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Sample records for mannose-resistant proteus-like fimbriae

  1. 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

  2. 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

  3. 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.

  4. 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

  5. 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.

  6. 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

  7. 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.

  8. 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

  9. 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

  10. 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.

  11. 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

  12. 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.

  13. 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

  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. 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.

  17. 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

  18. 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.

  19. 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.

  20. 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

  1. 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

  2. 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.

  3. 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.

  4. 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

  5. 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.

  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. 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

  8. 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.

  9. 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

  10. 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.

  11. 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.

  12. 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

  13. 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

  14. 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.

  15. 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.

  16. 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

  17. 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.

  18. 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

  19. 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

  20. 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

  1. 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

  2. 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.

  3. 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.

  4. 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.

  5. 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

  6. 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

  7. 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.

  8. 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.

  9. 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

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  11. 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

  12. 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.

  13. 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.

  14. 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

  15. 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.

  16. 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.

  17. 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

  18. 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.

  19. 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

  20. Regulation of type 1 fimbriae synthesis and biofilm formation by the transcriptional regulator LrhA of Escherichia coli.

    PubMed

    Blumer, Caroline; Kleefeld, Alexandra; Lehnen, Daniela; Heintz, Margit; Dobrindt, Ulrich; Nagy, Gábor; Michaelis, Kai; Emödy, Levente; Polen, Tino; Rachel, Reinhard; Wendisch, Volker F; Unden, Gottfried

    2005-10-01

    Type 1 fimbriae of Escherichia coli facilitate attachment to the host mucosa and promote biofilm formation on abiotic surfaces. The transcriptional regulator LrhA, which is known as a repressor of flagellar, motility and chemotaxis genes, regulates biofilm formation and expression of type 1 fimbriae. Whole-genome expression profiling revealed that inactivation of lrhA results in an increased expression of structural components of type 1 fimbriae. In vitro, LrhA bound to the promoter regions of the two fim recombinases (FimB and FimE) that catalyse the inversion of the fimA promoter, and to the invertible element itself. Translational lacZ fusions with these genes and quantification of fimE transcript levels by real-time PCR showed that LrhA influences type 1 fimbrial phase variation, primarily via activation of FimE, which is required for the ON-to-OFF transition of the fim switch. Enhanced type 1 fimbrial expression as a result of lrhA disruption was confirmed by mannose-sensitive agglutination of yeast cells. Biofilm formation was stimulated by lrhA inactivation and completely suppressed upon LrhA overproduction. The effects of LrhA on biofilm formation were exerted via the changed levels of surface molecules, most probably both flagella and type 1 fimbriae. Together, the data show a role for LrhA as a repressor of type 1 fimbrial expression, and thus as a regulator of the initial stages of biofilm development and, presumably, bacterial adherence to epithelial host cells also.

  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. 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.

  3. 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

  4. 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

  5. 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.

  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. 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.

  9. 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

  10. 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.

  11. 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.

  12. 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.

  13. 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

  14. 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.

  15. 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

  16. 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.

  17. 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

  18. 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.

  19. 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.

  20. 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

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  2. 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.

  3. 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.

  4. 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

  5. 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.

  6. 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.

  7. 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.

  8. 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.

  9. 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.

  10. 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.

  11. 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

  12. 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.

  13. 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

  14. 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.

  15. 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.

  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. 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.

  18. 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

  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. 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.

  3. 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.

  4. 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

  5. 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

  6. 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

  7. 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

  8. 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.

  9. 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.

  10. 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

  11. 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.

  12. 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.

  13. 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.

  14. 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

  15. 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

  16. 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.

  17. 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.

  18. 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.

  19. 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

  20. 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.

  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. 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

  5. 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.

  6. 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

  7. 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

  8. 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.

  9. 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.

  10. 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

  11. 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

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  13. Identification and Characterization of a Gene Cluster Mediating Enteroaggregative Escherichia Coli Aggregative Adherence Fimbria I Biogenesis

    DTIC Science & Technology

    1994-08-01

    adherent E. coli ( DAEC ). respectively. The LA ties to other known fimbrial biogenesis systems of pathogenic pattern is typified by the formation of...agg gene cluster is configured similarly to 60 to 80% of DAEC strains share relatedness with F1845 the determinants of members of the Dr adhesin

  14. 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

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  16. 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

    protein (MBP) fusions. For routine propagation and protein expression, bacteria were grown in Luria- Bertani medium (47) or in rich medium (10 g of...tryptone, 5 g of yeast extract, 5 g of NaCl, and 2 g of glucose per liter). For hemagglutination and tissue culture adherence assays, cultures were...grown on CFA agar (8) with or without the addition of 1.5 g of Bacto bile salts no. 3 (Difco, Detroit, Mich.) per liter. Ampicillin (62.5 g /ml) and

  17. 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.

  18. 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

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  20. 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

  1. 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

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  3. 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

  4. 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

  5. [Cloning and expression of F18 fimbrial operon gene clusters from enterotoxigenic Escherichia coli and their bioactivity].

    PubMed

    Zhang, Jian-Jun; Zhu, Guo-Qiang

    2007-10-01

    The fed operon gene clusters with each size of 5.6kb, encoding the F18ab or F18ac fimbriae, was amplified respectively by high fidelity PCR using the genomic DNA templates from F18 fimbriae E. coli strains 107/86 or 2134P. The PCR products with the restriction enzyme sites at each end were digested and then cloned into the vector pET-22b (+), the recombinant plamids with the inserts of both type of fed gene clusters were constructed and screened, further confirmed by the means of combination with restriction endonuclease analysis and sequencing. The both types of fimbriae F18ab and F18ac were expressed efficiently in the E. coli BL21 (DE3) after proper concentration of IPTG induction. Expressed fimbriae were revealed and confirmed by transmissible electromicroscope observation. The both fimbriae F18ab and F18ac were isolated and purified from the recombinant E. coli, and only a single major band of protein with size of approximately 15kDa was visualized in Coomassie blue-stained gels after SDS-PAGE. The rabbits sera with high titer of anti-F18 fimbriae were detected after being immunized with the purified F18ab or F18ac fimbriae. The results of combination of agglutination assay with Western blotting showed that the sera directed against both fimbriae F18ab and F18ac reacted positively with the F18 fimbriae from both wild E. coli 107/86 and 2134P. Small intestine epithelial cells with F18 fimbriae receptors, which were from post-weaning piglets with the genotypes of FUT1 gene both M307(GG) and M307(AG), were prepared and tested for the adherence of E. coli expressing F18 fimbriae under the microscopic examination. Adhesion and adhesion inhibition test showed both of the recombinant E. coli expressing F18ab or F18ac fimbriae respectively could adhere to the jejunal epithelial cells in vitro as E. coli 107/86 and 2134p did. The both of anti-sera directed against fimbriae F18ab or F18ac respectively can efficiently inhibit the fimbriae-mediated post-weaning piglet

  6. 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

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  8. 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.

  9. 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.

  10. 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

  11. Flagella from F18+Escherichia coli play a role in adhesion to pig epithelial cell lines.

    PubMed

    Duan, Qiangde; Zhou, Mingxu; Zhu, Xiaofang; Yang, Yang; Zhu, Jun; Bao, Wenbin; Wu, Shenglong; Ruan, Xiaosai; Zhang, Weiping; Zhu, Guoqiang

    2013-02-01

    F18 fimbriae and toxins produced by F18 fimbriae-carrying Escherichia coli (E. coli) strains are known virulence factors responsible for post-weaning diarrhea (PWD) and edema disease (ED). In this study, we showed that fliC isogenic mutants constructed in two reference wild-type F18 fimbriae (F18+) E. coli were markedly impaired in adherence in vitro cell models (p < 0.05). Flagella purified from F18+E. coli could directly bind to cultured piglet epithelial cells and block adherence of F18+E. coli to cells when pre-incubated. In addition, the F18+E. coli fliC deletion mutants up-regulated the expression of type I fimbriae produced by F18+E. coli strains. These results demonstrated that expression of flagella is essential for the adherence of F18+E. coli in vitro.

  12. 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

  13. 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

  14. 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

  15. 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

  16. 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

  17. 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

  18. 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

  19. 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.

  20. 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.

  1. [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.

  2. [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.

  3. Shear-enhanced Binding of Intestinal Colonization Factor Antigen l of Enterotoxigenic Escherichia coli

    DTIC Science & Technology

    2010-01-01

    cells Caco-2 cells were seeded in 12-well plates loaded with tissue culture-treated glass slides and grown for 14-16 days to differentiate in DMEM...express- ing CFA/1 fimbriae with different CfaE adhesins (BL21-SI with various pMAM2’s) were seeded from fresh LBON agar plates (no NaCI to prevent...adhesive fimbriae of enterotoxigenic Escheri- chia coli. Infect /mmun 72: 7190-7201. Anderson , B.N., Ding, A.M., Nilsson, L.M. , Kusuma, K. , Tchesnokova

  4. 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

  5. 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.

  6. 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

  7. 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.

  8. Virulence and metabolic characteristics of Salmonella Enteritidis sefD variants in hens.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  9. Media composition and incubation temperature affect Congo red dye affinity of Shiga toxin-producing Escherichia coli

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  10. Growth media and temperature effects on biofilm formation by serotype O157:H7 and non-O157 Shiga toxin-producing Escherichia coli

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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 ...

  11. 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.

  12. 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

  13. Complete Genome Sequences of Curli-Negative and Curli-Positive Isolates of Foodborne Escherichia coli O157:H7 Strain 86-24

    PubMed Central

    Bayles, Darrell O.; Alt, David P.; Looft, Torey

    2016-01-01

    Escherichia coli O157:H7 strain 86-24 does not produce curli fimbriae, but gives rise to curli-positive isolates at a variable frequency. Here, we report the complete genome sequences of curli-negative and curli-positive isolates of strain 86-24. PMID:27979932

  14. Curli variants of enterohemorrhagic Escherichia coli O157:H7 display distinct survival fitness

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  15. 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

  16. A FaeG-FedF-LT192 fusion elicits protective anti-adhesin and antitoxin antibodies against porcine enterotoxigenic Escherichia coli

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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 ...

  17. 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

  18. Reduction of Salmonella Enteritidis in the spleens of hens by bacterins that vary in fimbrial protein SefD

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  19. Stx1 prophage excision in Escherichia coli strain PA20 confers strong curli and biofilm formation by restoring native mlrA

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  20. 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

  1. RcsB contributes to the distinct stress fitness between Escherichia coli O157:H7 curli variants of 1993 hamburger-associated outbreak strains

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  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. 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.

  4. 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

  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. 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.

  7. 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.

  8. 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.

  9. 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.

  10. 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.

  11. [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.

  12. [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.

  13. 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.

  14. [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.

  15. 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

  16. 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

  17. 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

  18. 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

  19. 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

  20. 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

  1. 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.

  2. 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

  3. 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

  4. [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.

  5. 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.

  6. 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.

  7. 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.

  8. 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

  9. 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.

  10. 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

  11. 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

  12. 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

  13. 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

  14. 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

  15. 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.

