrome operon identified a novel S. typhimurium fimbrial operon with a high level of similarity to sequences encoding Proteus mirabilis mannose-resistant fimbriae. The novel fimbrial operon was absent from the S. typhi genome. The absence of specific genes may have allowed S. typhi to evolve as a highly invasive, systemic human pathogen. PMID:10496884

  18. Antibiotic resistance and putative virulence factors of Serratia marcescens with respect to O and K serotypes.

    PubMed

    Aucken, H M; Pitt, T L

    1998-12-01

    Serratia marcescens serotypes O6:K14, O8:K14 and O28:K28 are common in the natural environment, but rare in hospitals. Serotypes O14:K14 and O27:K14 predominate among clinical strains, but not in the environment, suggesting that the latter serotypes may be more suited for survival in the clinical setting. Consequently, 469 epidemiologically distinct strains of S. marcescens were tested for various putative virulence factors and analysed for associations with serotype. The factors positively associated with serotype O14:K14 were agglutination of five different species of red blood cells and expression of type 1 fimbriae. These were found in 63% and 53% of O14:K14 strains, respectively, compared with 7% and 12% of the three 'environmental serotypes'. Almost a quarter of the collection expressed the mannose-resistant haemagglutinin indicative of type 3 fimbriae, but this was not associated with any serotype. The production of DNAase, haemolysin, lipase, lecithinase, proteases and siderophores was almost universal and showed no serotype correlations. Almost half of the strains (46%) were resistant to serum and serotypes O27:K14 and O6:K14 were strongly associated with this characteristic. Serotype O27:K14 was also associated with higher proportions of antibiotic-resistant strains than other serotypes, but the same was not true of serotype O14:K14. All three 'environmental serotypes' were associated with low frequencies of antibiotic resistance; <12% were resistant to gentamicin, carbenicillin or piperacillin, or any combination of these three, compared with 20-25% of O14:K14 strains and >42-51% of O27:K14 strains. Pigment production was strongly associated with serotype. None of the O14:K14 or O27:K14 strains produced prodigiosin, but frequencies for the three 'environmental serotypes' ranged from 31% of O28:K28 strains to 85% of O6:K14 strains. The results of this study suggest that the adherence capability of S. marcescens strains may play a role in the colonisation

  19. Evolutionary and Functional Relationships of Colonization Factor Antigen I and Other Class 5 Adhesive Fimbriae of Enterotoxigenic Escherichia coli

    DTIC Science & Technology

    2004-12-01

    review and discussions of the manuscript. REFERENCES 1. Altboum, Z., M. M. Levine, J. E. Galen, and E. M. Barry . 2003. Genetic characterization and...generation of protein database search programs. Nucleic Acids Res. 25:3389–3402. 3. Barry , E. M., Z. Altboum, G. Losonsky, and M. M. Levine. 2003...R. Thomson, D. Pickard, J. Wain , C. Churcher, K. L. Mungall, S. D. Bentley, M. T. Holden, M. Sebaihia, S. Baker, D. Basham, K. Brooks, T

  20. Hha Represses Biofilm Formation in Escherichia coli O157:H7 by Affecting the Expression of Flagella and Curli Fimbriae

    USDA-ARS?s Scientific Manuscript database

    Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is a zoonotic pathogen that produces a broad-spectrum of diarrheal illnesses in infected humans. Although the genetic and molecular mechanisms enabling EHEC O157:H7 to produce characteristic adherence on epithelial cells are well characterized, the g...

  1. Volunteer Challenge With Enterotoxigenic Escherichia coli That Express Intestinal Colonization Factor Fimbriae CS17 and CS19

    DTIC Science & Technology

    2011-07-01

    Levine MM, Merson MM. Serologic differentiation between antitoxin responses to infection with Vibrio cholerae and enterotoxin-producing Escherichia coli...prototype cholera B subunit-colonization factor antigen cnterotoxigenic Escherichia coli vaccine. Vaccine 1993; 1[:929-34. 15. Levine MM, Nalin DR

  2. The role of Type 1, P and S fimbriae in binding of Escherichia coli to the canine endometrium.

    PubMed

    Krekeler, N; Marenda, M S; Browning, G F; Holden, K M; Charles, J A; Wright, P J

    2013-06-28

    Escherichia coli (E. coli) is the most commonly isolated infectious agent causing pyometra in bitches. Many E. coli strains isolated from the uteri of infected dogs carry several adhesin genes (fimH, papGIII and sfa). The objective of this study was to investigate the role of each adhesin gene product, acting alone or expressed in combination, in the bacterial binding to canine endometrium. E. coli strain P3, which was isolated from a uterus of a bitch naturally affected with pyometra, was shown by PCR to carry all three known fimbrial adhesin genes fimH, papGIII and sfa. Knockout (KO) mutants of this wildtype (P3-wt) strain were generated using insertional inactivation. Adhesion assays on anoestrous uteri of three post-pubertal bitches were undertaken. Overall, the number of bacteria adhering to canine endometrial biopsies was comparable between strains and no significant difference in the number of bound bacteria was found between the P3-wt strain and the single or double KO-strains. However, the triple knockout strain displayed less binding to the canine endometrium compared with the P3-wt strain. This study shows that a pathogenic E. coli strain (P3) isolated from the uterus of a bitch with pyometra was able to fully compensate for the loss of two of its three known adhesin genes. It was necessary to inactivate all three known adhesin genes in order to see a significant decrease in binding to canine endometrium.

  3. Glycoprotein glycans that inhibit adhesion of Escherichia coli mediated by K99 fimbriae: treatment of experimental colibacillosis.

    PubMed Central

    Mouricout, M; Petit, J M; Carias, J R; Julien, R

    1990-01-01

    Calf diarrhea due to infection by enterotoxigenic Escherichia coli was treated by administration of glycoprotein glycans derived from bovine plasma. The glycan moieties of the nonimmunoglobulin fraction of plasma mimicked the oligosaccharide moiety of intestinal receptors recognized by K99 pili. These glycoprotein glycans inhibited adhesion of E. coli K99+ ST+ to erythrocyte glycoconjugates in vitro, and they protected colostrum-deprived newborn calves against lethal doses of enterotoxigenic E. coli (10(10) bacteria). Adhesion of bacteria to the intestines (duodenum, jejunum, and ileum) was significantly reduced (by 2 orders of magnitude) in treated calves. PMID:2403535

  4. Effect of dietary taurine supplementation on growth, feed efficiency, and nutrient composition of juvenile sablefish (Anoplopoma fimbria)

    USDA-ARS?s Scientific Manuscript database

    Juvenile sablefish were fed a low taurine, basal feed with seven graded levels of supplemental taurine to determine taurine requirements for growth and feed efficiency. The basal feed was plant based, formulated primarily with soy and corn proteins with a minimal (9%) amount of fishmeal. The unsuppl...

  5. S-Fimbria-Encoding Determinant sfaI Is Located on Pathogenicity Island III536 of Uropathogenic Escherichia coli Strain 536

    PubMed Central

    Dobrindt, Ulrich; Blum-Oehler, Gabriele; Hartsch, Thomas; Gottschalk, Gerhard; Ron, Eliora Z.; Fünfstück, Reinhard; Hacker, Jörg

    2001-01-01

    The sfaI determinant encoding the S-fimbrial adhesin of uropathogenic Escherichia coli strains was found to be located on a pathogenicity island of uropathogenic E. coli strain 536. This pathogenicity island, designated PAI III536, is located at 5.6 min of the E. coli chromosome and covers a region of at least 37 kb between the tRNA locus thrW and yagU. As far as it has been determined, PAI III536 also contains genes which code for components of a putative enterochelin siderophore system of E. coli and Salmonella spp. as well as for colicin V immunity. Several intact or nonfunctional mobility genes of bacteriophages and insertion sequence elements such as transposases and integrases are present on PAI III536. The presence of known PAI III536 sequences has been investigated in several wild-type E. coli isolates. The results demonstrate that the determinants of the members of the S-family of fimbrial adhesins may be located on a common pathogenicity island which, in E. coli strain 536, replaces a 40-kb DNA region which represents an E. coli K-12-specific genomic island. PMID:11401961

  6. A Video Demonstration of Preserved Piloting by Scent Tracking but Impaired Dead Reckoning After Fimbria-Fornix Lesions in the Rat

    PubMed Central

    Whishaw, Ian Q.; Gorny, Boguslaw P.

    2009-01-01

    Piloting and dead reckoning navigation strategies use very different cue constellations and computational processes (Darwin, 1873; Barlow, 1964; O’Keefe and Nadel, 1978; Mittelstaedt and Mittelstaedt, 1980; Landeau et al., 1984; Etienne, 1987; Gallistel, 1990; Maurer and Séguinot, 1995). Piloting requires the use of the relationships between relatively stable external (visual, olfactory, auditory) cues, whereas dead reckoning requires the integration of cues generated by self-movement. Animals obtain self-movement information from vestibular receptors, and possibly muscle and joint receptors, and efference copy of commands that generate movement. An animal may also use the flows of visual, auditory, and olfactory stimuli caused by its movements. Using a piloting strategy an animal can use geometrical calculations to determine directions and distances to places in its environment, whereas using an dead reckoning strategy it can integrate cues generated by its previous movements to return to a just left location. Dead reckoning is colloquially called "sense of direction" and "sense of distance." Although there is considerable evidence that the hippocampus is involved in piloting (O’Keefe and Nadel, 1978; O’Keefe and Speakman, 1987), there is also evidence from behavioral (Whishaw et al., 1997; Whishaw and Maaswinkel, 1998; Maaswinkel and Whishaw, 1999), modeling (Samsonovich and McNaughton, 1997), and electrophysiological (O’Mare et al., 1994; Sharp et al., 1995; Taube and Burton, 1995; Blair and Sharp, 1996; McNaughton et al., 1996; Wiener, 1996; Golob and Taube, 1997) studies that the hippocampal formation is involved in dead reckoning. The relative contribution of the hippocampus to the two forms of navigation is still uncertain, however. Ordinarily, it is difficult to be certain that an animal is using a piloting versus a dead reckoning strategy because animals are very flexible in their use of strategies and cues (Etienne et al., 1996; Dudchenko et al., 1997; Martin et al., 1997; Maaswinkel and Whishaw, 1999). The objective of the present video demonstrations was to solve the problem of cue specification in order to examine the relative contribution of the hippocampus in the use of these strategies. The rats were trained in a new task in which they followed linear or polygon scented trails to obtain a large food pellet hidden on an open field. Because rats have a proclivity to carry the food back to the refuge, accuracy and the cues used to return to the home base were dependent variables (Whishaw and Tomie, 1997). To force an animal to use a a dead reckoning strategy to reach its refuge with the food, the rats were tested when blindfolded or under infrared light, a spectral wavelength in which they cannot see, and in some experiments the scent trail was additionally removed once an animal reached the food. To examine the relative contribution of the hippocampus, fimbria–fornix (FF) lesions, which disrupt information flow in the hippocampal formation (Bland, 1986), impair memory (Gaffan and Gaffan, 1991), and produce spatial deficits (Whishaw and Jarrard, 1995), were used. PMID:19398947

  7. Pathogenicity of an Escherichia coli O115:K"V165" mutant negative for F165(1) fimbriae in septicemia of gnotobiotic pigs.

    PubMed Central

    Ngeleka, M; Jacques, M; Martineau-Doizé, B; Daigle, F; Harel, J; Fairbrother, J M

    1993-01-01

    To evaluate the role of the F165(1) fimbrial system in the pathogenesis of septicemia, 2-day-old germfree pigs were inoculated intragastrically with Escherichia coli O115:K"V165":F165 wild-type strain 5131, its F165(1)-negative TnphoA mutant M48, or E. coli O115:K(-):F165(-) wild-type strain 862B. Pigs were sacrificed at different times (3, 6, 12, 24, 48, and 96 h) postinfection (p.i.). Pigs inoculated with strain 5131 developed clinical signs (anorexia, lameness, reluctance to move, or lack of motor coordination) and were moribund within 48 h p.i., and, at necropsy, infecting bacteria were isolated in various extraintestinal organs. Strain 5131 was isolated as early as 6 h p.i. from the blood of inoculated pigs. Pigs inoculated with mutant M48 developed only mild clinical signs at 96 h p.i. Mutant M48 colonized extraintestinal organs of pigs but to a lesser extent than the parent strain did. In contrast to the parent strain, this mutant was not isolated in the blood of inoculated pigs. Pigs inoculated with strain 862B remained normal during the experiment. All of the strains colonized the mucus layer of the intestine, but no histological changes of intestinal mucosa were observed by either light or electron microscopy. The parent strain, but not the mutant M48, expressed F165(1) in vivo. In a competitive study in which the parent strain and its afimbrial mutant were inoculated simultaneously, clinical signs of septicemia developed 24 h after inoculation, and only the parent strain 5131 was isolated from the blood of inoculated pigs. Our results suggest that the F165(1) fimbrial system of E. coli O115:K"V165" strains may play an important role in the ability of the bacteria to survive in the blood and spread systemically through the porcine host. Images PMID:8094383

  8. Contributions of EspA filaments and curli fimbriae in cellular adherence and biofilm formation of enterohemorrhagic Escherichia coli O157:H7

    USDA-ARS?s Scientific Manuscript database

    In Escherichia coli O157:H7 (O157), the filamentous structure of the type III secretion system is produced from the polymerization of the EspA protein. EspA filaments are essential for O157 adherence to epithelial cells. In previous studies, we demonstrated that O157 hha deletion mutants showed incr...

  9. Isopycnic separation of Escherichia coli cultures possessing colonization factor antigen I.

    PubMed Central

    Giesa, F R; Zajac, I; Bartus, H F; Actor, P

    1982-01-01

    A culture of Escherichia coli possessing colonization factor antigen I was subjected to isopycnic separation on Percoll gradients. The results demonstrated successful division of the culture into two populations: (i) bacteria which cause mannose-resistant hemagglutination and (ii) bacteria which lack the ability to hemagglutinate in the presence of mannose. PMID:6286714

  10. [Cloning, expression and activity of K99 fimbrial operon gene from enterotoxigenic Escherichia coli].

    PubMed

    Yang, Yang; Hou, Huayan; Yu, Lei; Zhu, Guoqiang

    2012-12-04

    To clone and express fan operon gene clusters of K99 fimbriae in enterotoxigenic Escherichia coli (ETEC) in vitro, and study the activity of the recombinant E. coli expressing K99 fimbriae. K99 fimbriae gene clusters were amplified by long-PCR method, using the genomic DNA of K99-fimbriae E. coli C8307 as the DNA template. The 5.7Kb PCR products were inserted into expressing vector pBR322 with restriction endonuclease, then positive clones were screened. The positive recombinant plasmid was transformed into non-fimbriae E. coli SE5000 strains, and pBR322 plasmid was also transformed into SE5000 for negative control strain. The recombination E. coli expressing K99 fimbriae was tested with agglutination assay, using monoclonal antibody serum and brush border vesicles from the piglet small intestinal epithelia cells. The expressed fimbriae on the surface of the recombinant E. coli SE5000 were observed by transmissible electromicroscope. Heat extraction method was employed to isolate and purify K99 fimbriae, which was exerted SDS-PAGE, and 18.5 kDa protein band was detected. The mouse sera produced from recombinant fimbriae was used to test K99-fimbriae strains C83907, C83914, C83260 with positive agglutination results, while negative results were found with E. coli contain other kinds of fimbriae. The assays of SDS-PAGE, Western blot, agglutination assay were used to evaluate antigenicity and biologic activity between C83907 and recombinant strain. Adhesion test with HeLa cell line demonstrated the recombinant strain and wild type have the similar adherence ability, and this adhesion can be inhibited with mouse serum containing polyclonal antibody against recombinant K99 fimbriae. This study has laid a good foundation for further study on bioactivity of K99.

  11. Binding Specificity of Piliated Strains of Escherichia coli and Salmonella typhimurium to Epithelial Cells, Saccharomyces cerevisiae Cells, and Erythrocytes

    PubMed Central

    Korhonen, Timo K.; Leffler, Hakon; Edén, Catharina Svanborg

    1981-01-01

    The binding to mammalian cells of piliated enteric bacteria and the inhibition of the binding by antibodies to purified pili were studied. The target cells were epithelial cells from human bucca and human and rat urinary tracts, erythrocytes from various species, and Saccharomyces cerevisiae cells. The strains were selected to represent the two main agglutination patterns of enteric bacteria: mannose-resistant agglutination of human and other erythrocytes and mannose-sensitive agglutination of guinea pig and other erythrocytes. Escherichia coli 3669 caused only mannose-resistant agglutination, E. coli 6013 caused only mannose-sensitive agglutination, and E. coli 3048 caused both types of agglutination simultaneously. Salmonella typhimurium SH6749 exhibited only mannose-sensitive hemagglutination and was included to allow comparison of its pili with those of E. coli strains. The range of epithelial cells to which the bacteria adhered was related to their agglutination patterns. All four strains attached to human buccal cells. Only E. coli strains 3669 and 3048, which caused mannose-resistant agglutination, adhered to human urinary tract epithelial cells, and only those strains that caused mannose-sensitive agglutination adhered to rat urinary tract epithelial cells. The binding of S. typhimurium SH6749, but not of the other strains with mannose-sensitive agglutination, was significantly inhibited by d-mannose. Globotetraosylceramide, a glycolipid present in the human urinary tract epithelium, inhibited attachment to human uroepithelial cells of the two strains with mannose-resistant hemagglutination. As tested by the enzyme-linked immunosorbent assay, cross-reactions between type 1 pili of the E. coli strains were strong, but those between S. typhimurium and E. coli mannose-sensitive pili were weak. The two pili that induced mannose-resistant hemagglutination on E. coli did not cross-react. Significant inhibition of adhesion of all four strains was obtained with the

  12. Complete Genome Sequences of Two Human Oral Microbiome Commensals, Streptococcus salivarius ATCC 25975 and S. salivarius ATCC 27945.

    PubMed

    Butler, Robert R; Soomer-James, Jahna T A; Frenette, Michel; Pombert, Jean-François

    2017-06-15

    Streptococcus salivarius strains are significant contributors to the human oral microbiome. Some possess unique fimbriae that give them the ability to coaggregate and colonize particular oral structures. We present here the complete genomes of Streptococcus salivarius Lancefield K(-)/K(+) strains ATCC 25975 and ATCC 27945, which can and cannot, respectively, produce fimbriae. Copyright © 2017 Butler et al.

  13. Transpositional inactivation of gadW enhances curli production and biofilm formation in Enterohemorrhagic Escherichia coli O157:H7

    USDA-ARS?s Scientific Manuscript database

    Enterohemorrhagic Escherichia coli (EHEC) O157:H7 has been shown to produce variants that either express or are repressed in the expression of curli fimbriae promoting bacterial attachment, aggregation, and biofilm formation. The variant expression of curli fimbriae in some instances could result fr...

  14. Designations F18ab and F18ac for the related fimbrial types F107, 2134P and 8813 of Escherichia coli isolated from porcine postweaning diarrhoea and from oedema disease.

    PubMed

    Rippinger, P; Bertschinger, H U; Imberechts, H; Nagy, B; Sorg, I; Stamm, M; Wild, P; Wittig, W

    1995-08-01

    The relatedness of the fimbriae produced by eight E. coli strains including type strains with F107 fimbriae, 2134P pili and colonization factor 8813 (preliminary F18), was examined. These strains had been isolated principally from pigs which were affected with postweaning diarrhoea or with oedema disease. The fimbriae were analyzed by means of electron microscopy, slide agglutination, immunofluorescence, immunogold labelling, immuno-diffusion, immunoelectrophoresis and western blot, molecular genetic techniques, and in vitro adhesion. The fimbriae of all the strains were long flexible filaments with a diameter not larger than 4.6 nm showing a zig-zag pattern. Results obtained by the serological techniques confirmed that the fimbriae possessed a common antigenic determinant designated 'a' in addition to a variant-specific determinant designated 'b' or 'c'. Immunoelectron microscopy demonstrated that the determinants 'a' and 'b' or 'a' and 'c' were localized along the same fimbrium. In immunoelectrophoresis, fimbrial extracts of selected strains yielded a single precipitation line towards the cathode. One single major subunit of approximately 15 kDa was recognised in western blots by antisera against the common antigenic determinant and the variant specific determinants. All strains possessed sequences related to gene fedA, coding for the major subunit of fimbriae F107. Two types of fedA-related subunit genes were differentiated, corresponding to the 'ab' and 'ac' types of fimbriae as defined by serological methods. The results demonstrated that F107 fimbriae, 2134P pili and colonization factor 8813 are related, and that two serological variants can be distinguished. We propose designations F18ab (for F107), and F18ac (for 2134P and 8813) in analogy to the nomenclature of F4 fimbriae.

  15. Phenotypic changes in nonfimbriated smooth strains of Aggregatibacter actinomycetemcomitans grown in low-humidity solid medium.

    PubMed

    Pei, Zhenhua; Niu, Zhongying; Shi, Shenggen; Shi, Liang; Tang, Chuhua

    2013-04-01

    Aggregatibacter actinomycetemcomitans is the primary etiologic agent of localized aggressive periodontitis. In vitro, it can undergo fimbriated rough to nonfimbriated smooth phenotypic transition, accompanied by an increase in invasive ability and a decrease in adhesive ability. No opposite direction phenotypic transition was reported. To better understand its pathogenicity, the authors studied the morphological changes of nonfimbriated smooth strains induced by growth environmental humidity. Transmission electron microscopy was used to identify fimbriae expression change. It was found that the lower medium humidity, the more fimbriae reexpressed. In conclusion, the smooth strain of A. actinomycetemcomitans can reexpress the fimbriae in lower humidity environment.

  16. Pilus Production, Hemagglutination, and Adhesion by Porcine Strains of Enterotoxigenic Escherichia coli Lacking K88, K99, and 987P Antigens

    PubMed Central

    Awad-Masalmeh, M.; Moon, H. W.; Runnels, P. L.; Schneider, R. A.

    1982-01-01

    Three strains of enterotoxigenic Escherichia coli which adhered, colonized intensively, and caused disease in pig intestine, but which did not produce pili of the K88, K99, or 987P antigen types were designated 3P− ETEC. The 3P− ETEC caused mannose-resistant hemagglutination, adhered to porcine intestinal epithelial cells in vitro, and produced pili. However, most bacteria taken directly from the intestine of pigs infected with 3P− ETEC appeared to be nonpiliated. Two preparations were isolated from the 3P− ETEC. One (material A) contained pili, caused mannose-sensitive hemagglutination, and did not inhibit adhesion of whole bacteria to epithelial cells in vitro. The other (material B) had no demonstrable pili, caused mannose-resistant hemagglutination, and blocked ahesion of bacteria to epithelial cells in vitro. Antiserum against an acapsular mutant (K−) of one 3P− ETEC strain was absorbed to remove antibodies directed against somatic (O) antigen. The absorbed antiserum agglutinated all three 3P− ETEC strains grown in the K− form at 37°C, but not when they were grown at 18°C. The absorbed antiserum blocked the hemagglutinating activity of material B, but not of material A. It also reacted (via indirect immunofluorescence) with all of the 3P− ETEC when they were grown in pig intestine. The results were interpreted to indicate that: (i) the epithelial adhesive and mannose-resistant hemagglutinating activities of the 3P− ETEC strains may be mediated by an antigen contained in material B; (ii) this antigen either is not pilus associated or is associated with pili that are not demonstrable by the methods used here; (iii) the 3P− ETEC strains produce type 1 pili which do not mediate their adhesion to intestinal epithelium of pigs. Images PMID:6119295

  17. Mechanism of adhesion of Alysiella bovis to glass surfaces.

    PubMed Central

    Irvin, R T; To, M; Costerton, J W

    1984-01-01

    Alysiella bovis adheres to surfaces by means of short, ruthenium red-staining, rod-like fimbriae. The fimbriae remain associated with the cell envelope of A. bovis, even when sonicated or exposed sequentially to toluene, Triton X-100, lysozyme, ribonuclease, and deoxyribonuclease. Adhesion of outer membrane-derived cell wall ghosts of A. bovis to glass was inhibited by IO4-, sodium dodecyl sulfate, urea, pronase, and trypsin. Protease treatment digested the fimbriae from the distal end, and exposure to sodium dodecyl sulfate depolymerized the fimbriae. Exposure of ghosts to 1% sodium dodecyl sulfate preferentially solubilized a 16,500-dalton protein which was subsequently purified by gel filtration and demonstrated to be a glycoprotein (ca. 17% carbohydrate). Antibodies raised against the 16,500-dalton glycoprotein agglutinated whole cells and inhibited adhesion of ghosts to glass. Images PMID:6209260

  18. Mechanism of attachment of swarm cells of Thiothrix nivea.

    PubMed Central

    Larkin, J M; Nelson, R

    1987-01-01

    Swarm cells of Thiothrix nivea were found to possess a group of fimbriae at one pole. The other pole either was bare or possessed from one to three fimbriae. By using this polarity as a marker, it was found that the initial step in attachment of swarm cells involves the fimbriated pole and that this initial step is followed by the production of holdfast material. Images PMID:2890625

  19. Adhesive threads of extraintestinal pathogenic Escherichia coli.

    PubMed

    Antão, Esther-Maria; Wieler, Lothar H; Ewers, Christa

    2009-12-10

    The ability to adhere to host surfaces is by far the most vital step in the successful colonization by microbial pathogens. Colonization begins with the attachment of the bacterium to receptors expressed by cells forming the lining of the mucosa. Long hair like extracellular appendages called fimbriae, produced by most Gram-negative pathogens, mediate specific attachment to the epithelial cell surface. Associated with the fimbriae is a protein called an adhesin, which directs high-affinity binding to specific cell surface components. In the last couple of years, an enormous amount of research has been undertaken that deals with understanding how bacterial pathogens adhere to host cells. E. coli in all probability is one of the best studied free-living organisms. A group of E. coli called Extraintestinal pathogenic E. coli (ExPEC) including both human and animal pathogens like Uropathogenic E. coli (UPEC), Newborn meningitic E. coli (NMEC) and Avian pathogenic E. coli (APEC), have been found to harbour many fimbriae including Type 1 fimbriae, P fimbriae, curli fibres, S fimbriae, F1C fimbriae, Dr fimbriae, afimbrial adhesins, temperature-sensitive haemagglutinin and many novel adhesin gene clusters that have not yet been characterized. Each of these adhesins is unique due to the recognition of an adhesin-specific receptor, though as a group these adhesins share common genomic organization. A newly identified putative adhesin temporarily termed ExPEC Adhesin I, encoded by gene yqi, has been recently found to play a significant role in the pathogenesis of APEC infection, thus making it an interesting candidate for future research. The aim of this review is to describe the role of ExPEC adhesins during extraintestinal infections known till date, and to suggest the idea of investigating their potential role in the colonization of the host gut which is said to be a reservoir for ExPEC.

  20. Antigen-43-Mediated Autoaggregation of Escherichia coli Is Blocked by Fimbriation

    PubMed Central

    Hasman, Henrik; Chakraborty, Trinad; Klemm, Per

    1999-01-01

    Antigen 43 (Ag43), the product of the flu gene, is a surface-displayed autotransporter protein of Escherichia coli. Ag43 is responsible for the autoaggregation and flocculation of static liquid cultures of many E. coli strains. The expression of Ag43 has been reported to be phase variable and controlled by the product of the oxyR gene. Type 1 fimbriae are thin adhesive thread-like surface organelles responsible for bacterial receptor recognition and tissue colonization. Like that of Ag43, the expression of type 1 fimbriae is phase variable. Interestingly, previous results have suggested that the expression of type 1 fimbriae and the expression of Ag43 are mutually exclusive. In the present report, we show, by use of well-defined mutants, that fimbriation abolishes Ag43-mediated autoaggregation but does not affect Ag43 expression. Autoaggregation is shown to require an intercellular Ag43-Ag43 interaction, and the physical presence of fimbriae on the cells seems to abrogate this interaction. The Ag43 or OxyR status does not appear to influence fimbria expression, and our results suggest that the expression of Ag43 and the expression of fimbriae are independent processes. PMID:10438752

  1. Humoral immune response to Bacteroides gingivalis fimbrial antigen in mice.

    PubMed Central

    Ogawa, T; Shimauchi, H; Kusumoto, Y; Hamada, S

    1990-01-01

    Bacteroides gingivalis fimbrial antigen incorporated into liposomes, but not in Tris-HCl buffer, significantly raised the levels of anti-fimbriae antibodies in serum, particularly of the IgG class, after oral primary and booster immunizations in BALB/c mice. An approximately linear relationship was observed between the dose of fimbrial antigen and the level of fimbriae-specific antibodies produced; antibody production reached its maximum at an immunization dosage of 500 micrograms of fimbriae per mouse. Fimbriae-specific antibody production was enhanced by use of a semi-synthetic adjuvant, a stearoyl derivative of sodium beta-N-acetylglucosaminyl-(1----4)-N-acetylmuramyl-L-alanyl-D-isoglutaminyl-(L) - stearoyl-(D)-meso-diamino-pimelic acid-(D)-amide-D-alanine (GM)-53) in liposomes. High anti-fimbriae antibody levels in serum and saliva were maintained for several months in the mice that had received two orally administered boosters of fimbrial antigen with GM-53 in liposomes. Salivary anti-fimbriae antibody levels, particularly of the IgA class, were markedly raised. PMID:1968885

  2. Correlation between esterase electrophoretic polymorphism and virulence-associated traits in extra-intestinal invasive strains of Escherichia coli.

    PubMed Central

    Goullet, P.; Picard, B.; Contrepois, M.; De Rycke, J.; Barnouin, J.

    1994-01-01

    The electrophoretic variations of carboxylesterase B and of esterases A, C and I, the presence of mannose resistant haemagglutinin, alpha-haemolysin, cytotoxic necrotizing factor type 1 (CNF1) and certain O antigens were compared in 150 strains of Escherichia coli responsible for extra-intestinal infections. Electrophoretic mobilities of outer membrane proteins (OMP) were also studied for strains belonging to O4, O6, O7, O8 and O75 serogroups. Fast migrating allozymes of carboxylesterase B (pattern B1) were correlated with slow migrating allozymes of esterase C, serogroups O7 and O8, lack of virulence factor, and particular OMP patterns, whereas slow migrating allozymes of carboxylesterase B (pattern B2) were correlated with fast migrating allozymes of esterase C, serogroups O2, O4, O6, O18 and O75, virulence factor production, and distinct OMP patterns. Allozymes of esterases A and I were not clearly correlated with the distribution of virulence factors. The pattern B2 was more strongly associated with CNF1 than with alpha-haemolysin and mannose resistant haemagglutinin. These results substantiate the view that the electrophoretic pattern B2 of carboxylesterase B identified most of the highly pathogenic strains implicated in extra-intestinal infection of humans. Images Fig. 2 PMID:7509755

  3. Spontaneous deletions and flanking regions of the chromosomally inherited hemolysin determinant of an Escherichia coli O6 strain.

    PubMed Central

    Hacker, J; Knapp, S; Goebel, W

    1983-01-01

    The hemolytic Escherichia coli strain 536 (O6) propagates spontaneous hemolysin-negative mutants at relatively high rates (10(-3) to 10(-4)). One type of mutant (type I) lacks both secreted (external) and periplasmic (internal) hemolysin activity (Hlyex-/Hlyin-) and in addition shows no mannose-resistant hemagglutination (Mrh-), whereas the other type (type II) is Hlyex-/Hlyin+ and Mrh+. The genetic determinants for hemolysin production (hly) and for mannose-resistant hemagglutination (mrh) of this strain are located on the chromosome. Hybridization experiments with DNA probes specific for various parts of the hly determinant reveal that mutants of type I have lost the total hly determinant, whereas those of type II lack only part of the hlyB that is essential for transport of hemolysin across the outer membrane. Using a probe that contains the end sequence of the plasmid pHly152-encoded hly determinant (adjacent to hlyB), we determined that a related sequence flanks also the hlyB-distal end of the chromosomal hly determinant of E. coli 536. In addition several other similar or even identical sequences are found in the vicinity of the hlyC- and the hlyB-distal ends of both the chromosomal and the plasmid hly determinants. Images PMID:6343344

  4. Porphyromonas (Bacteroides) gingivalis fimbrillin: size, amino-terminal sequence, and antigenic heterogeneity.

    PubMed Central

    Lee, J Y; Sojar, H T; Bedi, G S; Genco, R J

    1991-01-01

    Bacterial fimbriae mediate cell adhesion and are important in colonization. Fimbrial proteins from strains of Porphyromonas (Bacteroides) gingivalis isolated from different individuals were compared for their size, amino-terminal sequence, and antigenic diversity. Two major protein components of the crude fimbrial preparations differed in apparent molecular mass, ranging from 41 to 49 kDa for the fimbrillin monomer and from 61 to 78 kDa for the other major protein. The amino-terminal sequence of the antigenically related group of proteins of the fimbrillin monomer in the 41- to 49-kDa range showed significant homology; however, minor sequence heterogeneity was observed, mainly in residues 4 to 6. One of the observed amino-terminal sequences, AFGVGDDESKVAKLTVMVYNG, resembled the deduced sequence of P. gingivalis 381 (D.P. Dickinson, M. K. Kubiniec, F. Yoshimura, and R.J. Genco, J. Bacteriol. 170:1658-1665, 1988). Fimbriae from all the strains of P. gingivalis showing this sequence contained a fimbrillin monomer of 43 kDa and showed a strong reaction with both polyclonal and monoclonal antibodies directed to the fimbriae from P. gingivalis 2561 (381). Fimbriae from strains showing amino-terminal sequence variations in residues 4 to 6 (i.e., substitution of VGD with either E or NAG) were more diverse in their molecular sizes. Most of these variant fimbriae showed weak reactions with the polyclonal antibodies and no reaction with the monoclonal antibodies induced to the fimbriae of strain 2561. No correlation could be established between the molecular size and immunological reactivity of the fimbrillin monomer of P. gingivalis strains. Strains 9-14K-1 and HG 564 not only showed markedly different sequences from the other three amino-terminal sequences but also did not react with either polyclonal or monoclonal antibodies to the fimbriae of strain 2561. Strains W50, W83, and AJW 5 failed to show any immunological reactivity with the antibodies to fimbrillin or fimbriae

  5. Structure of the fimbrial protein Mfa4 from Porphyromonas gingivalis in its precursor form: implications for a donor-strand complementation mechanism

    PubMed Central

    Kloppsteck, Patrik; Hall, Michael; Hasegawa, Yoshiaki; Persson, Karina

    2016-01-01

    Gingivitis and periodontitis are chronic inflammatory diseases that can lead to tooth loss. One of the causes of these diseases is the Gram-negative Porphyromonas gingivalis. This periodontal pathogen is dependent on two fimbriae, FimA and Mfa1, for binding to dental biofilm, salivary proteins, and host cells. These fimbriae are composed of five proteins each, but the fimbriae assembly mechanism and ligands are unknown. Here we reveal the crystal structure of the precursor form of Mfa4, one of the accessory proteins of the Mfa1 fimbria. Mfa4 consists of two β-sandwich domains and the first part of the structure forms two well-defined β-strands that run over both domains. This N-terminal region is cleaved by gingipains, a family of proteolytic enzymes that encompass arginine- and lysine-specific proteases. Cleavage of the N-terminal region generates the mature form of the protein. Our structural data allow us to propose that the new N-terminus of the mature protein may function as a donor strand in the polymerization of P. gingivalis fimbriae. PMID:26972441

  6. Biofilm formation as a virulence determinant of uropathogenic Escherichia coli Dr+ strains.

    PubMed

    Zalewska-Piatek, Beata M; Wilkanowicz, Sabina I; Piatek, Rafał J; Kur, Józef W

    2009-01-01

    Urinary tract infections are the most common health problem affecting millions of people each year. Uropathogenic Escherichia coli (UPEC) strains are the major factor causing lower and upper urinary tract infections. UPEC produce several virulence factors among which are surface exposed adhesive organelldes (pili/fimbriae) responsible for colonization, invasion and amplification within uroepithelial cells. The virulence of the uropathogenic E. coli Dr IH11128 is associated with Dr fimbriae belonging to the Dr family of adhesins (associated with diarrhea and urinary tract infections) and a DraD protein capping the linear fiber at the bacterial cell surface. In this study we revealed that biofilm development can be another urovirulence determinant allowing pathogenic E. coli Dr+ to survive within the urinary tract. E. coli strains were grown in rich or minimal media, allowed to adhere to abiotic surfaces and analyzed microscopically by staining of cells with cristal violet. We found that both Dr fimbriae and DraD, exposed at the cell surface in two forms, fimbria-associated or fimbria non-associated, (DraE+/DraD+, DraE+/DraD- or DraE-/DraD+ E. coli strains) are required for biofilm formation. Additionally, we demonstrated the biofilm formation capacity of E. coli strains deficient in the surface secretion or production of the DraE adhesin.