  16. Glycolipid receptors for uropathogenic Escherichia coli on human erythrocytes and uroepithelial cells.

    PubMed Central

    Leffler, H; Svanborg-Edén, C

    1981-01-01

    A specific family of glycolipids, the globoseries, was shown to act as receptors on human uroepithelial cells and erythrocytes for the majority of uropathogenic Escherichia coli strains attaching to or hemagglutinating those cells. This was demonstrated in three different ways: (i) correlation between the natural presence of glycolipid in the target cell (erythrocytes of different species) and binding of bacteria; (ii) inhibition of attachment to human uroepithelial cells by preincubation of bacteria and glycolipid; and (iii) induction of binding to unreactive cells by coating of these cells with glycolipid. Strains reacting with the receptor agglutinated guinea pig erythrocytes in a mannose-resistant way after, but not before, coating of the cells with globotetraosylceramide. Unrelated glycolipids were not recognized. The reaction was made independent of simultaneous occurrence of mannose-sensitive adhesions on the strains by addition of D-mannose. The receptor-coated cells were used as a tool to screen for prevalence of receptor recognition in a collection of 453 E. coli strains isolated from patients with urinary tract infection or from the stools of healthy children. Of 150 strains attaching to human uroepithelial cells and agglutinating human erythrocytes, 121 bound to globotetraosylceramide (81%). Globoside recognition was especially frequent among pyelonephritis strains (74/81). The glycolipid composition of the urogenital epithelium and kidney tissue and the ability of uropathogenic E. coli to bind to these glycolipids may be a determinant in host-parasite interaction leading to urinary tract infection. PMID:7037645

  17. 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

  18. 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.

  19. 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

  20. Breast cancer risk and clinical implications for germline PTEN mutation carriers.

    PubMed

    Ngeow, Joanne; Sesock, Kaitlin; Eng, Charis

    2015-12-23

    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.

  1. 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

  2. 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.

  3. 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.

  4. 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.

  5. 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

  6. 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.

  7. 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

  8. 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.

  9. 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)

  10. 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.

  11. 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.

  12. 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

  13. 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

  14. 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.

  15. 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

  16. 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

  17. Asynchronous division by non-ring FtsZ in the gammaproteobacterial symbiont of Robbea hypermnestra.

    PubMed

    Leisch, Nikolaus; Pende, Nika; Weber, Philipp M; Gruber-Vodicka, Harald R; Verheul, Jolanda; Vischer, Norbert O E; Abby, Sophie S; Geier, Benedikt; den Blaauwen, Tanneke; Bulgheresi, Silvia

    2016-10-10

    The reproduction mode of uncultivable microorganisms deserves investigation as it can largely diverge from conventional transverse binary fission. Here, we show that the rod-shaped gammaproteobacterium thriving on the surface of the Robbea hypermnestra nematode divides by FtsZ-based, non-synchronous invagination of its poles-that is, the host-attached and fimbriae-rich pole invaginates earlier than the distal one. We conclude that, in a naturally occurring animal symbiont, binary fission is host-oriented and does not require native FtsZ to polymerize into a ring at any septation stage.

  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. 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

  20. 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.

  1. 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

  2. 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

  3. 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

  4. 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.

  5. 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.

  6. 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

  7. 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.

  8. 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

  9. 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.

  10. 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.

  11. 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.

  12. 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

  13. 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

  14. 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.

  15. 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.

  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. 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.

  18. 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.

  19. 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.

  20. 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

  1. 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

  2. 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

  3. 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.

  4. 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

  5. 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.

  6. 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

  7. 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.

  8. 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

  9. 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.

  10. 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

  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. 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.

    PubMed

    Rabban, Joseph T; Krasik, Ellen; Chen, Lee-May; Powell, Catherine B; Crawford, Beth; Zaloudek, Charles J

    2009-12-01

    Risk-reducing salpingo-oophorectomy (RRSO) significantly lowers the incidence of ovarian, tubal, peritoneal, and breast cancer in women who carry BRCA1 or BRCA2 germline mutations. A minority of RRSO specimens from these women will contain occult early-stage carcinoma. Most occult cancer is localized in the fallopian tube fimbriae and is as small as 1 mm in size. Pathologic detection is dependent on thoroughness of tissue examination. Recommended protocols to maximize tumor detection emphasize the role of thinly slicing the tubes and ovaries and embedding the entire specimen for microscopic examination. Additional multistep level sections of tubal fimbriae tissue blocks could theoretically increase detection of occult tubal carcinoma but the value of level sections has not been formally evaluated. This study tests the diagnostic utility of multistep level sections in RRSO specimens from 102 women with BRCA germline mutations. The original diagnoses were based on a single section from each block of thinly sliced (2 to 3 mm intervals) tissues of the entire RRSO specimen. Three multistep level sections were retrospectively obtained from each block containing tubal fimbriae. Clinically occult carcinoma ranging in size from 1 to 13 mm was initially detected in 11 of 102 women (5 in tubal fimbriae only, 1 in tubal isthmus only, 2 in fimbriae and ovary, and 3 in ovary only). Diagnoses in the original fimbrial slides and their level sections were concordant in all cases. All tubal cancers were detected in both the original sections and in the multistep level sections. None of the tubal carcinomas that were noninvasive on the original slides showed invasive growth on additional level sections. No tubal carcinoma was identified in the level sections of any case originally classified as benign. Clinical follow-up among women with benign RRSO findings revealed that 2 women subsequently developed peritoneal carcinomatosis at 22 and 62 months postoperatively. Retrospective

  13. Phenotypic and genotypic characteristics associated with biofilm formation in clinical isolates of atypical enteropathogenic Escherichia coli (aEPEC) strains

    PubMed Central

    2014-01-01

    Background Biofilm formation by enteropathogenic Escherichia coli (EPEC) have been recently described in the prototype typical EPEC E2348/69 strain and in an atypical EPEC O55:H7 strain. In this study, we sought to evaluate biofilm formation in a collection of 126 atypical EPEC strains isolated from 92 diarrheic and 34 nondiarrheic children, belonging to different serotypes. The association of biofilm formation and adhesin-related genes were also investigated. Results Biofilm formation occurred in 37 (29%) strains of different serotypes, when the assays were performed at 26°C and 37°C for 24 h. Among these, four strains (A79, A87, A88, and A111) formed a stronger biofilm than did the others. The frequency of biofilm producers was higher among isolates from patients compared with isolates from controls (34.8% vs 14.7%; P = 0.029). An association was found between biofilm formation and expression of type 1 fimbriae and curli (P < 0.05). Unlike the previously described aEPEC O55:H7, one aEPEC O119:HND strain (A111) formed a strong biofilm and pellicle at the air-liquid interface, but did not express curli. Transposon mutagenesis was used to identify biofilm-deficient mutants. Transposon insertion sequences of six mutants revealed similarity with type 1 fimbriae (fimC, fimD, and fimH), diguanylate cyclase, ATP synthase F1, beta subunit (atpD), and the uncharacterized YjiC protein. All these mutants were deficient in biofilm formation ability. Conclusion This study showed that the ability to adhere to abiotic surfaces and form biofilm is present in an array of aEPEC strains. Moreover, it seems that the ability to form biofilms is associated with the presence of type 1 fimbriae and diguanylate cyclase. Characterization of additional biofilm formation mutants may reveal other mechanisms involved in biofilm formation and bring new insights into aEPEC adhesion and pathogenesis. PMID:25012525

  14. Antibody responses to Bordetella pertussis Fim2 or Fim3 following immunization with a whole-cell, two-component, or five-component acellular pertussis vaccine and following pertussis disease in children in Sweden in 1997 and 2007.

    PubMed

    Hallander, Hans; Advani, Abdolreza; Alexander, Frances; Gustafsson, Lennart; Ljungman, Margaretha; Pratt, Catherine; Hall, Ian; Gorringe, Andrew R

    2014-02-01

    Bordetella pertussis fimbriae (Fim2 and Fim3) are components of a five-component acellular pertussis vaccine (diphtheria-tetanus-acellular pertussis vaccine [DTaP5]), and antibody responses to fimbriae have been associated with protection. We analyzed the IgG responses to individual Fim2 and Fim3 in sera remaining from a Swedish placebo-controlled efficacy trial that compared a whole-cell vaccine (diphtheria-tetanus-whole-cell pertussis vaccine [DTwP]), a two-component acellular pertussis vaccine (DTaP2), and DTaP5. One month following three doses of the Fim-containing vaccines (DTwP or DTaP5), anti-Fim2 geometric mean IgG concentrations were higher than those for anti-Fim3, with a greater anti-Fim2/anti-Fim3 IgG ratio elicited by DTaP5. We also determined the responses in vaccinated children following an episode of pertussis. Those who received DTaP5 showed a large rise in anti-Fim2 IgG, reflecting the predominant Fim2 serotype at the time. In contrast, those who received DTwP showed an equal rise in anti-Fim2 and anti-Fim3 IgG concentrations, indicating that DTwP may provide a more efficient priming effect for a Fim3 response following contact with B. pertussis. Anti-Fim2 and anti-Fim3 IgG concentrations were also determined in samples from two seroprevalence studies conducted in Sweden in 1997, when no pertussis vaccine was used and Fim2 isolates predominated, and in 2007, when either DTaP2 or DTaP3 without fimbriae was used and Fim3 isolates predominated. Very similar distributions of anti-Fim2 and anti-Fim3 IgG concentrations were obtained in 1997 and 2007, except that anti-Fim3 concentrations in 1997 were lower. This observation, together with the numbers of individuals with both anti-Fim2 and anti-Fim3 IgG concentrations, strongly suggests that B. pertussis expresses both Fim2 and Fim3 during infection.

  15. Antibody Responses to Bordetella pertussis Fim2 or Fim3 following Immunization with a Whole-Cell, Two-Component, or Five-Component Acellular Pertussis Vaccine and following Pertussis Disease in Children in Sweden in 1997 and 2007

    PubMed Central

    Hallander, Hans; Advani, Abdolreza; Alexander, Frances; Gustafsson, Lennart; Ljungman, Margaretha; Pratt, Catherine; Hall, Ian

    2014-01-01

    Bordetella pertussis fimbriae (Fim2 and Fim3) are components of a five-component acellular pertussis vaccine (diphtheria–tetanus–acellular pertussis vaccine [DTaP5]), and antibody responses to fimbriae have been associated with protection. We analyzed the IgG responses to individual Fim2 and Fim3 in sera remaining from a Swedish placebo-controlled efficacy trial that compared a whole-cell vaccine (diphtheria-tetanus-whole-cell pertussis vaccine [DTwP]), a two-component acellular pertussis vaccine (DTaP2), and DTaP5. One month following three doses of the Fim-containing vaccines (DTwP or DTaP5), anti-Fim2 geometric mean IgG concentrations were higher than those for anti-Fim3, with a greater anti-Fim2/anti-Fim3 IgG ratio elicited by DTaP5. We also determined the responses in vaccinated children following an episode of pertussis. Those who received DTaP5 showed a large rise in anti-Fim2 IgG, reflecting the predominant Fim2 serotype at the time. In contrast, those who received DTwP showed an equal rise in anti-Fim2 and anti-Fim3 IgG concentrations, indicating that DTwP may provide a more efficient priming effect for a Fim3 response following contact with B. pertussis. Anti-Fim2 and anti-Fim3 IgG concentrations were also determined in samples from two seroprevalence studies conducted in Sweden in 1997, when no pertussis vaccine was used and Fim2 isolates predominated, and in 2007, when either DTaP2 or DTaP3 without fimbriae was used and Fim3 isolates predominated. Very similar distributions of anti-Fim2 and anti-Fim3 IgG concentrations were obtained in 1997 and 2007, except that anti-Fim3 concentrations in 1997 were lower. This observation, together with the numbers of individuals with both anti-Fim2 and anti-Fim3 IgG concentrations, strongly suggests that B. pertussis expresses both Fim2 and Fim3 during infection. PMID:24307240

  16. Porphyromonas gingivalis genes isolated by screening for epithelial cell attachment.

    PubMed Central

    Duncan, M J; Emory, S A; Almira, E C

    1996-01-01

    Porphyromonas gingivalis is associated with chronic and severe periodontitis in adults. P. gingivalis and the other periodontal pathogens colonize and interact with gingival epithelial cells, but the genes and molecular mechanisms involved are unknown. To dissect the first steps in these interactions, a P. gingivalis expression library was screened for clones which bound human oral epithelial cells. Insert DNA from the recombinant clones did not contain homology to the P. gingivalis fimA gene, encoding fimbrillin, the subunit protein of fimbriae, but showed various degrees of homology to certain cysteine protease-hemagglutinin genes. The DNA sequence of one insert revealed three putative open reading frames which appeared to be in an operon. The relationship between P. gingivalis attachment to epithelial cells and the activities identified by the screen is discussed. PMID:8751909

  17. Appendages of the cyanobacterial cell.

    PubMed

    Schuergers, Nils; Wilde, Annegret

    2015-03-04

    Extracellular non-flagellar appendages, called pili or fimbriae, are widespread in gram-negative bacteria. They are involved in many different functions, including motility, adhesion, biofilm formation, and uptake of DNA. Sequencing data for a large number of cyanobacterial genomes revealed that most of them contain genes for pili synthesis. However, only for a very few cyanobacteria structure and function of these appendages have been analyzed. Here, we review the structure and function of type IV pili in Synechocystis sp. PCC 6803 and analyze the distribution of type IV pili associated genes in other cyanobacteria. Further, we discuss the role of the RNA-chaperone Hfq in pilus function and the presence of genes for the chaperone-usher pathway of pilus assembly in cyanobacteria.

  18. Expression and purification of the mannose recognition domain of the FimH adhesin.

    PubMed

    Schembri, M A; Hasman, H; Klemm, P

    2000-07-15

    Type 1 fimbriae have been shown to be specifically required for Escherichia coli colonisation and pathogenesis of the urinary tract. These structural organelles mediate specific adhesion to alpha-D-mannosides by virtue of the FimH adhesin. FimH is a two-domain protein in which the N-terminal domain contains the receptor-binding site and the C-terminal domain is required for organelle integration. To date, FimH has only been isolated as a complex with the system-specific chaperone FimC. Here we report that a functional form of the FimH receptor-binding domain can be readily isolated and characterised by replacing the C-terminal domain with a histidine tag.

  19. Purification of the outer membrane usher protein and periplasmic chaperone-subunit complexes from the P and type 1 pilus systems.

    PubMed

    Henderson, Nadine S; Thanassi, David G

    2013-01-01

    Understanding molecular mechanisms of protein secretion by bacteria requires the purification of secretion machinery components and the isolation of complexes between the secretion machinery and substrate proteins. Here, we describe methods for the purification of proteins from the chaperone/usher pathway, which is a conserved secretion pathway dedicated to the assembly of polymeric surface fibers termed pili or fimbriae in gram-negative bacteria. Specifically, we describe the isolation of the PapC and FimD usher proteins from the bacterial outer membrane, and the purification of PapD-PapG and FimC-FimH chaperone--subunit complexes from the periplasm. These Pap and Fim proteins belong to the P and type 1 pilus systems of uropathogenic Escherichia coli, respectively.

  20. [Oral ciprofloxacin therapy in salpingitis].

    PubMed

    De Wilde, R

    1988-01-01

    Thirty patients with clinical pelvic inflammatory disease were studied. Diagnosis was confirmed by laparoscopy. To determine the microbiological etiology, swab specimen for detection of aerobic, anaerobic and chlamydial infections were obtained from the endocervix, fimbriae and cul-de-sac. In 22 of 30 patients, microorganisms were detected. The patients received ciprofloxacin 2 X 750 mg p.o. daily at 12 hours interval for 10 days. During and after therapy, bacteriological examinations of the endocervix were performed. Based on the microbiological evaluation and clinical aspects, the bacteriological response and clinical efficacy were 86%. Laboratory analysis showed no alterations of blood values. As side effects we noticed gastrointestinal complaints (6/30), candidiasis vulvovaginalis (6/30), allergic exanthema (5/30) and non-bacterial cystitis (2/30). Oral ciprofloxacin-monotherapy proved to be safe and effective in pelvic inflammatory disease.

  1. Potential virulence factors of Proteus bacilli.

    PubMed Central

    Rózalski, A; Sidorczyk, Z; Kotełko, K

    1997-01-01

    The object of this review is the genus Proteus, which contains bacteria considered now to belong to the opportunistic pathogens. Widely distributed in nature (in soil, water, and sewage), Proteus species play a significant ecological role. When present in the niches of higher macroorganisms, these species are able to evoke pathological events in different regions of the human body. The invaders (Proteus mirabilis, P. vulgaris, and P. penneri) have numerous factors including fimbriae, flagella, outer membrane proteins, lipopolysaccharide, capsule antigen, urease, immunoglobulin A proteases, hemolysins, amino acid deaminases, and, finally, the most characteristic attribute of Proteus, swarming growth, enabling them to colonize and survive in higher organisms. All these features and factors are described and commented on in detail. The questions important for future investigation of these facultatively pathogenic microorganisms are also discussed. PMID:9106365

  2. Folding of the C-terminal bacterial binding domain in statherin upon adsorption onto hydroxyapatite crystals

    PubMed Central

    Goobes, Gil; Goobes, Rivka; Schueler-Furman, Ora; Baker, David; Stayton, Patrick S.; Drobny, Gary P.

    2006-01-01

    Statherin is an enamel pellicle protein that inhibits hydroxyapatite (HAP) nucleation and growth, lubricates the enamel surface, and is recognized by oral bacteria in periodontal diseases. We report here from solid-state NMR measurements that the protein's C-terminal region folds into an α-helix upon adsorption to HAP crystals. This region contains the binding sites for bacterial fimbriae that mediate bacterial cell adhesion to the surface of the tooth. The helical segment is shown through long-range distance measurements to fold back onto the intermediate region (residues Y16–P28) defining the global fold of the protein. Statherin, previously shown to be unstructured in solution, undergoes conformation selection on its substrate mineral surface. This surface-induced folding of statherin can be related to its functionality in inhibiting HAP crystal growth and can explain how oral pathogens selectively recognize HAP-bound statherin. PMID:17060618

  3. Development of an IgY-based rocket-immunoelectrophoresis for identity monitoring of Pertussis vaccines.

    PubMed

    Matheis, Walter; Schade, Rüdiger

    2011-06-30

    An important step in vaccine production and quality control is the analysis of identity of different lots. For that purpose chicken was immunized with acellular Pertussis components (Pertussis toxoid, Filamenteous haemagglutinin, Pertactin, Fimbriae 2/3 antigen). The resulting antibodies (IgY) were non-invasive extracted from egg yolk and used for rocket immunoelectrophoresis (RIE). We demonstrated that the Ab reacted with characteristic peaks ("rockets") with the corresponding antigen. The shape of the peaks varied depending on the manufacturer and the nature of antigen (adsorbed or non-adsorbed). The coefficients of variation was about 20% during a year period. In summary, our data illustrate that an IgY-based RIE is not only a cost-effective method but also proficient for monitoring Pertussis vaccines.

  4. [Adhesion of corynebacterium diphtheriae: the role of surface structures and formation mechanism].

    PubMed

    Kharseeva, G G; Alieva, A A

    2014-01-01

    The paper is devoted to the study of surface structures including pili (fimbriae) 67-72p surface protein, DIP 1281 surface protein, lipoarabinomannan CdiLAM and their role in the adhesion and colonization of the mucous membrane of the throat by Corynebacterium diphtheriae. A description is offered for the main stages in the adhesion process of diphtheria causative agent and the ability of its adhesins to stimulate the effect of innate and acquired immunity factors. The paper stresses prospectiveness of the development of vaccines forming immunoprotection of the organism against adhesive activity of C. diphtheriae and also preventing their colonization and reproduction. That would facilitate a solution for the problem of diphtheria carrier state, which cannot be solved using the existing means of preventive vaccination.

  5. High molecular weight bioemulsifiers, main properties and potential environmental and biomedical applications.

    PubMed

    Mnif, Inès; Ghribi, Dhouha

    2015-05-01

    High molecular weight bioemulsifiers are amphipathic polysaccharides, proteins, lipopolysaccharides, lipoproteins, or complex mixtures of these biopolymers, produced by a wide variety of microorganisms. They are characterized by highly structural diversity and have the ability to decrease the surface and interfacial tension at the surface and interface respectively and/or emulsify hydrophobic compounds. Emulsan, fatty acids, phospholipids, neutral lipids, exopolysaccharides, vesicles and fimbriae are among the most popular high molecular weight bioemulsifiers. They have great physic-chemical properties like tolerance to extreme conditions of pH, temperature and salinity, low toxicity and biodegradability. Owing their emulsion forming and breaking capacities, solubilization, mobilization and dispersion activities and their viscosity reduction activity; they possess great environmental application as enhancer of hydrocarbon biodegradation and for microbial enhanced oil recovery. Besides, they are applied in biomedical fields for their antimicrobial and anti-adhesive activities and involvement in immune responses.

  6. The role of quorum sensing in Escherichia coli (ETEC) virulence factors.

    PubMed

    Sturbelle, Régis Tuchtenhagen; de Avila, Luciana Farias da Costa; Roos, Talita Bandeira; Borchardt, Jéssica Lopes; da Conceição, Rita de Cássia dos Santos; Dellagostin, Odir Antonio; Leite, Fábio Pereira Leivas

    2015-11-18

    Quorum sensing (QS) is a signaling system among bacteria mediated by auto-inducer substances (AI). Whenever the concentration of these molecules reaches a threshold corresponding to a high cell density or quorum, the whole population starts a coordinated expression of specific genes. Studies have shown that epinephrine is also responsible for activating specific bacterial genes. This work aimed to investigate the role of conditioned medium (containing AI), epinephrine and their association on growth, motility, F4 fimbriae and heat-labile toxin (LT) expression on enterotoxigenic Escherichia coli (ETEC, E68). A significant increase in motility, F4 and LT expression, was observed in the ETEC culture supplemented with conditioned medium and epinephrine. These findings suggest that ETEC uses some components of conditioned medium (e.g., AI molecules), host molecules (epinephrine), and their association to modulate the expression of important virulence genes.