  7. Oxygen-Limiting Conditions Enrich for Fimbriate Cells of Uropathogenic Proteus mirabilis and Escherichia coli▿

    PubMed Central

    Lane, M. Chelsea; Li, Xin; Pearson, Melanie M.; Simms, Amy N.; Mobley, Harry L. T.

    2009-01-01

    MR/P fimbriae of uropathogenic Proteus mirabilis undergo invertible element-mediated phase variation whereby an individual bacterium switches between expressing fimbriae (phase ON) and not expressing fimbriae (phase OFF). Under different conditions, the percentage of fimbriate bacteria within a population varies and could be dictated by either selection (growth advantage of one phase) or signaling (preferentially converting one phase to the other in response to external signals). Expression of MR/P fimbriae increases in a cell-density dependent manner in vitro and in vivo. However, rather than the increased cell density itself, this increase in fimbrial expression is due to an enrichment of fimbriate bacteria under oxygen limitation resulting from increased cell density. Our data also indicate that the persistence of MR/P fimbriate bacteria under oxygen-limiting conditions is a result of both selection (of MR/P fimbrial phase variants) and signaling (via modulation of expression of the MrpI recombinase). Furthermore, the mrpJ transcriptional regulator encoded within the mrp operon contributes to phase switching. Type 1 fimbriae of Escherichia coli, which are likewise subject to phase variation via an invertible element, also increase in expression during reduced oxygenation. These findings provide evidence to support a mechanism for persistence of fimbriate bacteria under oxygen limitation, which is relevant to disease progression within the oxygen-restricted urinary tract. PMID:19114498

  8. Mannanoligosaccharide agglutination by Salmonella enterica strains isolated from carrier pigs

    PubMed Central

    Borowsky, Luciane; Corção, Gertrudes; Cardoso, Marisa

    2009-01-01

    Type-1 fimbriae are associated with most Salmonella enterica serovars and are an essential factor for host colonization. Mannanoligosaccharides (MOS), a prebiotic that is agglutinated by type-1 fimbriae, are proposed for the control of enterobacteria colonization and may be an alternative to Salmonella control in pigs. The aim of this study was to evaluate the capability of porcine Salmonella strains to adhere to MOS in vitro. A total of 108 strains of Salmonella sp. isolated from carrier pigs were evaluated for the amplification of fimA and fimH genes, agglutination of MOS and hemagglutination. In all tested strains, amplicons of expected size were detected for both fimA and fimH gene. In the hemagglutination assays, 31 (28.7%) strains presented mannose–sensitive agglutination of erythrocytes, indicating that the strains were expressing type-1 fimbriae. Considering only strains expressing the type-1 fimbriae, 23 (74.2%) presented a strong agglutination of MOS, 3 (9.6%) a weak reaction and 5 (16.2%) none. The results indicate that Salmonella enterica strains expressing type-1 fimbriae can agglutinate effectively in vitro to MOS. PMID:24031388

  9. Identification and sequence analysis of lpfABCDE, a putative fimbrial operon of Salmonella typhimurium.

    PubMed Central

    Bäumler, A J; Heffron, F

    1995-01-01

    A chromosomal region present in Salmonella typhimurium but absent from related species was identified by hybridization. A DNA probe originating from 78 min on the S. typhimurium chromosome hybridized with DNA from Salmonella enteritidis, Salmonella heidelberg, and Salmonella dublin but not with DNA from Salmonella typhi, Salmonella arizonae, Escherichia coli, and Shigella serotypes. Cloning and sequence analysis revealed that the corresponding region of the S. typhimurium chromosome encodes a fimbrial operon. Long fimbriae inserted at the poles of the bacterium were observed by electron microscopy when this fimbrial operon was introduced into a nonpiliated E. coli strain. The genes encoding these fimbriae were therefore termed lpfABCDE, for long polar fimbriae. Genetically, the lpf operon was found to be most closely related to the fim operon of S. typhimurium, both in gene order and in conservation of the deduced amino acid sequences. PMID:7721701

  10. Isolation and characterisation of dog uropathogenic Proteus mirabilis strains.

    PubMed

    Gaastra, W; van Oosterom, R A; Pieters, E W; Bergmans, H E; van Dijk, L; Agnes, A; ter Huurne, H M

    1996-01-01

    Proteus mirabilis strains isolated from the urine of dogs with urinary tract infections, were characterised with respect to the production of haemolysin and fimbriae. In contrast to healthy dogs, P. mirabilis was also isolated in high numbers from the faeces of dogs suffering from recurrent urinary tract infections. Production of fimbriae was demonstrated by electron microscopy and the presence of genes for two different types of major fimbrial subunits (MR/P-like or UCA-like) was demonstrated by Southern hybridisation. These genes were absent in the Proteus vulgaris, Providentia rettgeri and Morganella morganii strains tested. All but one P. mirabilis strains were haemolytic and most strains produced fimbriae albeit in different amounts. The UCA fimbrial subunits from dog and human isolates have identical molecular weights and N-terminal sequences and are immunologically cross reactive. It was concluded that dog uropathogenic P. mirabilis strains are very similar to human uropathogenic P. mirabilis strains.

  11. The role of genomic islands in Escherichia coli K1 interactions with intestinal and kidney epithelial cells.

    PubMed

    Yousuf, Farzana Abubakar; Rafiq, Sahar; Siddiqui, Ruqaiyyah; Khan, Naveed Ahmed

    2016-04-01

    The completion of Escherichia coli K1 genome has identified several genomic islands that are present in meningitis-causing E. coli RS218 but absent in the non-pathogenic E. coli MG1655. In this study, the role of various genomic islands in E. coli K1 interactions with intestinal epithelial cells (Caco-2) and kidney epithelial cells (MA104) was determined. Using association assays, invasion assays, and intracellular survival assays, the findings revealed that the genomic island deletion mutants of RS218 related to P fimbriae, S fimbriae, F17-like fimbriae, non-fimbrial adhesins, Hek and hemagglutinin, protein secretion system (T1SS for hemolysin; T2SS; T5SS for antigen 43), Iro system and hmu system), invasins (CNF1, IbeA), toxins (α-hemolysin), K1 capsule biosynthesis, metabolism (d-serine catabolism, dihydroxyacetone, glycerol, and glyoxylate metabolism), prophage genes, showed reduced interactions with both cell types. Next, we determined the role of various genomic islands in E. coli K1 resistance to serum. When exposed to the normal human serum, the viability of the genomic island deletion mutants related to adhesins such as S fimbriae, P fimbriae, F17-like fimbriae, non-fimbrial adhesins, Hek and hemagglutinin, antigen 43 and T5SS for antigen 43, T2SS, and T1SS for hemolysin, Iro system and hmu system, prophage genes, metabolism (sugar metabolism and d-serine catabolism), K1 capsule biosynthesis, and invasins such as CNF1 was affected, suggesting their role in bacteremia. The characterization of these genomic islands should reveal mechanisms of E. coli K1 pathogenicity that could be of value as therapeutic targets.

  12. [Avian Escherichia coli virulence factors associated with coli septicemia in broiler chickens].

    PubMed

    Ramirez Santoyo, R M; Moreno Sala, A; Almanza Marquez, Y

    2001-01-01

    In order to detect phenotypic characteristics associated with pathogenicity, 25 strains of Escherichia coli, isolated from clinical cases of colisepticemia in broiler chickens, were examined to determine the following properties: colicinogenicity, colicin V production, type 1 fimbriae, hemolysin expression and motility. Colicinogenicity occurred in 72% of the strains, 56% of all strains produced colicin V, 84% were positive for type 1 fimbriae and 80% were positive for motility. None of the strains had hemolytic activity; however, all of them, expressed at least one of the other characteristics studied. These results suggest that the diversity of phenotypes detected partially explain the multifactorial nature of avian colisepticemia.

  13. [Study on virulence factors associated with biofilm formation and phylogenetic groupings in Escherichia coli strains isolated from patients with cystitis].

    PubMed

    Tiba, Monique Ribeiro; Nogueira, Gustavo Prado; Leite, Domingos da Silva

    2009-01-01

    Escherichia coli samples isolated from female patients with cystitis were characterized with regard to the presence of virulence factors associated with biofilm formation and phylogenetic groupings. Polymerase chain reaction results demonstrated that all the samples were positive for the gene fimH (type 1 fimbriae), 91 for fliC (flagellins), 50 for papC (P fimbriae), 44 for kpsMTII (capsules) and 36 for flu (antigen 43). The results from assays to quantify the biofilm formation demonstrated that 44 samples produced biofilm on polystyrene microplates and 56 samples produced weak or no biofilm. We also confirmed that Escherichia coli samples were present in phylogenetic groups B2 and D.

  14. [Update on current care guidelines: ovarian cancer].

    PubMed

    Leminen, Arto; Auranen, Annika; Bützow, Ralf; Hietanen, Sakari; Komulainen, Marja; Kuoppala, Tapio; Mäenpää, Johanna; Puistola, Ulla; Vuento, Maarit; Vuorela, Piia; Yliskoski, Merja

    2012-01-01

    Ovarian cancer is the most lethal gynaecological cancer. It appears that seemingly ovarian or primary peritoneal carcinomas, in fact, originate from fimbriae. BRCA1/2 mutation carriers are recommended for the removal of ovaries and fimbriae, to reduce the risk of cancer. Treatment of epithelial ovarian cancer is based on the combination of surgery and chemotherapy. The residual tumour volume at the primary operation is the most important predictive factor of survival. The best response at the primary treatment is observed with combination chemotherapy with taxane and platinum. Adding bevacitzumab to first line chemotherapy may improve survival.

  15. Haemagglutinins and adherence properties to HeLa and intestine 407 cells of Klebsiella pneumoniae and Klebsiella oxytoca isolates.

    PubMed

    Podschun, R; Heineken, P; Sonntag, H G

    1987-03-01

    The occurrence of haemagglutination (HA) and adherence properties were examined in 50 strains of K. pneumoniae and 17 K. oxytoca strains isolated from humans. All isolates except three exhibited HA activity. Mannose-sensitive haemagglutinins (MSHA) were expressed by the majority of K. pneumoniae strains, but only by one K. oxytoca isolate. Mannose-resistant haemagglutination (MRHA) to human or guinea pig erythrocytes could not be detected; haemagglutinins of the MR/K type were found in both species with similar frequencies. Adhesive properties could be demonstrated in K. pneumoniae as often as in K. oxytoca: About half of the strains adhered to two human cell lines: HeLa and Intestine 407. The incidence of HA activity was similar in adhering and nonadhering strains. A correlation between MSHA, MR/K-HA and adherence to tissue-cultured cells could not be detected.

  16. Drug resistance and adherence to human intestines of enteroaggregative Escherichia coli.

    PubMed

    Yamamoto, T; Echeverria, P; Yokota, T

    1992-04-01

    Clinical isolates of enteroaggregative Escherichia coli (EAggEC) were tested for their in vitro susceptibilities to 27 antimicrobial agents. Marked drug resistance was observed with sulfamethoxazole, ampicillin, and chloramphenicol in contrast to such antimicrobial agents as cefixime, sparfloxacin, and ciprofloxacin. One of the EAggEC strains carried a plasmid that conferred on its host resistance to ampicillin, tetracycline, sulfamethoxazole, streptomycin, and spectinomycin and an ability to adhere to child ileal villi or HeLa cells in the characteristic aggregative pattern. This plasmid also mediated D-mannose-resistant hemagglutinin production and bacterial clump formation (autoagglutination). The data demonstrate appearance of marked drug resistance and an intestine-adherence and drug-resistance plasmid in the newest category of diarrheagenic E. coli.

  17. Correlation between uropathogenic properties of Escherichia coli from urinary tract infections and the antibody-coated bacteria test and comparison with faecal strains.

    PubMed Central

    Brooks, H. J.; Benseman, B. A.; Peck, J.; Bettelheim, K. A.

    1981-01-01

    Strains of Escherichia coli isolated from adult females with symptomatic urinary tract infection were found to possess the following properties significantly more frequently than faecal strains: (i) high K-antigen titre: (ii) haemolysin; (iii) type 1 pili; (iv) mannose-resistant haemagglutination; (v) fermentation of dulcitol and salicin; (vi) O serotype 2, 6 and 75; (vii) H serotype 1. E. coli isolated form urine specimens containing significant numbers of antibody-coated bacteria were richer in these seven properties than strains from urines without detectable antibody coated bacteria. The O and H serotypes of E. coli obtained from patients with urinary tract infection in two New Zealand cities were compared with those reported in the world literature and found to be similar. PMID:6114119

  18. Virulence Properties of Escherichia coli 83972, a Prototype Strain Associated with Asymptomatic Bacteriuria

    PubMed Central

    Hull, Richard A.; Rudy, Delbert C.; Donovan, William H.; Wieser, Inge E.; Stewart, Colleen; Darouiche, Rabih O.

    1999-01-01

    Little is known about bacteria associated with asymptomatic bacteriuria (ABU) with regard to urinary tract colonization mechanisms. In this study, virulence properties of Escherichia coli 83972, a strain that was isolated from a clinical ABU episode, were examined. The genetic potential for expression of P and type 1 pili was demonstrated, and DNA sequences related to type 1C and G (UCA) pilus genes were also detected. However, E. coli 83972 did not express d-mannose-resistant or d-mannose-sensitive hemagglutination after growth under standard conditions in vitro or upon isolation from the urine of colonized test subjects. Limited uroepithelial cell adherence was observed in vivo, and weak d-mannose-sensitive hemagglutination was detected after extended growth in urine in vitro. PMID:9864249

  19. Susceptibility of porcine intestine to pilus-mediated adhesion by some isolates of piliated enterotoxigenic Escherichia coli increases with age.

    PubMed Central

    Nagy, B; Casey, T A; Whipp, S C; Moon, H W

    1992-01-01

    Two porcine isolates of enterotoxigenic Escherichia coli (ETEC) (serogroup O157 and O141) derived from fatal cases of postweaning diarrhea and lacking K88, K99, F41, and 987P pili (4P- ETEC) were tested for adhesiveness to small-intestinal epithelia of pigs of different ages. Neither strain adhered to isolated intestinal brush borders of newborn (1-day-old) pigs in the presence of mannose. However, mannose-resistant adhesion occurred when brush borders from 10-day- and 3- and 6-week-old pigs were used. Electron microscopy revealed that both strains produced fine (3.5-nm) and type 1 pili at 37 degrees C but only type 1 pili at 18 degrees C. Mannose-resistant in vitro adhesion to brush borders of older pigs correlated with the presence of fine pili. These strains produced predominantly fine pili in ligated intestinal loops of both older and newborn pigs, but adherence was greater in loops in older pigs. Immunoelectron microscopic studies, using antiserum raised against piliated bacteria and absorbed with nonpiliated bacteria, of samples from brush border adherence studies revealed labelled appendages between adherent bacteria and intestinal microvilli. Orogastric inoculation of pigs weaned at 10 and 21 days of age indicated significantly (P less than 0.001) higher levels of adhesion by the ETEC to the ileal epithelia of older pigs than to that of younger ones. We suggest that small-intestinal adhesion and colonization by these ETEC isolates is dependent on receptors that develop progressively with age during the first 3 weeks after birth. Furthermore, our data are consistent with the hypothesis that the fine pili described mediate intestinal adhesion by the 4P- ETEC strains studied. Images PMID:1347758

  20. Susceptibility of porcine intestine to pilus-mediated adhesion by some isolates of piliated enterotoxigenic Escherichia coli increases with age.

    PubMed

    Nagy, B; Casey, T A; Whipp, S C; Moon, H W

    1992-04-01

    Two porcine isolates of enterotoxigenic Escherichia coli (ETEC) (serogroup O157 and O141) derived from fatal cases of postweaning diarrhea and lacking K88, K99, F41, and 987P pili (4P- ETEC) were tested for adhesiveness to small-intestinal epithelia of pigs of different ages. Neither strain adhered to isolated intestinal brush borders of newborn (1-day-old) pigs in the presence of mannose. However, mannose-resistant adhesion occurred when brush borders from 10-day- and 3- and 6-week-old pigs were used. Electron microscopy revealed that both strains produced fine (3.5-nm) and type 1 pili at 37 degrees C but only type 1 pili at 18 degrees C. Mannose-resistant in vitro adhesion to brush borders of older pigs correlated with the presence of fine pili. These strains produced predominantly fine pili in ligated intestinal loops of both older and newborn pigs, but adherence was greater in loops in older pigs. Immunoelectron microscopic studies, using antiserum raised against piliated bacteria and absorbed with nonpiliated bacteria, of samples from brush border adherence studies revealed labelled appendages between adherent bacteria and intestinal microvilli. Orogastric inoculation of pigs weaned at 10 and 21 days of age indicated significantly (P less than 0.001) higher levels of adhesion by the ETEC to the ileal epithelia of older pigs than to that of younger ones. We suggest that small-intestinal adhesion and colonization by these ETEC isolates is dependent on receptors that develop progressively with age during the first 3 weeks after birth. Furthermore, our data are consistent with the hypothesis that the fine pili described mediate intestinal adhesion by the 4P- ETEC strains studied.

  1. Heterogeneity among Strains of Diffusely Adherent Escherichia coli Isolated in Brazil

    PubMed Central

    Lopes, Lucia M.; Fabbricotti, Sandra H.; Ferreira, Antonio J. P.; Kato, Maria A. M. F.; Michalski, Jane; Scaletsky, Isabel C. A.

    2005-01-01

    One hundred twelve diffusely adherent Escherichia coli strains isolated from children in a case control study were evaluated for virulence-associated characteristics, serotyping, antibiotic resistance, and plasmid profiles. Half of the strains hybridized with the probes for icuA (aerobactin) and fimH (type 1 pili); daaE (F1845 fimbriae), afa (afimbrial Dr adhesin), agg-3A (aggregative adhesion fimbria type III fimbriae), pap (P fimbriae), astA (EAST1 toxin), and shET1 (Shigella enterotoxin 1) sequences were present in <20% of the strains. The shET1 gene was noted most frequently in strains isolated from patients. A minority (7%) of the strains produced hemolysin or colicin or showed cytotoxic effects on Vero cells. Forty-five different serotypes were found. The majority (70%) of the strains presented multiple antibiotic resistance. Antibiotic resistance and diffuse adherence were located on the same conjugative plasmids. These results suggest that the transfer of these potential virulence markers could be important in the epidemiology of diffusely adherent E. coli. PMID:15815034

  2. Proceedings of the Edgewood Chemical Biological Center In-House Laboratory Independent Research and Surface Science Initiative Programs FY12

    DTIC Science & Technology

    2012-01-01

    provide fundamental knowledge regarding extracellular proteins produced by Gram -negative bacteria, specifically those components important for bacterial...resistance. To address this possibility, we are characterizing the extracellular, fimbriae, and whole cell proteins produced by the pathogenic Gram ...function in survival mechanisms such as host immune system modulation [1] and biofilm formation.𔃼’ Additionally, Gram -negative bacteria release a

  3. Complete genome sequences of curli-negative and curli-positive isolates of foodborne Escherichia coli O157:H7 strain 86-24

    USDA-ARS?s Scientific Manuscript database

    Escherichia coli O157:H7 strain 86-24 does not produce curli fimbriae, but can give rise to curli-positive isolates at a variable frequency. Here, we report the whole-genome sequences of curli-negative and curli-positive isolates of strain 86-24....

  4. A FaeG-FedF-LT192 fusion elicits protective anti-adhesin and antitoxin antibodies against porcine enterotoxigenic Escherichia coli

    USDA-ARS?s Scientific Manuscript database

    Enterotoxigenic Escherichia coli (ETEC) strains expressing K88 or F18 fimbriae and heat-labile (LT) and/or heat-stable (ST) toxins are the major cause of diarrhea in young pigs. Effective vaccines inducing anti-adhesin (anti-K88 & anti-F18) and antitoxin (anti-LT & anti-ST) 5 immunity would provide ...

  5. Stx1 prophage excision in Escherichia coli strain PA20 confers strong curli and biofilm formation by restoring native mlrA

    USDA-ARS?s Scientific Manuscript database

    Prophage insertions in Escherichia coli O157:H7 mlrA contribute to the low expression of curli fimbriae and biofilm observed in many clinical isolates. Varying levels of CsgD-dependent curli/biofilm expression are restored to strains bearing prophage insertions in mlrA by mutation of regulatory gene...

  6. Media composition and incubation temperature affect Congo red dye affinity of Shiga toxin-producing Escherichia coli

    USDA-ARS?s Scientific Manuscript database

    Background: Escherichia coli biofilm formation is dependent on curli fimbriae and cellulose, and the expression of both varies among Shiga toxin-producing E. coli (STEC). Curli and cellulose expression are often identified by their affinity for Congo red dye (CR) but media composition and incubation...

  7. Growth media and temperature effects on biofilm formation by serotype O157:H7 and non-O157 Shiga toxin-producing Escherichia coli

    USDA-ARS?s Scientific Manuscript database

    Biofilm formation in most Escherichia coli strains is dependent on curli fimbriae and cellulose, and the expression of both varies widely among pathogenic strains. Curli and cellulose expression are often identified by their affinity for Congo red dye (CR). However, media composition and incubation ...

  8. Virulence patterns and long-range genetic mapping of extraintestinal Escherichia coli K1, K5, and K100 isolates: use of pulsed-field gel electrophoresis.

    PubMed Central

    Ott, M; Bender, L; Blum, G; Schmittroth, M; Achtman, M; Tschäpe, H; Hacker, J

    1991-01-01

    A total of 127 extraintestinal Escherichia coli strains of the capsule serotypes K1, K5, and K100 from human and animal sources were analyzed for DNA sequences specific for the genes for various adhesins (P fimbriae [pap] and P-related sequences [prs], S fimbriae [sfa]/F1C fimbriae [foc], and type I fimbriae [fim]), aerobactin (aer), and hemolysin (hly). The expression of corresponding virulence factors was also tested. Twenty-four selected strains were analyzed by long-range DNA mapping to evaluate their genetic relationships. DNA sequences for the adhesins were often found in strains not expressing them, while strains with hemolysin and aerobactin genes usually did express them. Different isolates of the same serotype often expressed different virulence patterns. The use of virulence-associated gene probes for Southern hybridization with genomic DNA fragments separated by pulsed-field gel electrophoresis revealed that a highly heterogeneous restriction fragment length and hybridization pattern existed even within strains of the same serotype. Long-range DNA mapping is therefore useful for the evaluation of genetic relatedness among individual isolates and facilitates the performance of precise molecular epidemiology. Images PMID:1677349

  9. A novel CsrA titration mechanism regulates fimbrial gene expression in Salmonella typhimurium

    PubMed Central

    Sterzenbach, Torsten; Nguyen, Kim T; Nuccio, Sean-Paul; Winter, Maria G; Vakulskas, Christopher A; Clegg, Steven; Romeo, Tony; Bäumler, Andreas J

    2013-01-01

    A hierarchical control of fimbrial gene expression limits laboratory grown cultures of Salmonella enterica serovar typhimurium (S. typhimurium) to the production of type I fimbriae encoded by the fimAICDHF operon. Here we show that an unlikely culprit, namely the 5′-untranslated region (5′-UTR) of a messenger (m)RNA, coordinated the regulation. Binding of CsrA to the 5′-UTR of the pefACDEF transcript was required for expression of plasmid-encoded fimbriae. The 5′-UTR of the fimAICDHF transcript cooperated with two small untranslated RNAs, termed CsrB and CsrC, in antagonizing the activity of the RNA binding protein CsrA. Through this post-transcriptional mechanism, the 5′-UTR of the fimAICDHF transcript prevented production of PefA, the major structural subunit of plasmid-encoded fimbriae. This regulatory mechanism limits the costly expression of plasmid-encoded fimbriae to host environments in a mouse model. Collectively, our data suggest that the 5′-UTR of an mRNA coordinates a hierarchical control of fimbrial gene expression in S. typhimurium. PMID:24056837

  10. A novel CsrA titration mechanism regulates fimbrial gene expression in Salmonella typhimurium.

    PubMed

    Sterzenbach, Torsten; Nguyen, Kim T; Nuccio, Sean-Paul; Winter, Maria G; Vakulskas, Christopher A; Clegg, Steven; Romeo, Tony; Bäumler, Andreas J

    2013-10-30

    A hierarchical control of fimbrial gene expression limits laboratory grown cultures of Salmonella enterica serovar typhimurium (S. typhimurium) to the production of type I fimbriae encoded by the fimAICDHF operon. Here we show that an unlikely culprit, namely the 5'-untranslated region (5'-UTR) of a messenger (m)RNA, coordinated the regulation. Binding of CsrA to the 5'-UTR of the pefACDEF transcript was required for expression of plasmid-encoded fimbriae. The 5'-UTR of the fimAICDHF transcript cooperated with two small untranslated RNAs, termed CsrB and CsrC, in antagonizing the activity of the RNA binding protein CsrA. Through this post-transcriptional mechanism, the 5'-UTR of the fimAICDHF transcript prevented production of PefA, the major structural subunit of plasmid-encoded fimbriae. This regulatory mechanism limits the costly expression of plasmid-encoded fimbriae to host environments in a mouse model. Collectively, our data suggest that the 5'-UTR of an mRNA coordinates a hierarchical control of fimbrial gene expression in S. typhimurium.

  11. Curli variants of enterohemorrhagic Escherichia coli O157:H7 display distinct survival fitness

    USDA-ARS?s Scientific Manuscript database

    Curli are adhesive fimbriae of Enterobactericaeae and are involved in surface attachment, cell aggregation and biofilm formation. They also mediate host cell invasion and are potent inducers of the host inflammatory response. Here we report that curli variants are distributed widely in Enterohemorrh...

  12. Gene duplication and promoter mutation expand the range csgD-dependent biofilm responses in a STEC population

    USDA-ARS?s Scientific Manuscript database

    Expression of Escherichia coli major biofilm components, curli fimbriae and cellulose, require the CsgD transcription factor. A complex regulatory network allows environmental control of csgD transcription and biofilm formation. However, most clinical serotype O157:H7 strains contain prophage inser...

  13. Reduction of Salmonella Enteritidis in the spleens of hens by bacterins that vary in fimbrial protein SefD

    USDA-ARS?s Scientific Manuscript database

    Gene sefD is part of operon sefABCD, and it is required for production of the SEF14 fimbria by Salmonella Enteritidis. We compared strains that varied in SefD content for their ability to reduce recovery of Salmonella Enteritidis from the spleens of hens infected by parenteral challenge. The two bac...

  14. A conserved fold for fimbrial components revealed by the crystal structure of a putative fimbrial assembly protein (BT1062) from Bacteroides thetaiotaomicron at 2.2 Å resolution

    PubMed Central

    Xu, Qingping; Abdubek, Polat; Astakhova, Tamara; Axelrod, Herbert L.; Bakolitsa, Constantina; Cai, Xiaohui; Carlton, Dennis; Chen, Connie; Chiu, Hsiu-Ju; Chiu, Michelle; Clayton, Thomas; Das, Debanu; Deller, Marc C.; Duan, Lian; Ellrott, Kyle; Farr, Carol L.; Feuerhelm, Julie; Grant, Joanna C.; Grzechnik, Anna; Han, Gye Won; Jaroszewski, Lukasz; Jin, Kevin K.; Klock, Heath E.; Knuth, Mark W.; Kozbial, Piotr; Krishna, S. Sri; Kumar, Abhinav; Marciano, David; McMullan, Daniel; Miller, Mitchell D.; Morse, Andrew T.; Nigoghossian, Edward; Nopakun, Amanda; Okach, Linda; Puckett, Christina; Reyes, Ron; Sefcovic, Natasha; Tien, Henry J.; Trame, Christine B.; van den Bedem, Henry; Weekes, Dana; Wooten, Tiffany; Yeh, Andrew; Zhou, Jiadong; Hodgson, Keith O.; Wooley, John; Elsliger, Marc-Andre; Deacon, Ashley M.; Godzik, Adam; Lesley, Scott A.; Wilson, Ian A.

    2010-01-01

    BT1062 from Bacteroides thetaiotaomicron is a homolog of Mfa2 (PGN0288 or PG0179), which is a component of the minor fimbriae in Porphyromonas gingivalis. The crystal structure of BT1062 revealed a conserved fold that is widely adopted by fimbrial components. PMID:20944223

  15. Virulence and metabolic characteristics of Salmonella Enteritidis sefD variants in hens.

    USDA-ARS?s Scientific Manuscript database

    Salmonella Enteritidis is one of a few pathogenic Salmonella enterica serotypes that have SEF14 fimbriae encoded by the sef operon, which consists of 4 co-transcribed genes sefABCD that are regulated by sefR. To explore the function of sefD within the infection pathway resulting in egg contamination...

  16. How methylglyoxal kills bacteria: An ultrastructural study.

    PubMed

    Rabie, Erika; Serem, June Cheptoo; Oberholzer, Hester Magdalena; Gaspar, Anabella Regina Marques; Bester, Megan Jean

    2016-01-01

    Antibacterial activity of honey is due to the presence of methylglyoxal (MGO), H2O2, bee defensin as well as polyphenols. High MGO levels in manuka honey are the main source of antibacterial activity. Manuka honey has been reported to reduce the swarming and swimming motility of Pseudomonas aeruginosa due to de-flagellation. Due to the complexity of honey it is unknown if this effect is directly due to MGO. In this ultrastructural investigation the effects of MGO on the morphology of bacteria and specifically the structure of fimbriae and flagella were investigated. MGO effectively inhibited Gram positive (Bacillus subtilis; MIC 0.8 mM and Staphylococcus aureus; MIC 1.2 mM) and Gram negative (P. aeruginosa; MIC 1.0 mM and Escherichia coli; MIC 1.2 mM) bacteria growth. The ultrastructural effects of 0.5, 1.0 and 2 mM MGO on B. substilis and E. coli morphology was then evaluated. At 0.5 mM MGO, bacteria structure was unaltered. For both bacteria at 1 mM MGO fewer fimbriae were present and the flagella were less or absent. Identified structures appeared stunted and fragile. At 2 mM MGO fimbriae and flagella were absent while the bacteria were rounded with shrinkage and loss of membrane integrity. Antibacterial MGO causes alterations in the structure of bacterial fimbriae and flagella which would limit bacteria adherence and motility.

  17. Draft Genome Sequence and Gene Annotation of the Uropathogenic Bacterium Proteus mirabilis Pr2921

    PubMed Central

    Giorello, F. M.; Romero, V.; Farias, J.; Scavone, P.; Umpiérrez, A.; Zunino, P.

    2016-01-01

    Here, we report the genome sequence of Proteus mirabilis Pr2921, a uropathogenic bacterium that can cause severe complicated urinary tract infections. After gene annotation, we identified two additional copies of ucaA, one of the most studied fimbrial protein genes, and other fimbriae related-proteins that are not present in P. mirabilis HI4320. PMID:27340058

  18. Serum Antibodies Protect against Intraperitoneal Challenge with Enterotoxigenic Escherichia coli

    PubMed Central

    Yang, Xinghong; Thornburg, Theresa; Holderness, Kathryn; Suo, Zhiyong; Cao, Ling; Lim, Timothy; Avci, Recep; Pascual, David W.

    2011-01-01

    To assess whether anticolonization factor antigen I (CFA/I) fimbriae antibodies (Abs) from enterotoxigenic Escherichia coli (ETEC) can protect against various routes of challenge, BALB/c mice were immunized with a live attenuated Salmonella vaccine vector expressing CFA/I fimbriae. Vaccinated mice elicited elevated systemic IgG and mucosal IgA Abs, unlike mice immunized with the empty Salmonella vector. Mice were challenged with wild-type ETEC by the oral, intranasal (i.n.), and intraperitoneal (i.p.) routes. Naïve mice did not succumb to oral challenge, but did to i.n. challenge, as did immunized mice; however, vaccinated mice were protected against i.p. ETEC challenge. Two intramuscular (i.m.) immunizations with CFA/I fimbriae without adjuvant conferred 100% protection against i.p. ETEC challenge, while a single 30 μg dose conferred 88% protection. Bactericidal assays showed that ETEC is highly sensitive to anti-CFA/I sera. These results suggest that parenteral immunization with purified CFA/I fimbriae can induce protective Abs and may represent an alternative method to elicit protective Abs for passive immunity to ETEC. PMID:22007145

  19. Variability of the fimA gene in Porphyromonas gingivalis isolated from periodontitis and non-periodontitis patients

    PubMed Central

    León, Rubén; Blanc, Vanessa; Herrera, David; Sanz, Mariano

    2013-01-01

    Objective: The goal of this study was to determine the genetic variability of the fimA gene in Porphyromonas gingivalis isolates from Spanish patients. Study Design: Pooled subgingival samples were taken, processed and cultured in non-selective blood agar medium. Pure cultures of one to six isolates per patient were obtained and PCR and PCR-RFLP were used for fimbrillin gene (fimA) type determination of the extracted genomic (DNA). Results: Two hundred and twenty four Porphyromonas gingivalis isolates from 65 patients were analyzed consisting of 15 non-periodontitis patients (66 isolates) and 50 with periodontitis (158 isolates). Genotype II was the most prevalent (50.9%), while the other types of fimbriae did not exceed fifteen percent of prevalence. Isolates with types II and IV of fimbriae were significantly more prevalent in periodontitis patients than isolates with genotype I. Co-infection was observed in 17.65% of the patients analyzed. Conclusion: The results suggest that in this population Porphyromonas gingivalis with type II of fimbriae are significantly more predominant in periodontitis patients than genotype I. Key words:Fimbriae, genotype, porphyromonas gingivalis, periodontitis. PMID:23229246

  20. RcsB contributes to the distinct stress fitness between Escherichia coli O157:H7 curli variants of 1993 hamburger-associated outbreak strains

    USDA-ARS?s Scientific Manuscript database

    Curli are adhesive fimbriae of Enterobactericaeae and are involved in surface attachment, cell aggregation and biofilm formation. We previously reported that natural curli variants of E. coli O157:H7 (EcO157) displayed distinct acid resistance; however, this difference was not linked to the curli fi...

  1. Biochemical characteristic of biofilm of uropathogenic Escherichia coli Dr(+) strains.

    PubMed

    Zalewska-Piątek, Beata; Wilkanowicz, Sabina; Bruździak, Piotr; Piątek, Rafał; Kur, Józef

    2013-07-19

    Urinary tract infections caused by Escherichia coli are very common health problem in the developed countries. The virulence of the uropathogenic E. coli Dr(+) IH11128 is determined by Dr fimbriae, which are homopolymeric structures composed of DraE subunits with the DraD protein capping the fiber. In this study, we have analyzed the structural and biochemical properties of biofilms developed by E. coli strains expressing Dr fimbriae with or without the DraD tip subunit and the surface-exposed DraD protein. We have also demonstrated that these E. coli strains form biofilms on an abiotic surface in a nutrient-dependent fashion. We present evidence that Dr fimbriae are necessary during the first stage of bacterial interaction with the abiotic surface. In addition, we reveal that the DraD alone is also sufficient for the initial surface attachment at an even higher level than Dr fimbriae and that chloramphenicol is able to reduce the normal attachment of the analyzed E. coli. The action of chloramphenicol also shows that protein synthesis is required for the early events of biofilm formation. Additionally, we have identified reduced exopolysaccharide coverage in E. coli that express only Dr fimbrial polyadhesins at the cell surface with or without the DraD capping subunit. Copyright © 2013 Elsevier GmbH. All rights reserved.