  7. Selection of indigenous lactic acid bacteria to reinforce the intestinal microbiota of newly hatched chicken: relevance of in vitro and ex vivo methods for strains characterization.

    PubMed

    Babot, Jaime D; Argañaraz-Martínez, Eloy; Saavedra, Lucila; Apella, María C; Perez Chaia, Adriana

    2014-08-01

    Based on the natural benefits of the indigenous microbiota, lactic acid bacteria (LAB) from poultry origin were isolated from hens and broilers intestine, and their probiotic potential was further studied. The tolerance to digestion, adhesion, capture of a mannose-binding lectin, absence of virulent factors and antibiotic resistances were studied. Different in vitro and ex vivo assays were performed to select tolerant and adherent strains because standardized protocols have not been defined. Fourteen strains highly tolerant to gastrointestinal digestion were genetically identified. Hydrophobic surfaces were not required for the bacterial adhesion and only nine strains adhered ex vivo to the intestinal mucosa. Three strains captured a lectin of the same specificity of Type-1 fimbriae. Virulence factors were absent but some strains evidenced multiple antibiotic resistances. These results provide bases for a future standardization of methods for the selection of probiotic strains intended to reinforce the microbiota of newly hatched chickens.

  8. Leishmania (Leishmania) chagasi interactions with Serratia marcescens: ultrastructural studies, lysis and carbohydrate effects.

    PubMed

    Moraes, Caroline S; Seabra, Sergio H; Castro, Daniele P; Brazil, Reginaldo P; de Souza, Wanderley; Garcia, Eloi S; Azambuja, Patrícia

    2008-04-01

    Studies on the lysis of L. chagasi caused by the bacteria Serratia marcescens were carried out. In vitro experiments demonstrated that S. marcescens variant SM 365, a prodigiosin pigment producer, lysed this species of Leishmania but variant DB11, a nonpigmented bacteria, was unable to lyse the parasite. High concentrations of d-mannose were found to protect L. chagasi markedly diminishing the lysis by S. marcescens SM 365. Promastigotes of L. chagasi bound the lectin Concanavalin A conjugated with FITC, the fluorescence was intensely found at the base of the flagellum (flagellar pocket). Scanning electron microscopy revealed that the bacteria adherence occurred mainly in the flagellar pocket. S. marcescens SM 365 formed filamentous structures, identified as biofilms, which connect the protozoan to the developing bacterial clusters, in low concentrations of bacteria after 30 min incubation time. We suggest that bacterial mannose-sensitive (MS) fimbriae are relevant to S. marcescens SM 365 in the lysis of L. chagasi.

  9. Evaluation of components of X-ray irradiated 7-valent pneumococcal conjugate vaccine and pneumococcal vaccine polyvalent and X-ray and gamma-ray irradiated acellular pertussis component of DTaP vaccine products

    NASA Astrophysics Data System (ADS)

    May, J. C.; Rey, L.; Lee, Chi-Jen; Arciniega, Juan

    2004-09-01

    Samples of pneumococcal vaccine polyvalent, 7-valent pneumococcal conjugate vaccine, and two different diphtheria and tetanus toxoids and acellular pertussis vaccines adsorbed were irradiated with X-rays and/or gamma-rays (Co-60). Mouse IgG and IgM antibody responses (ELISA) for types 9V, 14, 18C, and 19F pneumococcal polysaccharides and conjugates indicated that the polysaccharides were more tolerant of the radiation than the conjugates. The mouse antibody response for the detoxified pertussis toxin (PT) antigen, filamentous hemagglutinin antigen (FHA), pertactin (PRN), and fimbriae types 2 and 3 (FIM) antigens for the appropriate vaccine type indicated that the antibody response was not significantly changed in the 25 kGy X-ray irradiated vaccines frozen in liquid nitrogen compared to the control vaccine.

  10. Pili in Gram-negative and Gram-positive bacteria - structure, assembly and their role in disease.

    PubMed

    Proft, T; Baker, E N

    2009-02-01

    Many bacterial species possess long filamentous structures known as pili or fimbriae extending from their surfaces. Despite the diversity in pilus structure and biogenesis, pili in Gram-negative bacteria are typically formed by non-covalent homopolymerization of major pilus subunit proteins (pilins), which generates the pilus shaft. Additional pilins may be added to the fiber and often function as host cell adhesins. Some pili are also involved in biofilm formation, phage transduction, DNA uptake and a special form of bacterial cell movement, known as 'twitching motility'. In contrast, the more recently discovered pili in Gram-positive bacteria are formed by covalent polymerization of pilin subunits in a process that requires a dedicated sortase enzyme. Minor pilins are added to the fiber and play a major role in host cell colonization.This review gives an overview of the structure, assembly and function of the best-characterized pili of both Gram-negative and Gram-positive bacteria.

  11. Molecular cloning and sequencing of the gene encoding the fimbrial subunit protein of Bacteroides gingivalis.

    PubMed Central

    Dickinson, D P; Kubiniec, M A; Yoshimura, F; Genco, R J

    1988-01-01

    The gene encoding the fimbrial subunit protein of Bacteroides gingivalis 381, fimbrilin, has been cloned and sequenced. The gene was present as a single copy on the bacterial chromosome, and the codon usage in the gene conformed closely to that expected for an abundant protein. The predicted size of the mature protein was 35,924 daltons, and the secretory form may have had a 10-amino-acid, hydrophilic leader sequence similar to the leader sequences of the MePhe fimbriae family. The protein sequence had no marked similarity to known fimbrial sequences, and no homologous sequences could be found in other black-pigmented Bacteroides species, suggesting that fimbrillin represents a class of fimbrial subunit protein of limited distribution. Images PMID:2895100

  12. Aggregatibacter Actinomycetemcomitans – A Tooth Killer?

    PubMed Central

    Ummer, Fajar; Dhivakar, C.P

    2014-01-01

    Strong evidence is available on Aggregatibacter actinomycetemcomitans (A.a) on its role as the causative agent of localised juvenile periodontitis (LJP), a disease characterised by rapid destruction of the tooth-supporting tissues. This organism possesses a large number of virulence factors with a wide range of activities which enable it to colonise the oral cavity, invade periodontal tissues, evade host defences, initiate connective tissue destruction and interfere with tissue repair. Adhesion to epithelial and tooth surfaces is dependent on the presence of surface proteins and structures such as microvesicles and fimbriae. Invasion has been demonstrated in vivo and in vitro. The organism has a number of means of evading host defences which include: (i) production of leukotoxin; (ii) producing immunosuppressive factors; (iv) secreting proteases capable of cleaving IgG; and (v) producing Fc-binding. PMID:25302290

  13. From pertussis to meningococcal disease and back.

    PubMed

    Gorringe, Andrew

    2011-04-01

    From pertussis to meningococcal disease and back represents nearly 30 years of research at Porton, first at the Centre for Applied Microbiology and Research and latterly as part of the Health Protection Agency. I joined the group lead by Andy Robinson developing an acellular pertussis vaccine and was part of an exciting period that encompassed basic antigen characterisation and pathogenesis studies with the development of an acellular vaccine containing fimbriae. Research then changed to focus on serogroup B meningococcal disease, studying the vaccine potential of iron-regulated proteins and then Neisseria lactamica. The resurgence of pertussis seen in some countries alerted me to the lack of understanding of protective immune responses to Bordetella pertussis infection and disease and this is now an active area of research.

  14. Fermentation and dry fractionation increase bioactivity of cloudberry (Rubus chamaemorus).

    PubMed

    Puupponen-Pimiä, Riitta; Nohynek, Liisa; Juvonen, Riikka; Kössö, Tuija; Truchado, Pilar; Westerlund-Wikström, Benita; Leppänen, Tiina; Moilanen, Eeva; Oksman-Caldentey, Kirsi-Marja

    2016-04-15

    Phenolic composition and bioactivity of cloudberry was modified by bioprocessing, and highly bioactive fractions were produced by dry fractionation of the press cake. During fermentation polymeric ellagitannins were partly degraded into ellagic acid derivatives. Phenolic compounds were differentially distributed in seed coarse and fine fractions after dry fractionation process. Tannins concentrated in fine fraction, and flavonol derivatives were mainly found in coarse fraction. Ellagic acid derivatives were equally distributed between the dry fractions. Fermentation and dry fractionation increased statistically significantly anti-adhesion and anti-inflammatory activity of cloudberry. The seed fine fraction showed significant inhibition of P fimbria-mediated haemagglutination assay of uropathogenic Escherichia coli. The seed coarse fraction significantly reduced NO and IL-6 production and iNOS expression in activated macrophages. Fermentation did not affect antimicrobial activity, but slight increase in activity was detected in dry fractions. The results indicate the potential of cloudberry in pharma or health food applications.

  15. Associative Nitrogen Fixation by Klebsiella spp.: Adhesion Sites and Inoculation Effects on Grass Roots

    PubMed Central

    Haahtela, Kielo; Laakso, Tuula; Korhonen, Timo K.

    1986-01-01

    Adhesion sites on grass roots for Klebsiella strains carrying type 3 or type 1 fimbriae or both were determined. Adhesion of the strains to the roots of Poa pratensis and Festuca rubra was highly localized; the bacteria adhered strongly to root hairs and with a markedly lower efficiency to the surface of the zone of elongation and to the root cap mucilage. No adhesion to the epidermal cells between root hairs was observed. The adhesion sites were identical for the type 3- and 1-fimbriated bacteria and for P. pratensis, F. rubra, and Trifolium pratense. Inoculation of P. pratensis seedlings with Klebsiella pneumoniae strain As resulted in morphological changes in plant roots. The roots of infected plants were heavily covered with root hairs, which often were deformed and branched. Images PMID:16347205

  16. Evolution of the Chaperone/Usher Assembly Pathway: Fimbrial Classification Goes Greek†

    PubMed Central

    Nuccio, Sean-Paul; Bäumler, Andreas J.

    2007-01-01

    Summary: Many Proteobacteria use the chaperone/usher pathway to assemble proteinaceous filaments on the bacterial surface. These filaments can curl into fimbrial or nonfimbrial surface structures (e.g., a capsule or spore coat). This article reviews the phylogeny of operons belonging to the chaperone/usher assembly class to explore the utility of establishing a scheme for subdividing them into clades of phylogenetically related gene clusters. Based on usher amino acid sequence comparisons, our analysis shows that the chaperone/usher assembly class is subdivided into six major phylogenetic clades, which we have termed α-, β-, γ-, κ-, π-, and σ-fimbriae. Members of each clade share related operon structures and encode fimbrial subunits with similar protein domains. The proposed classification system offers a simple and convenient method for assigning newly discovered chaperone/usher systems to one of the six major phylogenetic groups. PMID:18063717

  17. The Main Aeromonas Pathogenic Factors

    PubMed Central

    Tomás, J. M.