  2. Fimbrial-mediated colonization of murine teeth by Actinomyces naeslundii.

    PubMed

    Beem, J E; Hurley, C G; Nesbitt, W E; Croft, D F; Marks, R G; Cisar, J O; Clark, W B

    1996-08-01

    Groups of mice fed diets high in sucrose or glucose were orally inoculated with 10(10), 10(9) or 10(8) colony-forming units of one of the following Actinomyces naeslundii strains possessing the type 1 (T1+) and/or the type 2 (T2+) fimbriae: T14VJ1 (T1+, T2+), 5519 (T1+), 5951 (T2+), and 147 (non-fimbriated). Ninety-six hours after inoculation their upper jaws were cultured to look at the implantation of each of these strains on the teeth. In mice fed a sucrose diet, regardless of the presence or absence of fimbriae, each bacterial strain colonized 100% of the mice at the highest inoculation doses of the infecting organism. But at a dose of 10(8), T14V-J1 was the only strain which colonized 100% (12/12) of the mice, 5519 colonized 10/11, 5951 colonized 9/11 and 147 colonized 7/11. These differences were not statistically significant. When mice were fed a high-glucose diet, 100% infection was achieved with strains T14V-J1, 5519 and 5951 only at the highest dose of 10(10) colony-forming units. Strain 147 colonized in 8/9 of the mice at that dosage. At lower dosages, no bacterial strain implanted in 100% of the mice. In the glucose experiment at a dose of 10(8), strains expressing the T1 fimbriae implanted significantly better than strains without the T1 fimbriae. At a dose of 10(9) colony-forming units, the parent strain T14V-J1 implanted significantly better than strains without the T1 fimbriae. Similarly, strain 5519 (T1+) implanted significantly better than 5951 and implanted better than 147, although the difference was not significant. These results suggest that while the presence of the T1 and T2 fimbriae may confer some advantage in the establishment of these organisms in vivo, even the strains without fimbriae were able to colonize. Strains T14VJ1 and 5519 were found to bind well to hydroxyapatite treated with mouse saliva, while strains 5951 and 147 did not. Only T2 fimbriated strains T14V-J1 and 5951 exhibited a lactose-reversible coaggreation with indigenous

  3. Genetic Characterization and Immunogenicity of Coli Surface Antigen 4 from Enterotoxigenic Escherichia coli when It Is Expressed in a Shigella Live-Vector Strain

    PubMed Central

    Altboum, Zeev; Levine, Myron M.; Galen, James E.; Barry, Eileen M.

    2003-01-01

    The genes that encode the enterotoxigenic Escherichia coli (ETEC) CS4 fimbriae, csaA, -B, -C, -E, and -D′, were isolated from strain E11881A. The csa operon encodes a 17-kDa major fimbrial subunit (CsaB), a 40-kDa tip-associated protein (CsaE), a 27-kDa chaperone-like protein (CsaA), a 97-kDa usher-like protein (CsaC), and a deleted regulatory protein (CsaD′). The predicted amino acid sequences of the CS4 proteins are highly homologous to structural and assembly proteins of other ETEC fimbriae, including CS1 and CS2, and to CFA/I in particular. The csaA, -B, -C, -E operon was cloned on a stabilized plasmid downstream from an osomotically regulated ompC promoter. pGA2-CS4 directs production of CS4 fimbriae in both E. coli DH5α and Shigella flexneri 2a vaccine strain CVD 1204, as detected by Western blot analysis and bacterial agglutination with anti-CS4 immune sera. Electron-microscopic examination of Shigella expressing CS4 confirmed the presence of fimbriae on the bacterial surface. Guinea pigs immunized with CVD 1204(pGA2-CS4) showed serum and mucosal antibody responses to both the Shigella vector and the ETEC fimbria CS4. Among the seven most prevalent fimbrial antigens of human ETEC, CS4 is the last to be cloned and sequenced. These findings pave the way for CS4 to be included in multivalent ETEC vaccines, including an attenuated Shigella live-vector-based ETEC vaccine. PMID:12595452

  4. Human decay-accelerating factor and CEACAM receptor-mediated internalization and intracellular lifestyle of Afa/Dr diffusely adhering Escherichia coli in epithelial cells.

    PubMed

    Guignot, Julie; Hudault, Sylvie; Kansau, Imad; Chau, Ingrid; Servin, Alain L

    2009-01-01

    We used transfected epithelial CHO-B2 cells as a model to identify the mechanism mediating internalization of Afa/Dr diffusely adhering Escherichia coli. We provide evidence that neither the alpha5 or beta1 integrin subunits nor alpha5beta1 integrin functioned as a receptor mediating the adhesion and/or internalization of Dr or Afa-III fimbria-positive bacteria. We also demonstrated that (i) whether or not the AfaD or DraD invasin subunits were present, there was no difference in the cell association and entry of bacteria and that (ii) DraE or AfaE-III adhesin subunits are necessary and sufficient to promote the receptor-mediated bacterial internalization into epithelial cells expressing human decay-accelerating factor (DAF), CEACAM1, CEA, or CEACAM6. Internalization of Dr fimbria-positive E. coli within CHO-DAF, CHO-CEACAM1, CHO-CEA, or CHO-CEACAM6 cells occurs through a microfilament-independent, microtubule-dependent, and lipid raft-dependent mechanism. Wild-type Dr fimbria-positive bacteria survived better within cells expressing DAF than bacteria internalized within CHO-CEACAM1, CHO-CEA, or CHO-CEACAM6 cells. In DAF-positive cells, internalized Dr fimbria-positive bacteria were located in vacuoles that contained more than one bacterium, displaying some of the features of late endosomes, including the presence of Lamp-1 and Lamp-2, and some of the features of CD63 proteins, but not of cathepsin D, and were acidic. No interaction between Dr fimbria-positive-bacterium-containing vacuoles and the autophagic pathway was observed.

  5. Investigation of engineered bacterial adhesins for opportunity to interface cells with abiotic materials

    NASA Astrophysics Data System (ADS)

    Terrell, Jessica L.; Dong, Hong; Holthoff, Ellen L.; Small, Meagan C.; Sarkes, Deborah A.; Hurley, Margaret M.; Stratis-Cullum, Dimitra N.

    2016-05-01

    The convenience of cellular genetic engineering has afforded the power to build `smart' synthetic biological tools with novel applications. Here, we have explored opportunities to hybridize engineered cells with inorganic materials toward the development of 'living' device-compatible systems. Cellular structural biology is engineerable based on the ability to rewrite genetic code to generate recombinant, foreign, or even unnatural proteins. With this capability on the biological end, it should be possible to achieve superior abio-compatibility with the inorganic materials that compose current microfabricated technology. This work investigated the hair-like appendages of Escherichia coli known as Type 1 fimbriae that enable natural adhesion to glycosylated substrates. Sequence alterations within the fimbrial gene cluster were found to be well-tolerated, evidenced by tagging the fimbriae with peptide-based probes. As a further development, fimbriae tips could be reconfigured to, in turn, alter cell binding. In particular, the fimbriae were fused with a genetically optimized peptide-for-inorganics to enable metal binding. This work established methodologies to systematically survey cell adhesion properties across a suite of fimbriae-modified cell types as well as to direct patterned cell adhesion. Cell types were further customized for added complexity including turning on secondary gene expression and binding to gold surfaces. The former demonstrates potential for programmable gene switches and the latter for interfacing biology with inorganic materials. In general, the incorporation of 'programmed' cells into devices can be used to provide the feature of dynamic and automated cell response. The outcomes of this study are foundational toward the critical feature of deliberate positioning of cells as configurable biocomponentry. Overall, cellular integration into bioMEMs will yield advanced sensing and actuation.

  6. The fimbrio-ovarian relation and its role on ovum picking in unexplained infertility: the fimbrio-ovarian accessibility tests.

    PubMed

    Ahmad-Thabet, S M

    2000-02-01

    To study the ovum pickup mechanism and the fimbrio-ovarian relation in fertile women and cases of unexplained infertility and to design a method for investigation and anticipation of the state of ovum pickup. Prospective clinical study. Department of Obstetrics and gynecology Kasr El Aini Hospital, Cairo University, Egypt. Fifty fertile and 25 cases of unexplained infertility. In all of cases the fimbrio-ovarian relation is studied by determining the length of the free ovarian border between the ovarian ligament and the fimbria ovarica that contains the corpus luteum (the ovulation border), the site of ovulation on that border and the length of the fimbria ovarica. It is also determined by a simple test "The fimbrio-ovarian accessibility test." The fimbriae are held by non-traumatizing grasper and their ability to reach and/or to cover the ovulation site are taken as a prove for successful ovum pickup. In addition, cases having suspected failed ovum picking are treated by controlled superovulation, also by drilling of the ovary at an accessible site to the fimbria ovarica and by freeing short fimbria ovarica with or without its advancement on the ovulation border. The ovulation site is found to be usually accessible to the fimbrial end of the tube (94%) in the fertile cases and not accessible (92%) in the infertile cases. Correction of the fimbrio-ovarian relation in the infertile cases was associated with a cumulative ongoing pregnancy rate of 68%. Failed ovum pickup should be considered one of the important causes of unexplained infertility. The suggested "fimbrio-ovarian accessibility" test may be useful for the elucidation of the state of ovum pickup in infertile cases. Correction of the fimbrio-ovarian relation, to make ovum pickup possible, should be considered a method for treating such cases.

  7. Characterization of F18 fimbrial genes fedE and fedF involved in adhesion and length of enterotoxemic Escherichia coli strain 107/86.

    PubMed

    Imberechts, H; Wild, P; Charlier, G; De Greve, H; Lintermans, P; Pohl, P

    1996-09-01

    Infection of susceptible weaned pigs with oedema disease strains of E. coli is associated with bacterial adhesion to the small intestine. F18 fimbria (previously named F107) was the first colonisation factor described on oedema disease strains, and its genetic determinant was cloned. In the present study, genes fedE and fedF were positioned in the F18 gene cluster, downstream of the major structural subunit gene fedA. Two fedE and two fedF mutants were identified that had lost their capacity to adhere to isolated porcine villi. Moreover, these mutants produced significantly longer fimbriae. In vitro adhesion tests, electron microscopy study, transcomplementation tests, and nucleotide sequence analysis indicated that proteins FedE and FedF are F18 minor subunits essential for fimbrial adhesion and effecting fimbrial length.

  8. Genetic Diversity among Escherichia coli Isolates Carrying f18 Genes from Pigs with Porcine Postweaning Diarrhea and Edema Disease

    PubMed Central

    Nagy, Béla; Wilson, Richard A.; Whittam, Thomas S.

    1999-01-01

    Multilocus enzyme electrophoresis was applied to detect allelic variation and multilocus genotypes (electrophoretic types [ETs]) among 43 Escherichia coli isolates from weaned pigs suffering from edema disease or from diarrhea. ETs were analyzed in relation to O serogroups and virulence genes (sta, stb, lt, stx2, and f18) by DNA hybridization. Genomic diversity was the lowest in serogroup O138, while virulence genes (stx2 and f18) were the most uniform in serogroup O139. In general, the serogroups or toxin and F18 fimbria types were not related to selected ETs, suggesting that the toxin and f18 fimbria genes in E. coli isolates from pigs with postweaning diarrhea or edema disease occur in a variety of chromosomal backgrounds. PMID:10203547

  9. Impact of cranberry on Escherichia coli cellular surface characteristics

    SciTech Connect

    Johnson, Brandy J.; Malanoski, Anthony P.; Ligler, Frances S.

    2008-12-19

    The anti-adhesive effects of cranberry have been attributed to both interactions of its components with the surface of bacterial cells and to inhibition of p-fimbriae expression. Previous reports also suggested that the presence of cranberry juice changed the Gram stain characteristics of Escherichia coli. Here, we show that the morphology of E. coli is changed when grown in the presence of juice or extract from Vaccinium macrocarpon (cranberry). Gene expression analysis indicates the down regulation of flagellar basal body rod and motor proteins. Consistent with this finding and previous reports, the SEM images indicate a decrease in the visible p-fimbriae. The iodine used in Gram-staining protocols was found to interact differently with the bacterial membrane when cells were cultured in spiked media. Slight alterations in the Gram stain protocol demonstrated that culturing in the presence of cranberry juice does not change the Gram stain characteristics contradicting other reports.

  10. [Investigation of hydrophobicity of Proteus vulgaris strains and ability of Proteus vulgaris and Proteus penneri strains to penetrate bladder membrane HCV T-29 cells ].

    PubMed

    Bartodziejska, Beata; Błaszczyk, Aleksandra; Wykrota, Marianna; Kwil, Iwona; Babicka, Dorota; Rózalski, Antoni

    2002-01-01

    Proteus bacilli play a particularly important role in urinary tract infections (UTI). Fimbriae and adherence ability and hemolysins production (HpmA, HlyA) are one of the factors of pathogenicity of these bacteria. In this paper we describe the invasion of HCV T-29 transitional bladder urothelial cells carcinoma strains of P. penneri, as well as P. vulgaris strains belonging to different serogroups. The cytotoxic effect was observed at 8 hour of incubation of the tested cells with P. vulgaris O21 and the same effect (complete lysis) at 6 hours by P. vulgaris O4 (this strain manifests maximal activity in the production of HlyA hemolysin). P. penneri strains, produce different types of fimbriae, expressed similar bacterial invasiveness. The hydrophobic properties of 25 P. vulgaris strains were also tested and only 3 strains occur to have hydrophobic cell surface.

  11. Detection of pap-, sfa- and afa-specific DNA sequences in Escherichia coli strains isolated from extraintestinal material.

    PubMed

    Bogyiová, E; Kmetová, M; Biros, E; Siegfried, L

    2002-01-01

    P-fimbriae, S-fimbriae and AFA-adhesins are virulence factors responsible for adherence of Escherichia coli strains to extraintestinal host-cell surface. Detection of pap-, sfa- and afa-specific sequences performed by PCR revealed 74% pap+, 65% sfa+, and 8.3% afa+ strains in a group of 84 extraintestinal E. coli isolates. Detection in a group of fecal strains showed 29% pap+, 21% sfa+ and 4% afa+ strains. pap together with sfa were found as the most frequent combination (56%) among extraintestinal isolates probably due to localization of pap- and sfa-operons on a common pathogenicity island. The occurrence of afa-specific sequence among 56 urine strains was 11%, although no afa+ strain was detected among 28 gynecological isolates. No strains with detected adhesin operons were found among twenty (24%) extraintestinal E. coli strains.

  12. Direct observation of type 1 fimbrial switching.

    PubMed

    Adiciptaningrum, Aileen M; Blomfield, Ian C; Tans, Sander J

    2009-05-01

    The defining feature of bacterial phase variation is a stochastic 'all-or-nothing' switching in gene expression. However, direct observations of these rare switching events have so far been lacking, obscuring possible correlations between switching events themselves, and between switching and other cellular events, such as division and DNA replication. We monitored the phase variation of type 1 fimbriae in individual Escherichia coli in real time and simultaneously tracked the chromosome replication process. We observed distinctive patterns of fim (fimbriae) expression in multiple genealogically related lineages. These patterns could be explained by a model that combines a single switching event with chromosomal fim replication, as well as the epigenetic inheritance of expressed fim protein and RNA, and their dilution by growth. Analysis of the moment of switching at sub-cell-cycle resolution revealed a correlation between fim switching and cell age, which challenges the traditional idea of phase variation as a random Poissonian phenomenon.

  13. Enhanced phagocytosis of group A streptococci M type 6 by oleic acid

    SciTech Connect

    Speert, D.P.; Quie, P.G.; Wannamaker, L.W.

    1981-04-01

    M protein, located on the surface fimbriae of group A streptococci, is antiphagocytic by unknown means. It is known that oleic acid kills group A streptococci and distorts the fimbriae. The effect of oleic acid on phagocytosis of group A streptococci was examined. Phagocytosis of a strain possessing M protein (M+) and its M- variant was assessed by uptake of radiolabeled bacteria and by chemiluminescence. The M- but not the M+ streptococci were well phagocytized and induced chemiluminescence. Oleic acid-killed and heat-killed streptococci (both M+ and M-) were readily phagocytized and induced sustained chemiluminescence. M+ streptococci killed by ultraviolet irradiation were inefficiently phagocytized and did not induce chemiluminescence. Oleic acid-killed M+ streptococci absorbed type-specific antibody. An extract of M protein reduced the bactericidal capacity of oleic acid. It is proposed that oleic acid may bind to and alter the M protein of group A streptococci and thereby enhance phagocytosis.

  14. [Bacterial adherence in pathogenesis of urinary tract infectious].

    PubMed

    Piédrola Angulo, Gonzalo

    2003-01-01

    The possibility of a colonization and later urinary infection is due to a first contact among a series of structures of the bacterium, denominated adhesins (fimbrica or no-fimbrica) and some receivers or ligands of the surface of the urinary epitelium. The bacterial fimbriae of Escherichia coli, of those that have been studied up to seven different types, are protean structures coded by the chromosomal DNA, being the most important those of type 1, in connection with the colonization of the low roads, and the type P, with the cystitis and pyelonephritis. They are studied with detail their different protean components and the very complex genetic regulation of their production, made of great interest in the pathogeny of these infections and in the possibility of their prevention. The receivers of each fimbriae type are also chemically different, and their knowledge would explain important clinical data.

  15. Crystal structure and analysis of HdaB: The enteroaggregative Escherichia coli AAF/IV pilus tip protein.

    PubMed

    Lee, Wei-Chao; Matthews, Steve; Garnett, James A

    2016-10-01

    Enteroaggregative Escherichia coli is the primary cause of pediatric diarrhea in developing countries. They utilize aggregative adherence fimbriae (AAFs) to promote initial adherence to the host intestinal mucosa, promote the formation of biofilms, and mediate host invasion. Five AAFs have been identified to date and AAF/IV is amongst the most prevalent found in clinical isolates. Here we present the X-ray crystal structure of the AAF/IV tip protein HdaB at 2.0 Å resolution. It shares high structural homology with members of the Afa/Dr superfamily of fimbriae, which are involved in host invasion. We highlight surface exposed residues that share sequence homology and propose that these may function in invasion and also non-conserved regions that could mediate HdaB specific adhesive functions. © 2016 The Authors Protein Science published by Wiley Periodicals, Inc. on behalf of The Protein Society.

  16. A two-component regulatory system modulates twitching motility in Dichelobacter nodosus.

    PubMed

    Kennan, Ruth M; Lovitt, Carrie J; Han, Xiaoyan; Parker, Dane; Turnbull, Lynne; Whitchurch, Cynthia B; Rood, Julian I

    2015-08-31

    Dichelobacter nodosus is the essential causative agent of footrot in sheep and type IV fimbriae-mediated twitching motility has been shown to be essential for virulence. We have identified a two-component signal transduction system (TwmSR) that shows similarity to chemosensory systems from other bacteria. Insertional inactivation of the gene encoding the response regulator, TwmR, led to a twitching motility defect, with the mutant having a reduced rate of twitching motility when compared to the wild-type and a mutant complemented with the wild-type twmR gene. The reduced rate of twitching motility was not a consequence of a reduced growth rate or decreased production of surface located fimbriae, but video microscopy indicated that it appeared to result from an overall loss of twitching directionality. These results suggest that a chemotactic response to environmental factors may play an important role in the D. nodosus-mediated disease process.

  17. Importance of Escherichia coli in young beef calves from northwestern Quebec.

    PubMed Central

    Ganaba, R; Bigras-Poulin, M; Fairbrother, J M; Bélanger, D

    1995-01-01

    The objectives of this study were: (i) to investigate the prevalence of Escherichia coli producing F5 (K99), F41, or F165 fimbriae and STa enterotoxin; (ii) to determine serum antibody levels against these fimbriae; (iii) and to examine the association between bacteriological and serological results and the presence of diarrhea, in beef calves from northwestern Quebec. A total of 373 live three to four week old calves and 27 dead calves were sampled between January and March 1991. No isolates positive for F5 were detected in live calves, and only one E. coli producing STa and F41 was isolated. Escherichia coli producing F41-like surface antigens or F165 fimbriae were isolated from 17.43% and 5.63% of live calves, respectively. Antibodies against F5, F41 and F165 were low. Escherichia coli isolates positive for F41-like surface antigen were most often observed in calves born between January and March. No association was found between bacteriological and serological findings, nor between these findings and diarrhea. Calves born from dams vaccinated against E. coli had higher median antibody levels than those born from unvaccinated dams. No E. coli positive for F5 or F41 fimbriae were isolated from dead calves. Escherichia coli with F41-like surface antigen or F165 were found in 55.56% and 11.11% of ileal samples; 4% and 16% of cecal samples, and 0% and 7.4% of colon samples, respectively. Escherichia coli positive for F41-like surface antigen were detected significantly more frequently in the ileum (chi (2)2df = 31.01, p < 0.001). PMID:7704838

  18. [Proteus bacilli: features and virulence factors].

    PubMed

    Rózalski, Antoni; Kwil, Iwona; Torzewska, Agnieszka; Baranowska, Magdalena; Staczek, Paweł

    2007-01-01

    In this article, different aspects of virulence factors of Proteus bacilii (P. mirabilis, P. vulgaris, P. penneri i P. hauseri) are presented. These are opportunistic pathogens that cause different kinds of infections, most frequently of the urinary tract. These bacteria have developed several virulence factors, such as adherence due to the presence of fimbriae or afimbrial adhesins, invasiveness, swarming phenomenon, hemolytic activity, urea hydrolysis, proteolysis, and endotoxicity. Below we focus on data concerning the molecular basis of the pathogenicity of Proteus bacilli.

  19. Generation of Salmonella ghost cells expressing fimbrial antigens of enterotoxigenic Escherichia coli and evaluation of their antigenicity in a murine model

    PubMed Central

    Kim, Chan Song; Hur, Jin; Eo, Seong Kug; Park, Sang-Youel; Lee, John Hwa

    2016-01-01

    Salmonella Typhimurium ghost cells expressing K88ab, K88ac, K99, and FasA fimbriae of enterotoxigenic Escherichia coli (ETEC) in their envelopes were constructed. The genes encoding the fimbriae were individually cloned into an expression plasmid, pMMP81, carrying the asd gene, which was subsequently electroporated into the Δasd S. Typhimurium mutant. Plasmid pJHLP99, carrying the phiX174 lysis gene E, was also subsequently electroporated into the Salmonella mutant. The presence of the individual fimbriae on the ghost cells was examined by Western blot analysis. Forty BALB/c mice were equally divided into 2 groups of 20 mice each. Group A mice were intramuscularly vaccinated with a mixture of the 4 ghost cells expressing the individual fimbriae. The group B mice were inoculated with sterile phosphate-buffered saline as a control. The antigen-specific serum IgG concentrations were significantly higher in group A than in group B from week 2 until week 6 after inoculation. In addition, the antigen-specific IgA concentrations in fecal samples were significantly higher in group A than in group B at week 2 after inoculation. A large difference between the groups in the number of antigen-specific IgA-secreting cells in the small intestine was observed by immunohistochemical study. Also, the splenic lymphocyte proliferative responses were significantly greater in group A than in the control mice. These results suggest that vaccination with our Salmonella ghost cells can induce both humoral and cell-mediated immune responses and that the increased number of antigen-specific IgA-secreting cells in the small intestine may be correlated with the elevated fecal IgA immune response. PMID:26733731

  20. In vitro adherence of type 1-fimbriated uropathogenic Escherichia coli to human ureteral mucosa.

    PubMed Central

    Fujita, K; Yamamoto, T; Yokota, T; Kitagawa, R

    1989-01-01

    Type 1-fimbriated Escherichia coli isolated from patients with urinary tract infections adhered in vitro to the epithelial cell surface of an excised human ureter. The bacteria also adhered to a mucous coating and to Formalin-fixed human ureteral mucosa. D-Mannose strongly inhibited such adherence. The bacteria in their nonfimbriated phase lacked the ability to adhere. We concluded that type 1 fimbriae play a role, at least in part, in upper urinary tract infections in humans. Images PMID:2568346

  1. Final Report for Contract N00014-86-K-0347 (University of California)

    DTIC Science & Technology

    1990-01-01

    2: 1177- 1184, 1989. Adult cholinergic interneurons of the neostriatum are not immunoreactive for monoclonal antibody to NGF receptor , whereas the... receptor such that many cholinergic interneurons became immunoreactive for NGF receptor . NGF infusion dramatically increased the size and choline...hippocampal formation. Specifically, fimbria- fornix lesions resulted in an increase in the GFAP-IR in the pyramidal and oriens area of the CA3 as well as

  2. Quantitative differences in adhesiveness of type 1 fimbriated Escherichia coli due to structural differences in fimH genes.

    PubMed Central

    Sokurenko, E V; Courtney, H S; Maslow, J; Siitonen, A; Hasty, D L

    1995-01-01

    Type 1 fimbriae are heteropolymeric surface organelles responsible for the D-mannose-sensitive (MS) adhesion of Escherichia coli. We recently reported that variation of receptor specificity of type 1 fimbriae can result solely from minor alterations in the structure of the gene for the FimH adhesin subunit. To further study the relationship between allelic variation of the fimH gene and adhesive properties of type 1 fimbriae, the fimH genes from five additional strains were cloned and used to complement the FimH deletion in E. coli KB18. When the parental and recombinant strains were tested for adhesion to immobilized mannan, a wide quantitative range in the ability of bacteria to adhere was noted. The differences in adhesion do not appear to be due to differences in the levels of fimbriation or relative levels of incorporation of FimH, because these parameters were similar in low-adhesion and high-adhesion strains. The nucleotide sequence for each of the fimH genes was determined. Analysis of deduced FimH sequences allowed identification of two sequence homology groups, based on the presence of Asn-70 and Ser-78 or Ser-70 and Asn-78 residues. The consensus sequences for each group conferred very low adhesion activity, and this low-adhesion phenotype predominated among a group of 43 fecal isolates. Strains isolated from a different host niche, the urinary tract, expressed type 1 fimbriae that conferred an increased level of adhesion. The results presented here strongly suggest that the quantitative variations in MS adhesion are due primarily to structural differences in the FimH adhesin. The observed differences in MS adhesion among populations of E. coli isolated from different host niches call attention to the possibility that phenotypic variants of FimH may play a functional role in populations dynamics. PMID:7601831

  3. Selection and phenotypic characterization of nonhemagglutinating mutants of Porphyromonas gingivalis.

    PubMed Central

    Chandad, F; Mayrand, D; Grenier, D; Hinode, D; Mouton, C

    1996-01-01

    To further investigate the relationship between fimbriae and the hemagglutinating adhesin HA-Ag2 of Porphyromonas gingivalis, three spontaneous mutants of the type strain ATCC 33277 were selected by a hemadsorption procedure. They were characterized for hemagglutination, trypsin-like and lectin-binding activities, and hydrophobicity and for the presence of fimbriae. The presence of the 42-kDa (the fimbrilin subunit) and the 43- and 49-kDa (the HA-Ag2 components) polypeptides was investigated by immunoblotting using polyclonal and monoclonal antibodies directed to fimbriae and to the hemagglutinating adhesin HA-Ag2. Cells from two of the three mutants (M1 and M2) exhibited no or little hemagglutination activity and very low trypsin-like activity and did not show the 43- and 49-kDa polypeptides. Abnormal fimbriation in M1 was deduced from the following observations of cells grown for 18 h: absence of the 42-kDa polypeptide and of a 14-kDa polypeptide and no fimbriae visible on electron micrographs. While the cells of mutant M2, irrespective of the age of the culture, were found to lack the 43- and 49-kDa polypeptides and hemagglutination activity, the supernatants of cultures grown for 72 h had high hemagglutination and trypsin-like activities and revealed the presence of the 42-, 43-, and 49-kDa polypeptides. This suggests that M2 may be missing some molecules which anchor the components to the cell surface. Mutant M3 showed levels of activities similar to those of the parental strain but lacked the 43-kDa polypeptide. Other pleiotropic effects observed for the mutants included loss of dark pigmentation and lower hydrophobicity. The data from this study fuel an emerging consensus whereby fimbriation, hemagglutination, and proteolytic activities, as well as other functions in P. gingivalis, are intricate. PMID:8641806

  4. Prevention of Ovarian High-Grade Serous Carcinoma by Elucidating Its Early Change

    DTIC Science & Technology

    2013-10-01

    HRT at time of surgery o Unknown menstrual cycle status at time of surgery The tissue microarray includes the following permutations (2 cores per...epithelium. As a result, a significant effort has been placed on determining the menstrual status of samples collected – this has included reviewing the...conundrum, it would be technically robust to use cryopreserved ampulla and fimbria from non-BRCA mutation carriers with known ovulation cycle status, using

  5. Role of Cell Appendages in Initial Attachment and Stability of E. coli on Silica Monitored by Nondestructive TIRF Microscopy.

    PubMed

    Wong, Kerwin K W; Olsson, Adam L J; Asadishad, Bahareh; van der Bruggen, Bart; Tufenkji, Nathalie

    2017-04-11

    Total internal reflection fluorescence (TIRF) microscopy was used to investigate initial attachment and stability of wild-type, curli-deficient (ΔcsgA), flagella-deficient (ΔflhDC), and type-1 fimbriae-deficient (Δfim) mutant E. coli strains. Suspended bacteria were injected into a flow cell where they deposited on a silica coverslip, and images were acquired over a 2 min period. TIRF microscope image analysis revealed that curli- and flagella-deficient mutants attached closer to the surface and required a longer time to find their equilibrium position (i.e., bond maturation) as compared to the wild-type and fimbriae-deficient mutants. Analysis of the change in bacterial surface area over the 2 min period also indicated that curli- and flagella-deficient mutants have less initial stability than the wild-type and fimbriae-deficient mutants, evidenced by their fluctuating position at equilibrium. TIRF observations at the microscopic level were complemented macroscopically using quartz crystal microbalance with dissipation (QCM-D) and sand-packed column experiments, which support the distinctive behavior observed at the microscopic scale. For each mutant strain, as fluorescence intensity increased in TIRF, the negative frequency shift in QCM-D (related to the attached mass of bacteria) also increased. Packed-column experiments indicated that curli- and flagella-deficient mutants exhibited a characteristically different attachment behavior and more retention as compared to the wild-type and fimbriae-deficient strains. This study utilized a new approach to understand bacterial attachment/detachment and provides new insights into the role of various appendages on initial attachment and stability.

  6. Fractional Analysis of Escherichia coli O157:H7 by Mass Spectrometry-Based Proteomics

    DTIC Science & Technology

    2012-10-01

    column with the Dionex UltiMate 3000 (Thermo Scientific Dionex , Sunnyvale, CA). The resolved peptides were electrosprayed into a linear ion trap MS... chromatography -tandem mass spectrometry, followed by biochemical pathway mapping using the Kyoto Encyclopedia of Genes and Genomes. The fimbriae-specific subset...15. SUBJECT TERMS 3T3 murine fibroblasts Cell toxicity Liquid chromatography Mass spectrometry LC-MS Ricin Ricinus communis

  7. Role of yqiC in the Pathogenicity of Salmonella and Innate Immune Responses of Human Intestinal Epithelium

    PubMed Central

    Wang, Ke-Chuan; Huang, Chih-Hung; Ding, Shih-Min; Chen, Ching-Kuo; Fang, Hsu-Wei; Huang, Ming-Te; Fang, Shiuh-Bin

    2016-01-01

    The yqiC gene of Salmonella enterica serovar Typhimurium (S. Typhimurium) regulates bacterial growth at different temperatures and mice survival after infection. However, the role of yqiC in bacterial colonization and host immunity remains unknown. We infected human LS174T, Caco-2, HeLa, and THP-1 cells with S. Typhimurium wild-type SL1344, its yqiC mutant, and its complemented strain. Bacterial colonization and internalization in the four cell lines significantly reduced on yqiC depletion. Post-infection production of interleukin-8 and human β-defensin-3 in LS174T cells significantly reduced because of yqiC deleted in S. Typhimurium. The phenotype of yqiC mutant exhibited few and short flagella, fimbriae on the cell surface, enhanced biofilm formation, upregulated type-1 fimbriae expression, and reduced bacterial motility. Type-1 fimbriae, flagella, SPI-1, and SPI-2 gene expression was quantified using real-time PCR. The data show that deletion of yqiC upregulated fimA and fimZ expression and downregulated flhD, fliZ, invA, and sseB expression. Furthermore, thin-layer chromatography and high-performance liquid chromatography revealed the absence of menaquinone in the yqiC mutant, thus validating the importance of yqiC in the bacterial electron transport chain. Therefore, YqiC can negatively regulate FimZ for type-1 fimbriae expression and manipulate the functions of its downstream virulence factors including flagella, SPI-1, and SPI-2 effectors. PMID:27777572

  8. PapX, a P fimbrial operon-encoded inhibitor of motility in uropathogenic Escherichia coli.

    PubMed

    Simms, Amy N; Mobley, Harry L T

    2008-11-01

    Motility and adherence are two integral aspects of bacterial pathogenesis. Adherence, often mediated by fimbriae, permits bacteria to attach to host cells and establish infection, whereas flagellum-driven motility allows bacteria to disseminate to sites more advantageous for colonization. Both fimbriae and flagella have been proven important for virulence of uropathogenic Escherichia coli (UPEC). Reciprocal regulation is one mechanism by which bacteria may reconcile the contradictory actions of adherence and motility. PapX, a P fimbrial gene product of UPEC strain CFT073, is a functional homolog of MrpJ of Proteus mirabilis; ectopic expression of papX in P. mirabilis reduces motility. To define the connection between P fimbria expression and motility in UPEC, the role of papX in the regulation of motility of strain CFT073 was examined. Overexpression of papX decreased motility of CFT073, which correlated with both a significant reduction in flagellin protein synthesized and flagella assembled on the cell surface. Conversely, an increase in motility and flagellin production was seen in an isogenic papX deletion mutant of CFT073. Microarray and quantitative reverse transcription-PCR analysis indicated that repression of motility of CFT073 by PapX appears to occur at the transcriptional level; expression of many motility-associated genes, including flhDC, the master regulator of motility, is decreased when papX is overexpressed. Transcription of motility genes is increased in the papX mutant compared to wild type. Electrophoretic mobility gel shift analysis revealed that PapX binds to the flhD promoter. We conclude that synthesis of P fimbriae regulates flagellum synthesis to repress motility via PapX.

  9. In Vivo Consumption of Cranberry Exerts ex Vivo Antiadhesive Activity against FimH-Dominated Uropathogenic Escherichia coli: A Combined in Vivo, ex Vivo, and in Vitro Study of an Extract from Vaccinium macrocarpon.

    PubMed

    Rafsanjany, Nasli; Senker, Jandirk; Brandt, Simone; Dobrindt, Ulrich; Hensel, Andreas

    2015-10-14

    For investigation of the molecular interaction of cranberry extract with adhesins of uropathogenic Escherichia coli (UPEC), urine from four volunteers consuming standardized cranberry extract (proanthocyanidin content = 1.24%) was analyzed within ex vivo experiments, indicating time-dependent significant inhibition of 40-50% of bacterial adhesion of UPEC strain NU14 to human T24 bladder cells. Under in vitro conditions a dose-dependent increase in bacterial adhesion was observed with proanthocyanidin-enriched cranberry Vaccinium macrocarpon extract (proanthocyanidin content = 21%). Confocal laser scanning microscopy and scanning electron microscopy proved that V.m. extract led to the formation of bacterial clusters on the outer plasma membrane of the host cells without subsequent internalization. This agglomerating activity was not observed when a PAC-depleted extract (V.m. extract(≠PAC)) was used, which showed significant inhibition of bacterial adhesion in cases where type 1 fimbriae dominated and mannose-sensitive UPEC strain NU14 was used. V.m. extract(≠PAC) had no inhibitory activity against P- and F1C-fimbriae dominated strain 2980. Quantitative gene expression analysis indicated that PAC-containing as well as PAC-depleted cranberry extracts increased the fimH expression in NU14 as part of a feedback mechanism after blocking FimH. For strain 2980 the PAC-containing extract led to up-regulation of P- and F1C-fimbriae, whereas the PAC-depleted extract had no influence on gene expression. V.m. and V.m. extract(≠PAC) did not influence biofilm and curli formation in UPEC strains NU14 and 2980. These data lead to the conclusion that also proanthocyanidin-free cranberry extracts exert antiadhesive activity by interaction with mannose-sensitive type 1 fimbriae of UPEC.

  10. [Cranberry (Vaccinium macrocarpon) and urinary tract infections: study model and review of literature].

    PubMed

    Lavigne, J-P; Bourg, G; Botto, H; Sotto, A

    2007-11-01

    Cranberries (Vaccinium macrocarpon) have long been the focus of interest for their beneficial effects in preventing urinary tract infections. Among cranberry compounds, a group of proanthocyanidins (PACs) with A-type linkages were isolated which exhibit bacterial anti-adhesion activity against uropathogenic Escherichia coli strains. These PAC inhibit P-fimbriae synthesis and induce a bacterial deformation. This activity was demonstrated on both antibiotic susceptible and resistant bacteria. This review focused on the last discoveries in the knowledge of cranberry effects.

  11. Lack of flagella disadvantages Salmonella enterica serovar Enteritidis during the early stages of infection in the rat.

    PubMed

    Robertson, Jeanette M C; McKenzie, Norma H; Duncan, Michelle; Allen-Vercoe, Emma; Woodward, Martin J; Flint, Harry J; Grant, George

    2003-01-01

    The roles of flagella and five fimbriae (SEF14, SEF17, SEF21, pef, lpf) in the early stages (up to 3 days) of Salmonella enterica serovar Enteritidis (S. Enteritidis) infection have been investigated in the rat. Wild-type strains LA5 and S1400 (fim+/fla+) and insertionally inactivated mutants unable to express the five fimbriae (fim-/fla+), flagella (fim+/fla-) or fimbriae and flagella (fim-/fla-) were used. All wild-type and mutant strains were able to colonize the gut and spread to the mesenteric lymph nodes, liver and spleen. There appeared to be little or no difference between the fim-/fla+ and wild-type (fim+/fla+) strains. In contrast, the numbers of aflagellate (fim+/fla- or fim-/fla-) salmonella in the liver and spleen were transiently reduced. In addition, fim+/fla- or fim-/fla- strains were less able to persist in the upper gastrointestinal tract and the inflammatory responses they elicited in the gut were less severe. Thus, expression of SEF14, SEF17, SEF21, pef and lpf did not appear to be a prerequisite for induction of S. Enteritidis infection in the rat. Deletion of flagella did, however, disadvantage the bacterium. This may be due to the inability to produce or release the potent immunomodulating protein flagellin.

  12. Conservation of an Actinomyces viscosus T14V type 1 fimbrial subunit homolog among divergent groups of Actinomyces spp.

    PubMed Central

    Yeung, M K

    1992-01-01

    The type 1 fimbrial subunit gene of the human Actinomyces viscosus T14V was used as a DNA probe in Southern analyses to detect related DNA sequences in 16 of 30 strains of Actinomyces spp. under conditions of high stringency. The organisms with homology to the DNA probe included two human and six nonhuman A. viscosus, three human and three nonhuman A. naeslundii, and two A. bovis isolates. Homologous DNA sequences were not detected in strains of A. odontolyticus and A. israelii examined in this study. Northern (RNA) blot analysis revealed expression of a transcript from each of the A. viscosus and A. naeslundii strains and from one A. bovis strain that was comparable in size to that detected from A. viscosus T14V. Cell surface fimbriae were observed on a majority of the strains that expressed the transcript. Various degrees of cross-immunoreactivities between these strains and antibodies specific for type 1 fimbriae of A. viscosus T14V were also observed by colony immunoassay. Thus, the data clearly demonstrate the existence in, and expression by, divergent Actinomyces groups of genomic sequences that are closely related to the type 1 fimbriae of A. viscosus T14V. Images PMID:1347285

  13. Proliferation of smooth muscle cells stimulated by Porphyromonas gingivalis is inhibited by apple polyphenol.

    PubMed

    Inaba, Hiroaki; Tagashira, Motoyuki; Kanda, Tomomasa; Amano, Atsuo

    2011-11-01

    Porphyromonas gingivalis (Pg) is thought to be involved in the progression of occlusive arterial lesions, whereas vascular smooth muscle cell (SMC) proliferation is considered to be involved in occlusive arterial disease. We previously showed that bacteremia caused by Pg infection induced proliferation of mouse aortic SMCs. Furthermore, human SMCs stimulated with human plasma incubated with Pg showed a marked transformation from the contractile to proliferative phenotype. In the present study, we examine the involvement of Pg gingipains and fimbriae in induction of the SMC transformation and proliferation, and effective inhibitors. Pg strains including gingipain- and fimbria-null mutants were incubated in human plasma, after which the bacteria were removed and the supernatants were added to cultured SMCs. To evaluate the effects of inhibitors, Pg organisms were incubated in plasma in the presence of apple polyphenol (AP), epigallocatechin gallate, KYT-1 (Arg-gingipain inhibitor), and KYT-36 (Lys-gingipain inhibitor). Plasma supernatants from wild-type and fimbria-mutant cultures markedly stimulated cellular proliferation, whereas those containing gingipain-null mutants showed negligible effects. SMC proliferation was also induced by plasma treated with trypsin. Furthermore, plasma supernatants cultured in the presence of KYT-1/KYT-36 and AP showed significant inhibitory effects on SMC proliferation, whereas cultures with epigallocatechin gallate did not. Our results suggest that Pg gingipains are involved in the induction of SMC transformation and proliferation, whereas this was inhibited by AP.

  14. Novel fimbrilin PGN_1808 in Porphyromonas gingivalis.

    PubMed

    Nagano, Keiji; Hasegawa, Yoshiaki; Yoshida, Yasuo; Yoshimura, Fuminobu

    2017-01-01

    Porphyromonas gingivalis, a periodontopathic gram-negative anaerobic bacterium, generally expresses two types of fimbriae, FimA and Mfa1. However, a novel potential fimbrilin, PGN_1808, in P. gingivalis strain ATCC 33277 was recently identified by an in silico structural homology search. In this study, we experimentally examined whether the protein formed a fimbrial structure. Anion-exchange chromatography showed that the elution peak of the protein was not identical to those of the major fimbrilins of FimA and Mfa1, indicating that PGN_1808 is not a component of these fimbriae. Electrophoretic analyses showed that PGN_1808 formed a polymer, although it was detergent and heat labile compared to FimA and Mfa1. Transmission electron microscopy showed filamentous structures (2‒3 nm × 200‒400 nm) on the cell surfaces of a PGN_1808-overexpressing P. gingivalis mutant (deficient in both FimA and Mfa1 fimbriae) and in the PGN_1808 fraction. PGN_1808 was detected in 81 of 84 wild-type strains of P. gingivalis by western blotting, suggesting that the protein is generally present in P. gingivalis.

  15. A low-pH medium in vitro or the environment within a macrophage decreases the transcriptional levels of fimA, fimZ and lrp in Salmonella enterica serovar Typhimurium.

    PubMed

    Wang, Ke-Chuan; Hsu, Yuan-Hsun; Huang, Yi-Ning; Chen, Ter-Hsin; Lin, Jiunn-Horng; Hsuan, Shih-Ling; Chien, Maw-Sheng; Lee, Wei-Cheng; Yeh, Kuang-Sheng

    2013-09-01

    Many Salmonella Typhimurium isolates produce type 1 fimbriae and exhibit fimbrial phase variation in vitro. Static broth culture favours the production of fimbriae, while solid agar medium inhibits the generation of these appendages. Little information is available regarding whether S. Typhimurium continues to produce type 1 fimbriae during in vivo growth. We used a type 1 fimbrial phase-variable strain S. Typhimurium LB5010 and its derivatives to infect RAW 264.7 macrophages. Following entry into macrophages, S. Typhimurium LB5010 gradually decreased the transcript levels of fimbrial subunit gene fimA, positive regulatory gene fimZ, and global regulatory gene lrp. A similar decrease in transcript levels was detected by RT-PCRwhen the pH of static brothmediumwas shifted frompH 7 to amore acidic pH 4. A fimA-deleted strain continued to multiply within macrophages as did the parental strain. An lrp deletion strain was unimpaired for in vitro growth at pH 7 or pH 4, while a strain harboring an lrp-containing plasmid exhibited impaired in vitro growth at pH 4. We propose that acidic medium, which resembles one aspect of the intracellular environment in a macrophage, inhibits type 1 fimbrial production by down-regulation of the expression of lrp, fimZ and fimA.

  16. Genotypic characterization of virulence factors in Escherichia coli strains from patients with cystitis.

    PubMed

    Tiba, Monique Ribeiro; Yano, Tomomasa; Leite, Domingos da Silva

    2008-01-01

    Adhesins (P-fimbriae, S-fimbriae, type 1 fimbriae and afimbrial adhesin), toxins (alpha-hemolysin and cytotoxic necrotizing factor type 1), iron acquisition systems (aerobactin) and host defense avoidance mechanisms (capsule or lipopolysaccharide) have been shown to be prevalent in Escherichia coli strains associated with urinary tract infections. In this work, 162 Uropathogenic Escherichia coli (UPEC) strains from patients with cystitis were genotypically characterized by polymerase chain reaction (PCR) assay. We developed three multiplex PCR assays for virulence-related genes papC, papE/F, papG alleles, fimH, sfa/foc, afaE, hly, cnf-1, usp, cdtB, iucD, and kpsMTII, all of them previously identified in UPEC strains. The PCR assay results identified 158 fimH (97.5%), 86 kpsMTII (53.1%), 53 papC/papEF/papG (32.7%), 45 sfa (27.8%), 42 iucD (25.9%), 41 hly (25.3%), 36 usp (22.2%), 30 cnf-1(18.5%) and 10 afa (6.2%) strains. No strain was positive for cdtB. In this work, we also demonstrated that adhesins may be multiple within a single strain and that several virulence genes can occur combined in association.

  17. Cytochemical Differences in Bacterial Glycocalyx

    NASA Astrophysics Data System (ADS)

    Krautgartner, Wolf Dietrich; Vitkov, Ljubomir; Hannig, Matthias; Pelz, Klaus; Stoiber, Walter

    2005-02-01

    To examine new cytochemical aspects of the bacterial adhesion, a strain 41452/01 of the oral commensal Streptococcus sanguis and a wild strain of Staphylococcus aureus were grown with and without sucrose supplementation for 6 days. Osmiumtetraoxyde (OsO4), uranyl acetate (UA), ruthenium red (RR), cupromeronic blue (CB) staining with critical electrolytic concentrations (CECs), and the tannic acid-metal salt technique (TAMST) were applied for electron microscopy. Cytochemically, only RR-positive fimbriae in S. sanguis were visualized. By contrast, some types of fimbriae staining were observed in S. aureus glycocalyx: RR-positive, OsO4-positive, tannophilic and CB-positive with ceasing point at 0.3 M MgCl2. The CB staining with CEC, used for the first time for visualization of glycoproteins of bacterial glycocalyx, also reveals intacellular CB-positive substances-probably the monomeric molecules, that is, subunits forming the fimbriae via extracellular assembly. Thus, glycosylated components of the biofilm matrix can be reliably related to single cells. The visualization of intracellular components by CB with CEC enables clear distinction between S. aureus and other bacteria, which do not produce CB-positive substances. The small quantities of tannophilic substances found in S. aureus makes the use of TAMST for the same purpose difficult. The present work protocol enables, for the first time, a partial cytochemical differentiation of the bacterial glycocalyx.

  18. Mapping the Binding Domain of the F18 Fimbrial Adhesin

    PubMed Central

    Smeds, A.; Pertovaara, M.; Timonen, T.; Pohjanvirta, T.; Pelkonen, S.; Palva, A.

    2003-01-01

    F18 fimbrial Esherichia coli strains are associated with porcine postweaning diarrhea and pig edema disease. Recently, the FedF subunit was identified as the adhesin of the F18 fimbriae. In this study, adhesion domains of FedF were further studied by constructing deletions within the fedF gene and expressing FedF proteins with deletions either together with the other F18 fimbrial subunits or as fusion proteins tagged with maltose binding protein. The region essential for adhesion to porcine intestinal epithelial cells was mapped between amino acid residues 60 and 109 of FedF. To map the binding domain even more closely, all eight charged amino acid residues within this region were independently replaced by alanine. Three of these single point mutants expressing F18 fimbriae exhibited significantly diminished capabilities to adhere to porcine epithelial cells in vitro. In addition, a triple point mutation and a double point mutation completely abolished receptor adhesiveness. The result further confirmed that the region between amino acid residues 60 and 109 is essential for the binding of F18 fimbriae to their receptor. In addition, the adhesion capability of the binding domain was eliminated after treatment with iodoacetamide, suggesting the formation of a disulfide bridge between Cys-63 and Cys-83, whereas Cys-111 and Cys-116 could be deleted without affecting the binding ability of FedF. PMID:12654838

  19. The major subunit, CfaB, of colonization factor antigen i from enterotoxigenic Escherichia coli is a glycosphingolipid binding protein.

    PubMed

    Jansson, Lena; Tobias, Joshua; Lebens, Michael; Svennerholm, Ann-Mari; Teneberg, Susann

    2006-06-01

    Bacterial adherence to mucosal surfaces is an important virulence trait of pathogenic bacteria. Adhesion of enterotoxigenic Escherichia coli (ETEC) to the intestine is mediated by a number of antigenically distinct colonization factors (CFs). One of the most common CFs is CFA/I. This has a fimbrial structure composed of a major repeating subunit, CfaB, and a single tip subunit, CfaE. The potential carbohydrate recognition by CFA/I was investigated by binding CFA/I-fimbriated bacteria and purified CFA/I fimbriae to a large number of variant glycosphingolipids separated on thin-layer chromatograms. For both fimbriated bacteria and purified fimbriae, specific interactions could be identified with a number of nonacid glycosphingolipids. These included glucosylceramide, lactosylceramide with phytosphingosine and/or hydroxy fatty acids, neolactotetraosylceramide, gangliotriaosylceramide, gangliotetraosylceramide, the H5 type 2 pentaglycosylceramide, the Lea-5 glycosphingolipid, the Lex-5 glycosphingolipid, and the Ley-6 glycosphingolipid. These glycosphingolipids were also recognized by recombinant E. coli expressing CFA/I in the absence of tip protein CfaE, as well as by purified fimbriae from the same strain. This demonstrates that the glycosphingolipid-binding capacity of CFA/I resides in the major CfaB subunit.

  20. Fimbrial types among respiratory isolates belonging to the family Enterobacteriaceae.

    PubMed

    Hornick, D B; Allen, B L; Horn, M A; Clegg, S

    1991-09-01

    Bacterial attachment is believed to be an early step in gram-negative nosocomial pneumonia. The frequency of fimbria-associated adhesins among respiratory pathogens has not been studied in detail. In this study isolates belonging to the family Enterobacteriaceae, prospectively obtained from intensive care unit patients who were suspected of having nosocomial pneumonia, were examined for fimbria-associated adhesins. Type 3, P, type 1, and other fimbrial phenotypes were identified by specific hemagglutination and electron microscopy. The Klebsiella type 3 fimbrial phenotype was further characterized by using a monoclonal antibody. Also, both type 3 and Escherichia coli P fimbrial genotypes were detected by using DNA colony blot assays. The frequencies of genera or species isolated were as follows: Enterobacter (38.6%), Klebsiella (26.8%), Serratia (17.7%), E. coli (13%), and Proteus (5.2%). Isolates of Klebsiella oxytoca, K. pneumoniae, and Enterobacter cloacae most commonly possessed the type 3 fimbrial phenotype and genotype. The phenotype and genotype for E. coli P fimbriae (46.2 and 50%, respectively), a known pathogenic determinant in the urinary tract, were detected more frequently than expected. In addition, a previously unspecified hemagglutinin that was specific for porcine erythrocytes was almost uniformly expressed among isolates of Enterobacter aerogenes. Finally, the expression of the type 1 fimbrial phenotype was widely detected among the isolates tested but notably absent among K. oxytoca and Proteus mirabilis isolates. The frequency of the various fimbrial types identified suggests a role for these bacterial organelles in adherence to respiratory epithelia.

  1. Bacterial Cyclic AMP-Phosphodiesterase Activity Coordinates Biofilm Formation

    PubMed Central

    Kalivoda, Eric J.; Brothers, Kimberly M.; Stella, Nicholas A.; Schmitt, Matthew J.; Shanks, Robert M. Q.

    2013-01-01

    Biofilm-related infections are a major contributor to human disease, and the capacity for surface attachment and biofilm formation are key attributes for the pathogenesis of microbes. Serratia marcescens type I fimbriae-dependent biofilms are coordinated by the adenylate cyclase, CyaA, and the cyclic 3′,5′-adenosine monophosphate (cAMP)-cAMP receptor protein (CRP) complex. This study uses S. marcescens as a model system to test the role of cAMP-phosphodiesterase activity in controlling biofilm formation. Herein we describe the characterization of a putative S. marcescens cAMP-phosphodiesterase gene (SMA3506), designated as cpdS, and demonstrated to be a functional cAMP-phosphodiesterase both in vitro and in vivo. Deletion of cpdS resulted in defective biofilm formation and reduced type I fimbriae production, whereas multicopy expression of cpdS conferred a type I fimbriae-dependent hyper-biofilm. Together, these results support a model in which bacterial cAMP-phosphodiesterase activity modulates biofilm formation. PMID:23923059

  2. Novel fimbrilin PGN_1808 in Porphyromonas gingivalis

    PubMed Central

    Hasegawa, Yoshiaki; Yoshida, Yasuo; Yoshimura, Fuminobu

    2017-01-01

    Porphyromonas gingivalis, a periodontopathic gram-negative anaerobic bacterium, generally expresses two types of fimbriae, FimA and Mfa1. However, a novel potential fimbrilin, PGN_1808, in P. gingivalis strain ATCC 33277 was recently identified by an in silico structural homology search. In this study, we experimentally examined whether the protein formed a fimbrial structure. Anion-exchange chromatography showed that the elution peak of the protein was not identical to those of the major fimbrilins of FimA and Mfa1, indicating that PGN_1808 is not a component of these fimbriae. Electrophoretic analyses showed that PGN_1808 formed a polymer, although it was detergent and heat labile compared to FimA and Mfa1. Transmission electron microscopy showed filamentous structures (2‒3 nm × 200‒400 nm) on the cell surfaces of a PGN_1808-overexpressing P. gingivalis mutant (deficient in both FimA and Mfa1 fimbriae) and in the PGN_1808 fraction. PGN_1808 was detected in 81 of 84 wild-type strains of P. gingivalis by western blotting, suggesting that the protein is generally present in P. gingivalis. PMID:28296909

  3. Pleiotropic roles of uvrY on biofilm formation, motility and virulence in uropathogenic Escherichia coli CFT073.

    PubMed

    Mitra, Arindam; Palaniyandi, Senthilkumar; Herren, Christopher D; Zhu, Xiaoping; Mukhopadhyay, Suman

    2013-01-01

    Urinary tract infections primarily caused by uropathogenic strains of Escherichia coli (E. coli) remain a significant public health problem in both developed and developing countries. An important virulence determinant in uropathogenesis is biofilm formation which requires expression of fimbriae, flagella, and other surface components such as lipopolysaccharides. In this study, we explored the regulation of uvrY and csrA genes in biofilm formation, motility and virulence determinants in uropathogenic E. coli. We found that mutation in uvrY suppressed biofilm formation on abiotic surfaces such as polyvinyl chloride, polystyrene and glass, and complementation of uvrY in the mutant restored the biofilm phenotype. We further evaluated the role of uvrY gene in expression of type 1 fimbriae, an important adhesin that facilitates adhesion to various abiotic surfaces. We found that phase variation of type 1 fimbriae between fimbriated and afimbriated mode was modulated by uvrY at its transcriptional level. Deletion mutant of uvrY lowered expression of fimbrial recombinase genes, such as fimB, fimE, and fimA, a gene encoding major fimbrial subunit. Furthermore, transcription of virulence specific genes such as papA, hlyB and galU was also reduced in the deletion mutant. Swarming motility and expression of flhD and flhC was also diminished in the mutant. Taken together, our findings unravel a possible mechanism in which uvrY facilitates biofilm formation, persistence and virulence of uropathogenic E. coli.

  4. Virulence characteristics of Escherichia coli isolates obtained from broiler breeders with salpingitis.

    PubMed

    Monroy, Maria A R; Knöbl, Terezinha; Bottino, José A; Ferreira, Claudete S Astolfi; Ferreira, Antonio J Piantino

    2005-01-01

    Thirty isolates of Escherichia coli from broiler breeders with salpingitis were studied. Using the slide agglutination test, the isolates were found to belong to serogroups O1, O2, O5, O36, O45, O53 and O78. Pathogenicity for day-old chicks was determined by air sac inoculation and isolates were categorized as having high, intermediate or low virulence. Growth on iron starvation medium was observed together with aerobactin production. Based on the results of in vitro adherence tests, attachment to oviduct epithelium from old birds was found to be superior to that observed using corresponding material from young birds. DNA hybridization testing for type 1, P, and S fimbriae revealed predominant expression of type 1, correlating with mannose-sensitive hemagglutination using guinea-pig erythrocytes. In this study, P and S fimbriae were not considered to be important adherence factors. Study findings would suggest that, as far as salpingitis is concerned, type 1 fimbriae can play an important role in E. coli infection in breeders. An interesting result to emerge from the study was the observation that E. coli isolates were completely resistant to serum from young breeders, whereas they were completely sensitive using serum from older breeders. Based on serogroups involved, pathogenicity for day-old chicks and virulence indicators, the salpingitis isolates were similar to those from cases of chronic respiratory disease.

  5. A randomized, double-blind, placebo-controlled trial to assess the bacterial anti-adhesion effects of cranberry extract beverages.

    PubMed

    Kaspar, Kerrie L; Howell, Amy B; Khoo, Christina

    2015-04-01

    In this study, we examined the ex vivo urinary anti-adhesion activity of low-calorie cranberry extract beverages in both a pilot study (n = 10) and a randomized, double-blind, placebo controlled clinical trial (n = 59). In the pilot study, subjects consumed a cranberry extract beverage (CEB) or a cranberry extract and juice beverage (CEJB), compared to placebo. Both cranberry beverages utilized a standardized cranberry extract powder at a level equivalent to low-calorie cranberry juice cocktail (LCJC) on a PAC content basis. Clean-catch urine samples collected at baseline and post intervention were tested for anti-adhesion activity utilizing a mannose-resistant human red blood cell hemagglutination assay specific for P-fimbriated E. coli. Results from the pilot study indicated that ex vivo anti-adhesion activity for both cranberry treatments were higher (p < 0.05) than placebo. In the clinical trial, we compared CEJB to LCJC and a placebo beverage. Post-consumption urine from both cranberry treatment groups showed significantly higher (p < 0.05) anti-adhesion activity compared to placebo. There were no differences observed in anti-adhesion activity between CJEB and LCJC, indicating similar bioactivity. Therefore, acute beverage consumption of cranberry extract and/or juice provides ex vivo anti-adhesion activity, which may help to improve urinary tract health.

  6. Lack of inhibition of adhesion of an enteropathogenic Escherichia coli by polycarbophil.

    PubMed

    Mack, D R; Blain-Nelson, P L; Mauger, J W

    1993-12-01

    Anionic polyacrylic acid polymers, such as polycarbophil, have a number of properties that would make them suitable carriers for sustained antibiotic release formulations in the intestinal tract. However, little is known with regards to possible microbial adhesion to polycarbophil. The aim of this study was to evaluate for such an interaction using the rabbit enteric pathogen Escherichia coli RDEC-1 (serotype O15:H-). RDEC-1 mediates attaching and effacing binding to intestinal epithelium in a manner morphologically identical to that observed in both human enteropathogenic E. coli and enterohemorrhagic E. coli infections. RDEC-1 bacteria were grown to promote the expression of the mannose-resistant AF/R1 adhesion pili. A nonpiliated mutant, strain M34, was used as a negative control. Using radioactive labeling of bacteria, we quantitated adhesion of piliated RDEC-1 in the presence of polycarbophil using an in vitro adhesion assay system. Binding of piliated RDEC-1 in the adhesion assay was greater than for nonpiliated M34 for all concentrations of bacteria greater than 10(9) (P < .05). Polycarbophil did not cause concentration-dependent inhibition of piliated RDEC-1 binding (P > .05). We conclude polycarbophil does not interfere with the AF/R1 adhesin ligand of RDEC-1. Use of this polymer as a mucoadhesive drug delivery vehicle for nonabsorbable antibiotics in the treatment of gastrointestinal infections would not be expected to interfere with the protective effects of intestinal mucins.

  7. Molecular cloning, expression, and sequence of the pilin gene from nontypeable Haemophilus influenzae M37.

    PubMed Central

    Coleman, T; Grass, S; Munson, R

    1991-01-01

    Nontypeable Haemophilus influenzae M37 adheres to human buccal epithelial cells and exhibits mannose-resistant hemagglutination of human erythrocytes. An isogenic variant of this strain which was deficient in hemagglutination was isolated. A protein with an apparent molecular weight of 22,000 was present in the sodium dodecyl sulfate-polyacrylamide gel profile of sarcosyl-insoluble proteins from the hemagglutination-proficient strain but was absent from the profile of the isogenic hemagglutination-deficient variant. A monoclonal antibody which reacts with the hemagglutination-proficient isolate but not with the hemagglutination-deficient isolate has been characterized. This monoclonal antibody was employed in an affinity column for purification of the protein as well as to screen a genomic library for recombinant clones expressing the gene. Several clones which contained overlapping genomic fragments were identified by reaction with the monoclonal antibody. The gene for the 22-kDa protein was subcloned and sequenced. The gene for the type b pilin from H. influenzae type b strain MinnA was also cloned and sequenced. The DNA sequence of the strain MinnA gene was identical to that reported previously for two other type b strains. The DNA sequence of the strain M37 gene is 77% identical to that of the type b pilin gene, and the derived amino acid sequence is 68% identical to that of the type b pilin. Images PMID:1673447

  8. Correlation between electrophoretic types B1 and B2 of carboxylesterase B and host-dependent factors in Escherichia coli septicaemia.

    PubMed Central

    Picard, B.; Goullet, P.

    1988-01-01

    Electrophoretic types B1 and B2 of carboxylesterase B produced by strains of Escherichia coli isolated from 100 septicaemia cases were correlated with alpha-haemolysin and mannose resistant haemagglutinin (MRHA) production and with clinical data including eventual underlying diseases, origin of septicaemia and evolution. Electrophoretic type B2 was phenotypically linked with alpha-haemolysin and MRHA production. The proportion of type B2 isolates varied significantly with occurrence of an underlying illness (45% for patients without an underlying disease and 22% for compromised patients) and with the site of origin of the septicaemia (40% for those of urinary origin and 18% for infection of digestive origin). In the former infections, type B2 isolates were obtained in the majority from male patients while type B1 isolates predominated in women. The septicaemias associated with type B1 were characterized by a lower proportion of isolates producing alpha-haemolysin and MRHA and by a greater frequency of septic shock and death than those associated with type B2. These facts emphasize the importance of host-dependent factors in E. coli septicaemia. PMID:3276540

  9. Characterization of a novel hemagglutinin of diarrhea-associated Escherichia coli that has characteristics of diffusely adhering E. coli and enteroaggregative E. coli.

    PubMed Central

    Yamamoto, T; Wakisaka, N; Nakae, T; Kamano, T; Serichantalergs, O; Echeverria, P

    1996-01-01

    Escherichia coli 73-1 (serotype O73:H33) and 5-2 (serotype O89:H-) isolated from patients with diarrhea adhered to tissue culture cells (HeLa and HEp-2) as well as coverslips (plastic and glass) in a diffuse pattern. Adherence of strain 73-1 was mediated by a 110-kbp plasmid designated pEDA1 and correlated with D-mannose-resistant hemagglutinin (MRHA) detected with bovine, sheep, or human erythrocytes. The MRHA region was duplicated on pEDA1 and mediated the production of the 57-kDa outer membrane protein whose N-terminal amino acid sequence was hydrophobic. In accordance with MRHA and adherence, the 57-kDa outer membrane protein was observed best at 37 degrees C and to a lesser extent at 25 degrees C. In human intestine, adherence to mucus and colonic epithelium was obvious. No detectable pili were observed. The enteroaggregative E. coli heat-stable enterotoxin 1 (EAST1) gene, whose nucleotide sequence was 99.1% homologous to that of enteroaggregative E. coli, was present adjacent to the MRHA region on pEDA1. Strain 5-2 also exhibited MRHA activities and adherence and had sequences corresponding to those of the MRHA region and EAST1 gene. The data suggest that strain 73-1 (and strain 5-2), which has characteristics of both diffusely adhering E. coli and enteroaggregative E. coli, possesses a novel hemagglutinin associated with diffuse adherence. PMID:8751919

  10. A novel cryohemagglutinin associated with adherence of enteroaggregative Escherichia coli.

    PubMed Central

    Yamamoto, T; Wakisaka, N; Nakae, T

    1997-01-01

    Strain O42 (serotype O44:H18) of enteroaggregative Escherichia coli (EAggEC) has been shown to be pathogenic in volunteer experiments. This strain exhibited plasmid (pO42)-encoded D-mannose-resistant hemagglutinating activity (MRHA) that was detected only at low temperatures (e.g., 0 degrees C) and only with human erythrocytes. The production of this cryogenic MRHA (cryo-MRHA) was observed when the bacteria were grown in liquid media and was strictly regulated by bacterial growth temperatures. Transposon-insertion mutagenesis revealed that this MRHA is associated with (i) bacterial clump formation in liquid cultures, (ii) bacterial adherence to HEp-2 cells as well as (Formalin-fixed) human colonic mucosa, and (iii) production of a 16-kDa outer membrane protein. The PCR designed on the basis of the determined cryo-MRHA-associated DNA sequence sharply distinguished strain O42 from eight other EAggEC strains whose MRHAs were detected at both cold and room temperatures to the same (or similar) extent. Strain O42 possessed a surface layer that may enhance the pO42-mediated adherence. The data suggest that a plasmid-encoded cryo-MRHA is a candidate for a major adhesin of EAggEC strain O42. PMID:9234817

  11. Differential adherence of Shiga toxin-producing Escherichia coli harboring saa to epithelial cells.

    PubMed

    Toma, Claudia; Nakasone, Noboru; Miliwebsky, Elizabeth; Higa, Naomi; Rivas, Marta; Suzuki, Toshihiko

    2008-10-01

    The majority of Shiga toxigenic Escherichia coli (STEC) strains isolated from severe STEC disease are those harboring the locus of enterocyte effacement (LEE), which encodes factors involved in adherence to epithelial cells. However, LEE-negative STEC are increasingly isolated from clinical cases. STEC autoagglutinating adhesin (Saa) is widely used as a marker of adhesin in the absence of LEE. In the present study, we compared the adherence of 32 saa-harboring STEC strains to cultured epithelial cells in the absence or presence of d-mannose. In the absence of d-mannose, 19 strains were adherent to HEp-2 and Caco-2 cells, while 12 were non-adherent. One strain showed detachment of epithelial cells. The adherence of 13 strains was sensitive to the presence of d-mannose. The saa mutant of strain T141, in which adherence was mannose resistant, did not show a significant decrease in adherence compared to the wild type, suggesting a Saa-independent mechanism of adherence. saa-harboring STEC exhibited differential binding properties to epithelial cells, which could not be attributed to the number of C-terminal repeats of Saa, or to the expression of Saa as detected by Western blotting. Our results suggest that multiple adherence mechanisms are present in saa-harboring STEC, implying a high degree of diversity in this group of STEC.

  12. Early detection of high-grade tubal serous carcinoma in women at low risk for hereditary breast and ovarian cancer syndrome by systematic examination of fallopian tubes incidentally removed during benign surgery.

    PubMed

    Rabban, Joseph T; Garg, Karuna; Crawford, Beth; Chen, Lee-may; Zaloudek, Charles J

    2014-06-01

    Early detection of sporadic pelvic serous carcinoma remains an elusive goal. In women at high risk for hereditary breast and ovarian cancer syndrome who undergo prophylactic salpingectomy, systematic pathologic examination of the fallopian tubes will detect occult tubal cancer, mostly in the fimbriae, of a minority of women. Such tubal cancers are the putative precursor to advanced-stage pelvic cancer. We hypothesized that early tubal cancer detection can also be accomplished in women at low risk using a similar approach. In this study, we performed complete and systematic examination of the fallopian tubes removed during surgery performed for benign indications. Among 522 women, 4 cases of serous tubal intraepithelial carcinoma (STIC) were identified. Three of these cases would have gone undetected using the current standard of care of sampling only a single random section of the tube. The fourth case was accompanied by occult ovarian carcinoma. The fimbriae contained STIC in 3 of the 4 cases and atypical mucosa in 1 case in which the STIC was in the nonfimbriated portion of the tube. The morphologic and immunohistochemical features (aberrant p53 and MIB-1) of these STICs were similar to those expected in high-risk women. All 4 patients with STIC underwent BRCA1 and BRCA2 gene testing; no germline mutations were identified in any patient. An additional 11 specimens contained atypical mucosal proliferations that fell short of morphologic and immunohistochemical criteria for STIC. Two of these 11 fulfilled criteria for a serous tubal intraepithelial lesion, and the remaining atypical proliferations exhibited normal p53 and MIB-1. For most specimens, the fimbriae could be completely submitted in 1 or 2 cassettes per tube. These results demonstrate that systematic examination of the tubal fimbriae can serve as a form of early detection of sporadic tubal cancer without incurring significant labor or cost. We propose that the tubal fimbriae should be completely examined

  13. Carbohydrates as future anti-adhesion drugs for infectious diseases.

    PubMed

    Sharon, Nathan

    2006-04-01

    Adhesion of pathogenic organisms to host tissues is the prerequisite for the initiation of the majority of infectious diseases. In many systems, it is mediated by lectins present on the surface of the infectious organism that bind to complementary carbohydrates on the surface of the host tissues. Lectin-deficient mutants often lack the ability to initiate infection. The bacterial lectins are typically in the form of elongated submicroscopic multi-subunit protein appendages, known as fimbriae (or pili). The best characterized of these are the mannose-specific type 1 fimbriae, the galabiose-specific P fimbriae and the N-acetylglucosamine-specific fimbriae of Escherichia coli. Soluble carbohydrates recognized by the bacterial surface lectins block the adhesion of the bacteria to animal cells in vitro. Aromatic alpha-mannosides are potent inhibitors of type 1 fimbriated E. coli, being up to 1000 times more active than MealphaMan, with affinities in the nanomolar range. This is due to the presence of a hydrophobic region next to the monosaccharide-binding site of the fimbriae, as recently demonstrated by X-ray studies. Polyvalent saccharides (e.g., neoglycoproteins or dendrimers) are also powerful inhibitors of bacterial adhesion in vitro. Very significantly, lectin-inhibitory saccharides have been shown to protect mice, rabbits, calves and monkeys against experimental infection by lectin-carrying bacteria. Since anti-adhesive agents do not act by killing or arresting the growth of the pathogens, it is very likely that strains resistant to such agents will emerge at a markedly lower rate than of strains that are resistant to antibiotics. Suitable sugars also inhibit the binding to cells of carbohydrate-specific toxins, among them those of Shigella dysenteriae Type 1, and of the homologous Verotoxins of E. coli, specific for galabiose. Appropriately designed polyvalent ligands are up to six orders of magnitude stronger inhibitors of toxin binding in vitro than the

  14. Breast cancer risk and clinical implications for germline PTEN mutation carriers.

    PubMed

    Ngeow, Joanne; Sesock, Kaitlin; Eng, Charis

    2017-08-01

    PTEN Hamartoma Tumor syndrome (PHTS) encompasses a clinical spectrum of heritable disorders including Cowden syndrome (CS), Bannayan-Riley-Ruvalcaba syndrome, and Proteus and Proteus-like syndrome that are associated with germline mutations in the PTEN tumor suppressor gene. Breast cancer risk estimates (67-85 %) for women with germline PTEN mutations are similar to those quoted for patients with germline mutations in the BRCA1/2 genes. With PTEN on several germline gene testing panels, finding PTEN mutations and variants have increased exponentially. PHTS can be differentiated from other hereditary cancer syndromes including Hereditary Breast Ovarian Cancer syndrome, Lynch syndrome, and hamartomatous polyposis syndromes based on personal as well as family history. However, many of the benign features of CS are common in the general population, making the diagnosis of CS challenging. Breast cancer patients with an identified germline PTEN mutation are at increased risk of endometrial, thyroid, renal, and colorectal cancers as well as a second breast cancer. Increased screening for the various component cancers as well as predictive testing in first-degree relatives is recommended. Prophylactic mastectomy may be considered especially if breast tissue is dense or if repeated breast biopsies have been necessary. Management of women with breast cancer suspected of CS who test negative for germline PTEN mutations should be managed as per a mutation carrier if she meets CS diagnostic criteria, and should be offered enrollment in research to identify other predisposition genes.