    2012-01-01

    The members of the Aeromonas genus are ubiquitous, water-borne bacteria. They have been isolated from marine waters, rivers, lakes, swamps, sediments, chlorine water, water distribution systems, drinking water and residual waters; different types of food, such as meat, fish, seafood, vegetables, and processed foods. Aeromonas strains are predominantly pathogenic to poikilothermic animals, and the mesophilic strains are emerging as important pathogens in humans, causing a variety of extraintestinal and systemic infections as well as gastrointestinal infections. The most commonly described disease caused by Aeromonas is the gastroenteritis; however, no adequate animal model is available to reproduce this illness caused by Aeromonas. The main pathogenic factors associated with Aeromonas are: surface polysaccharides (capsule, lipopolysaccharide, and glucan), S-layers, iron-binding systems, exotoxins and extracellular enzymes, secretion systems, fimbriae and other nonfilamentous adhesins, motility and flagella. PMID:23724321

  18. Identification of antigenically related polypeptides at centrioles and basal bodies.

    PubMed

    Lin, W; Fung, B; Shyamala, M; Kasamatsu, H

    1981-04-01

    An antigen localized at the centriolar region has been identified by indirect immunofluorescence studies in African green monkey kidney, human, hamster, rat, and mouse cells. The antigen consists of two polypeptides of 14,000 and 17,000 daltons. A related antigen is also present at the basal body region in ciliated cells from chicken, cat, mouse, pig, steer, and rabbit trachea and from rabbit fimbria. Immunoelectron microscopy shows that the immunoreactive antigen is indeed located in the region around the basal bodies of ciliated cat tracheal cells. Thus, we have found an antigen that is common to a variety of cell types from many different animal sources and is specifically associated with both centrioles and basal bodies. The possible role of the antigen in differentiation is discussed.

  19. Identification of antigenically related polypeptides at centrioles and basal bodies.

    PubMed Central

    Lin, W; Fung, B; Shyamala, M; Kasamatsu, H

    1981-01-01

    An antigen localized at the centriolar region has been identified by indirect immunofluorescence studies in African green monkey kidney, human, hamster, rat, and mouse cells. The antigen consists of two polypeptides of 14,000 and 17,000 daltons. A related antigen is also present at the basal body region in ciliated cells from chicken, cat, mouse, pig, steer, and rabbit trachea and from rabbit fimbria. Immunoelectron microscopy shows that the immunoreactive antigen is indeed located in the region around the basal bodies of ciliated cat tracheal cells. Thus, we have found an antigen that is common to a variety of cell types from many different animal sources and is specifically associated with both centrioles and basal bodies. The possible role of the antigen in differentiation is discussed. Images PMID:6166008

  20. Escherichia coli pili as possible mediators of attachment to human urinary tract epithelial cells.

    PubMed Central

    Edén, C S; Hansson, H A

    1978-01-01

    Presence of pili of fimbriae on Escherichia coli bacteria isolated from the urine of patients with urinary tract infection was related to the ability of the bacteria to attach to human uroepithelial cells. Piliated E. coli strains agglutinated guinea pig erythrocytes. D-Mannose and alpha-methyl-D-mannopyranoside inhibited this agglutination with all but one of the 12 strains tested. D-Mannose, D-galactose, alpha-methyl-D-mannopyranoside, and L-fucose did not afect attachment of piliated strains to uroepithelial cells. Heating as well as washing of piliated strains caused a parallel decrease of piliation and adhesive ability. Growth in glucose-enriched medium increased capsule formation but decreased piliation and adhesion. Capsulated strains retained their adhesive ability provided that pili extended outside the capsule. Images PMID:361565

  1. The alginate regulator AlgR and an associated sensor FimS are required for twitching motility in Pseudomonas aeruginosa.

    PubMed Central

    Whitchurch, C B; Alm, R A; Mattick, J S

    1996-01-01

    Mucoid strains of Pseudomonas aeruginosa isolated from the lungs of cystic fibrosis patients produce large amounts of the exopolysaccharide alginate. AlgR has long been considered a key regulator of alginate production, but its cognate sensor has not been identified. Here we show that AlgR is required for twitching motility, which is a form of bacterial surface translocation mediated by type 4 fimbriae. Adjacent to algR we have identified a sensor gene (fimS), which is also required for twitching motility. However, FimS does not appear to be required for alginate production in mucoid strains. FimS and AlgR are representative of a new subclass of two-component transmitter-receiver regulatory systems. The alternative sigma factor AlgU also affects both alginate production and twitching motility. Therefore, these two virulence determinants appear to be closely associated and coordinately regulated. Images Fig. 1 Fig. 2 Fig. 3 PMID:8790418

  2. Neuropathology and structural changes in hydrocephalus.

    PubMed

    Del Bigio, Marc R

    2010-01-01

    In the context of spina bifida, hydrocephalus is usually caused by crowding of the posterior fossa with obstruction to cerebrospinal fluid flow from the forth ventricle, and less often by malformation of the cerebral aqueduct. Enlargement of the cerebral ventricles causes gradual destruction of periventricular white matter axons. Motor, sensory, visual, and memory systems may be disturbed through involvement of the long projection axons, periventricular structures including the corpus callosum, and the fimbria-fornix pathway. Secondary changes occur in neuronal cell bodies and synapses, but there is minimal death of neurons. The clinical syndrome of hydrocephalic brain dysfunction is thus due to subcortical disconnection. Some of the brain dysfunction is reversible by shunting, probably through restoration of cerebral blood flow and normalization of the extracellular environment. However, destroyed axons cannot be restored.

  3. Antibiotic resistance is linked to carriage of papC and iutA virulence genes and phylogenetic group D background in commensal and uropathogenic Escherichia coli from infants and young children.

    PubMed

    Karami, N; Wold, A E; Adlerberth, I

    2017-04-01

    P fimbriae, enabling adherence to colonic and urinary epithelium, and aerobactin, an iron sequestering system, are both colonization factors in the human colon and virulence factors for urinary tract infection. The colonic microbiota is suggested to be a site suitable for the transfer of antibiotic resistance genes. We investigated whether phenotypic resistance to antibiotics in commensal and uropathogenic Escherichia coli from infants and young children is associated with carriage of virulence genes and to phylogenetic group origin and, in the case of fecal strains, to persistence in the gut and fecal population levels. The commensal strains (n = 272) were derived from a birth cohort study, while the urinary isolates (n = 205) were derived from outpatient clinics. Each strain was assessed for phenotypic antibiotic resistance and for carriage of virulence genes (fimA, papC, sfaD/E, hlyA, iutA, kfiC, and neuB), phylogenetic group (A, B1, B2, or D), and markers of particular virulent clones (CGA-D-ST69, O15:H1-D-ST393, and O25b:H4-B2-ST131). Resistance to ampicillin, tetracycline, and trimethoprim was most prevalent. Multivariate analysis showed that resistance to any antibiotic was significantly associated with carriage of genes encoding P fimbriae (papC) and aerobactin (iutA), and a phylogenetic group D origin. Neither fecal population numbers nor the capacity for long-term persistence in the gut were related to antibiotic resistance among fecal strains. Our study confirms the importance of phylogenetic group D origin for antibiotic resistance in E. coli and identifies the virulence genes papC and iutA as determinants of antibiotic resistance. The reason for the latter association is currently unclear.

  4. Sequencing of IncX-plasmids suggests ubiquity of mobile forms of a biofilm-promoting gene cassette recruited from Klebsiella pneumoniae.

    PubMed

    Burmølle, Mette; Norman, Anders; Sørensen, Søren J; Hansen, Lars Hestbjerg

    2012-01-01

    Plasmids are a highly effective means with which genetic traits that influence human health, such as virulence and antibiotic resistance, are disseminated through bacterial populations. The IncX-family is a hitherto sparsely populated group of plasmids that are able to thrive within Enterobacteriaceae. In this study, a replicon-centric screening method was used to locate strains from wastewater sludge containing plasmids belonging to the IncX-family. A transposon aided plasmid capture method was then employed to transport IncX-plasmids from their original hosts (and co-hosted plasmids) into a laboratory strain (Escherichia coli Genehogs®) for further study. The nucleotide sequences of the three newly isolated IncX-plasmids (pLN126_33, pMO17_54, pMO440_54) and the hitherto un-sequenced type-plasmid R485 revealed a remarkable occurrence of whole or partial gene cassettes that promote biofilm-formation in Klebsiella pneumonia or E. coli, in all four instances. Two of the plasmids (R485 and pLN126_33) were shown to directly induce biofilm formation in a crystal violet retention assay in E. coli. Sequence comparison revealed that all plasmid-borne forms of the type 3 fimbriae encoding gene cassette mrkABCDF were variations of a composite transposon Tn6011 first described in the E. coli IncX plasmid pOLA52. In conclusion, IncX-plasmids isolated from Enterobacteriaceae over almost 40 years and on three different continents have all been shown to carry a type 3 fimbriae gene cassette mrkABCDF stemming from pathogenic K. pneumoniae. Apart from contributing general knowledge about IncX-plasmids, this study also suggests an apparent ubiquity of a mobile form of an important virulence factor and is an illuminating example of the recruitment, evolution and dissemination of genetic traits through plasmid-mediated horizontal gene transfer.

  5. Non-granule PSA-NCAM immunoreactive neurons in the rat hippocampus.

    PubMed

    Nacher, Juan; Blasco-Ibáñez, José M; McEwen, Bruce S

    2002-03-15

    The polysialylated form of the neural cell adhesion molecule (PSA-NCAM) continues to be expressed in the adult hippocampus, mainly in a subset of neurons located in the innermost portion of the granule cell layer. PSA-NCAM immunoreactive neurons have also been described outside this layer in humans, where they are severely reduced in schizophrenic brains. Given this important clinical implication, we were interested in finding whether similar neurons existed in the adult rat hippocampus and to characterize their distribution, morphology and phenotype. PSA-NCAM immunocytochemistry reveals labeled neurons in the subiculum, fimbria, alveus, hilus, and stratum oriens, lucidum and radiatum of CA3 and CA1. They are mainly distributed in the ventral hippocampus, and have polygonal or fusiform somata with multipolar or bipolar morphology. These neurons show long straight dendrites, which reach several strata and even enter the fimbria and the alveus. These dendrites are often varicose, appear devoid of excrescences and apparently do not show spines. Most of these neurons display GABA immunoreactivity and further analysis has shown that a subpopulation expresses calretinin, but not somatostatin, neuropeptide Y, parvalbumin, calbindin or NADPH diaphorase. Our study demonstrates that there is an important subpopulation of PSA-NCAM immunoreactive neurons, many of which can be considered interneurons, outside the rat granule cell layer, probably homologous to those described in the human hippocampus. The presence of the polysialylated form of NCAM in these neurons could indicate that they are undergoing continuous remodeling during adulthood and may have an important role in hippocampal structural plasticity.

  6. Chronological recognition by chicken of antigenic polypeptides in Salmonella enteritidis with different plasmid profiles: relationship to infection rate.