  15. Positive autoregulation of mrkHI by the cyclic di-GMP-dependent MrkH protein in the biofilm regulatory circuit of Klebsiella pneumoniae.

    PubMed

    Tan, Jason W H; Wilksch, Jonathan J; Hocking, Dianna M; Wang, Nancy; Srikhanta, Yogitha N; Tauschek, Marija; Lithgow, Trevor; Robins-Browne, Roy M; Yang, Ji; Strugnell, Richard A

    2015-05-01

    Klebsiella pneumoniae is an important cause of nosocomial infections, primarily through the formation of surface-associated biofilms to promote microbial colonization on host tissues. Expression of type 3 fimbriae by K. pneumoniae facilitates surface adherence, a process strongly activated by the cyclic di-GMP (c-di-GMP)-dependent transcriptional activator MrkH. In this study, we demonstrated the critical importance of MrkH in facilitating K. pneumoniae attachment on a variety of medically relevant materials and demonstrated the mechanism by which bacteria activate expression of type 3 fimbriae to colonize these materials. Sequence analysis revealed a putative MrkH recognition DNA sequence ("MrkH box"; TATCAA) located in the regulatory region of the mrkHI operon. Mutational analysis, electrophoretic mobility shift assay, and quantitative PCR experiments demonstrated that MrkH binds to the cognate DNA sequence to autoregulate mrkHI expression in a c-di-GMP-dependent manner. A half-turn deletion, but not a full-turn deletion, between the MrkH box and the -35 promoter element rendered MrkH ineffective in activating mrkHI expression, implying that a direct interaction between MrkH and RNA polymerase exists. In vivo analyses showed that residues L260, R265, N268, C269, E273, and I275 in the C-terminal domain of the RNA polymerase α subunit are involved in the positive control of mrkHI expression by MrkH and revealed the regions of MrkH required for DNA binding and transcriptional activation. Taken together, the data suggest a model whereby c-di-GMP-dependent MrkH recruits RNA polymerase to the mrkHI promoter to autoactivate mrkH expression. Increased MrkH production subsequently drives mrkABCDF expression when activated by c-di-GMP, leading to biosynthesis of type 3 fimbriae and biofilm formation. Bacterial biofilms can cause persistent infections that are refractory to antimicrobial treatments. This study investigated how a commonly encountered hospital

  16. Positive Autoregulation of mrkHI by the Cyclic Di-GMP-Dependent MrkH Protein in the Biofilm Regulatory Circuit of Klebsiella pneumoniae

    PubMed Central

    Tan, Jason W. H.; Hocking, Dianna M.; Wang, Nancy; Srikhanta, Yogitha N.; Tauschek, Marija; Lithgow, Trevor; Robins-Browne, Roy M.; Yang, Ji; Strugnell, Richard A.

    2015-01-01

    ABSTRACT Klebsiella pneumoniae is an important cause of nosocomial infections, primarily through the formation of surface-associated biofilms to promote microbial colonization on host tissues. Expression of type 3 fimbriae by K. pneumoniae facilitates surface adherence, a process strongly activated by the cyclic di-GMP (c-di-GMP)-dependent transcriptional activator MrkH. In this study, we demonstrated the critical importance of MrkH in facilitating K. pneumoniae attachment on a variety of medically relevant materials and demonstrated the mechanism by which bacteria activate expression of type 3 fimbriae to colonize these materials. Sequence analysis revealed a putative MrkH recognition DNA sequence (“MrkH box”; TATCAA) located in the regulatory region of the mrkHI operon. Mutational analysis, electrophoretic mobility shift assay, and quantitative PCR experiments demonstrated that MrkH binds to the cognate DNA sequence to autoregulate mrkHI expression in a c-di-GMP-dependent manner. A half-turn deletion, but not a full-turn deletion, between the MrkH box and the −35 promoter element rendered MrkH ineffective in activating mrkHI expression, implying that a direct interaction between MrkH and RNA polymerase exists. In vivo analyses showed that residues L260, R265, N268, C269, E273, and I275 in the C-terminal domain of the RNA polymerase α subunit are involved in the positive control of mrkHI expression by MrkH and revealed the regions of MrkH required for DNA binding and transcriptional activation. Taken together, the data suggest a model whereby c-di-GMP-dependent MrkH recruits RNA polymerase to the mrkHI promoter to autoactivate mrkH expression. Increased MrkH production subsequently drives mrkABCDF expression when activated by c-di-GMP, leading to biosynthesis of type 3 fimbriae and biofilm formation. IMPORTANCE Bacterial biofilms can cause persistent infections that are refractory to antimicrobial treatments. This study investigated how a commonly

  17. Bacteroides gingivalis-specific serum IgG and IgA subclass antibodies in periodontal diseases.

    PubMed Central

    Ogawa, T; Kusumoto, Y; Hamada, S; McGhee, J R; Kiyono, H

    1990-01-01

    The level of serum IgM, IgG and IgA antibodies including IgG1, IgG2, IgG3, IgG4, IgA1 and IgA2 subclass-specific antibodies to Bacteroides (Porphyromonas) gingivalis fimbriae and to lipopolysaccharide (LPS) were analysed in patients with different forms of periodontal disease (PD) and control subjects by ELISA. Among PD subjects, sera obtained from adult periodontitis (AP), rapidly progressive periodontitis (RPP) and gingivitis contained high titres of fimbriae-specific IgG antibodies (7500-15,000 ELISA units) followed by IgA (90-700 units) and IgM (30-90 units). In contrast, sera from localized juvenile periodontitis (LJP) subjects exhibited much lower titres of fimbriae-specific IgG (89 +/- 11 units), IgA (31 +/- 5 units) and IgM (17 +/- 3 units) antibodies. A similar response pattern was also seen in sera from normal subjects aged 35-41 years who practice normal oral hygiene, while sera of younger adults (aged 18-24) with superior hygiene did not have any antigen-specific antibodies. Analysis of IgG subclass anti-fimbriae responses revealed that the major response was IgG3 followed by IgG1, IgG2 and IgG4 in AP, RPP and gingivitis. Although lower, a similar pattern of IgG subclass titre was seen in LJP and normal subjects aged 35-41 years. When IgA subclass responses were measured in AP and RPP, higher titres of the fimbriae-specific response were noted with IgA1 when compared with IgA2. However, lower but approximately equal levels of fimbriae-specific IgA1 and IgA2 titres were seen in other PD groups. When anti-B. gingivalis LPS-specific responses were measured, the sera of AP patients contained high levels of IgG antibodies (2265 +/- 224 units) followed by IgA (411 +/- 90 units) and IgM (214 +/- 56 units). Further, IgG anti-LPS responses were mainly IgG2 followed by IgG4, IgG3 and IgG1. For IgA subclass responses, higher titres of anti-LPS-specific antibodies were noted in IgA2 subclass over IgA1. These results showed that higher anti-B. gingivalis antibody

  18. Pilot trials in Australia on eradication of footrot by flock specific vaccination.

    PubMed

    Dhungyel, O P; Lehmann, D R; Whittington, R J

    2008-12-10

    Footrot is a contagious disease of ruminants requiring strains of Dichelobacter nodosus that possess virulence factors including proteases and fimbriae. Sheep can be immunised against footrot using vaccine-containing fimbriae, either native or recombinant. The fimbriae are responsible for the serological K-agglutination reaction, which has been used to classify field isolates into nine major serogroups. The range of protection conferred by vaccination is largely restricted to the serogroup involved, but antigenic competition precludes effective vaccination with multivalent vaccines that contain all serogroups. However, vaccination with specific bivalent recombinant fimbrial vaccine led to eradication of virulent footrot from small ruminants in Nepal and the same result was obtained in Bhutan using a specific whole cell vaccine. In the study reported here two pilot trials have been conducted in Australian sheep flocks, one with a virulent form of footrot caused by a single serogroup F, and the other with an intermediate form also caused by a single serogroup C. In trial 1 pre-vaccination prevalence of clinical footrot in a group of randomly selected animals was 44%. This reduced to 2% at 3 months and 0.5% at 4 months, and there were no clinical cases at 5 months or at 16 months post-vaccination in the whole flock. Similarly in trial 2 pre-vaccination whole flock prevalence was 8.5%, while it was 2% at 3 months, 0.3% at 6 months and zero at 18 months post-vaccination. Use of flock specific monovalent whole cell vaccines over whole flocks for only one season and culling of the few non-responders has been a successful approach in eradication of the disease from both these flocks. This is the first study to report the successful use of specific vaccine for the intermediate form of footrot.

  19. Vaccines for preventing enterotoxigenic Escherichia coli infections in farm animals.

    PubMed

    Moon, H W; Bunn, T O

    1993-01-01

    Fimbrial vaccines are routinely given parenterally to pregnant cattle, sheep and swine to protect suckling newborn calves, lambs and pigs against enterotoxigenic Escherichia coli (ETEC) infections. Such vaccines are practical and effective because (1) most fatal ETEC infections in farm animals occur in the early neonatal period when the antibody titres in colostrum and milk are highest; (2) more than 90% of the ETEC in farm animals belong to a small family of fimbrial antigen types; (3) fimbriae consist of good protein antigens on the bacterial surface where they are readily accessible to antibody; (4) fimbriae are required for a critical step (adhesion-colonization) early in the pathogenesis of the disease. ETEC infections continue to be a significant clinical problem in farm animals in spite of extensive use of fimbriae-based vaccines. Definitive data on the efficacy of the commercial vaccines in field use are not available. The prevailing perception among animal health professionals is that the vaccines are effective, that the problem occurs chiefly among non-vaccinated animals, and that in some herds vaccination moves peak prevalence of disease from the first to the second or third week after birth, when mortality is lower. It has been suggested that extensive use of vaccines will rapidly select for the emergence of novel or previously low prevalence fimbrial antigen types. There is no evidence that this has happened after a decade of routine vaccine use in the United States. However, there is no active direct surveillance for such emergence.(ABSTRACT TRUNCATED AT 250 WORDS)

  20. Acute White Matter Tract Damage after Frontal Mild Traumatic Brain Injury.

    PubMed

    Herrera, Juan J; Bockhorst, Kurt; Kondraganti, Shakuntala; Stertz, Laura; Quevedo, João; Narayana, Ponnada A

    2017-01-15

    Our understanding of mild traumatic brain injury (mTBI) is still in its infancy and to gain a greater understanding, relevant animal models should replicate many of the features seen in human mTBI. These include changes to diffusion tensor imaging (DTI) parameters, absence of anatomical lesions on conventional neuroimaging, and neurobehavioral deficits. The Maryland closed head TBI model causes anterior-posterior plus sagittal rotational acceleration of the brain, frequently observed with motor vehicle and sports-related TBI injuries. The injury reflects a concussive injury model without skull fracture. The goal of our study was to characterize the acute (72 h) pathophysiological changes occurring following a single mTBI using magnetic resonance imaging (MRI), behavioral assays, and histology. We assessed changes in fractional anisotropy (FA), mean (MD), longitudinal (LD), and radial (RD) diffusivities relative to pre-injury baseline measures. Significant differences were observed in both the longitudinal and radial diffusivities in the fimbria compared with baseline. A significant difference in radial diffusivity was also observed in the splenium of the corpus callosum compared with baseline. The exploratory activity of the mTBI animals was also assessed using computerized activity monitoring. A significant decrease was observed in ambulatory distance, average velocity, stereotypic counts, and vertical counts compared with baseline. Histological examination of the mTBI brain sections indicated a significant decrease in the expression of myelin basic protein in the fimbria, splenium, and internal capsule. Our findings demonstrate the vulnerability of the white matter tracts, specifically the fimbria and splenium, and the ability of DTI to identify changes to the integrity of the white matter tracts following mTBI.

  1. The Role of Fibronectin in the Adherence and Inflammatory Response Induced by Enteroaggregative Escherichia coli on Epithelial Cells

    PubMed Central

    Yáñez, Dominique; Izquierdo, Mariana; Ruiz-Perez, Fernando; Nataro, James P.; Girón, Jorge A.; Vidal, Roberto M.; Farfan, Mauricio J.

    2016-01-01

    Enteroaggregative Escherichia coli (EAEC) infections are still one of the most important etiologic pathogens of diarrhea in children worldwide. EAEC pathogenesis comprises three stages: adherence and colonization, production of toxins, and diarrhea followed by inflammation. Previous studies have demonstrated that EAEC strains have the ability to bind to fibronectin (FN); however, the role this extracellular matrix protein plays in the inflammatory response induced by EAEC remains unknown. In this study, we postulated that FN-mediated adherence of EAEC strains to epithelial cells increases the expression of pro-inflammatory genes. To verify this hypothesis, we infected HEp-2 and HT-29 cells, in both the presence and absence of FN, with EAEC reference strain 042. We quantified IL-8 secretion and the relative expression of a set of genes regulated by the NF-κB pathway. Although FN increased EAEC adherence, no changes in IL-8 protein secretion or IL8 gene expression were observed. Similar observations were found in HEp-2 cells transfected with FN-siRNA and infected with EAEC. To evaluate the involvement of AAF/II fimbriae, we infected HEp-2 and HT-29 cells, in both the presence and absence of FN, with an EAEC 042aafA mutant strain transformed with a plasmid harboring the native aafA gene with a site-directed mutation in Lys72 residue (K72A and K72R strains). No changes in IL-8 secretion were observed. Finally, SEM immunogold assay of cells incubated with FN and infected with EAEC revealed that AAF fimbriae can bind to cells either directly or mediated by FN. Our data suggests that FN participates in AAF/II fimbriae-mediated adherence of EAEC to epithelial cells, but not in the inflammatory response of cells infected by this pathogen. PMID:28008386

  2. Porphyromonas gingivalis evasion of autophagy and intracellular killing by human myeloid dendritic cells involves DC-SIGN-TLR2 crosstalk.

    PubMed

    El-Awady, Ahmed R; Miles, Brodie; Scisci, Elizabeth; Kurago, Zoya B; Palani, Chithra D; Arce, Roger M; Waller, Jennifer L; Genco, Caroline A; Slocum, Connie; Manning, Matthew; Schoenlein, Patricia V; Cutler, Christopher W

    2015-02-01

    Signaling via pattern recognition receptors (PRRs) expressed on professional antigen presenting cells, such as dendritic cells (DCs), is crucial to the fate of engulfed microbes. Among the many PRRs expressed by DCs are Toll-like receptors (TLRs) and C-type lectins such as DC-SIGN. DC-SIGN is targeted by several major human pathogens for immune-evasion, although its role in intracellular routing of pathogens to autophagosomes is poorly understood. Here we examined the role of DC-SIGN and TLRs in evasion of autophagy and survival of Porphyromonas gingivalis in human monocyte-derived DCs (MoDCs). We employed a panel of P. gingivalis isogenic fimbriae deficient strains with defined defects in Mfa-1 fimbriae, a DC-SIGN ligand, and FimA fimbriae, a TLR2 agonist. Our results show that DC-SIGN dependent uptake of Mfa1+P. gingivalis strains by MoDCs resulted in lower intracellular killing and higher intracellular content of P. gingivalis. Moreover, Mfa1+P. gingivalis was mostly contained within single membrane vesicles, where it survived intracellularly. Survival was decreased by activation of TLR2 and/or autophagy. Mfa1+P. gingivalis strain did not induce significant levels of Rab5, LC3-II, and LAMP1. In contrast, P. gingivalis uptake through a DC-SIGN independent manner was associated with early endosomal routing through Rab5, increased LC3-II and LAMP-1, as well as the formation of double membrane intracellular phagophores, a characteristic feature of autophagy. These results suggest that selective engagement of DC-SIGN by Mfa-1+P. gingivalis promotes evasion of antibacterial autophagy and lysosome fusion, resulting in intracellular persistence in myeloid DCs; however TLR2 activation can overcome autophagy evasion and pathogen persistence in DCs.

  3. Porphyromonas gingivalis Evasion of Autophagy and Intracellular Killing by Human Myeloid Dendritic Cells Involves DC-SIGN-TLR2 Crosstalk

    PubMed Central

    El-Awady, Ahmed R.; Miles, Brodie; Scisci, Elizabeth; Kurago, Zoya B.; Palani, Chithra D.; Arce, Roger M.; Waller, Jennifer L.; Genco, Caroline A.; Slocum, Connie; Manning, Matthew; Schoenlein, Patricia V.; Cutler, Christopher W.

    2015-01-01

    Signaling via pattern recognition receptors (PRRs) expressed on professional antigen presenting cells, such as dendritic cells (DCs), is crucial to the fate of engulfed microbes. Among the many PRRs expressed by DCs are Toll-like receptors (TLRs) and C-type lectins such as DC-SIGN. DC-SIGN is targeted by several major human pathogens for immune-evasion, although its role in intracellular routing of pathogens to autophagosomes is poorly understood. Here we examined the role of DC-SIGN and TLRs in evasion of autophagy and survival of Porphyromonas gingivalis in human monocyte-derived DCs (MoDCs). We employed a panel of P. gingivalis isogenic fimbriae deficient strains with defined defects in Mfa-1 fimbriae, a DC-SIGN ligand, and FimA fimbriae, a TLR2 agonist. Our results show that DC-SIGN dependent uptake of Mfa1+P. gingivalis strains by MoDCs resulted in lower intracellular killing and higher intracellular content of P. gingivalis. Moreover, Mfa1+P. gingivalis was mostly contained within single membrane vesicles, where it survived intracellularly. Survival was decreased by activation of TLR2 and/or autophagy. Mfa1+P. gingivalis strain did not induce significant levels of Rab5, LC3-II, and LAMP1. In contrast, P. gingivalis uptake through a DC-SIGN independent manner was associated with early endosomal routing through Rab5, increased LC3-II and LAMP-1, as well as the formation of double membrane intracellular phagophores, a characteristic feature of autophagy. These results suggest that selective engagement of DC-SIGN by Mfa-1+P. gingivalis promotes evasion of antibacterial autophagy and lysosome fusion, resulting in intracellular persistence in myeloid DCs; however TLR2 activation can overcome autophagy evasion and pathogen persistence in DCs. PMID:25679217

  4. Nucleotide sequences of two fimbrial major subunit genes, pmpA and ucaA, from canine-uropathogenic Proteus mirabilis strains.

    PubMed

    Bijlsma, I G; van Dijk, L; Kusters, J G; Gaastra, W

    1995-06-01

    Proteus mirabilis strains were isolated from dogs with urinary tract infection (UTI) and fimbriae were prepared from two strains. The N-terminal amino acid sequences of the major fimbrial subunits were determined and both sequences appeared identical to the N-terminal amino acid sequence of a urinary cell adhesin (UCA) (Wray, S. K., Hull, S. I., Cook, R. G., Barrish, J. & Hull, R. A., 1986, Infect Immun 54, 43-49). The genes of two different major fimbrial subunits were cloned using oligonucleotide probes that were designed on the basis of the N-terminal UCA sequence. Nucleotide sequencing revealed the complete ucaA gene of 540 bp (from strain IVB247) encoding a polypeptide of 180 amino acids, including a 22 amino acid signal sequence peptide, and the pmpA (P. mirabilis P-like pili) gene of 549 bp (from strain IVB219) encoding a polypeptide of 183 amino acids, including a 23 amino acid signal sequence. Hybridization experiments gave clear indications of the presence of both kinds of fimbriae in many UTI-related canine P. mirabilis isolates. However, the presence of these fimbriae could not be demonstrated in P. vulgaris or other Proteus-related species. Database analysis of amino acid sequences of major subunit proteins revealed that the UcaA protein shares about 56% amino acid identity with the F17A and F111A major fimbrial subunits from bovine enterotoxigenic Escherichia coli. In turn, the PmpA protein more closely resembled the pyelonephritis-associated pili (Pap)-like major subunit protein from UTI-related E. coli. The evolutionary relationship of UcaA, PmpA and various other fimbrial subunit proteins is presented in a phylogenetic tree.

  5. Exopolysaccharides Synthesized by Lactobacillus reuteri Protect against Enterotoxigenic Escherichia coli in Piglets

    PubMed Central

    Chen, Xiao Yan; Woodward, Adrienne; Zijlstra, Ruurd T.

    2014-01-01

    Enterotoxigenic Escherichia coli (ETEC) is a major cause of diarrhea in piglets; ETEC cells colonize the intestinal mucosa with adhesins and deliver toxins that cause fluid loss. This study determined the antiadhesive properties of bacterial exopolysaccharides (reuteran and levan) and related glycans (dextran and inulin) in a small intestinal segment perfusion (SISP) model. The SISP model used 10 jejunal segments from 5-week-old piglets. Five segments were infected with ETEC expressing K88 fimbriae (ETEC K88), while five segments were treated with saline. Every two segments (ETEC and non-ETEC infected) were infused with 65 ml of 10 g liter−1 of glycans or saline (control) for 8 h. High-resolution melting-curve (HRM) quantitative PCR (qPCR) indicated that E. coli is the dominant bacterium in infected segments, while other bacteria were predominant in noninfected segments. Infection by ETEC K88 was also verified by qPCR; gene copy numbers of K88 fimbriae and the heat-labile toxin (LT) in mucosal scrapings and outflow fluid of infected segments were significantly higher than those in noninfected segments. Genes coding for K88 fimbriae and LT were also detected in noninfected segments. LT amplicons from infected and noninfected segments were 99% identical over 481 bp, demonstrating the presence of autochthonous ETEC K88. All glycans reduced fluid loss caused by ETEC K88 infection. Reuteran tended (P = 0.06) to decrease ETEC K88 levels in mucosal scraping sample, as judged by qPCR. Fluorescent in situ hybridization analysis demonstrated that reuteran significantly (P = 0.012) decreased levels of adherent ETEC K88. Overall, reuteran may prevent piglet diarrhea by reducing adhesion of ETEC K88. PMID:25015886

  6. Insights into a Multidrug Resistant Escherichia coli Pathogen of the Globally Disseminated ST131 Lineage: Genome Analysis and Virulence Mechanisms

    PubMed Central

    Sarkar, Sohinee; Phan, Minh-Duy; Petty, Nicola K.; Bachmann, Nathan; Szubert, Marek; Sidjabat, Hanna E.; Paterson, David L.; Upton, Mathew; Schembri, Mark A.

    2011-01-01

    Escherichia coli strains causing urinary tract infection (UTI) are increasingly recognized as belonging to specific clones. E. coli clone O25b:H4-ST131 has recently emerged globally as a leading multi-drug resistant pathogen causing urinary tract and bloodstream infections in hospitals and the community. While most molecular studies to date examine the mechanisms conferring multi-drug resistance in E. coli ST131, relatively little is known about their virulence potential. Here we examined E. coli ST131 clinical isolates from two geographically diverse collections, one representing the major pathogenic lineages causing UTI across the United Kingdom and a second representing UTI isolates from patients presenting at two large hospitals in Australia. We determined a draft genome sequence for one representative isolate, E. coli EC958, which produced CTX-M-15 extended-spectrum β-lactamase, CMY-23 type AmpC cephalosporinase and was resistant to ciprofloxacin. Comparative genome analysis indicated that EC958 encodes virulence genes commonly associated with uropathogenic E. coli (UPEC). The genome sequence of EC958 revealed a transposon insertion in the fimB gene encoding the activator of type 1 fimbriae, an important UPEC bladder colonization factor. We identified the same fimB transposon insertion in 59% of the ST131 UK isolates, as well as 71% of ST131 isolates from Australia, suggesting this mutation is common among E. coli ST131 strains. Insertional inactivation of fimB resulted in a phenotype resembling a slower off-to-on switching for type 1 fimbriae. Type 1 fimbriae expression could still be induced in fimB-null isolates; this correlated strongly with adherence to and invasion of human bladder cells and bladder colonisation in a mouse UTI model. We conclude that E. coli ST131 is a geographically widespread, antibiotic resistant clone that has the capacity to produce numerous virulence factors associated with UTI. PMID:22053197

  7. An electron microscope survey of the surface structures and hydrophobicity of oral and non-oral species of the bacterial genus Bacteroides.

    PubMed

    Handley, P S; Tipler, L S

    1986-01-01

    Seventeen strains of Bacteroides representing 10 species were examined by negative staining; the majority were from the mouth but a few non-oral strains were included. Seven species had peritrichously-arranged, non-flagellar appendages which could be divided by morphology and ultrastructure into two subgroups, fibrils and fimbriae. Bacteroides asaccharolyticus strains B536 and B537 and Bacteroides gingivalis strains W50, W83, WPH15 and WPH35 had fimbriae with mean width of 4.4 nm and 0.5 to 6.0 microns long depending on the strain. The fimbrial length within each strain also varied. Fibrils were present on two fresh oral isolates of Bacteroides melaninogenicus, Bacteroides intermedius strains T588 and W09, Bacteroides corporis ATCC 33547, Bacteroides oralis ATCC 33269 and Bacteroides buccae ATCC 33574. Fibrils consistently clumped into bundles of variable thickness and formed a fringe around the cell periphery, ranging from 0.27 to 1.2 microns long depending on the strain. Fibril lengths of each strain were uniform. Fibrils had no measurable width and the clumps tapered towards the free ends. Bacteroides loeschii VPI 9085, Bacteroides pentosaceus strains NP333 and J1 and Bacteroides capillosus 29799 had no detectable surface appendages. Fimbriate strains had a layer outside the outer membrane, with a mean thickness of between 17.8 and 28.6 nm. Both fibrillar and fimbriate strains produced many small membranous vesicles budding from the outer membrane. There were two morphological forms of vesicles, ones with either fimbriae or fibrils (species-dependent) and ones with no attached appendages. Of eleven strains tested for cell-surface hydrophobicity by partitioning between hexadecane and buffer, all but one was non-hydrophobic.

  8. New Aspects of RpoE in Uropathogenic Proteus mirabilis

    PubMed Central

    Liu, Ming-Che; Kuo, Kuan-Ting; Chien, Hsiung-Fei; Tsai, Yi-Lin

    2014-01-01

    Proteus mirabilis is a common human pathogen causing recurrent or persistent urinary tract infections (UTIs). The underlying mechanisms for P. mirabilis to establish UTIs are not fully elucidated. In this study, we showed that loss of the sigma factor E (RpoE), mediating extracytoplasmic stress responses, decreased fimbria expression, survival in macrophages, cell invasion, and colonization in mice but increased the interleukin-8 (IL-8) expression of urothelial cells and swarming motility. This is the first study to demonstrate that RpoE modulated expression of MR/P fimbriae by regulating mrpI, a gene encoding a recombinase controlling the orientation of MR/P fimbria promoter. By real-time reverse transcription-PCR, we found that the IL-8 mRNA amount of urothelial cells was induced significantly by lipopolysaccharides extracted from rpoE mutant but not from the wild type. These RpoE-associated virulence factors should be coordinately expressed to enhance the fitness of P. mirabilis in the host, including the avoidance of immune attacks. Accordingly, rpoE mutant-infected mice displayed more immune cell infiltration in bladders and kidneys during early stages of infection, and the rpoE mutant had a dramatically impaired ability of colonization. Moreover, it is noteworthy that urea (the major component in urine) and polymyxin B (a cationic antimicrobial peptide) can induce expression of rpoE by the reporter assay, suggesting that RpoE might be activated in the urinary tract. Altogether, our results indicate that RpoE is important in sensing environmental cues of the urinary tract and subsequently triggering the expression of virulence factors, which are associated with the fitness of P. mirabilis, to build up a UTI. PMID:25547796

  9. New aspects of RpoE in uropathogenic Proteus mirabilis.

    PubMed

    Liu, Ming-Che; Kuo, Kuan-Ting; Chien, Hsiung-Fei; Tsai, Yi-Lin; Liaw, Shwu-Jen

    2015-03-01

    Proteus mirabilis is a common human pathogen causing recurrent or persistent urinary tract infections (UTIs). The underlying mechanisms for P. mirabilis to establish UTIs are not fully elucidated. In this study, we showed that loss of the sigma factor E (RpoE), mediating extracytoplasmic stress responses, decreased fimbria expression, survival in macrophages, cell invasion, and colonization in mice but increased the interleukin-8 (IL-8) expression of urothelial cells and swarming motility. This is the first study to demonstrate that RpoE modulated expression of MR/P fimbriae by regulating mrpI, a gene encoding a recombinase controlling the orientation of MR/P fimbria promoter. By real-time reverse transcription-PCR, we found that the IL-8 mRNA amount of urothelial cells was induced significantly by lipopolysaccharides extracted from rpoE mutant but not from the wild type. These RpoE-associated virulence factors should be coordinately expressed to enhance the fitness of P. mirabilis in the host, including the avoidance of immune attacks. Accordingly, rpoE mutant-infected mice displayed more immune cell infiltration in bladders and kidneys during early stages of infection, and the rpoE mutant had a dramatically impaired ability of colonization. Moreover, it is noteworthy that urea (the major component in urine) and polymyxin B (a cationic antimicrobial peptide) can induce expression of rpoE by the reporter assay, suggesting that RpoE might be activated in the urinary tract. Altogether, our results indicate that RpoE is important in sensing environmental cues of the urinary tract and subsequently triggering the expression of virulence factors, which are associated with the fitness of P. mirabilis, to build up a UTI.

  10. Adherence properties of an mrkD-negative mutant of Klebsiella pneumoniae.

    PubMed Central

    Hornick, D B; Thommandru, J; Smits, W; Clegg, S

    1995-01-01

    The role of the mrkD gene in attachment by a type 3 fimbriate Klebsiella pneumoniae strain was further characterized. A clinical isolate, K. pneumoniae IA565, was found to contain two copies of the gene encoding the fimbrial subunit, mrkA, and one copy of the gene encoding the adhesin subunit, mrkD. One copy of mrkA was located on the bacterial chromosome, and the other copy was associated with mrkD and located on a plasmid. The plasmid-borne mrk gene cluster was lost when K. pneumoniae IA565 was subcultured serially in broth at 44 degrees C. The resulting mrkD-negative strain, designated K. pneumoniae IApc35, did not exhibit the following adherence characteristics associated with K. pneumoniae possessing MrkD-positive fimbriae: agglutination of tannic acid-treated human erythrocytes and attachment to trypsinized human buccal cells. However, K. pneumoniae IApc35 produced type 3 fimbriae that were composed of the characteristic 21.5-kDa major fimbrial subunit, were reactive with specific serum, and were visualized specifically by immunoelectron microscopy. K. pneumoniae IApc35 retained a copy of the mrkA gene on its chromosome. This mrkA-containing gene cluster could be complemented by a recombinant plasmid carrying only the mrkD gene, resulting in restoration of the K. pneumoniae IA565-like adhesive phenotype and demonstration of type 3 filament-associated MrkD subunits by using colloidal gold labeling and immunoelectron microscopy. These data indicate that K. pneumoniae may contain multiple copies of the mrk genes which may be present simultaneously on both plasmid and chromosomal DNAs and which may encode fimbriae with different binding specificities. PMID:7729917

  11. Plant-synthesized E. coli CFA/I fimbrial protein protects Caco-2 cells from bacterial attachment.

    PubMed

    Lee, Jin-Yong; Yu, Jie; Henderson, David; Langridge, William H R

    2004-11-25

    A DNA fragment encoding the cholera toxin A2 subunit (CTA2) linked to the enterotoxigenic Escherichia coli (ETEC) colony forming fimbrial antigen CFA/I was inserted into a plant expression vector containing the cholera toxin B subunit (CTB) fused to the rotavirus enterotoxin 22 amino acid epitope NSP422. Anti-CFA/I antibodies recognized a single band of approximately 72-kDa in transformed potato tuber tissue consistent with CFA/I-CTA2 and CTB-NSP4 fusion protein assembly into a cholera holotoxin-like structure. Enzyme-linked immunosorbent assay (GM1 ELISA) indicated that the CFA/I-CTA2 fusion protein bound specific GM1 ganglioside membrane receptors and made up approximately 0.002% of the total soluble tuber protein. Oral immunization of BALB/c mice with transformed tuber tissues generated anti-CFA/I serum and intestinal IgG and IgA secretory antibodies. Attachment of ETEC H10407 to enterocyte-like Caco-2 human colon carcinoma cells incubated with antiserum from immunized mice was reduced by 15% in comparison with Caco-2 cells incubated with serum from unimmunized mice. Immunogold staining of bacterial preparations revealed deposition of gold particles on E. coli H10407 fimbria incubated with immune serum but not on fimbria treated with sera from unimmunized mice demonstrating the specificity of antibodies in the immune serum for binding to CFA/I protein containing fimbria. The protection against toxic E. coli binding to Caco-2 cells generated by antisera from mice immunized with plant-synthesized CFA/I antigen demonstrates the feasibility of plant-based multi-component vaccine protection against enterotoxigenic E. coli, rotavirus and cholera, three enteric diseases that together exert the highest levels of child morbidity and mortality in economically emerging countries.

  12. Regulatory T-cell vaccination independent of auto-antigen.

    PubMed

    Pascual, David W; Yang, Xinghong; Holderness, Kathryn; Jun, SangMu; Maddaloni, Massimo; Kochetkova, Irina

    2014-03-14

    To date, efforts to treat autoimmune diseases have primarily focused on the disease symptoms rather than on the cause of the disease. In large part, this is attributed to not knowing the responsible auto-antigens (auto-Ags) for driving the self-reactivity coupled with the poor success of treating autoimmune diseases using oral tolerance methods. Nonetheless, if tolerogenic approaches or methods that stimulate regulatory T (Treg) cells can be devised, these could subdue autoimmune diseases. To forward such efforts, our approach with colonization factor antigen I (CFA/I) fimbriae is to establish bystander immunity to ultimately drive the development of auto-Ag-specific Treg cells. Using an attenuated Salmonella vaccine expressing CFA/I fimbriae, fimbriae-specific Treg cells were induced without compromising the vaccine's capacity to protect against travelers' diarrhea or salmonellosis. By adapting the vaccine's anti-inflammatory properties, it was found that it could also dampen experimental inflammatory diseases resembling multiple sclerosis (MS) and rheumatoid arthritis. Because of this bystander effect, disease-specific Treg cells are eventually induced to resolve disease. Interestingly, this same vaccine could elicit the required Treg cell subset for each disease. For MS-like disease, conventional CD25(+) Treg cells are stimulated, but for arthritis CD39(+) Treg cells are induced instead. This review article will examine the potential of treating autoimmune diseases without having previous knowledge of the auto-Ag using an innocuous antigen to stimulate Treg cells via the production of transforming growth factor-β and interleukin-10.

  13. Identification and Whole Genome Sequencing of the First Case of Kosakonia radicincitans Causing a Human Bloodstream Infection

    PubMed Central

    Bhatti, Micah D.; Kalia, Awdhesh; Sahasrabhojane, Pranoti; Kim, Jiwoong; Greenberg, David E.; Shelburne, Samuel A.

    2017-01-01

    The taxonomy of Enterobacter species is rapidly changing. Herein we report a bloodstream infection isolate originally identified as Enterobacter cloacae by Vitek2 methodology that we found to be Kosakonia radicincitans using genetic means. Comparative whole genome sequencing of our isolate and other published Kosakonia genomes revealed these organisms lack the AmpC β-lactamase present on the chromosome of Enterobacter sp. A fimbriae operon primarily found in Escherichia coli O157:H7 isolates was present in our organism and other available K. radicincitans genomes. This is the first report of a Kosakonia species, which are typically associated with plants, causing a human infection. PMID:28174569

  14. Monoclonal antibodies against enterotoxigenic Escherichia coli colonization factor antigen I (CFA/I) that cross-react immunologically with heterologous CFAs.

    PubMed Central

    Rudin, A; McConnell, M M; Svennerholm, A M

    1994-01-01

    Enterotoxigenic Escherichia coli binds to enterocytes in the small intestine by means of antigenically distinct colonization factors (CFs), usually termed colonization factor antigens (CFAs), coli surface antigens (CS), or putative colonization factor antigens (PCFs). To explore the immunological relationship between different CFs, we dissociated CFA/I fimbriae into subunits and produced monoclonal antibodies (MAbs) against these subunits. We selected three MAbs that cross-reacted immunologically with a number of different, whole purified CFs in a dot blot test and with the corresponding subunits in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. One of the MAbs, i.e., subunit CFA/I 17:8 (S-CFA/I 17:8), reacted more strongly with subunits of CFA/I than with whole purified fimbriae. This MAb cross-reacted with whole purified fimbriae and subunits of CS4, PCFO166, CS1, and CS2. Moreover, it bound strongly to a peptide of 25 amino acids corresponding to the N-terminal end of CFA/I. The other two MAbs, i.e., S-CFA/I 5:6 and S-CFA/I 8:11, cross-reacted with CS1, CS2, CS4, PCFO166, and CS17 fimbriae but reacted only slightly or not at all with the CFA/I peptide. MAbs S-CFA/I 17:8 and S-CFA/I 5:6 were shown to inhibit hemagglutination by bacterial strains that express either CFA/I, CS1, or CS4. In addition, the binding of enterotoxigenic E. coli strains expressing CFA/I, CS2, CS4, and PCFO166 to enterocyte-like cell-line Caco-2 was inhibited by both MAbs. These results show that several antigenically different CFs have common epitopes and that among these at least one is located in the N-terminal end of the subunit protein. Moreover, antibodies against the common epitopes seem to block binding of the bacterial strains that express different CFs to both erythrocytes and Caco-2 cells. Images PMID:7927693

  15. Pilicides inhibit the FGL chaperone/usher assisted biogenesis of the Dr fimbrial polyadhesin from uropathogenic Escherichia coli

    PubMed Central

    2013-01-01

    Background The global spread of bacterial resistance has given rise to a growing interest in new anti-bacterial agents with a new strategy of action. Pilicides are derivatives of ring-fused 2-pyridones which block the formation of the pili/fimbriae crucial to bacterial pathogenesis. They impair by means of a chaperone-usher pathway conserved in the Gram-negative bacteria of adhesive structures biogenesis. Pili/fimbriae of this type belong to two subfamilies, FGS and FGL, which differ in the details of their assembly mechanism. The data published to date have shown that pilicides inhibit biogenesis of type 1 and P pili of the FGS type which are encoded by uropathogenic E. coli strains. Results We evaluated the anti-bacterial activity of literature pilicides as blockers of the assembly of a model example of FGL-type adhesive structures, – the Dr fimbriae encoded by a dra gene cluster of uropathogenic Escherichia coli strains. In comparison to the strain grown without pilicide, the Dr+ bacteria cultivated in the presence of the 3.5 mM concentration of pilicides resulted in a reduction of 75 to 87% in the adherence properties to CHO cells expressing Dr fimbrial DAF receptor protein. Using quantitative assays, we determined the amount of Dr fimbriae in the bacteria cultivated in the presence of 3.5 mM of pilicides to be reduced by 75 to 81%. The inhibition effect of pilicides is concentration dependent, which is a crucial property for their use as potential anti-bacterial agents. The data presented in this article indicate that pilicides in mM concentration effectively inhibit the adherence of Dr+ bacteria to the host cells, – the crucial, initial step in bacterial pathogenesis. Conclusions Structural analysis of the DraB chaperone clearly showed it to be a model of the FGL subfamily of chaperones. This permits us to conclude that analyzed pilicides in mM concentration are effective inhibitors of the assembly of adhesins belonging to the Dr family, and more

  16. Bilateral Ovarian Teratoma: One Parasitic Twisted In-situ and Another Parasitic at the Hepato Renal Space.

    PubMed

    Chitrakar, N S; Suwal, S; Neupane, S

    2015-01-01

    Parasitic ovarian dermoid cysts are very rare. We report a rare case of bilateral ovarian dermoid cysts with parasitic teratoma at the hepato renal space measuring 11x11x6 cm while the other was twisted measuring 10x6x5 cm.Right ovary and tube were absent. The mass found at the hepato renal space was surrounded by and adherent to the omentum with viable tubal fimbria like structure at upper surface. Histopathologically both masses were confirmed as mature ovarian teratoma. The etiology of parasitic teratoma would be due to torsion followed by autoamputation and reimplantation of the right ovarian dermoid cyst.

  17. Glycan-functionalized diamond nanoparticles as potent E. coli anti-adhesives

    NASA Astrophysics Data System (ADS)

    Barras, Alexandre; Martin, Fernando Ariel; Bande, Omprakash; Baumann, Jean-Sébastien; Ghigo, Jean-Marc; Boukherroub, Rabah; Beloin, Christophe; Siriwardena, Aloysius; Szunerits, Sabine

    2013-02-01

    Bacterial attachment and subsequent biofilm formation on biotic surfaces or medical devices is an increasing source of infections in clinical settings. A large proportion of these biofilm-related infections are caused by Escherichia coli, a major nosocomial pathogen, in which the major adhesion factor is the FimH adhesin located at the tip of type 1 fimbriae. Inhibition of FimH-mediated adhesion has been identified as an efficient antibiotic-alternative strategy to potentially reduce E. coli-related infections. In this article we demonstrate that nanodiamond particles, covently modified with mannose moieties by a ``click'' chemistry approach, are able to efficiently inhibit E. coli type 1 fimbriae-mediated adhesion to eukaryotic cells with relative inhibitory potency (RIP) of as high as 9259 (bladder cell adhesion assay), which is unprecedented when compared with RIP values previously reported for alternate multivalent mannose-functionalized nanostructures designed to inhibit E. coli adhesion. Also remarkable is that these novel mannose-modified NDs reduce E. coli biofilm formation, a property previously not observed for multivalent glyco-nanoparticles and rarely demonstrated for other multivalent or monovalent mannose glycans. This work sets the stage for the further evaluation of these novel NDs as an anti-adhesive therapeutic strategy against E. coli-derived infections.Bacterial attachment and subsequent biofilm formation on biotic surfaces or medical devices is an increasing source of infections in clinical settings. A large proportion of these biofilm-related infections are caused by Escherichia coli, a major nosocomial pathogen, in which the major adhesion factor is the FimH adhesin located at the tip of type 1 fimbriae. Inhibition of FimH-mediated adhesion has been identified as an efficient antibiotic-alternative strategy to potentially reduce E. coli-related infections. In this article we demonstrate that nanodiamond particles, covently modified with

  18. The location of four fimbrin-encoding genes, agfA, fimA, sefA and sefD, on the Salmonella enteritidis and/or S. typhimurium XbaI-BlnI genomic restriction maps.

    PubMed

    Collinson, S K; Liu, S L; Clouthier, S C; Banser, P A; Doran, J L; Sanderson, K E; Kay, W W

    1996-02-22

    Four fimbrin-encoding genes, fimA (type-1 or SEF21 fimbriae), agfA (thin aggregative or SEF17 fimbriae), sefA (SEF14 fimbriae and sefD (SEF18 fimbriae) from Salmonella enteritidis (Se) 27655-3b were located onto the XbaI-BlnI genomic restriction maps of Salmonella typhimurium (St) LT2 and Se strains SSU7998 and 27655-3b. The XbaI or BlnI genomic fragments carrying these genes were identified by hybridization with labeled oligodeoxyribonucleotides or fimbrin-encoding genes. The fimbrin-encoding genes were not encoded by the virulence plasmids, but were located on chromosomal DNA fragments. The position of each gene on a given XbaI fragment was determined by hybridization of a series of XbaI-digested genomic DNA samples from previously characterized Tn10 mutants of Se and St with its respective probe. The fimA gene mapped near 13 centisomes (Cs) between purE884::Tn10 at 12.6 Cs (11.8 min) and apeE2::Tn10 at 12.8 Cs (12.3 min) beside the first XbaI site at 13.0 Cs in St or between purE884::Tn10 at 12.6 Cs and the XbaI site at 13.6 Cs in Se. The agfA gene mapped near 26 Cs between putA::Tn10 and pyrC691::Tn10 in St, but near 40 Cs between pncX::Tn10 and the XbaI site at 43.3 Cs in Se. This difference in map position was due to the location of agfA near one end of the 815-kb chromosomal fragment inverted between Se and St. The sefA and sefD genes mapped precisely at 97.6 Cs in Se, but were absent from the genome of St LT2. To verify the mapping procedures used herein, tctC was also mapped in both Salmonella serovars. As expected, tctC mapped near 60 Cs in both St and Se, thereby confirming previous studies.

  19. An unwritten anatomy lesson: The influence of Roman clothing on neuroanatomical terminology: In memoriam Albert L. Rhoton, Jr. (1932-2016).

    PubMed

    Turliuc, Dana Mihaela; Turliuc, Serban; Cucu, Andrei Ionut; Sava, Anca; Dumitrescu, Gabriela Florenta; Cărăuleanu, Alexandru; Buzdugă, Cătălin; Trandafir, Daniela; Costea, Claudia Florida

    2016-09-01

    Throughout the centuries, anatomists attempting to denominate the new structures they discovered have found inspiration in the civilization of ancient Rome and the clothing worn by its citizens. This aricle presents the origins of seven neuroanatomical terms, fimbria, velum, funiculus, lemniscus, corona, splenium, and cingulum, inspired by the clothing and jewellery of Roman women and the military attire of Roman soldiers. Thus, through their apparel, the Romans influenced the Terminologia Anatomica and "clothed" the structures of the brain and spinal cord, making them immortal. Clin. Anat. 29:685-690, 2016. © 2016 Wiley Periodicals, Inc.

  20. Moraxella bovis hemagglutinins: effect of carbohydrates, heating and erythrocytes.

    PubMed Central

    Gil-Turnes, C; Ribeiro, G A

    1985-01-01

    Several properties of the adhesins of eight isolates of Moraxella bovis recovered from cattle suffering from infectious keratoconjunctivitis, were studied. Adhesions were detected through autoagglutination in saline and hemagglutination. Autoagglutinating strains agglutinated red blood cells of the chicken, rabbit, sheep and swine, but not those of the guinea pig. The adhesins were not inhibited by D-mannose or D-galactose and resisted heating at 100 degrees C for 15 minutes. Magnesium chloride at a final concentration of 10% inhibited autoagglutination and hemagglutination. The value of the hemagglutination test for monitoring synthesis of fimbriae by M. bovis, is discussed. PMID:3986674

  1. Subcortical deafferentation impairs behavioral reinforcement of long-term potentiation in the dentate gyrus of freely moving rats.

    PubMed

    Almaguer-Melian, W; Rosillo, J C; Frey, J U; Bergado, J A

    2006-01-01

    Long-term potentiation is a form of neural functional plasticity which has been related with memory formation and recovery of function after brain injury. Previous studies have shown that a transient early-long-term potentiation can be prolonged by direct stimulation of distinct brain areas, or behavioral stimuli with a high motivational content. The basolateral amygdala and other subcortical structures, like the medial septum and the locus coeruleus, are involved in mediating the reinforcing effect. We have previously shown that the lesion of the fimbria-fornix--the main entrance of subcortical afferents to the hippocampus--abolishes the reinforcing basolateral amygdala-effects on long-term potentiation in the dentate gyrus in vivo. It remains to be investigated, however, if such subcortical afferents may also be important for behavioral reinforcement of long-term potentiation. Young-adult (8 weeks) Sprague-Dawley male rats were fimbria-fornix-transected under anesthesia, and electrodes were implanted at the dentate gyrus and the perforant path. One week after surgery the freely moving animals were studied. Fimbria-fornix-lesion reduced the ability of the animals to develop long-term potentiation when a short pulse duration was used for tetanization (0.1 ms per half-wave of a biphasic stimulus), whereas increasing the pulse duration to 0.2 ms per half-wave during tetanization resulted in a transient early-long-term potentiation lasting about 4 h in the lesioned animals, comparable to that obtained in non-lesioned or sham-operated control rats. In water-deprived (24 h) control animals, i.e. in non-lesioned and sham-operated rats, early-long-term potentiation could be behaviorally reinforced by drinking 15 min after tetanization. However, in fimbria-fornix-lesioned animals long-term potentiation-reinforcement by drinking was not detected. This result indicates that the effect of behavioral-motivational stimuli to reinforce long-term potentiation is mediated by

  2. Virulence factors of uropathogenic Escherichia coli.

    PubMed

    Emody, L; Kerényi, M; Nagy, G

    2003-10-01

    Virulence factors of Escherichia coli are of two main types; those produced on the surface of the cell and those produced within the cell and then exported to the site of action. Those on the surface include different sorts of fimbriae that have a role in adhesion to the surface of host cells but may also have additional roles such as tissue invasion, biofilm formation or cytokine induction. The activities of cell wall components are discussed and several exported virulence factors are described that have anti host cell activities. Others virulence factors enable the bacteria to grow in an environment of iron restriction.

  3. pH6 antigen (PsaA protein) of Yersinia pestis, a novel bacterial Fc-receptor.

    PubMed

    Zav'yalov, V P; Abramov, V M; Cherepanov, P G; Spirina, G V; Chernovskaya, T V; Vasiliev, A M; Zav'yalova, G A

    1996-05-01

    It was found that recombinant pH6 antigen (rPsaA protein) forming virulence-associated fimbriae on the surface of Yersinia pestis at pH 6.7 in host macrophage phagolysosomes or extracellularly in abscesses such as buboes, is a novel bacterial Fc-receptor. rPsaA protein displays reactivity with human IgG1, IgG2 and IgG3 subclasses but does not react with rabbit, mouse and sheep IgG.

  4. Vaccines for Proteus mirabilis in urinary tract infection.

    PubMed

    Li, Xin; Mobley, Harry L T

    2002-06-01

    Proteus mirabilis is a documented cause of urinary tract infection (UTI) in the complicated urinary tract. Urease-mediated urea hydrolysis is responsible for both virulence of the organism and the ability to cause urolithiasis. A urease-negative mutant of P. mirabilis is unable to initiate stone formation and colonizes the kidney at a significantly lower rate. The considerable pathology caused by P. mirabilis warrants the development of a vaccine. We have initiated the advancement of vaccine studies and have determined that the MR/P fimbria, a surface adhesin of P. mirabilis, is a promising vaccine candidate. Successful vaccination would be expected both to prevent colonization by P. mirabilis and urolithiasis.

  5. A new method for the extraction and purification of K99 pili from enterotoxigenic Escherichia coli and their characterization.

    PubMed Central

    Altmann, K; Pyliotis, N A; Mukkur, T K

    1982-01-01

    It was found that K99 pili from enterotoxigenic Escherichia coli (of bovine origin) could be extracted by treatment with 3M-KSCN solution. The K99 pili were purified by preparative isoelectric focusing to apparent homogeneity as judged by the presence of a single band on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis; the molecular weight of this component was calculated to be 12 600 +/- 300. This indicated that the K99 pili were composed of a single subunit. On analytical ultracentrifugation, a single boundary with an s20,w of 12.2 S at a concentration of 0.42 mg/ml was observed. The average length of purified pili at zero concentration was approx. 160 nm and the diameter was 7.4 +/- 0.6 nm. Amino acid analysis of the purified K99 pili revealed that sulphur-containing amino acids, cysteine and methionine, were absent. Aromatic amino acids, phenylalanine and tyrosine, previously reported to be absent [Isaacson (1977) Infect. Immun. 15. 272-279], constituted 7.14% of the total amino acid residues present. On immunoelectrophoresis, purified K99 pili migrated towards the cathode and caused mannose-resistant haemagglutination of horse, but not of sheep or guinea-pig, red blood cells. Pili from enterotoxigenic E. coli of porcine and human origin and from another bacterial species, namely Fusiformis nodosus, could also be extracted by the treatment of respective micro-organisms with 3 M-KSCN. Images PLATE 1 Fig. 3. Fig. 5. PMID:6124240

  6. Structures of the activator of K. pneumonia biofilm formation, MrkH, indicates PilZ domains involved in c-di-GMP and DNA binding

    PubMed Central

    Schumacher, Maria A.; Zeng, Wenjie

    2016-01-01

    The pathogenesis of Klebsiella pneumonia is linked to the bacteria’s ability to form biofilms. Mannose-resistant Klebsiella-like (Mrk) hemagglutinins are critical for K. pneumonia biofilm development, and the expression of the genes encoding these proteins is activated by a 3′,5′-cyclic diguanylic acid (c-di-GMP)–regulated transcription factor, MrkH. To gain insight into MrkH function, we performed structural and biochemical analyses. Data revealed MrkH to be a monomer with a two-domain architecture consisting of a PilZ C-domain connected to an N domain that unexpectedly also harbors a PilZ-like fold. Comparison of apo- and c-di-GMP–bound MrkH structures reveals a large 138° interdomain rotation that is induced by binding an intercalated c-di-GMP dimer. c-di-GMP interacts with PilZ C-domain motifs 1 and 2 (RxxxR and D/NxSxxG) and a newly described c-di-GMP–binding motif in the MrkH N domain. Strikingly, these c-di-GMP–binding motifs also stabilize an open state conformation in apo MrkH via contacts from the PilZ motif 1 to residues in the C-domain motif 2 and the c-di-GMP–binding N-domain motif. Use of the same regions in apo structure stabilization and c-di-GMP interaction allows distinction between the states. Indeed, domain reorientation by c-di-GMP complexation with MrkH, which leads to a highly compacted structure, suggests a mechanism by which the protein is activated to bind DNA. To our knowledge, MrkH represents the first instance of specific DNA binding mediated by PilZ domains. The MrkH structures also pave the way for the rational design of inhibitors that target K. pneumonia biofilm formation. PMID:27551088

  7. Structures of the activator of K. pneumonia biofilm formation, MrkH, indicates PilZ domains involved in c-di-GMP and DNA binding.

    PubMed

    Schumacher, Maria A; Zeng, Wenjie

    2016-09-06

    The pathogenesis of Klebsiella pneumonia is linked to the bacteria's ability to form biofilms. Mannose-resistant Klebsiella-like (Mrk) hemagglutinins are critical for K pneumonia biofilm development, and the expression of the genes encoding these proteins is activated by a 3',5'-cyclic diguanylic acid (c-di-GMP)-regulated transcription factor, MrkH. To gain insight into MrkH function, we performed structural and biochemical analyses. Data revealed MrkH to be a monomer with a two-domain architecture consisting of a PilZ C-domain connected to an N domain that unexpectedly also harbors a PilZ-like fold. Comparison of apo- and c-di-GMP-bound MrkH structures reveals a large 138° interdomain rotation that is induced by binding an intercalated c-di-GMP dimer. c-di-GMP interacts with PilZ C-domain motifs 1 and 2 (RxxxR and D/NxSxxG) and a newly described c-di-GMP-binding motif in the MrkH N domain. Strikingly, these c-di-GMP-binding motifs also stabilize an open state conformation in apo MrkH via contacts from the PilZ motif 1 to residues in the C-domain motif 2 and the c-di-GMP-binding N-domain motif. Use of the same regions in apo structure stabilization and c-di-GMP interaction allows distinction between the states. Indeed, domain reorientation by c-di-GMP complexation with MrkH, which leads to a highly compacted structure, suggests a mechanism by which the protein is activated to bind DNA. To our knowledge, MrkH represents the first instance of specific DNA binding mediated by PilZ domains. The MrkH structures also pave the way for the rational design of inhibitors that target K pneumonia biofilm formation.

  8. Detection of pap, sfa and afa adhesin-encoding operons in uropathogenic Escherichia coli strains: relationship with expression of adhesins and production of toxins.

    PubMed

    Blanco, M; Blanco, J E; Alonso, M P; Mora, A; Balsalobre, C; Muñoa, F; Juárez, A; Blanco, J

    1997-12-01

    A total of 243 Escherichia coli strains isolated from patients with urinary tract infections (UTI) were investigated for the presence of pap, sfa and afa adhesin-encoding operons by using the polymerase chain reaction. It was found that 54%, 53% and 2% of the strains exhibited the pap, sfa and afa genotypes, respectively. Pap+ and/or sfa+ strains were more frequent in cases of acute pyelonephritis (94%) than in cases of cystitis (67%) (P < 0.001) and asymptomatic bacteriuria (57%) (P < 0.001). The pap and/or sfa operons were found in 90% of strains expressing mannose-resistant haemagglutination (MRHA) versus 37% of MRHA-negative strains (P < 0.001). The presence of pap and sfa operons was especially significant in strains belonging to MRHA types III (100%) (without P adhesins) and IVa (97%) (expressing the specific Gal-Gal binding typical of P adhesins). Both pap and sfa operons were closely associated with toxigenic E. coli producing alpha-haemolysin (Hly+) and/or the cytotoxic necrotizing factor type 1. There was an apparent correlation between the pap and sfa operons and the O serogroups of the strains. Thus, 93% of strains belonging to O1, O2, O4, O6, O7, O14, O15, O18, O22, O75 and O83 possessed pap and/or sfa operons, versus only 32% of strains belonging to other serogroups (P < 0.001). The results obtained in this study confirm the usefulness of our MRHA typing system for presumptive identification of pathogenic E. coli exhibiting different virulence factors. Thus, 85% of strains that possessed both pap and sfa adhesin-encoding operons showed MRHA types III or IVa previously associated with virulence of E. coli strains that cause UTI and bacteraemia.

  9. Virulence factors of lactose-negative Escherichia coli strains isolated from children with diarrhea in Somalia.

    PubMed Central

    Nicoletti, M; Superti, F; Conti, C; Calconi, A; Zagaglia, C

    1988-01-01

    Lactose-negative Escherichia coli strains were isolated at high frequency from children with diarrhea in Somalia during a 2-year study on diarrheal diseases. Sixty-four of these strains, considered to be a representative sample, were characterized for virulence factors, plasmid profiles, and antibiotic resistance. Of these strains, 5 were recognized as enteroinvasive E. coli (they were serotyped as O135:K-:H-), 6 belonged to classical enteropathogenic E. coli serotypes, 9 were able to adhere to tissue culture cells (of these, 4 showed a pattern of localized adherence and 1 was an enteropathogenic strain), 18 were both adherent and hemolytic, and 8 were simply hemolytic. None hybridized with 32P-labeled heat-labile or heat-stable (a and b) enterotoxin gene probes or produced moderate or high-level cytotoxic effects on HeLa cells. Of the 64 strains examined, 24 produced mannose-resistant hemagglutination with human, chicken, and monkey erythrocytes. One of these was serotyped as O4:K-:H8, and a rabbit O antiserum raised against this strain allowed us to establish that 23 strains had the same O antigen. The 23 O4 strains were hemolytic and were not enterotoxic for rabbit ileal loops, and intact bacteria were able to destroy tissue culture cell monolayers very rapidly. The uniformity of the antibiotic resistance pattern and of the plasmid DNA content, together with the fact that they were isolated in different years and in different children, suggests that the O4 strains must be epidemiologically relevant in Somalia. A possible diarrheagenic role for the adherent-hemolytic E. coli strains is also discussed. Images PMID:3281977

  10. Frequency distribution of virulence factors in uropathogenic Escherichia coli isolated from Kermanshah in 2011-2012

    PubMed Central

    Mohajeri, Parviz; Khademi, Hosna; Ebrahimi, Roya; Farahani, Abbas; Rezaei, Mansour

    2014-01-01

    Background: Uropathogenic Escherichia coli (UPEC) can cause urinary tract infection (UTI). To prevent urine flow lavage, UPEC has acquired several virulence factors called adhesins. These adhesins are expressed and controlled by different genes. Aim: This study was aimed to determine some of the most important genes that control virulence factors of UPEC (pyelonephritis associated pili [pap], S fimbrial adhesion [sfa] and A fimbrial adhesion [afa] genes), which code for adhesins and phenotypic factors. Materials and Methods: In total, 205 UPEC isolates from in- and out-patients with UTI were obtained. Polymerase chain reaction was used for gene amplification. One drop of bacterial suspension, one of red blood cells and one of peripheral blood smear were mixed for hemagglutination (HA). Formation of a clump was considered to be positive. Bacteria were grown on blood agar to determine hemolysis. Surface hydrophobicity was determined using the SAT test. Result: Frequencies of pap, afa and sfa were 42 (20.5%), 17 (8.3%) and 44 (21.5%), respectively. Frequencies of HA, hemolysis and hydrophobicity were 138 (67.3%), 56 (27.3%) and 39 (19%), respectively. Among HA-positive bacteria, 103 (74.6%) were mannose resistant. Our results highlight higher frequency of HA than that of other virulence factors, indicating a crucial role of this virulence factor in UPEC. Discussion: We concluded that major differences exist in the prevalence of virulence factors among different UPEC isolated from different countries. The association observed between pathogenicity and virulence factors may promote UPEC survival and growth within the urinary tract. Detecting these genes as the primary controllers of UPEC virulence factors may aid in better management of related infections. PMID:25143887

  11. Detection of diarrheagenic Escherichia coli strains isolated from dogs and cats in Brazil.

    PubMed

    Puño-Sarmiento, Juan; Medeiros, Leonardo; Chiconi, Carolina; Martins, Fernando; Pelayo, Jacinta; Rocha, Sérgio; Blanco, Jorge; Blanco, Miguel; Zanutto, Marcelo; Kobayashi, Renata; Nakazato, Gerson

    2013-10-25

    Escherichia coli are gut microbiota bacteria that can cause disease in some humans and other animals, including dogs and cats that humans often keep as pets. Diarrheagenic E. coli (DEC) strains are classified into six categories: enteropathogenic (EPEC), enterotoxigenic (ETEC), Shiga toxin-producing (STEC), enteroinvasive (EIEC), enteroaggregative (EAEC), and diffuse-adhering E. coli (DAEC). In this study 144 and 163 E. coli colonies were isolated from the fecal samples of 50 dogs and 50 cats, respectively, with and without diarrhea from a Veterinary Hospital (clinical isolates). The virulence factors were determined using multiplex Polymerase Chain Reaction. Adherence assays, antibacterial susceptibility and serotyping (somatic or flagellar antigens) were performed on DEC isolates. We found 25 (17.4%) and 4 (2.5%) DEC strains isolated from dogs and cats, respectively. Only the EPEC and EAEC pathotypes were found in both animals. Meanwhile, genes from other pathotypes (STEC, EIEC, and ETEC) were not found in these clinical isolates. All of the DEC strains showed mannose-resistant adherence to HEp-2 and HeLa cells, and aggregative adherence was predominant in these isolates. Multiresistant strains to antimicrobials were found in most DEC strains including usual and unusual antimicrobials in veterinary practices. The serotypes of these DEC isolates were variable. The ONT serotype was predominant in these isolates. Some serotypes found in our study were described to human DEC. Here, we demonstrate that pets carry virulent DEC genes, which are mainly strains of EPECs and EAECs. The presence of these virulence factors in isolates from animals without diarrhea suggests that pets can act as a reservoir for human infection. Copyright © 2013 Elsevier B.V. All rights reserved.

  12. Identification of Novel Components Influencing Colonization Factor Antigen I Expression in Enterotoxigenic Escherichia coli

    PubMed Central

    Haines, Sara; Gautheron, Sylviane; Nasser, William; Renauld-Mongénie, Geneviève

    2015-01-01

    Colonization factors (CFs) mediate early adhesion of Enterotoxigenic Escherichia coli (ETEC) in the small intestine. Environmental signals including bile, glucose, and contact with epithelial cells have previously been shown to modulate CF expression in a strain dependent manner. To identify novel components modulating CF surface expression, 20 components relevant to the intestinal environment were selected for evaluation. These included mucin, bicarbonate, norepinephrine, lincomycin, carbon sources, and cations. Effects of individual components on surface expression of the archetype CF, CFA/I, were screened using a fractional factorial Hadamard matrix incorporating 24 growth conditions. As most CFs agglutinate erythrocytes, surface expression was evaluated by mannose resistant hemagglutination. Seven components, including porcine gastric mucin, lincomycin, glutamine, and glucose were found to induce CFA/I surface expression in vitro in a minimal media while five others were inhibitory, including leucine and 1,10-phenanthroline. To further explore the effect of components positively influencing CFA/I surface expression, a response surface methodology (RSM) was designed incorporating 36 growth conditions. The optimum concentration for each component was identified, thereby generating a novel culture media, SP1, for CFA/I expression. CFs closely related to CFA/I, including CS4 and CS14 were similarly induced in SP1 media. Other epidemiologically relevant CFs were also induced when compared to the level obtained in minimal media. These results indicate that although CF surface expression is complex and highly variable among strains, the CF response can be predicted for closely related strains. A novel culture media inducing CFs in the CF5a group was successfully identified. In addition, mucin was found to positively influence CF expression in strains expressing either CFA/I or CS1 and CS3, and may function as a common environmental cue. PMID:26517723

  13. Identification of Novel Components Influencing Colonization Factor Antigen I Expression in Enterotoxigenic Escherichia coli.

    PubMed

    Haines, Sara; Gautheron, Sylviane; Nasser, William; Renauld-Mongénie, Geneviève

    2015-01-01

    Colonization factors (CFs) mediate early adhesion of Enterotoxigenic Escherichia coli (ETEC) in the small intestine. Environmental signals including bile, glucose, and contact with epithelial cells have previously been shown to modulate CF expression in a strain dependent manner. To identify novel components modulating CF surface expression, 20 components relevant to the intestinal environment were selected for evaluation. These included mucin, bicarbonate, norepinephrine, lincomycin, carbon sources, and cations. Effects of individual components on surface expression of the archetype CF, CFA/I, were screened using a fractional factorial Hadamard matrix incorporating 24 growth conditions. As most CFs agglutinate erythrocytes, surface expression was evaluated by mannose resistant hemagglutination. Seven components, including porcine gastric mucin, lincomycin, glutamine, and glucose were found to induce CFA/I surface expression in vitro in a minimal media while five others were inhibitory, including leucine and 1,10-phenanthroline. To further explore the effect of components positively influencing CFA/I surface expression, a response surface methodology (RSM) was designed incorporating 36 growth conditions. The optimum concentration for each component was identified, thereby generating a novel culture media, SP1, for CFA/I expression. CFs closely related to CFA/I, including CS4 and CS14 were similarly induced in SP1 media. Other epidemiologically relevant CFs were also induced when compared to the level obtained in minimal media. These results indicate that although CF surface expression is complex and highly variable among strains, the CF response can be predicted for closely related strains. A novel culture media inducing CFs in the CF5a group was successfully identified. In addition, mucin was found to positively influence CF expression in strains expressing either CFA/I or CS1 and CS3, and may function as a common environmental cue.

  14. Serratia marcescens.

    PubMed

    Hejazi, A; Falkiner, F R

    1997-11-01

    Over the last 30 years, Serratia marcescens has become an important cause of nosocomial infection. There have been many reports concerning the identification, antibiotic susceptibility, pathogenicity, epidemiological investigations and typing of this organism. Accurate identification is important in defining outbreaks. The API 20E system has been used widely, but is not individually satisfactory. The growth of S. marcescens in the environment has been investigated in relation to water, disinfectants and plastics such as blood bags. Certain extracellular products are unique to S. marcescens. Pigment (prodigiosin) biosynthesis by S. marcescens has been investigated fully since the emergence of the organism as a cause of infection. Many other aspects of the pathogenicity and virulence of S. marcescens have been studied, including adherence and hydrophobicity, lipopolysaccharide (LPS) and extracellular products. Two modes of adhesion to host epithelial surfaces have been suggested. These are mannose-resistant (MR) pili and mannose-sensitive (MS) pili. LPS, which is responsible for the biological activity of endotoxin, has been investigated fully and 24 somatic antigens have been described. The production of different enzymes by S. marcescens as virulence factors has also been reported, including chitinase, lipase, chloroperoxidase and an extracellular protein, HasA. Antibiotics used to treat serratia infection include beta-lactam agents, aminoglycosides and fluoroquinolones and a variety of different resistance mechanisms have been demonstrated. Typing methods used to study the epidemiology of S. marcescens include biotyping, bacteriocin typing, phage typing, plasmid analysis, polymerase chain reaction amplification of enterobacterial repetitive intergenic consensus sequences (ERIC-PCR) and ribotyping. Serological typing has also been used and this method seems to be a suitable first-line typing method for S. marcescens, although some strains remain untypable. RAPD

  15. Consumption of sweetened dried cranberries versus unsweetened raisins for inhibition of uropathogenic Escherichia coli adhesion in human urine: a pilot study.

    PubMed

    Greenberg, James A; Newmann, Sara J; Howell, Amy B

    2005-10-01

    The aim of this study was to determine whether consumption of sweetened dried cranberries elicits urinary anti-adherence properties against Escherichia coli as previously demonstrated with cranberry juice and/or sweetened cranberry juice cocktail, compared to unsweetened raisins. Uropathogenic E. coli isolates were obtained from five women with culture-confirmed urinary tract infections (UTIs). Four urine samples were collected from each subject. The first urine sample was collected before any study intervention. The second urine sample was collected 2-5 hours after consumption of one box (42.5 g) of raisins. The third urine sample was collected 5-7 days later. The final urine sample was collected 2-5 hours after consumption of approximately 42.5 g of dried cranberries. E. coli isolates were incubated separately in each of the four urine samples collected from the five subjects. Bacteria were harvested from the urine and tested for the ability to prevent adhesion of P-fimbriated E. coli bacteria using a mannose-resistant hemagglutination assay with human red blood cells (A1, Rh+). Of the urine samples collected after dried cranberry consumption, one demonstrated 50% antiadherence activity, two demonstrated 25% activity, and two did not show any increased activity. None of the control urine samples and none of the postraisin consumption samples demonstrated any inhibitory activity. Data from this pilot study on only five subjects suggest that consumption of a single serving of sweetened dried cranberries may elicit bacterial antiadhesion activity in human urine, whereas consumption of a single serving of raisins does not. Further studies are needed to verify the antiadhesion effect of sweetened dried cranberries. In addition, dose-response and pharmacokinetics of the active compounds in the dried cranberries need to be determined. If clinical research is positive, dried cranberries could potentially be a viable alternative to cranberry juice consumption for

  16. Dosage effect on uropathogenic Escherichia coli anti-adhesion activity in urine following consumption of cranberry powder standardized for proanthocyanidin content: a multicentric randomized double blind study

    PubMed Central

    2010-01-01

    Background Ingestion of cranberry (Vaccinium macrocarpon Ait.) has traditionally been utilized for prevention of urinary tract infections. The proanthocyanidins (PACs) in cranberry, in particular the A-type linkages have been implicated as important inhibitors of primarily P-fimbriated E. coli adhesion to uroepithelial cells. Additional experiments were required to investigate the persistence in urine samples over a broader time period, to determine the most effective dose per day and to determine if the urinary anti-adhesion effect following cranberry is detected within volunteers of different origins. Methods Two separate bioassays (a mannose-resistant hemagglutination assay and an original new human T24 epithelial cell-line assay) have assessed the ex-vivo urinary bacterial anti-adhesion activity on urines samples collected from 32 volunteers from Japan, Hungary, Spain and France in a randomized, double-blind versus placebo study. An in vivo Caenorhabditis elegans model was used to evaluate the influence of cranberry regimen on the virulence of E. coli strain. Results The results indicated a significant bacterial anti-adhesion activity in urine samples collected from volunteers that consumed cranberry powder compared to placebo (p < 0.001). This inhibition was clearly dose-dependent, prolonged (until 24 h with 72 mg of PAC) and increasing with the amount of PAC equivalents consumed in each cranberry powder regimen. An in vivo Caenorhabditis elegans model showed that cranberry acted against bacterial virulence: E. coli strain presented a reduced ability to kill worms after a growth in urines samples of patients who took cranberry capsules. This effect is particularly important with the regimen of 72 mg of PAC. Conclusions Administration of PAC-standardized cranberry powder at dosages containing 72 mg of PAC per day may offer some protection against bacterial adhesion and virulence in the urinary tract. This effect may offer a nyctohemeral protection. PMID

  17. Proteus cibarius sp. nov., a swarming bacterium from Jeotgal, a traditional Korean fermented seafood, and emended description of the genus Proteus.