    PubMed

    Barbour, E K; El Jurdi, L H; Faroo, O M; Daghir, N J; Bouljihad, M

    2000-06-01

    The antigenic polypeptides in Salmonella Enteritidis (SE) were chronologically recognized by the chicken immune system, using Western immunoblotting. Broiler chicks challenged at three days of age with SE strain carrying the most prevalent plasmid profile of 14.1 and about (approximately) 50 kb were bled at 17, 24, 31, 38 and 45 days of age. Pooled sera of blood collected at each age was reacted by Western immunoblotting with banded polypeptides of three predominant SE strains that acquired the following respective plasmid profiles: 14.1 kb; 14.1 and approximately 50.0 kb; and 1.8, 14.1 and approximately 50.0 kb. The immunoblots of each pooled sera collected at a specific age against the three SE strains were similar. More specifically, the early immune response at 17 days of age had antibodies recognizing only one polypeptide in the three SE strains namely, the 35.8 kDa. At 24 or 31 days of age, the acquired immunity to infection had antibodies recognizing five similar polypeptides in the three SE strains namely, the 14.4 (fimbriae protein), 21.5 (fimbriae protein), 30.5, 35.8, and 66.2 kDa. At 38 and 45 days of age, the antibodies recognized additional polypeptides namely, the 41.5 and 55.6 kDa, respectively. The recognition of the 41.5 and 55.6 kDa polypeptides at 45 days of age was associated with higher invasiveness of SE to spleens and livers (15.6%) and in higher cecal colonization (59.4%) in comparison to absence of recognition to the two polypeptides at 31 days of age associated with low infectivity to spleens (0%), livers (3.1%), and ceca (9.4%).

  7. The exopolysaccharide gene cluster Bcam1330-Bcam1341 is involved in Burkholderia cenocepacia biofilm formation, and its expression is regulated by c-di-GMP and Bcam1349.

    PubMed

    Fazli, Mustafa; McCarthy, Yvonne; Givskov, Michael; Ryan, Robert P; Tolker-Nielsen, Tim

    2013-02-01

    In Burkholderia cenocepacia, the second messenger cyclic diguanosine monophosphate (c-di-GMP) has previously been shown to positively regulate biofilm formation and the expression of cellulose and type-I fimbriae genes through binding to the transcriptional regulator Bcam1349. Here, we provide evidence that cellulose and type-I fimbriae are not involved in B. cenocepacia biofilm formation in flow chambers, and we identify a novel Bcam1349/c-di-GMP-regulated exopolysaccharide gene cluster which is essential for B. cenocepacia biofilm formation. Overproduction of Bcam1349 in trans promotes wrinkly colony morphology, pellicle, and biofilm formation in B. cenocepacia. A screen for transposon mutants unable to respond to the overproduction of Bcam1349 led to the identification of a 12-gene cluster, Bcam1330-Bcam1341, the products of which appear to be involved in the production of a putative biofilm matrix exopolysaccharide and to be essential for flow-chamber biofilm formation. We demonstrate that Bcam1349 binds to the promoter region of genes in the Bcam1330-Bcam1341 cluster and that this binding is enhanced by the presence of c-di-GMP. Furthermore, we demonstrate that overproduction of both c-di-GMP and Bcam1349 leads to increased transcription of these genes, indicating that c-di-GMP and Bcam1349 functions together in regulating exopolysaccharide production from the Bcam1330-Bcam1341 gene cluster. Our results suggest that the product encoded by the Bcam1330-Bcam1341 gene cluster is a major exopolysaccharide that provides structural stability to the biofilms formed by B. cenocepacia, and that its production is regulated by c-di-GMP through binding to and promotion of the activity of the transcriptional regulator Bcam1349.

  8. Secondary Lymphoid Organ Homing Phenotype of Human Myeloid Dendritic Cells Disrupted by an Intracellular Oral Pathogen

    PubMed Central

    Miles, Brodie; Zakhary, Ibrahim; El-Awady, Ahmed; Scisci, Elizabeth; Carrion, Julio; O'Neill, John C.; Rawlings, Aaron; Stern, J. Kobi; Susin, Cristiano

    2014-01-01

    Several intracellular pathogens, including a key etiological agent of chronic periodontitis, Porphyromonas gingivalis, infect blood myeloid dendritic cells (mDCs). This infection results in pathogen dissemination to distant inflammatory sites (i.e., pathogen trafficking). The alteration in chemokine-chemokine receptor expression that contributes to this pathogen trafficking function, particularly toward sites of neovascularization in humans, is unclear. To investigate this, we utilized human monocyte-derived DCs (MoDCs) and primary endothelial cells in vitro, combined with ex vivo-isolated blood mDCs and serum from chronic periodontitis subjects and healthy controls. Our results, using conditional fimbria mutants of P. gingivalis, show that P. gingivalis infection of MoDCs induces an angiogenic migratory profile. This profile is enhanced by expression of DC-SIGN on MoDCs and minor mfa-1 fimbriae on P. gingivalis and is evidenced by robust upregulation of CXCR4, but not secondary lymphoid organ (SLO)-homing CCR7. This disruption of SLO-homing capacity in response to respective chemokines closely matches surface expression of CXCR4 and CCR7 and is consistent with directed MoDC migration through an endothelial monolayer. Ex vivo-isolated mDCs from the blood of chronic periodontitis subjects, but not healthy controls, expressed a similar migratory profile; moreover, sera from chronic periodontitis subjects expressed elevated levels of CXCL12. Overall, we conclude that P. gingivalis actively “commandeers” DCs by reprogramming the chemokine receptor profile, thus disrupting SLO homing, while driving migration toward inflammatory vascular sites. PMID:24126519

  9. Maternal Vaccination with a Fimbrial Tip Adhesin and Passive Protection of Neonatal Mice against Lethal Human Enterotoxigenic Escherichia coli Challenge

    PubMed Central

    Luiz, Wilson B.; Rodrigues, Juliana F.; Crabb, Joseph H.

    2015-01-01

    Globally, enterotoxigenic Escherichia coli (ETEC) is a leading cause of childhood and travelers' diarrhea, for which an effective vaccine is needed. Prevalent intestinal colonization factors (CFs) such as CFA/I fimbriae and heat-labile enterotoxin (LT) are important virulence factors and protective antigens. We tested the hypothesis that donor strand-complemented CfaE (dscCfaE), a stabilized form of the CFA/I fimbrial tip adhesin, is a protective antigen, using a lethal neonatal mouse ETEC challenge model and passive dam vaccination. For CFA/I-ETEC strain H10407, which has been extensively studied in volunteers, an inoculum of 2 × 107 bacteria resulted in 50% lethal doses (LD50) in neonatal DBA/2 mice. Vaccination of female DBA/2 mice with CFA/I fimbriae or dscCfaE, each given with a genetically attenuated LT adjuvant (LTK63) by intranasal or orogastric delivery, induced high antigen-specific serum IgG and fecal IgA titers and detectable milk IgA responses. Neonates born to and suckled by dams antenatally vaccinated with each of these four regimens showed 78 to 93% survival after a 20× LD50 challenge with H10407, compared to 100% mortality in pups from dams vaccinated with sham vaccine or LTK63 only. Crossover experiments showed that high pup survival rates after ETEC challenge were associated with suckling but not birthing from vaccinated dams, suggesting that vaccine-specific milk antibodies are protective. In corroboration, preincubation of the ETEC inoculum with antiadhesin and antifimbrial bovine colostral antibodies conferred a dose-dependent increase in pup survival after challenge. These findings indicate that the dscCfaE fimbrial tip adhesin serves as a protective passive vaccine antigen in this small animal model and merits further evaluation. PMID:26371126

  10. Metagenomic Analysis Revealed Methylamine and Ureide Utilization of Soybean-Associated Methylobacterium

    PubMed Central

    Minami, Tomoyuki; Anda, Misue; Mitsui, Hisayuki; Sugawara, Masayuki; Kaneko, Takakazu; Sato, Shusei; Ikeda, Seishi; Okubo, Takashi; Tsurumaru, Hirohito; Minamisawa, Kiwamu

    2016-01-01

    Methylobacterium inhabits the phyllosphere of a large number of plants. We herein report the results of comparative metagenome analyses on methylobacterial communities of soybean plants grown in an experimental field in Tohoku University (Kashimadai, Miyagi, Japan). Methylobacterium was identified as the most dominant genus (33%) among bacteria inhabiting soybean stems. We classified plant-derived Methylobacterium species into Groups I, II, and III based on 16S rRNA gene sequences, and found that Group I members (phylogenetically close to M. extorquens) were dominant in soybean-associated Methylobacterium. By comparing 29 genomes, we found that all Group I members possessed a complete set of genes for the N-methylglutamate pathway for methylamine utilization, and genes for urea degradation (urea carboxylase, urea amidolyase, and conventional urease). Only Group I members and soybean methylobacterial isolates grew in a culture supplemented with methylamine as the sole carbon source. They utilized urea or allantoin (a urea-related compound in legumes) as the sole nitrogen source; however, group III also utilized these compounds. The utilization of allantoin may be crucial in soybean-bacterial interactions because allantoin is a transported form of fixed nitrogen in legume plants. Soybean-derived Group I strain AMS5 colonized the model legume Lotus japonicus well. A comparison among the 29 genomes of plant-derived and other strains suggested that several candidate genes are involved in plant colonization such as csgG (curli fimbriae). Genes for the N-methylglutamate pathway and curli fimbriae were more abundant in soybean microbiomes than in rice microbiomes in the field. Based on these results, we discuss the lifestyle of Methylobacterium in the legume phyllosphere. PMID:27431374

  11. Metagenomic Analysis Revealed Methylamine and Ureide Utilization of Soybean-Associated Methylobacterium.

    PubMed

    Minami, Tomoyuki; Anda, Misue; Mitsui, Hisayuki; Sugawara, Masayuki; Kaneko, Takakazu; Sato, Shusei; Ikeda, Seishi; Okubo, Takashi; Tsurumaru, Hirohito; Minamisawa, Kiwamu

    2016-09-29

    Methylobacterium inhabits the phyllosphere of a large number of plants. We herein report the results of comparative metagenome analyses on methylobacterial communities of soybean plants grown in an experimental field in Tohoku University (Kashimadai, Miyagi, Japan). Methylobacterium was identified as the most dominant genus (33%) among bacteria inhabiting soybean stems. We classified plant-derived Methylobacterium species into Groups I, II, and III based on 16S rRNA gene sequences, and found that Group I members (phylogenetically close to M. extorquens) were dominant in soybean-associated Methylobacterium. By comparing 29 genomes, we found that all Group I members possessed a complete set of genes for the N-methylglutamate pathway for methylamine utilization, and genes for urea degradation (urea carboxylase, urea amidolyase, and conventional urease). Only Group I members and soybean methylobacterial isolates grew in a culture supplemented with methylamine as the sole carbon source. They utilized urea or allantoin (a urea-related compound in legumes) as the sole nitrogen source; however, group III also utilized these compounds. The utilization of allantoin may be crucial in soybean-bacterial interactions because allantoin is a transported form of fixed nitrogen in legume plants. Soybean-derived Group I strain AMS5 colonized the model legume Lotus japonicus well. A comparison among the 29 genomes of plant-derived and other strains suggested that several candidate genes are involved in plant colonization such as csgG (curli fimbriae). Genes for the N-methylglutamate pathway and curli fimbriae were more abundant in soybean microbiomes than in rice microbiomes in the field. Based on these results, we discuss the lifestyle of Methylobacterium in the legume phyllosphere.

  12. Changes in the oviducal epithelium during the estrous cycle in the marsupial Monodelphis domestica

    PubMed Central

    Kress, Annetrudi; Morson, Gianni

    2007-01-01

    The Monodelphis oviduct can be divided into four anatomical segments: preampulla (comprising fimbriae and infundibulum), ampulla, isthmus with crypts and uterotubal junction. Ovaries are enclosed in a periovarial sac, the bursa, and in some specimens tubules of an epoophoron could be identified. In both structures non-ciliated cells develop small translucent vesicles, which accumulate in the cell apices and presumably produce fluid as often seen in the bursa and in the tubules of the epooophoron. These vesicles do not stain with Alcian blue or PAS. The same applies also to the non-ciliated cells of the fimbriae. The oviducal epithelium of ampulla and the surface epithelium of the isthmus consisting of ciliated and non-ciliated, secretory cells undergo considerable changes during the estrous cycle. Proestrus shows low numbers of ciliated cells, some are in the process of neo-ciliogenesis, non-ciliated cells carry solitary cilia and few remnant secretory granules from the previous cycle may be found. At estrus the amount of ciliated cells in ampulla and isthmus has increased, most non-cililated cells lost the solitary cilia, developed longer microvilli and formed numerous secretory granules in their cell apices. At postestrus secretory products, often surrounded by membranes, are extruded into the oviducal lumen and contribute towards egg coat formation. First signs of deciliation processes are apparent. Solitary cilia reappear. At metestrus only few secretory cells are left with some secretory material. The lumen is often filled with shed cilia and cell apices. Proliferation of basal bodies within non-secretory cells indicate the formation of new ciliated cells. The non-ciliated epithelial cells of the isthmic crypts form no secretory granules but accumulate a great number of translucent vesicles, which in contrast to the secretory granules do not stain with Alcian blue or PAS. PMID:17883438

  13. Immunogenicity of a prototype enterotoxigenic Escherichia coli adhesin vaccine in mice and nonhuman primates.

    PubMed

    Sincock, Stephanie A; Hall, Eric R; Woods, Colleen M; O'Dowd, Aisling; Poole, Steven T; McVeigh, Annette L; Nunez, Gladys; Espinoza, Nereyda; Miller, Milagros; Savarino, Stephen J

    2016-01-04

    Enterotoxigenic Escherichia coli (ETEC) are the most common cause of bacterial diarrhea in young children in developing countries and in travelers. Efforts to develop an ETEC vaccine have intensified in the past decade, and intestinal colonization factors (CFs) are somatic components of most investigational vaccines. CFA/I and related Class 5 fimbrial CFs feature a major stalk-forming subunit and a minor, antigenically conserved tip adhesin. We hypothesized that the tip adhesin is critical for stimulating antibodies that specifically inhibit ETEC attachment to the small intestine. To address this, we compared the capacity of donor strand complemented CfaE (dscCfaE), a stabilized form of the CFA/I fimbrial tip adhesin, and CFA/I fimbriae to elicit anti-adhesive antibodies in mice, using hemagglutination inhibition (HAI) as proxy for neutralization of intestinal adhesion. When given with genetically attenuated heat-labile enterotoxin LTR192G as adjuvant by intranasal (IN) or orogastric (OG) vaccination, dscCfaE exceeded CFA/I fimbriae in eliciting serum HAI titers and anti-CfaE antibody titers. Based on these findings, we vaccinated Aotus nancymaae nonhuman primates (NHP) with dscCfaE alone or admixed with one of two adjuvants, LTR192G and cholera toxin B-subunit, by IN and OG administration. Only IN vaccination with dscCfaE with either adjuvant elicited substantial serum HAI titers and IgA and IgG anti-adhesin responses, with the latter detectable a year after vaccination. In conclusion, we have shown that dscCfaE elicits robust HAI and anti-adhesin antibody responses in both mice and NHPs when given with adjuvant by IN vaccination, encouraging further evaluation of an ETEC adhesin-based vaccine approach.

  14. Fur Represses Adhesion to, Invasion of, and Intracellular Bacterial Community Formation within Bladder Epithelial Cells and Motility in Uropathogenic Escherichia coli.

    PubMed

    Kurabayashi, Kumiko; Agata, Tomohiro; Asano, Hirofumi; Tomita, Haruyoshi; Hirakawa, Hidetada

    2016-11-01

    Uropathogenic Escherichia coli (UPEC) is a major pathogen that causes urinary tract infections (UTIs). This bacterium adheres to and invades the host cells in the bladder, where it forms biofilm-like polymicrobial structures termed intracellular bacterial communities (IBCs) that protect UPEC from antimicrobial agents and the host immune systems. Using genetic screening, we found that deletion of the fur gene, which encodes an iron-binding transcriptional repressor for iron uptake systems, elevated the expression of type I fimbriae and motility when UPEC was grown under iron-rich conditions, and it led to an increased number of UPEC cells adhering to and internalized in bladder epithelial cells. Consequently, the IBC colonies that the fur mutant formed in host cells were denser and larger than those formed by the wild-type parent strain. Fur is inactivated under iron-restricted conditions. When iron was depleted from the bacterial cultures, wild-type UPEC adhesion, invasion, and motility increased, similar to the case with the fur mutant. The purified Fur protein bound to regions upstream of fimA and flhD, which encode type I fimbriae and an activator of flagellar expression that contributes to motility, respectively. These results suggest that Fur is a repressor of fimA and flhD and that its repression is abolished under iron-depleted conditions. Based on our in vitro experiments, we conclude that UPEC adhesion, invasion, IBC formation, and motility are suppressed by Fur under iron-rich conditions but derepressed under iron-restricted conditions, such as in patients with UTIs.

  15. The flagella of F18ab Escherichia coli is a virulence factor that contributes to infection in a IPEC-J2 cell model in vitro.

    PubMed

    Duan, Qiangde; Zhou, Mingxu; Zhu, Xiaofang; Bao, Wenbin; Wu, Shenglong; Ruan, Xiaosai; Zhang, Weiping; Yang, Yang; Zhu, Jun; Zhu, Guoqiang

    2012-11-09

    Bacterial flagella contribute to pathogen virulence; however, the role of flagella in the pathogenesis of F18ab E. coli-mediated swine edema disease (ED) is not currently known. We therefore evaluated the role of flagella in F18ab E. coli adhesion, invasion, biofilm formation, and IL-8 production using an in vitro cell infection model approach with gene-deletion mutant and complemented bacterial strains. We demonstrated that the flagellin-deficient fliC mutant had a marked decrease in the ability to adhere to and invade porcine epithelial IPEC-J2 cells. Surprisingly, there was no difference in adhesion between the F18 fimbriae-deficient ΔfedA mutant and its parent strain. In addition, both the ΔfedA and double ΔfliCΔfedA mutants exhibited an increased ability to invade IPEC-J2 cells compared to the wild-type strain, although this may be due to increased expression of other adhesins following the loss of F18ab fimbriae and flagella. Compared to the wild-type strain, the ΔfliC mutant showed significantly reduced ability to form biofilm, whereas the ΔfedA mutant increased biofilm formation. Although ΔfliC, ΔfedA, and ΔfliCΔfedA mutants had a reduced ability to stimulate IL-8 production from infected Caco-2 cells, the ΔfliC mutant impaired this ability to a greater extent than the ΔfedA mutant. The results from this study clearly demonstrate that flagella are required for efficient F18ab E. coli adhesion, invasion, biofilm formation, and IL-8 production in vitro.

  16. Changes in the oviducal epithelium during the estrous cycle in the marsupial Monodelphis domestica.

    PubMed

    Kress, Annetrudi; Morson, Gianni

    2007-10-01

    The Monodelphis oviduct can be divided into four anatomical segments: preampulla (comprising fimbriae and infundibulum), ampulla, isthmus with crypts and uterotubal junction. Ovaries are enclosed in a periovarial sac, the bursa, and in some specimens tubules of an epoophoron could be identified. In both structures non-ciliated cells develop small translucent vesicles, which accumulate in the cell apices and presumably produce fluid as often seen in the bursa and in the tubules of the epooophoron. These vesicles do not stain with Alcian blue or PAS. The same applies also to the non-ciliated cells of the fimbriae. The oviducal epithelium of ampulla and the surface epithelium of the isthmus consisting of ciliated and non-ciliated, secretory cells undergo considerable changes during the estrous cycle. Proestrus shows low numbers of ciliated cells, some are in the process of neo-ciliogenesis, non-ciliated cells carry solitary cilia and few remnant secretory granules from the previous cycle may be found. At estrus the amount of ciliated cells in ampulla and isthmus has increased, most non-cililated cells lost the solitary cilia, developed longer microvilli and formed numerous secretory granules in their cell apices. At postestrus secretory products, often surrounded by membranes, are extruded into the oviducal lumen and contribute towards egg coat formation. First signs of deciliation processes are apparent. Solitary cilia reappear. At metestrus only few secretory cells are left with some secretory material. The lumen is often filled with shed cilia and cell apices. Proliferation of basal bodies within non-secretory cells indicate the formation of new ciliated cells. The non-ciliated epithelial cells of the isthmic crypts form no secretory granules but accumulate a great number of translucent vesicles, which in contrast to the secretory granules do not stain with Alcian blue or PAS.

  17. Salmonella can reach tomato fruits on plants exposed to aerosols formed by rain.

    PubMed

    Cevallos-Cevallos, Juan M; Gu, Ganyu; Danyluk, Michelle D; Dufault, Nicholas S; van Bruggen, Ariena H C

    2012-08-17

    Outbreaks of Salmonella enterica have been associated with tomatoes and traced back to production areas but the spread of Salmonella in agricultural fields is still poorly understood. Post-rain Salmonella transfer from a point source to the air and then to tomato plants was evaluated. GFP-labeled kanamycin-resistant S. enterica serovar Typhimurium (10(8)CFU/mL) with and without expression of the rdar morphotype (rough colonies; cells with fimbriae and cellulose) was used as the point source in the center of a rain simulator. Rain intensities of 60 and 110 mm/h were applied for 5, 10, 20, and 30 min. Petri dishes with lactose broth and tomato plants with fruit (50-80 cm high) were placed in the simulator after the rain had ceased. Salmonella recovery from air was maximum (300 CFU/plate) after a rain episode of 60 mm/h for 10 min at distances of at least 85.5 cm above the source and when the rdar morphotype strain was used. Small scale experiments showed that the smooth-colony strain without fimbriae precipitated from the air in significantly higher numbers than the rdar strain. Transfer of aerial Salmonella with the rdar morphotype to tomato fruits on plants followed a beta distribution (2.5950, 4.7393) within the generalized range from 0 to 30 min of rain. Results show for the first time that Salmonella may transfer from rain to the air and contaminate tomato fruits at levels that could possibly be infectious to humans.

  18. Characterization of monoclonal antibodies against putative colonization factors of enterotoxigenic Escherichia coli and their use in an epidemiological study.