    PubMed

    Hyun, Dong-Wook; Jung, Mi-Ja; Kim, Min-Soo; Shin, Na-Ri; Kim, Pil Soo; Whon, Tae Woong; Bae, Jin-Woo

    2016-06-01

    A novel Proteus-like, Gram-stain-negative, facultatively anaerobic, rod-shaped bacterium, designated strain JS9T, was isolated from Korean fermented seafood, Jeotgal. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that strain JS9T belonged to the genus Proteus in the family Enterobacteriaceae. The highest 16S rRNA gene sequence similarity of strain JS9T was to Proteus vulgaris KCTC 2579T (98.98 %) and the genomic DNA G+C content is 39.0 mol%. DNA-DNA hybridization values were measured and strain JS9T showed <20.8 % genomic relatedness with closely-related members of the genus Proteus. The isolate showed bacterial motility and swarming activity similar to those of pathogenic Proteus mirabilis but distinct from those of other species of the genus Proteus. The isolate grows optimally at 30 °C, at pH 7, and in the presence of 2 % (w/v) NaCl. The main respiratory quinones are ubiquinone Q-8 and Q-10, and the major cellular fatty acids are C16 : 0, summed feature 3 and summed feature 8. The polar lipids comprise phosphatidylglycerol, phosphatidylethanolamine, diphosphatidylglycerol, an unidentified amino lipid, two unidentified amino-phospholipids, and three unidentified lipids. Based on phylogenetic, phenotypic, chemotaxonomic and genotypic analyses, strain JS9T represents a novel species of the genus Proteus, for which the name Proteus cibarius sp. nov. is proposed. The type strain is JS9T (=KACC 18404T=JCM 30699T). An emended description of the genus Proteus is also provided.

  18. [Bacterial urovirulence factors and their association with functional and anatomical abnormalities and recurrence of urinary tract infections in children].

    PubMed

    Lagos, R; Herrera, P; Sepúlveda, A; Muñoz, A; Benavente, C; Bravo, I; Lamberg, T

    1996-10-01

    Urinary tract infections in children are associated with functional and anatomical abnormalities of the urinary tract, they tend to recur and can cause permanent kidney damage. To study in children with urinary tract infections, microbiological factors associated to recurrence, functional and anatomical abnormalities of the urinary tract. A prospective sample of children was incorporated into a follow-up protocol after their first episode of bacteriologically-demonstrated urinary tract infection. In all patients an abdominal ultrasound examination and a mictional urethrocystography were done and the presence of fimbriae was studied in isolated strains of Escherichia coli. Two hundred fifteen cases bad an adequate adherence to the study protocol, 190 caused by E coli. Fimbriated E coli strains were isolated with greater frequency from children with pyelonephritis than from those with a low urinary tract infection (50 and 28% respectively). The absence of fimbriae in E coli strains was associated with a higher risk of recurrent infections (odds ratio = 3, confidence intervals = 2-9.2) and an abnormal urethrocystography (odds ratio = 3, confidence intervals = 1.1-10.2). These data are consistent with foreign reports and support the need to study adhesins in E coli strains isolated from children with urinary tract infections.

  19. Multiplex polymerase chain reaction assay for detection of nonserotypable Shiga toxin-producing Escherichia coli strains of serogroup O147.

    PubMed

    DebRoy, Chitrita; Roberts, Elisabeth; Davis, Michael; Bumbaugh, Alyssa

    2010-11-01

    Nonserotypable Shiga toxin-producing Escherichia coli (STEC) strains (n = 72) from the collection of the E. coli Reference Center were O typed by a polymerase chain reaction (PCR)-restriction fragment length polymorphism method, and those that exhibited similar profiles (n = 17) were chosen for the study. These isolates, derived from pigs, carried genes for Shiga toxin variant 2e (100%), heat stable enterotoxins STa and STb (70% and 76%, respectively), and F107 (F18) fimbriae (82%). DNA sequencing and analysis of the O-antigen gene cluster of one of the nonserotypable strains exhibited 100% homology with O-antigen cluster of E. coli O147 although the lipopolysaccharide profiles differed significantly between the nonserotypable strains and O147 reference control strain normally used for antibody production. Scanning electron micrographs of the nonserotypable strains showed altered morphology as compared to E. coli O147. Therefore, nonserotypable strains may share 100% homology with O-antigen gene cluster of a certain serogroup but may not express that specific O-antigen. Highly specific multiplex PCR for detecting the nonserotypable STEC of serogroup O147 was developed targeting virulence genes stx2, stb, and fedA encoding for F107 fimbriae, and wzx and wzy of the O147 O-antigen cluster genes. The multiplex PCR method will allow identifying potentially pathogenic subgroup of STEC important in porcine and human health.

  20. The F18 fimbrial adhesin FedF is highly conserved among F18+Escherichia coli isolates.

    PubMed

    Tiels, P; Verdonck, F; Smet, A; Goddeeris, B; Cox, E

    2005-10-31

    F18+Escherichia coli cause postweaning diarrhoea and oedema disease in newly weaned piglets. Protection against these diseases can be established by preventing the fimbrial adhesion of these bacteria to the enterocytes of the porcine intestine. To test a vaccine against F18+E. coli consisting of the adhesin of F18 fimbriae, FedF, the conservation of the FedF subunit had to be examined. Therefore, the fedF sequence of 37 F18+E. coli isolates from different countries was determined and compared to the fedF gene of the F18ab reference strain F107/86. The amino acid sequence of the mature FedF from the individual F18+E. coli isolates was 96-100% identical to that from E. coli F107/86, but the overall homology was 90.4%. Hyper variable regions were not found in the FedF sequence. The FedF sequence was conserved over the different countries and between the two antigenic variants, F18ab and F18ac, suggesting that F18ab and F18ac strains have the same receptor. Furthermore, the conserved C-terminal region in the FedF adhesin suggests that the F18 fimbriae, in analogy with type 1 and P pili, are assembled by a donor strand mechanism. In conclusion, the reported conservation of FedF supports the usefulness of the fimbrial adhesin as a subunit vaccine against F18+E. coli infection.

  1. VEJ{phi}, a novel filamentous phage of Vibrio cholerae able to transduce the cholera toxin genes.

    PubMed

    Campos, Javier; Martínez, Eriel; Izquierdo, Yovanny; Fando, Rafael

    2010-01-01

    A novel filamentous bacteriophage, designated VEJphi, was isolated from strain MO45 of Vibrio cholerae of the O139 serogroup. A molecular characterization of the phage was carried out, which included sequencing of its whole genome, study of the genomic structure, identification of the phage receptor, and determination of the function of some of the genes, such as those encoding the major capsid protein and the single-stranded DNA-binding protein. The genome nucleotide sequence of VEJphi, which consists of 6842 bp, revealed that it is organized in modules of functionally related genes in an array that is characteristic of the genus Inovirus (filamentous phages). VEJphi is closely related to other previously described filamentous phages of V. cholerae, including VGJphi, VSK and fs1. Like these phages, VEJphi uses as a cellular receptor the type IV fimbria called the mannose-sensitive haemagglutinin (MSHA). It was also demonstrated that VEJphi, like phage VGJphi, is able to transmit the genome of phage CTXphi, and therefore the genes encoding the cholera toxin (CT), horizontally among populations of V. cholerae expressing the MSHA receptor fimbria. This suggests that the variety of phages implicated in the horizontal transmission of the CT genes could be more diverse than formerly thought.

  2. Two autonomous structural modules in the fimbrial shaft adhesin FimA mediate Actinomyces interactions with streptococci and host cells during oral biofilm development

    SciTech Connect

    Mishra, Arunima; Devarajan, Bharanidharan; Reardon, Melissa E.; Dwivedi, Prabhat; Krishnan, Vengadesan; Cisar, John O.; Das, Asis; Narayana, Sthanam V.L.; Ton-That, Hung

    2011-09-06

    By combining X-ray crystallography and modelling, we describe here the atomic structure of distinct adhesive moieties of FimA, the shaft fimbrillin of Actinomyces type 2 fimbriae, which uniquely mediates the receptor-dependent intercellular interactions between Actinomyces and oral streptococci as well as host cells during the development of oral biofilms. The FimA adhesin is built with three IgG-like domains, each of which harbours an intramolecular isopeptide bond, previously described in several Gram-positive pilins. Genetic and biochemical studies demonstrate that although these isopeptide bonds are dispensable for fimbrial assembly, cell-cell interactions and biofilm formation, they contribute significantly to the proteolytic stability of FimA. Remarkably, FimA harbours two autonomous adhesive modules, which structurally resemble the Staphylococcus aureus Cna B domain. Each isolated module can bind the plasma glycoprotein asialofetuin as well as the polysaccharide receptors present on the surface of oral streptococci and epithelial cells. Thus, FimA should serve as an excellent paradigm for the development of therapeutic strategies and elucidating the precise molecular mechanisms underlying the interactions between cellular receptors and Gram-positive fimbriae.

  3. Monoclonal antibody passive hemagglutination and capture enzyme-linked immunosorbent assays for direct detection and quantitation of F41 and K99 fimbrial antigens in enterotoxigenic Escherichia coli.

    PubMed Central

    Raybould, T J; Crouch, C F; Acres, S D

    1987-01-01

    Production of diarrhea in neonatal calves by enterotoxigenic Escherichia coli depends on its ability to attach to the epithelial cells of the intestine via surface adhesins called pili or fimbriae and to secrete enterotoxins. The most important of these fimbriae are designated K99 and F41. We produced and characterized a murine monoclonal antibody specific to F41. This monoclonal antibody and a K99-specific monoclonal antibody were used to develop sensitive and specific passive hemagglutination and capture enzyme-linked immunosorbent assays (ELISAs) for detection and quantitation of F41 and K99 antigens in E. coli cultures and culture supernatants. The capture ELISA systems exhibited excellent sensitivity and specificity, whereas the passive hemagglutination systems appeared to be oversensitive. The ability of the capture ELISAs to detect K99 and F41 fimbrial antigens in fecal specimens from calves was evaluated. Fimbrial antigens were detected in six of six specimens from scouring calves but not in four of four specimens from nonscouring calves. PMID:2880866

  4. Relationship of immunogenicity to protective potency in acellular pertussis vaccines.

    PubMed

    Xing, Dorothy; Asokanathan, Catpagavalli; Xu, Ying Hua; Bolgiano, Barbara; Douglas-Bardsley, Alex; Zhang, Shumin; Wang, Junzhi; Corbel, Michael

    2014-01-01

    Comparison of the immunogenicity response and resistance to challenge in the modified intracerebral challenge assay induced by various acellular pertussis vaccines showed that these were not closely linked. The immunogenicity assay was effective for confirming the presence of specific antigenic components and was invaluable for detecting minor components present in co-purified vaccines. However, the magnitude of antibody responses was not consistently related to antigen concentration nor did it correlate with protection in the modified intracerebral challenge assay. The immunogenicity assay detected degradation of pertussis toxin and pertactin components but not of filamentous haemagglutinin or fimbriae 2 and 3 in denatured acellular pertussis vaccines. The modified intracerebral challenge assay was effective in detecting antigen degradation in all types of acellular pertussis vaccines including those of European/North American origin but was dominated by the response to pertussis toxin. Aerosol challenge was more sensitive in detecting denaturation of filamentous haemagglutinin or fimbriae. The modified intracerebral challenge assay was the only assay that provided a quantitative indication of protective activity. Both immunogenicity and challenge assays provided useful data on acellular pertussis vaccine properties but were complementary and not alternatives.

  5. Relationship of immunogenicity to protective potency in acellular pertussis vaccines

    PubMed Central

    Xing, Dorothy; Asokanathan, Catpagavalli; Xu, Ying Hua; Bolgiano, Barbara; Douglas-Bardsley, Alex; Zhang, Shumin; Wang, Junzhi; Corbel, Michael

    2014-01-01

    Comparison of the immunogenicity response and resistance to challenge in the modified intracerebral challenge assay induced by various acellular pertussis vaccines showed that these were not closely linked. The immunogenicity assay was effective for confirming the presence of specific antigenic components and was invaluable for detecting minor components present in co-purified vaccines. However, the magnitude of antibody responses was not consistently related to antigen concentration nor did it correlate with protection in the modified intracerebral challenge assay. The immunogenicity assay detected degradation of pertussis toxin and pertactin components but not of filamentous haemagglutinin or fimbriae 2 and 3 in denatured acellular pertussis vaccines. The modified intracerebral challenge assay was effective in detecting antigen degradation in all types of acellular pertussis vaccines including those of European/North American origin but was dominated by the response to pertussis toxin. Aerosol challenge was more sensitive in detecting denaturation of filamentous haemagglutinin or fimbriae. The modified intracerebral challenge assay was the only assay that provided a quantitative indication of protective activity. Both immunogenicity and challenge assays provided useful data on acellular pertussis vaccine properties but were complementary and not alternatives. PMID:25424817

  6. Adhesion of K88ab fimbriated E. coli in piglet small intestines in relation with iron transport molecules.

    PubMed

    Grange, P; Védrine, B; Mouricout, M

    1997-01-01

    Enteropathogenic K88 fimbricated E. coli colonize the piglet small intestine. In swine, it has been previously established that some pigs lack intestinal receptors for K88 lectins and that these animals are resistant to infections by K88-positive E. coli. The receptor is inherited as a simple mendelian character. The interactions established between the glycoconjugate receptors of pig brush borders and K88 lectins are mediated by O- and N-linked glycoproteins which differ between adhesive and non-adhesive piglets. In this study the adhesion of E. coli K88+ in crossbred F2 (LW x MS) x (LW x MS) populations. By using in vitro brush border test, we observed modulation of the adhesion of K88 fimbriae and distinguished high and low affinity receptors. Furthermore, we correlated the attachment with glycoprotein pattern of epithelial cells and mucus. Epithelial cells and mucus contained several glycopeptides (from 42 to 74 kDa) recognized by K88ab fimbriae. The 74 kDa glycoprotein was characteristic of adhesive phenotype and was a mucosal transferrin (iTf). It appeared that iTf was more abundant in adhesive intestines than in non-adhesive ones, suggesting that susceptibility/resistance phenotype could be related to iron metabolism in the intestinal tract. Furthermore, we visualized the intestinal transferrin receptors on the brush border membrane of epithelial cells, probably implicated in iron absorption.

  7. Effect of sub-minimum inhibitory concentrations of ciprofloxacin, amikacin and colistin on biofilm formation and virulence factors of Escherichia coli planktonic and biofilm forms isolated from human urine

    PubMed Central

    Wojnicz, Dorota; Tichaczek-Goska, Dorota

    2013-01-01

    The aim of this study was to determine the effect of subinhibitory concentrations (sub-MICs) of ciprofloxacin, amikacin and colistin on biofilm formation, motility, curli fimbriae formation by planktonic and biofilm cells of E. coli strains isolated from the urine of patients with various urinary system infections. Quantification of biofilm formation was carried out using a microtiter plate assay and a spectrophotometric method. Bacterial enumeration was used to assess the viability of bacteria in the biofilm. Curli expression was determined by using YESCA agar supplemented with congo red. Using motility agar the ability to move was examined. All the antibiotics used at sub-MICs reduced biofilm formation in vitro, decreased the survival of bacteria, but had no effect on the motility of planktonic as well as biofilm cells. The inhibitory effect of sub-MICs of antimicrobial agents on curli fimbriae formation was dependent on the form in which the bacteria occurred, incubation time and antibiotic used. Our results clearly show that all the three antibiotics tested reduce biofilm production, interfere with curli expression but do not influence motility. This study suggests that ciprofloxacin, amikacin and colistin may be useful in the treatment of biofilm-associated infections caused by E. coli strains. PMID:24159313

  8. Impact of a mutator phenotype on motility and cell adherence in Salmonella Heidelberg.

    PubMed

    Le Bars, Hervé; Le Gall-David, Sandrine; Renoux, Virginie Madeleine; Bonnaure-Mallet, Martine; Jolivet-Gougeon, Anne; Bousarghin, Latifa

    2012-09-14

    In this study, we investigated adherence and motility of the hypermutator Salmonella enterica Heidelberg B182 bovine strain related to a 12bp deletion in mutS. This mutator phenotype was associated with increased adherence to epithelial cells and with high expression of fimA as shown by real-time RT-PCR. Motility studies showed that fliC were up-regulated in the B182 strain, while fljA and fljB were down-regulated. In order to determine if mutated mutS is implicated in this genes expression, isogenic strains, derived from a WT strain, containing the 12bp deletion in mutS (Δ12bpmutS) or an inactivated mutS (ΔmutS) were generated. Δ12bpmutS and ΔmutS strains showed a spontaneous mutation rate similar to the environmental strain B182, but exhibited lower adherence capacity and fimA expression. In contrast to the fimbriae genes, in Δ12bpmutS, fliC expression was up-regulated, but fljA and fljB expression were decreased, as in the B182 strain. Only fljB expression was increased in ΔmutS mutants. Taken together, our data suggest that mutS alteration does not influence fimbriae expression but can impact flagella genes.

  9. Effects of ciprofloxacin, norfloxacin, and ofloxacin on in vitro adhesion and survival of Pseudomonas aeruginosa AK1 on urinary catheters.

    PubMed Central

    Reid, G; Sharma, S; Advikolanu, K; Tieszer, C; Martin, R A; Bruce, A W

    1994-01-01

    Pretreatment of urinary silicone latex catheters in vitro with 0.1 and 0.5 microgram of ciprofloxacin per ml for 1, 24, and 48 h significantly reduced the adhesion and survival of the clinical isolate Pseudomonas aeruginosa AK1. UV spectroscopy and high-performance liquid chromatography confirmed the presence of ciprofloxacin adsorbed onto the catheters and showed that up to 56% of the drug leached into the surrounding fluid within 24 h. Scanning electron microscopy demonstrated that the adherent organisms were malformed and elongated after exposure to ciprofloxacin. Transmission electron microscopy showed the presence of fimbriae on the bacterial surfaces, but there was no conclusive evidence of changes in the fimbriae upon exposure to ciprofloxacin. It was found that a significant eradication of 24-h Pseudomonas biofilms could be achieved with ciprofloxacin as well as with ofloxacin and norfloxacin. Preincubation of catheters with 50- and 100-micrograms/ml concentrations of ciprofloxacin resulted in up to a 99% reduction in the number of adherent bacteria in comparison with the reduction on control catheters. In addition, adherent biofilms were eradicated by 24 h of challenge with 50 and 100 micrograms of ciprofloxacin per ml at pH 7.0 and 5.5. Results of these in vitro studies suggest that there could be a clinical role for fluoroquinolones in preventing and treating urinary tract infections associated with P. aeruginosa adherence to prosthetic devices. Images PMID:7979277

  10. Adjuvant effect of Gantrez®AN nanoparticles during oral vaccination of piglets against F4+enterotoxigenic Escherichia coli.

    PubMed

    Vandamme, Katrien; Melkebeek, Vesna; Vesna, Melkebeek; Cox, Eric; Eric, Cox; Remon, Jean Paul; Paul, Remon Jean; Vervaet, Chris; Chris, Vervaet

    2011-02-15

    In this study, the adjuvanticity of methylvinylether-co-maleic anhydride (Gantrez(®)AN) nanoparticles (NP) was investigated in an oral immunisation experiment of pigs against F4+enterotoxigenic Escherichia coli (F4+ETEC). In addition, Wheat Germ Agglutinin (WGA)-coating of the nanoparticles was tested for enterocyte-targeting. Pigs were either vaccinated with F4 fimbriae, F4 encapsulated in Gantrez(®)AN NP, F4 encapsulated in Gantrez(®)AN NP coated with WGA or F4 fimbriae mixed with empty Gantrez(®)AN NP. Only vaccination with the combination of F4 mixed with empty Gantrez(®)AN NP improved protection against F4+ETEC infection. In addition, vaccination with this formulation also resulted in an F4-specific serum antibody response prior to F4+ETEC challenge. Encapsulation of F4 in Gantrez(®)AN NP only raised the serum antibody response after F4+ETEC challenge compared to soluble F4, but did not improve protection, whereas WGA-coating almost completely abolished the serum antibody response. These data indicate that nanoparticle effects after F4 encapsulation were of lesser importance for the adjuvant effect of Gantrez(®)AN NP, contrarily to the reactivity of the Gantrez(®)AN polymer used to prepare the nanoparticles. Copyright © 2010 Elsevier B.V. All rights reserved.

  11. Essential Oils and Eugenols Inhibit Biofilm Formation and the Virulence of Escherichia coli O157:H7

    PubMed Central

    Kim, Yong-Guy; Lee, Jin-Hyung; Gwon, Giyeon; Kim, Soon-Il; Park, Jae Gyu; Lee, Jintae

    2016-01-01

    Enterohemorrhagic Escherichia coli O157:H7 (EHEC) has caused foodborne outbreaks worldwide and the bacterium forms antimicrobial-tolerant biofilms. We investigated the abilities of various plant essential oils and their components to inhibit biofilm formation by EHEC. Bay, clove, pimento berry oils and their major common constituent eugenol at 0.005% (v/v) were found to markedly inhibit EHEC biofilm formation without affecting planktonic cell growth. In addition, three other eugenol derivatives isoeugenol, 2-methoxy-4-propylphenol, and 4-ethylguaiacol had antibiofilm activity, indicating that the C-1 hydroxyl unit, the C-2 methoxy unit, and C-4 alkyl or alkane chain on the benzene ring of eugenol play important roles in antibiofilm activity. Interestingly, these essential oils and eugenol did not inhibit biofilm formation by three laboratory E. coli K-12 strains that reduced curli fimbriae production. Transcriptional analysis showed that eugenol down-regulated 17 of 28 genes analysed, including curli genes (csgABDFG), type I fimbriae genes (fimCDH) and ler-controlled toxin genes (espD, escJ, escR, and tir), which are required for biofilm formation and the attachment and effacement phenotype. In addition, biocompatible poly(lactic-co-glycolic acid) coatings containing clove oil or eugenol exhibited efficient biofilm inhibition on solid surfaces. In a Caenorhabditis elegans nematode model, clove oil and eugenol attenuated the virulence of EHEC. PMID:27808174

  12. LINGO-1 antibody ameliorates myelin impairment and spatial memory deficits in experimental autoimmune encephalomyelitis mice.

    PubMed

    Sun, Jun-Jun; Ren, Qing-Guo; Xu, Lin; Zhang, Zhi-Jun

    2015-09-18

    More than 50% of multiple sclerosis patients develop cognitive impairment. However, the underlying mechanisms are still unclear, and there is no effective treatment. LINGO-1 (LRR and Ig domain containing NOGO receptor interacting protein 1) has been identified as an inhibitor of oligodendrocyte differentiation and myelination. Using the experimental autoimmune encephalomyelitis (EAE) mouse model, we assessed cognitive function at early and late stages of EAE, determined brain expression of myelin basic protein (MBP) and investigated whether the LINGO-1 antibody could restore deficits in learning and memory and ameliorate any loss of MBP. We found that deficits in learning and memory occurred in late EAE and identified decreased expression of MBP in the parahippocampal cortex (PHC) and fimbria-fornix. Moreover, the LINGO-1 antibody significantly improved learning and memory in EAE and partially restored MBP in PHC. Furthermore, the LINGO-1 antibody activated the AKT/mTOR signaling pathway regulating myelin growth. Our results suggest that demyelination in the PHC and fimbria-fornix might contribute to cognitive deficits and the LINGO-1 antibody could ameliorate these deficits by promoting myelin growth in the PHC. Our research demonstrates that LINGO-1 antagonism may be an effective approach to the treatment of the cognitive impairment of multiple sclerosis patients.

  13. Ongoing Horizontal and Vertical Transmission of Virulence Genes and papA Alleles among Escherichia coli Blood Isolates from Patients with Diverse-Source Bacteremia

    PubMed Central

    Johnson, James R.; O'Bryan, Timothy T.; Kuskowski, Michael; Maslow, Joel N.

    2001-01-01

    The phylogenetic distributions of multiple putative virulence factors (VFs) and papA (P fimbrial structural subunit) alleles among 182 Escherichia coli blood isolates from patients with diverse-source bacteremia were defined. Phylogenetic correspondence among these strains, the E. coli Reference (ECOR) collection, and other collections of extraintestinal pathogenic E. coli (ExPEC) was assessed. Although among the 182 bacteremia isolates phylogenetic group B2 predominated, exhibited the greatest concentration of individual VFs, and contained the largest number of familiar virulent clones, other phylogenetic groups exhibited greater concentrations of certain VFs than did group B2 and included several additional virulent clones. Certain of the newly detected VF genes, e.g., fyuA (yersiniabactin; 76%) and focG (F1C fimbriae; 25%), were as prevalent or more prevalent than their more familiar traditional counterparts, e.g., iut (aerobactin; 57%) and sfaS (S fimbriae; 14%), thus possibly offering additional useful targets for preventive interventions. Considerable diversity of VF profiles was observed at every level within the phylogenetic tree, including even within individual lineages. This suggested that many different pathways can lead to extraintestinal virulence in E. coli and that the evolution of ExPEC, which involves extensive horizontal transmission of VFs and continuous remodeling of pathogenicity-associated islands, is a highly active, ongoing process. PMID:11500406

  14. Regulation of Expression of Uropathogenic Escherichia coli Nonfimbrial Adhesin TosA by PapB Homolog TosR in Conjunction with H-NS and Lrp

    PubMed Central

    Engstrom, Michael D.

    2016-01-01

    Urinary tract infections (UTIs) are a major burden to human health. The overwhelming majority of UTIs are caused by uropathogenic Escherichia coli (UPEC) strains. Unlike some pathogens, UPEC strains do not have a fixed core set of virulence and fitness factors but do have a variety of adhesins and regulatory pathways. One such UPEC adhesin is the nonfimbrial adhesin TosA, which mediates adherence to the epithelium of the upper urinary tract. The tos operon is AT rich, resides on pathogenicity island aspV, and is not expressed under laboratory conditions. Because of this, we hypothesized that tosA expression is silenced by H-NS. Lrp, based on its prominent function in the regulation of other adhesins, is also hypothesized to contribute to tos operon regulation. Using a variety of in vitro techniques, we mapped both the tos operon promoter and TosR binding sites. We have now identified TosR as a dual regulator of the tos operon, which could control the tos operon in association with H-NS and Lrp. H-NS is a negative regulator of the tos operon, and Lrp positively regulates the tos operon. Exogenous leucine also inhibits Lrp-mediated tos operon positive regulation. In addition, TosR binds to the pap operon, which encodes another important UPEC adhesin, P fimbria. Induction of TosR synthesis reduces production of P fimbria. These studies advance our knowledge of regulation of adhesin expression associated with uropathogen colonization of a host. PMID:26755158

  15. Differences in Virulence Factors among Clinical Isolates of Escherichia coli Causing Cystitis and Pyelonephritis in Women and Prostatitis in Men

    PubMed Central

    Ruiz, Joaquim; Simon, Karine; Horcajada, Juan P.; Velasco, Maria; Barranco, Margarita; Roig, Gloria; Moreno-Martínez, Antonio; Martínez, Jose A.; Jiménez de Anta, Teresa; Mensa, Josep; Vila, Jordi

    2002-01-01

    Differences in the presence of nine urovirulence factors among clinical isolates of Escherichia coli causing cystitis and pyelonephritis in women and prostatitis in men have been studied. Hemolysin and necrotizing factor type 1 occur significantly more frequently among isolates causing prostatitis than among those causing cystitis (P < 0.0001) or pyelonephritis (P < 0.005). Moreover, the papGIII gene occurred more frequently in E. coli isolates associated with prostatitis (27%) than in those associated with pyelonephritis (9%) (P < 0.05). Genes encoding aerobactin and PapC occurred significantly less frequently in isolates causing cystitis than in those causing prostatitis (P < 0.01 and P < 0.0001, respectively) and pyelonephritis (P < 0.01 and P < 0.0001, respectively). No differences in the presence of Sat or type 1 fimbriae were found. Finally, AAFII and Bfp fimbriae are no longer considered uropathogenic virulence factors since they were not found in any of the strains analyzed. Overall, the results showed that clinical isolates producing prostatitis need greater virulence than isolates producing pyelonephritis in women or, in particular, cystitis in women (P < 0.05). Overall, the results suggest that clinical isolates producing prostatitis are more virulent that those producing pyelonephritis or cystitis in women. PMID:12454134

  16. Cinnamaldehyde, Carvacrol and Organic Acids Affect Gene Expression of Selected Oxidative Stress and Inflammation Markers in IPEC-J2 Cells Exposed to Salmonella typhimurium.

    PubMed

    Burt, Sara A; Adolfse, Simone J M; Ahad, Dina S A; Tersteeg-Zijderveld, Monique H G; Jongerius-Gortemaker, Betty G M; Post, Jan A; Brüggemann, Holger; Santos, Regiane R

    2016-12-01

    Essential oils and organic acids are used as feed additives to improve health status and reduce colonization with pathogens. Although bactericidal in vitro, concentrations achieved in the animal gut are probably not lethal to pathogens. The aim of this study was to investigate the effects of cinnamaldehyde, carvacrol and cinnamic, lactic and propionic acids on the ability of Salmonella typhimurium ATCC 14028 (ST) to invade intestinal epithelial cells (IPEC-J2) and on the expression levels of immune related genes in the cells. The minimum inhibitory concentration (MIC) and non-inhibitory concentration (NIC) were determined and influence on the invasion capacity of ST was investigated. The structure of fimbriae and flagella was analysed by electron microscopy, and expression levels of HSP70, IkBa, IL-8 and IL-10 in the IPEC-J2 cells were carried out by q-PCR. Cinnamaldehyde, carvacrol and cinnamic and propionic acids inhibited ST invasion but not cell viability, bacterial viability and motility or the development of flagella. Propionic acid and cinnamaldehyde in combination with cinnamic acid caused structural impairment of fimbriae. Cinnamaldehyde up-regulated expression of HSP70 irrespective of the presence of organic acids or ST; exposure to carvacrol induced HSP70 only in the presence of propionic acid and ST. © 2016 The Authors. Phytotherapy Research published by John Wiley & Sons Ltd.

  17. Virulence factors and genetic variability of uropathogenic Escherichia coli isolated from dogs and cats in Italy.

    PubMed

    Tramuta, Clara; Nucera, Daniele; Robino, Patrizia; Salvarani, Sara; Nebbia, Patrizia

    2011-03-01

    In this study, the association between virulence genotypes and phylogenetic groups among Escherichia (E.) coli isolates obtained from pet dogs and cats with cystitis was detected, and fingerprinting methods were used to explore the relationship among strains. Forty uropathogenic E. coli (UPEC) isolated from dogs (n = 30) and cats (n = 10) in Italy were analysed by polymerase chain reaction (PCR) for the presence of virulence factors and their classification into phylogenetic groups. The same strains were characterized by repetitive extragenic palindromic (REP)- and enterobacterial repetitive intergenic consensus (ERIC)-PCR techniques. We found a high number of virulence factors such as fimbriae A, S fimbriae (sfa) and cytotoxic necrotizing factor 1 (cnf1) significantly associated with phylogenetic group B2. We demonstrated a high correlation between α-hemolysin A and pyelonephritis C, sfa, and cnf1 operons, confirming the presence of pathogenicity islands in these strains. In addition, UPEC belonging to group B2 harboured a greater number of virulence factors than strains from phylogenetic groups A, B1, and D. REP- and ERIC-PCR grouped the UPEC isolates into two major clusters, the former grouping E. coli strains belonging to phylogenetic group B2 and D, the latter grouping those belonging to groups A and B1. Given the significant genetic variability among the UPEC strains found in our study, it can be hypothesized that no specific genotype is responsible for cystitis in cats or dogs.

  18. The sialylated fraction of milk oligosaccharides is partially responsible for binding to enterotoxigenic and uropathogenic Escherichia coli human strains.

    PubMed

    Martín-Sosa, Samuel; Martín, María-Jesús; Hueso, Pablo

    2002-10-01

    Milk oligosaccharides can act as soluble receptors that block bacterial adhesion to the different epithelia. Colonization factor antigens (CFA)/I- and CFA/II-expressing enterotoxigenic Escherichia coli (ETEC) strains constitute one of the main causes of diarrhea in infants. Here, the inhibition of hemagglutination mediated by these strains by milk oligosaccharides was tested. Human milk oligosaccharides showed a strong inhibitory capacity, which decreased when the oligosaccharides were desialylated. Because milk oligosaccharides also are present in the urine of neonates receiving mothers' milk, their ability to bind two uropathogenic Escherichia coli (UPEC) strains was also examined. UPEC strains expressing P (Pap) and P-like (Prs) fimbriae are responsible for infections of the urinary tract such as pyelonephritis and cystitis. The hemagglutination mediated by these strains was inhibited by human milk oligosaccharides. The sialylated fraction was partially responsible for this inhibition in the case of the UPEC expressing the P-like fimbria because differences were found after desialylation. Although bovine milk oligosaccharides were less efficient at inhibiting the hemagglutination of ETEC strains, they were still quite good inhibitors of UPEC strains.

  19. Carcinosarcoma of the fallopian tube with disappearance of carcinoma cells by neoadjuvant chemotherapy: case study.

    PubMed

    Takemoto, Y; Ota, T; Aoki, Y; Ogura, K; Ogishima, D; Matsumoto, T

    2015-01-01

    The authors report a case of carcinosarcoma (CS) of the fimbria of the fallopian tube in which carcinoma cells disappeared with neoadjuvant chemotherapy (NAC). A 74-year-old woman visited the present hospital with a large pelvic mass and pleural effusion. A magnetic resonance image of the tumor was highly suggestive of ovarian carcinoma. Due to the presence of both serous.adenocarcinoma cells in pleural effusion and pulmonary thrombosis, the patient was given NAC consisting of carboplatin plus paclitaxel (TC) and anticoagulant therapy with warfarin potassium. With six courses of NAC, the pleural effusion and pulmonary thrombosis disappeared, and the tumor decreased 36.2% in greatest diameter. Maximum debulking surgery was then performed. The tumor was found to be located in the fimbria of the right fallopian tube. Hysterectomy and bilateral salpingo-oophorectomy were performed, and histologic examination revealed chondrosarcoma with the presence of necrotic epithelial cells. The necrotic areas were interspersed with papillary structures, and immunohistochemical study showed positivity for CK7 and negativity for CK20, p53, and estrogen receptor (ER), indicating serous adenocarcinoma. Thus, heterologous CS with disappearance of viable carcinoma cells by NAC was diagnosed. The patient was given adjuvant chemotherapy consisting of three courses of TC, and there has been no evidence of disease for 20 months. The authors' experience in this case of gynecologic CS indicates that a serous adenocarcinomatous component of tubal CS can be well cured by TC-based NAC.

  20. BRCA, the oviduct, and the space and time continuum of pelvic serous carcinogenesis.

    PubMed

    Crum, Christopher P; McKeon, Frank D; Xian, Wa

    2012-05-01

    In recent years, the distal oviduct has emerged as a critical organ in the pathogenesis of pelvic ("ovarian") serous cancer. Studies have uncovered early serous tubal intraepithelial carcinomas in approximately 8% of asymptomatic women with germline BRCA1 or BRCA2 mutations, linked serous tubal intraepithelial carcinomas to one half of serous cancers irrespective of genetic risk, and described a precursor lesion in the distal tube with early alterations in p53 function (the p53 signature). This work has established a linear serous carcinogenic sequence in a single focus within the fimbria. In addition, a more broadly distributed array of gene alterations has been discovered in the oviduct, manifested as secretory (or stem) cell outgrowths that are increased in frequency as a function of older age and serous cancer status. These "surrogate precursors" expand the existing model beyond the fimbria, implying that the molecular events leading to serous cancer are distributed over space and time. The potential promise of these discoveries is "targeted prevention" by discovering of multiple pathways integral to carcinogenesis and successfully preventing malignancy by interrupting one or a few of these pathways.

  1. Adhesion of type A Pasteurella mulocida to rabbit pharyngeal cells and its possible role in rabbit respiratory tract infections.

    PubMed Central

    Glorioso, J C; Jones, G W; Rush, H G; Pentler, L J; Darif, C A; Coward, J E

    1982-01-01

    Pasteurella multocida serotype A was found in association with the mucosal epithelium of the nasopharynges of rabbits with respiratory tract infections. The bacteria specifically attached to squamous epithelial cells of the pharyngeal mucosa both in vivo and in vitro and to some tissue culture cell lines such as HeLa. All strains with serotype A capsules were adhesive. With the exception of one serotype D strain, strains with capsular serotypes B, D, and E were at least 10-fold less adhesive. Bacterial adhesiveness was much reduced after pronase digestion, heat treatment, and homogenization, but removal of the hyaluronic acid capsule increased adhesion. Electron microscopy revealed that fimbriae were produced by an adhesive pasteurella strain, but not by two nonadherent strains. The attachment of the former strain to pharyngeal and HeLa cells was inhibited by N-acetyl-D-glucosamine. Together, these findings suggest that this amino sugar may be a component of the receptor on both animal cell surfaces and that the fimbriae may be the adhesions. It is proposed that bacterial attachment has a role in colonization and infection of rabbit upper respiratory mucosae. Images PMID:7068213

  2. BolA Is a Transcriptional Switch That Turns Off Motility and Turns On Biofilm Development

    PubMed Central

    Dressaire, Clémentine; Moreira, Ricardo Neves; Barahona, Susana; Alves de Matos, António Pedro

    2015-01-01

    ABSTRACT Bacteria are extremely versatile organisms that rapidly adapt to changing environments. When bacterial cells switch from planktonic growth to biofilm, flagellum formation is turned off and the production of fimbriae and extracellular polysaccharides is switched on. BolA is present in most Gram-negative bacteria, and homologues can be found from proteobacteria to eukaryotes. Here, we show that BolA is a new bacterial transcription factor that modulates the switch from a planktonic to a sessile lifestyle. It negatively modulates flagellar biosynthesis and swimming capacity in Escherichia coli. Furthermore, BolA overexpression favors biofilm formation, involving the production of fimbria-like adhesins and curli. Our results also demonstrate that BolA is a protein with high affinity to DNA and is able to regulate many genes on a genome-wide scale. Moreover, we show that the most significant targets of this protein involve a complex network of genes encoding proteins related to biofilm development. Herein, we propose that BolA is a motile/adhesive transcriptional switch, specifically involved in the transition between the planktonic and the attachment stage of biofilm formation. PMID:25691594

  3. Oral Immunization with Recombinant Streptococcus gordonii Expressing Porphyromonas gingivalis FimA Domains

    PubMed Central

    Sharma, Ashu; Honma, Kiyonobu; Evans, Richard T.; Hruby, Dennis E.; Genco, Robert J.

    2001-01-01

    Porphyromonas gingivalis, a gram-negative anaerobe, is implicated in the etiology of adult periodontitis. P. gingivalis fimbriae are one of several critical surface virulence factors involved in both bacterial adherence and inflammation. P. gingivalis fimbrillin (FimA), the major subunit protein of fimbriae, is considered an important antigen for vaccine development against P. gingivalis-associated periodontitis. We have previously shown that biologically active domains of P. gingivalis fimbrillin can be expressed on the surface of the human commensal bacterium Streptococcus gordonii. In this study, we examined the effects of oral coimmunization of germfree rats with two S. gordonii recombinants expressing N (residues 55 to 145)- and C (residues 226 to 337)-terminal epitopes of P. gingivalis FimA to elicit FimA-specific immune responses. The effectiveness of immunization in protecting against alveolar bone loss following P. gingivalis infection was also evaluated. The results of this study show that the oral delivery of P. gingivalis FimA epitopes via S. gordonii vectors resulted in the induction of FimA-specific serum (immunoglobulin G [IgG] and IgA) and salivary (IgA) antibody responses and that the immune responses were protective against subsequent P. gingivalis-induced alveolar bone loss. These results support the potential usefulness of the S. gordonii vectors expressing P. gingivalis fimbrillin as a mucosal vaccine against adult periodontitis. PMID:11292708

  4. Inactivation of the virulence factors from 2,3-butanediol-producing Klebsiella pneumoniae.

    PubMed

    Huynh, Duyen Thi Ngoc; Kim, Ah-Young; Seol, In-Hye; Jung, Samuel; Lim, Min-Cheol; Lee, Jeong-A; Jo, Mi-Rae; Choi, Soo-Jin; Kim, Borim; Lee, Jinwon; Kim, Wooki; Kim, Young-Rok

    2015-11-01

    The microbiological production of 2,3-butanediol (2,3-BDO) has attracted considerable attention as an alternative way to produce high-value chemicals from renewable sources. Among the number of 2,3-BDO-producing microorganisms, Klebsiella pneumoniae has been studied most extensively and is known to produce large quantity of 2,3-BDO from a range of substrates. On the other hand, the pathogenic characteristics of the bacteria have limited its industrial applications. In this study, two major virulence traits, outer core LPS and fimbriae, were removed through homologous recombination from 2,3-BDO-producing K. pneumoniae 2242 to expand its uses to the industrial scale. The K. pneumoniae 2242 ∆wabG mutant strain was found to have an impaired capsule, which significantly reduced its ability to bind to the mucous layer and evade the phagocytic activity of macrophage. The association with the human ileocecal epithelial cell, HCT-8, and the bladder epithelial cell, T-24, was also reduced dramatically in the K. pneumoniae 2242 ∆fimA mutant strain that was devoid of fimbriae. However, the growth rate and production yield for 2,3-BDO were unaffected. The K. pneumoniae strains developed in this study, which are devoid of the major virulence factors, have a high potential for the efficient and sustainable production of 2,3-BDO.

  5. Detection of pap, sfa, afa, foc, and fim Adhesin-Encoding Operons in Uropathogenic Escherichia coli Isolates Collected From Patients With Urinary Tract Infection

    PubMed Central

    Rahdar, Masoud; Rashki, Ahmad; Miri, Hamid Reza; Rashki Ghalehnoo, Mehdi

    2015-01-01

    Background: Uropathogenic Escherichia coli (UPEC) with its virulence factors is the most prevalent cause of urinary tract infection (UTI). Objectives; This study aimed to determine the occurrence of fim, pap, sfa, and afa genes among 100 UPEC isolates collected from patients diagnosed with UTI. Materials and Methods A total of 100 UPEC isolates were obtained from urine samples of patients with UTI. The prevalence of 5 virulence genes encoding type 1 fimbriae (fimH), pili associated with pyelonephritis (pap), S and F1C fimbriae (sfa and foc) and afimbrial adhesins (afa) were determined through PCR method. We also investigated the phylogenetic background of all isolates. In addition, the distribution of adhesin-encoding operons between the phylogroups was assessed. Results: The prevalence of genes encoding for fimbrial adhesive systems was 95% for fim, 57% for pap, 16% for foc, and 81% for sfa. The operons encoding for afa afimbrial adhesins were identified in 12% of isolates. The various combinations of detected genes were designated as virulence patterns. The fim gene, which occurred in strains from all phylogenetic groups (A, B1, B2, and D) was evaluated and no significant differences were found among these groups. Conversely, significant differences were observed in relation to pap, afa, foc, and sfa operons. Conclusions: These results indicate that the PCR method is a powerful genotypic assay for the detection of adhesin-encoding operons. Thus, this assay can be recommended for clinical use to detect virulent urinary E. coli strains, as well as epidemiological studies. PMID:26464770

  6. Characterization of urinary Escherichia coli O75 strains.

    PubMed Central

    Nimmich, W; Voigt, W; Seltmann, G

    1997-01-01

    Forty-four Escherichia coli O75 strains from patients with urinary tract infections were characterized by a variety of methods to obtain evidence of their clonal distribution and uropathogenic properties. By K and H antigen typing, the strains were divided into the following serotypes: O75:K5:H- (18 strains), O75:K95:H- (10 strains), O75:K95:H5 (7 strains), O75:K100:H5 (4 strains), and O75:K-:H55 (5 strains). Generally, biotyping proved to be of no discriminative value. With two exceptions the strains were found to be sensitive to the bactericidal effect of normal human serum. As shown by multilocus enzyme electrophoresis, the whole-cell protein profile (WCPP), and the patterns of the outer membrane proteins and lipopolysaccharides, all but the five O75:H55 strains were genetically closely related to each other and could be classified into one clonal group. The O75:K-:H55 strains proved to be quite different and lacked type 1 fimbriae. All 17 K95 (H-, H5) strains produced hemolysin and P fimbriae. Five of the O75:K5:H- strains were different from the other K5 strains by showing hemagglutinating properties, on the basis of the presence of the OX adhesin. The last two groups are suggested to be uropathogenic and are proposed to represent separate clonal groups or subgroups. PMID:9114391

  7. Shear-enhanced binding of intestinal colonization factor antigen I of enterotoxigenic Escherichia coli

    PubMed Central

    Tchesnokova, Veronika; McVeigh, Annette L.; Kidd, Brian; Yakovenko, Olga; Thomas, Wendy E.; Sokurenko, Evgeni V.; Savarino, Stephen J.

    2010-01-01

    SUMMARY In the intestine, enterotoxigenic Escherichia coli works against peristaltic forces, adhering to the epithelium via the CFA/I fimbrial adhesin CfaE. The CfaE adhesin is similar in localization and tertiary (but not primary) structure to FimH, the type 1 fimbrial adhesin of uropathogenic Escherichia coli, which shows shear-dependent binding to epithelial receptors by an allosteric catch-bond mechanism. Thus, we speculated that CfaE is also capable of shear-enhanced binding. Indeed, bovine erythrocytes coursing over immobilized CFA/I fimbriae in flow-chambers exhibited low accumulation levels and fast rolling at low shear, but an 80-fold increase in accumulation and 3-fold decrease in rolling velocity at elevated shear. This effect was reversible and abolished by pre-incubation of fimbriae with anti-CfaE antibody. Erythrocytes bound to whole CfaE in the same shear-enhanced manner, but to CfaE adhesin domain in a shear-inhibitable fashion. Residue replacements designed to disrupt CfaE interdomain interaction decreased the shear-dependency of adhesion and increased binding under static conditions to human intestinal epithelial cells. These findings indicate that close interaction between adhesive and anchoring pilin domains of CfaE keeps the former in a low-affinity state that toggles into a high-affinity state upon separation of two domains, all consistent with an allosteric catch-bond mechanism of CfaE binding. PMID:20345656

  8. Turn Up the Heat—Food and Clinical Escherichia coli Isolates Feature Two Transferrable Loci of Heat Resistance

    PubMed Central

    Boll, Erik J.; Marti, Roger; Hasman, Henrik; Overballe-Petersen, Søren; Stegger, Marc; Ng, Kim; Knøchel, Susanne; Krogfelt, Karen A.; Hummerjohann, Joerg; Struve, Carsten

    2017-01-01

    Heat treatment is a widely used process to reduce bacterial loads in the food industry or to decontaminate surfaces, e.g., in hospital settings. However, there are situations where lower temperatures must be employed, for instance in case of food production such as raw milk cheese or for decontamination of medical devices such as thermo-labile flexible endoscopes. A recently identified locus of heat resistance (LHR) has been shown to be present in and confer heat resistance to a variety of Enterobacteriaceae, including Escherichia coli isolates from food production settings and clinical ESBL-producing E. coli isolates. Here, we describe the presence of two distinct LHR variants within a particularly heat resistant E. coli raw milk cheese isolate. We demonstrate for the first time in this species the presence of one of these LHRs on a plasmid, designated pFAM21805, also encoding type 3 fimbriae and three bacteriocins and corresponding self-immunity proteins. The plasmid was highly transferable to other E. coli strains, including Shiga-toxin-producing strains, and conferred LHR-dependent heat resistance as well as type 3 fimbriae-dependent biofilm formation capabilities. Selection for and acquisition of this “survival” plasmid by pathogenic organisms, e.g., in food production environments, may pose great concern and emphasizes the need to screen for the presence of LHR genes in isolates. PMID:28439262

  9. Oral vaccination with attenuated Salmonella enterica strains encoding T-cell epitopes from tumor antigen NY-ESO-1 induces specific cytotoxic T-lymphocyte responses.

    PubMed

    Meng, Jia-Zi; Dong, Yu-Jun; Huang, He; Li, Shuang; Zhong, Yi; Liu, Shu-Lin; Wang, Yue-Dan

    2010-06-01

    Bacterial fimbriae can accept foreign peptides and display them on the cell surface. A highly efficient gene replacement method was used to generate peptide vaccines based on Salmonella enterica serovar Typhimurium SL3261. The T-cell epitopes (NY-ESO-1 p157-165 and p157-167) from NY-ESO-1, which is a promising target antigen in patients for the specific immune recognition of cancer, were incorporated into the gene encoding AgfA (the major subunit protein of thin aggregative fimbriae of Salmonella) by replacing an equal length of the DNA segment. To improve cytotoxic T-lymphocyte recognition, both termini of the peptide were flanked by double alanine (AA) residues. Immunofluorescence microscopy with AgfA-specific antiserum verified the expression of chimeric AgfA, which was also proved by a Congo red binding assay. Oral immunizations of HLA-A*0201 transgenic mice with recombinant SL3261 strains encoding NY-ESO-1 p157-165 or p157-167 induced NY-ESO-1 p157-165-specific CD8(+) T cells, detected by an HLA-A*0201 pentamer, and induced a T-cell response detected by an enzyme-linked immunospot assay. The Salmonella fimbrial display system was efficient at the induction of an antitumor cellular immune response in vivo, providing a new strategy for the development of efficient cancer vaccinations.

  10. Salmonella expressing a T-cell epitope from Sendai virus are able to induce anti-infection immunity.

    PubMed

    Huang, He; Wang, Ye-Jun; White, Aaron P; Meng, Jia-Zi; Liu, Gui-Rong; Liu, Shu-Lin; Wang, Yue-Dan

    2009-09-01

    Bacterial fimbriae can accept foreign peptides and display them on the cell surface. A highly efficient gene replacement method was used to generate peptide vaccines based on Salmonella enterica subsp. enterica serovar Typhimurium LT2. DNA encoding an epitope from Sendai virus, SV9 (Sendai virus nucleoprotein peptide 324-332, FAPGNYPAL), which is known to induce cytotoxic T lymphocytes, was incorporated into the gene encoding AgfA (the major subunit protein of thin aggregative fimbriae of Salmonella) by replacing an equal length DNA segment. To improve cytotoxic T lymphocyte recognition, both termini of the peptide were flanked by double alanine (AA) or arginine (RR) residues. Western blotting and immunofluorescence microscopy using AgfA-specific antiserum verified the expression of chimeric AgfA; expression was also proved by a Congo red binding assay. Oral immunizations of C57BL/6 mice with the four strains induced an epitope-specific T-cell response (detected by enzyme-linked immunosorbent spot assay). When the mice were challenged with the Sendai virus, the magnitude of the infection was significantly reduced in the immunized groups compared with the controls. The Salmonella fimbrial display system efficiently induces a cellular immune response and anti-infection immunity in vivo, providing a new strategy for the development of efficient peptide vaccination.

  11. Differentiation of Crohn’s Disease-Associated Isolates from Other Pathogenic Escherichia coli by Fimbrial Adhesion under Shear Force

    PubMed Central

    Szunerits, Sabine; Zagorodko, Oleksandr; Cogez, Virginie; Dumych, Tetiana; Chalopin, Thibaut; Alvarez Dorta, Dimitri; Sivignon, Adeline; Barnich, Nicolas; Harduin-Lepers, Anne; Larroulet, Iban; Yanguas Serrano, Aritz; Siriwardena, Aloysius; Pesquera, Amaia; Zurutuza, Amaia; Gouin, Sébastien G.; Boukherroub, Rabah; Bouckaert, Julie

    2016-01-01

    Shear force exerted on uropathogenic Escherichia coli adhering to surfaces makes type-1 fimbriae stretch out like springs to catch on to mannosidic receptors. This mechanism is initiated by a disruption of the quaternary interactions between the lectin and the pilin of the two-domain FimH adhesin and transduces allosterically to the mannose-binding pocket of FimH to increase its affinity. Mannose-specific adhesion of 14 E. coli pathovars was measured under flow, using surface plasmon resonance detection on functionalized graphene-coated gold interfaces. Increasing the shear had important differential consequences on bacterial adhesion. Adherent-invasive E. coli, isolated from the feces and biopsies of Crohn’s disease patients, consistently changed their adhesion behavior less under shear and displayed lower SPR signals, compared to E. coli opportunistically infecting the urinary tract, intestines or loci of knee and hip prostheses. We exemplified this further with the extreme behaviors of the reference strains UTI89 and LF82. Whereas their FimA major pilins have identical sequences, FimH of LF82 E. coli is marked by the Thr158Pro mutation. Positioned in the inter-domain region known to carry hot spots of mutations in E. coli pathotypes, residue 158 is indicated to play a structural role in the allosteric regulation of type-1 fimbriae-mediated bacterial adhesion. PMID:27043645

  12. A novel competence gene, comP, is essential for natural transformation of Acinetobacter sp. strain BD413.

    PubMed Central

    Porstendörfer, D; Drotschmann, U; Averhoff, B

    1997-01-01

    Acinetobacter sp. strain BD413 (= ATCC 33305), a nutritionally versatile bacterium, has an extremely efficient natural transformation system. Here we describe the generation of eight transformation-affected mutants of Acinetobacter sp. strain BD413 by insertional mutagenesis. These mutants were found by Southern blot analysis and complementation studies to result from single nptII marker insertions at different chromosomal loci. DNA binding and uptake studies with one mutant, T205, revealed that the transformation deficiency of this mutant results from a complete lack of DNA binding and, therefore, uptake activity. A novel competence gene essential for natural transformation, named comP, was cloned by complementation of mutant T205. The nucleotide sequence of comP was determined, and its deduced 15-kDa polypeptide displays significant similarities to type IV pilins. Analysis of the ultrastructure of a transformation-deficient comP mutant and the transformation-competent wild-type strain revealed that both are covered with bundle-forming thin fimbriae (3 to 4 nm in diameter) and individual thick fimbriae (6 nm in diameter). These results provide evidence that the pilinlike ComP is unrelated to the piluslike structures of strain BD413. Taking all data into account, we propose that ComP functions as a major subunit of an organelle acting as a channel or pore mediating DNA binding and/or uptake in Acinetobacter sp. strain BD413. PMID:9361398

  13. Denervated hippocampus provides a favorable microenvironment for neuronal differentiation of endogenous neural stem cells

    PubMed Central

    Zhang, Lei; Han, Xiao; Cheng, Xiang; Tan, Xue-feng; Zhao, He-yan; Zhang, Xin-hua

    2016-01-01

    Fimbria-fornix transection induces both exogenous and endogenous neural stem cells to differentiate into neurons in the hippocampus. This indicates that the denervated hippocampus provides an environment for neuronal differentiation of neural stem cells. However, the pathways and mechanisms in this process are still unclear. Seven days after fimbria fornix transection, our reverse transcription polymerase chain reaction, western blot assay, and enzyme linked immunosorbent assay results show a significant increase in ciliary neurotrophic factor mRNA and protein expression in the denervated hippocampus. Moreover, neural stem cells derived from hippocampi of fetal (embryonic day 17) Sprague-Dawley rats were treated with ciliary neurotrophic factor for 7 days, with an increased number of microtubule associated protein-2-positive cells and decreased number of glial fibrillary acidic protein-positive cells detected. Our results show that ciliary neurotrophic factor expression is up-regulated in the denervated hippocampus, which may promote neuronal differentiation of neural stem cells in the denervated hippocampus. PMID:27212920

  14. Lgr5 marks stem/progenitor cells in ovary and tubal epithelia.

    PubMed

    Ng, Annie; Tan, Shawna; Singh, Gurmit; Rizk, Pamela; Swathi, Yada; Tan, Tuan Zea; Huang, Ruby Yun-Ju; Leushacke, Marc; Barker, Nick

    2014-08-01

    The ovary surface epithelium (OSE) undergoes ovulatory tear and remodelling throughout life. Resident stem cells drive such tissue homeostasis in many adult epithelia, but their existence in the ovary has not been definitively proven. Lgr5 marks stem cells in multiple epithelia. Here we use reporter mice and single-molecule fluorescent in situ hybridization to document candidate Lgr5(+) stem cells in the mouse ovary and associated structures. Lgr5 is broadly expressed during ovary organogenesis, but becomes limited to the OSE in neonate life. In adults, Lgr5 expression is predominantly restricted to proliferative regions of the OSE and mesovarian-fimbria junctional epithelia. Using in vivo lineage tracing, we identify embryonic and neonate Lgr5(+) populations as stem/progenitor cells contributing to the development of the OSE cell lineage, as well as epithelia of the mesovarian ligament and oviduct/fimbria. Adult Lgr5(+) populations maintain OSE homeostasis and ovulatory regenerative repair in vivo. Thus, Lgr5 marks stem/progenitor cells of the ovary and tubal epithelia.

  15. Nanoscale surface modification favors benign biofilm formation and impedes adherence by pathogens.

    PubMed

    Trautner, Barbara W; Lopez, Analette I; Kumar, Amit; Siddiq, Danish M; Liao, Kershena S; Li, Yan; Tweardy, David J; Cai, Chengzhi

    2012-04-01

    We have found in vitro that a biofilm of benign Escherichia coli 83972 interferes with urinary catheter colonization by pathogens, and in human studies E. coli 83972-coated urinary catheters are associated with lower rates of catheter-associated urinary tract infections. We hypothesized that modifying surfaces to present mannose ligands for the type 1 fimbriae of E. coli would promote formation of dense E. coli 83972 biofilms, thereby interfering with surface colonization by Enterococcus faecalis, a common uropathogen. We covalently immobilized mannose on silicon substrates by attaching amino-terminated mannose derivative to carboxylic acid-terminated monolayers via amidation. Fluorescence microscopy showed that E. coli 83972 adherence to mannose-modified surfaces increased 4.4-fold compared to unmodified silicon surfaces. Pre-exposing mannose-modified surfaces to E. coli 83972 established a protective biofilm that reduced E. faecalis adherence by 83-fold. Mannose-fimbrial interactions were essential for the improved E. coli 83927 adherence and interference effects. From the Clinical Editor: Recurrent urinary tract infections remain major adverse events associated with catheter use. The authors report that modifying catheter surface to present mannose ligands for the type 1 fimbriae of benign Escherichia coli 83972 promotes formation of dense E. coli biofilms, which 100-fold reduces urinary catheter colonization of uropathogens. Future application of this technology is expected to result in substantial UTI risk reduction in catheter users. Published by Elsevier Inc.

  16. Does the use of a uterine manipulator with an intrauterine balloon in total laparoscopic hysterectomy facilitate tumor cell spillage into the peritoneal cavity in patients with endometrial cancer?

    PubMed

    Lim, S; Kim, H S; Lee, K B; Yoo, C W; Park, S Y; Seo, S S

    2008-01-01

    The objective of this study was to determine if total laparoscopic hysterectomy using a uterine manipulator with an intrauterine balloon increases the risk of positive peritoneal washings in patients with endometrial cancer. Three sets of peritoneal washings were obtained during surgery from 46 women with endometrial cancer at the Center for Uterine Cancer, National Cancer Center, Korea, between May 2004 and July 2006: the first before the insertion of the uterine manipulator (premanipulator), the second after clipping the fallopian tubes and inserting the uterine manipulator (postmanipulator), and the third after the removal of the uterus through the vagina (posthysterectomy). The cytology samples were examined by the same cytopathologist for the presence of malignant cells. Two of 46 (4.3%) patients were upstaged to IIIA disease due to positive cytology conversion after the insertion of the uterine manipulator, one after the insertion of the uterine manipulator, and the other after the hysterectomy. However, during the follow-up for 3-28 months (median 18), neither of the 2 patients experienced a tumor recurrence. In conclusion, using a uterine manipulator with an intrauterine balloon during the laparoscopic surgery for endometrial cancer might be associated with positive cytologic conversion. Possible explanations are retrograde seeding of tumor cells into the peritoneal cavity, the pressure effect of the inflatable manipulator tip, and spillage of preexited tumor cells between the isthmus and the fimbriae. More effective preventive methods such as distal tubal clipping or coagulation of the fimbriae may be necessary in treating women with endometrial cancer.

  17. Native flagellin does not protect mice against an experimental Proteus mirabilis ascending urinary tract infection and neutralizes the protective effect of MrpA fimbrial protein.

    PubMed

    Scavone, Paola; Umpiérrez, Ana; Rial, Analía; Chabalgoity, José A; Zunino, Pablo

    2014-06-01

    Proteus mirabilis expresses several virulence factors including MR/P fimbriae and flagella. Bacterial flagellin has frequently shown interesting adjuvant and protective properties in vaccine formulations. However, native P. mirabilis flagellin has not been analyzed so far. Native P. mirabilis flagellin was evaluated as a protective antigen and as an adjuvant in co-immunizations with MrpA (structural subunit of MR/P fimbriae) using an ascending UTI model in the mouse. Four groups of mice were intranasally treated with either MrpA, native flagellin, both proteins and PBS. Urine and blood samples were collected before and after immunization for specific antibodies determination. Cytokine production was assessed in immunized mice splenocytes cultures. Mice were challenged with P. mirabilis, and bacteria quantified in kidneys and bladders. MrpA immunization induced serum and urine specific anti-MrpA antibodies while MrpA coadministered with native flagellin did not. None of the animals developed significant anti-flagellin antibodies. Only MrpA-immunized mice showed a significant decrease of P. mirabilis in bladders and kidneys. Instead, infection levels in MrpA-flagellin or flagellin-treated mice showed no significant differences with the control group. IL-10 was significantly induced in splenocytes of mice that received native flagellin or MrpA-flagellin. Native P. mirabilis flagellin did not protect mice against an ascending UTI. Moreover, it showed an immunomodulatory effect, neutralizing the protective role of MrpA. P. mirabilis flagellin exhibits particular immunological properties compared to other bacterial flagellins.

  18. Two-component regulatory systems in Pseudomonas aeruginosa: an intricate network mediating fimbrial and efflux pump gene expression

    PubMed Central

    Sivaneson, Melissa; Mikkelsen, Helga; Ventre, Isabelle; Bordi, Christophe; Filloux, Alain

    2011-01-01

    Pseudomonas aeruginosa is responsible for chronic and acute infections in humans. Chronic infections are associated with production of fimbriae and the formation of a biofilm. The two-component system Roc1 is named after its role in the regulation of cup genes, which encode components of a machinery allowing assembly of fimbriae. A non-characterized gene cluster, roc2, encodes components homologous to the Roc1 system. We show that cross-regulation occurs between the Roc1 and Roc2 signalling pathways. We demonstrate that the sensors RocS2 and RocS1 converge on the response regulator RocA1 to control cupC gene expression. This control is independent of the response regulator RocA2. Instead, we show that these sensors act via the RocA2 response regulator to repress the mexAB-oprM genes. These genes encode a multidrug efflux pump and are upregulated in the rocA2 mutant, which is less susceptible to antibiotics. It has been reported that in cystic fibrosis lungs, in which P. aeruginosa adopts the biofilm lifestyle, most isolates have an inactive MexAB-OprM pump. The concomitant RocS2-dependent upregulation of cupC genes (biofilm formation) and downregulation of mexAB-oprM genes (antibiotic resistance) is in agreement with this observation. It suggests that the Roc systems may sense the environment in the cystic fibrosis lung. PMID:21205015

  19. Essential Oils and Eugenols Inhibit Biofilm Formation and the Virulence of Escherichia coli O157:H7.

    PubMed

    Kim, Yong-Guy; Lee, Jin-Hyung; Gwon, Giyeon; Kim, Soon-Il; Park, Jae Gyu; Lee, Jintae

    2016-11-03

    Enterohemorrhagic Escherichia coli O157:H7 (EHEC) has caused foodborne outbreaks worldwide and the bacterium forms antimicrobial-tolerant biofilms. We investigated the abilities of various plant essential oils and their components to inhibit biofilm formation by EHEC. Bay, clove, pimento berry oils and their major common constituent eugenol at 0.005% (v/v) were found to markedly inhibit EHEC biofilm formation without affecting planktonic cell growth. In addition, three other eugenol derivatives isoeugenol, 2-methoxy-4-propylphenol, and 4-ethylguaiacol had antibiofilm activity, indicating that the C-1 hydroxyl unit, the C-2 methoxy unit, and C-4 alkyl or alkane chain on the benzene ring of eugenol play important roles in antibiofilm activity. Interestingly, these essential oils and eugenol did not inhibit biofilm formation by three laboratory E. coli K-12 strains that reduced curli fimbriae production. Transcriptional analysis showed that eugenol down-regulated 17 of 28 genes analysed, including curli genes (csgABDFG), type I fimbriae genes (fimCDH) and ler-controlled toxin genes (espD, escJ, escR, and tir), which are required for biofilm formation and the attachment and effacement phenotype. In addition, biocompatible poly(lactic-co-glycolic acid) coatings containing clove oil or eugenol exhibited efficient biofilm inhibition on solid surfaces. In a Caenorhabditis elegans nematode model, clove oil and eugenol attenuated the virulence of EHEC.

  20. Intrinsic apoptotic pathways of gingival epithelial cells modulated by Porphyromonas gingivalis.

    PubMed

    Mao, Song; Park, Yoonsuk; Hasegawa, Yoshiaki; Tribble, Gena D; James, Chlöe E; Handfield, Martin; Stavropoulos, M Franci; Yilmaz, Ozlem; Lamont, Richard J

    2007-08-01

    Porphyromonas gingivalis can inhibit chemically induced apoptosis in primary cultures of gingival epithelial cells through blocking activation of the effector caspase-3. The anti-apoptotic phenotype of P. gingivalis is conserved across strains and does not depend on the presence of fimbriae, as fimbriae-deficient mutants and a naturally occurring non-fimbriated strain were able to impede apoptosis. To dissect the survival pathways modulated by P. gingivalis, protein and gene expression of a number of components of apoptotic death pathways were investigated. P. gingivalis infection of epithelial cells resulted in the phosphorylation of JAK1 and Stat3. Quantitative real-time reverse transcription polymerase chain reaction showed that expression of Survivin and Stat3 itself, targets of activated Stat3, were elevated in P. gingivalis-infected cells. siRNA knockdown of JAK1, in combination with knockdown of Akt, abrogated the ability of P. gingivalis to block apoptosis. In contrast, cIAP-1 and cIAP-2 were not differentially regulated at either the protein or mRNA levels by P. gingivalis. One mechanism by which P. gingivalis can block apoptotic pathways in gingival epithelial cells therefore is through manipulation of the JAK/Stat pathway that controls the intrinsic mitochondrial cell death pathways. Induction of a pro-survival phenotype may prevent programmed host cell death and aid survival of P. gingivalis within gingival epithelial cells.

  1. Intrinsic apoptotic pathways of gingival epithelial cells modulated by Porphyromonas gingivalis

    PubMed Central

    Mao, Song; Park, Yoonsuk; Hasegawa, Yoshiaki; Tribble, Gena D.; James, Chlöe E.; Handfield, Martin; Stavropoulos, M. Franci; Yilmaz, özlem; Lamont, Richard J.

    2010-01-01

    Summary Porphyromonas gingivalis can inhibit chemically induced apoptosis in primary cultures of gingival epithelial cells through blocking activation of the effector caspase-3. The anti-apoptotic phenotype of P. gingivalis is conserved across strains and does not depend on the presence of fimbriae, as fimbriae-deficient mutants and a naturally occurring non-fimbriated strain were able to impede apoptosis. To dissect the survival pathways modulated by P. gingivalis, protein and gene expression of a number of components of apoptotic death pathways were investigated. P. gingivalis infection of epithelial cells resulted in the phosphorylation of JAK1 and Stat3. Quantitative real-time reverse transcription polymerase chain reaction showed that expression of Survivin and Stat3 itself, targets of activated Stat3, were elevated in P. gingivalis-infected cells. siRNA knockdown of JAK1, in combination with knockdown of Akt, abrogated the ability of P. gingivalis to block apoptosis. In contrast, cIAP-1 and cIAP-2 were not differentially regulated at either the protein or mRNA levels by P. gingivalis. One mechanism by which P. gingivalis can block apoptotic pathways in gingival epithelial cells therefore is through manipulation of the JAK/Stat pathway that controls the intrinsic mitochondrial cell death pathways. Induction of a pro-survival phenotype may prevent programmed host cell death and aid survival of P. gingivalis within gingival epithelial cells. PMID:17419719

  2. ANIMAL ENTEROTOXIGENIC ESCHERICHIA COLI

    PubMed Central

    Dubreuil, J. Daniel; Isaacson, Richard E.; Schifferli, Dieter M.

    2016-01-01

    Enterotoxigenic Escherichia coli (ETEC) is the most common cause of E. coli diarrhea in farm animals. ETEC are characterized by the ability to produce two types of virulence factors; adhesins that promote binding to specific enterocyte receptors for intestinal colonization and enterotoxins responsible for fluid secretion. The best-characterized adhesins are expressed in the context of fimbriae, such as the F4 (also designated K88), F5 (K99), F6 (987P), F17 and F18 fimbriae. Once established in the animal small intestine, ETEC produces enterotoxin(s) that lead to diarrhea. The enterotoxins belong to two major classes; heat-labile toxin that consist of one active and five binding subunits (LT), and heat-stable toxins that are small polypeptides (STa, STb, and EAST1). This chapter describes the disease and pathogenesis of animal ETEC, the corresponding virulence genes and protein products of these bacteria, their regulation and targets in animal hosts, as well as mechanisms of action. Furthermore, vaccines, inhibitors, probiotics and the identification of potential new targets identified by genomics are presented in the context of animal ETEC. PMID:27735786

  3. A systematic review and meta-analysis of the epidemiology of pathogenic Escherichia coli of calves and the role of calves as reservoirs for human pathogenic E. coli.

    PubMed

    Kolenda, Rafał; Burdukiewicz, Michał; Schierack, Peter

    2015-01-01

    Escherichia coli bacteria are the most common causes of diarrhea and septicemia in calves. Moreover, calves form a major reservoir for transmission of pathogenic E. coli to humans. Systematic reviews and meta-analyses of publications on E. coli as calf pathogens and the role of calves as reservoir have not been done so far. We reviewed studies between 1951 and 2013 reporting the presence of virulence associated factors (VAFs) in calf E. coli and extracted the following information: year(s) and country of sampling, animal number, health status, isolate number, VAF prevalence, serotypes, diagnostic methods, and biological assays. The prevalence of VAFs or E. coli pathotypes was compared between healthy and diarrheic animals and was analyzed for time courses. Together, 106 papers with 25,982 E. coli isolates from 27 countries tested for VAFs were included. F5, F17, and F41 fimbriae and heat-stable enterotoxin (ST) - VAFs of enterotoxigenic E. coli (ETEC) were significantly associated with calf diarrhea. On the contrary, ETEC VAF F4 fimbriae and heat-labile enterotoxin as well as enteropathogenic (EPEC), Shiga toxin-producing (STEC), and enterohemorrhagic E. coli (EHEC) were not associated with diarrhea. The prevalence increased overtime for ST-positive isolates, but decreased for F5- and STEC-positive isolates. Our study provides useful information about the history of scientific investigations performed in this domain so far, and helps to define etiological agents of calf disease, and to evaluate calves as reservoir hosts for human pathogenic E. coli.

  4. Cell surface attachment structures contribute to biofilm formation and xylem colonization by Erwinia amylovora.

    PubMed

    Koczan, Jessica M; Lenneman, Bryan R; McGrath, Molly J; Sundin, George W

    2011-10-01

    Biofilm formation plays a critical role in the pathogenesis of Erwinia amylovora and the systemic invasion of plant hosts. The functional role of the exopolysaccharides amylovoran and levan in pathogenesis and biofilm formation has been evaluated. However, the role of biofilm formation, independent of exopolysaccharide production, in pathogenesis and movement within plants has not been studied previously. Evaluation of the role of attachment in E. amylovora biofilm formation and virulence was examined through the analysis of deletion mutants lacking genes encoding structures postulated to function in attachment to surfaces or in cellular aggregation. The genes and gene clusters studied were selected based on in silico analyses. Microscopic analyses and quantitative assays demonstrated that attachment structures such as fimbriae and pili are involved in the attachment of E. amylovora to surfaces and are necessary for the production of mature biofilms. A time course assay indicated that type I fimbriae function earlier in attachment, while type IV pilus structures appear to function later in attachment. Our results indicate that multiple attachment structures are needed for mature biofilm formation and full virulence and that biofilm formation facilitates entry and is necessary for the buildup of large populations of E. amylovora cells in xylem tissue.

  5. Multistep level sections to detect occult fallopian tube carcinoma in risk-reducing salpingo-oophorectomies from women with BRCA mutations: implications for defining an optimal specimen dissection protocol.