    PubMed Central

    Viboud, G I; Binsztein, N; Svennerholm, A M

    1993-01-01

    Monoclonal antibodies (MAbs) against five putative colonization factors (PCFs), i.e., colonization factor antigen (CFA)/III, coli surface antigen (CS)7 and CS17, PCFO159, and PCFO166 of enterotoxigenic Escherichia coli (ETEC), were produced. Hybridomas (one each) producing specific antibodies against the respective PCFs were selected. All the MAbs reacted with the corresponding fimbriae but not with CFA/I, CFA/II, or CFA/IV or the heterologous PCFs in bacterial agglutination and enzyme-linked immunosorbent assays (ELISAs). In immunoelectron microscopy these MAbs bound along the fimbriae, and they also reacted with the corresponding subunits in immunoblots. The five MAbs were used to evaluate the prevalence of CFA/III, CS7, CS17, PCFO159, and PCFO166 in ETEC strains isolated from children with diarrhea in Argentina. One hundred five ETEC isolates negative for CFA/I, CFA/II, and CFA/IV were tested in slide agglutination or in a dot blot test for spontaneously agglutinating strains; positive results were confirmed by inhibition ELISAs. It was found that 27% of the CFA-negative ETEC strains carried one of the PCFs. The sensitivity of slide agglutination with these MAbs was similar to that with specific polyclonal antisera; however, the specificity was higher. PCFO166 was found in 9.5% of the strains tested, mainly in ETEC of serogroup O78 producing heat-stable toxin alone. CS17 and CS7 were identified in 6.7 and 5.7%, respectively, of strains producing heat-labile toxin only, most of which belonged to serogroup O114. PCFO159 was found in 3.8% of the isolates tested, whereas CFA/III was detected in only one ETEC strain. Images PMID:8096215

  19. Avirulent K88 (F4)+ Escherichia coli strains constructed to express modified enterotoxins protect young piglets from challenge with a virulent enterotoxigenic Escherichia coli strain that expresses the same adhesion and enterotoxins.

    PubMed

    Santiago-Mateo, Kristina; Zhao, Mojun; Lin, Jun; Zhang, Weiping; Francis, David H

    2012-10-12

    Virulence of enterotoxigenic Escherichia coli (ETEC) is associated with fimbrial adhesins and enterotoxins such as heat-labile (LT) and/or heat-stable (ST) enterotoxins. Previous studies using a cell culture model suggest that exclusion of ETEC from attachment to epithelial cells requires expression of both an adhesin such as K88 (F4) fimbriae, and LT. To test the ability of non-pathogenic E. coli constructs to exclude virulent ETEC sufficiently to prevent clinical disease, we utilized a piglet ETEC challenge model. Thirty-nine 5-day-old piglets were inoculated with a placebo (control), or with either of the three K88(+)E. coli strains isogenic with regard to modified LT expression: 8017 (pBR322 plasmid vector control), non-toxigenic mutant 8221 (LT(R192G)) in pBR322, or 8488, with the LT gene fused to the STb gene in pBR322 (LT(R192G)-STb). Piglets were challenged with virulent ETEC Strain 3030-2 (K88(+)/LT/STb) 24h post-inoculation. K88ac receptor-positive piglets in the control group developed diarrhea and became dehydrated 12-24h post-challenge. Piglets inoculated with 8221 or 8488 did not exhibit clinical signs of ETEC disease; most piglets inoculated with 8017 showed diarrhea. Control pigs exhibited significant weight loss, increased blood total protein, and higher numbers of colony-forming units of 3030-2 E. coli in washed ileum and jejunum than treated pigs. This study shows for the first time that pre-inoculation with an avirulent strain expressing adhesive fimbriae and a non-toxic form of LT provides significant short term protection from challenge with a virulent ETEC strain that expresses the same fimbrial adhesion and enterotoxin.

  20. Incidental nonuterine high-grade serous carcinomas arise in the fallopian tube in most cases: further evidence for the tubal origin of high-grade serous carcinomas.

    PubMed

    Gilks, C Blake; Irving, Julie; Köbel, Martin; Lee, Chenghan; Singh, Naveena; Wilkinson, Nafisa; McCluggage, W Glenn

    2015-03-01

    Most nonuterine high-grade serous carcinomas (HGSCs) in women with hereditary breast and ovarian cancer syndrome, due to germline BRCA1/2 mutation, arise in the fimbria of the fallopian tube. However, the site of origin of sporadic HGSC, which is usually widely disseminated at presentation, is not well established. We sought to characterize cases of HGSC discovered incidentally in patients not known to be at high risk, in order to determine the site distribution and possible origin of sporadic HGSC. Incidental microscopic, non-mass-forming cases of serous tubal intraepithelial carcinoma or HGSC in salpingo-oophorectomy specimens in which the tubes and ovaries had been extensively examined were identified. No patients were known or suspected BRCA1/2 mutation carriers. Twenty-one cases were identified (mean age: 57 y). Surgery was for benign disease (n=15), uterine endometrioid adenocarcinoma or atypical hyperplasia (n=3), bladder carcinoma (n=1), or ovarian serous borderline tumor (n=2). In 16 of 21 cases, the lesion was confined to the fallopian tube (unilateral in 14 cases, bilateral in 2). There was serous tubal intraepithelial carcinoma in all cases and invasive HGSC into the underlying lamina propria in 8 of these 16 cases; the invasive focus measured 1.3 cm or less in every case. In the remaining 5 cases, there was fallopian tube mucosal and ovarian involvement; in 2 of these cases, there was also microscopic peritoneal involvement. Sporadic cases of nonuterine HGSC arise in the fallopian tube fimbria in a large majority of cases, providing further evidence for the tubal origin of these neoplasms.

  1. A Comparison of Diets Supplemented with a Feed Additive Containing Organic Acids, Cinnamaldehyde and a Permeabilizing Complex, or Zinc Oxide, on Post-Weaning Diarrhoea, Selected Bacterial Populations, Blood Measures and Performance in Weaned Pigs Experimentally Infected with Enterotoxigenic E. coli.

    PubMed

    Stensland, Ingunn; Kim, Jae Cheol; Bowring, Bethany; Collins, Alison M; Mansfield, Josephine P; Pluske, John R

    2015-11-23

    The effects of feeding a diet supplemented with zinc oxide (ZnO) or a blend of organic acids, cinnamaldehyde and a permeabilizing complex (OACP) on post-weaning diarrhoea (PWD) and performance in pigs infected with enterotoxigenic E. coli (ETEC) were examined. Additionally, changes in selected bacterial populations and blood measures were assessed. A total of 72 pigs weaned at 22 d of age and weighing 7.2 ± 1.02 kg (mean ± SEM) was used. Treatments were: base diet (no antimicrobial compounds); base diet + 3 g ZnO/kg; base diet + 1.5 g OACP/kg. Dietary treatments started on the day of weaning and were fed ad libitum for 3 weeks. All pigs were infected with an F4 ETEC on d 4, 5 and 6 after weaning. The incidence of PWD was lower in pigs fed ZnO ( p = 0.026). Overall, pigs fed ZnO grew faster ( p = 0.013) and ate more ( p = 0.004) than the base diet-fed pigs, with OACP-fed pigs performing the same ( p > 0.05) as both the ZnO- and base diet-fed pigs. Feed conversion ratio was similar for all diets ( p > 0.05). The percentage of E. coli with F4 fimbriae was affected a day by treatment interaction ( p = 0.037), with more E. coli with F4 fimbriae found in pigs fed ZnO on d 11 ( p = 0.011) compared to base diet-fed pigs. Only significant time effects ( p < 0.05) occurred for blood measures. Under the conditions of this study, inclusion of OACP gave statistically similar production responses to pigs fed ZnO, however pigs fed ZnO had less PWD compared to OACP- and the base diet-fed pigs.

  2. Putative glycoprotein and glycolipid polymorphonuclear leukocyte receptors for the Actinomyces naeslundii WVU45 fimbrial lectin.

    PubMed Central

    Sandberg, A L; Ruhl, S; Joralmon, R A; Brennan, M J; Sutphin, M J; Cisar, J O

    1995-01-01

    Recognition of receptors on sialidase-treated polymorphonuclear leukocytes (PMNs) by the Gal/GalNAc lectin associated with the type 2 fimbriae of certain strains of actinomyces results in activation of the PMNs, phagocytosis, and destruction of the bacteria. In the present study, plant lectins were utilized as probes to identify putative PMN receptors for the actinomyces lectin. The Gal-reactive lectin from Ricinus communis (RCAI), the Gal/GalNAc-reactive lectins from R. communis (RCAII) and Bauhinia purpurea (BPA), as well as the Gal beta 1-3GalNAc-specific lectins from Arachis hypogaea (PNA) and Agaricus bisporus (ABA) inhibited killing of Actinomyces naeslundii WVU45 by sialidase-treated PMNs. These five lectins detected a 130-kDa surface-labeled glycoprotein on nitrocellulose transfers of PMN extracts separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This glycoprotein was revealed only after treatment of the transfers with sialidase, a condition analogous to the sialidase dependence of the lectin-mediated biological responses of the PMNs to the actinomyces. The mannose-reactive lectin concanavalin A did not inhibit killing of the actinomyces and failed to detect the 130-kDa glycoprotein but did block PMN-dependent killing of Escherichia coli B, a bacterium that possesses mannose-sensitive fimbriae. Therefore, the PMN glycoprotein receptor for A. naeslundii is clearly distinct from those recognized by E. coli. Two major putative glycolipid receptors were also identified by actinomyces and RCAI overlays on sialidase-treated thin-layer chromatograms of PMN gangliosides. Thus, both a 130-kDa glycoprotein and certain gangliosides are implicated in the attachment of the actinomyces to PMNs. PMID:7790078

  3. Adhesion of Type 1-Fimbriated Escherichia coli to Abiotic Surfaces Leads to Altered Composition of Outer Membrane Proteins

    PubMed Central

    Otto, Karen; Norbeck, Joakim; Larsson, Thomas; Karlsson, Karl-Anders; Hermansson, Malte

    2001-01-01

    Phenotypic differences between planktonic bacteria and those attached to abiotic surfaces exist, but the mechanisms involved in the adhesion response of bacteria are not well understood. By the use of two-dimensional (2D) polyacrylamide gel electrophoresis, we have demonstrated that attachment of Escherichia coli to abiotic surfaces leads to alteration in the composition of outer membrane proteins. A major decrease in the abundance of resolved proteins was observed during adhesion of type 1-fimbriated E. coli strains, which was at least partly caused by proteolysis. Moreover, a study of fimbriated and nonfimbriated mutants revealed that these changes were due mainly to type 1 fimbria-mediated surface contact and that only a few changes occurred in the outer membranes of nonfimbriated mutant strains. Protein synthesis and proteolytic degradation were involved to different extents in adhesion of fimbriated and nonfimbriated cells. While protein synthesis appeared to affect adhesion of only the nonfimbriated strain, proteolytic activity mostly seemed to contribute to adhesion of the fimbriated strain. Using matrix-assisted laser desorption ionization–time of flight mass spectrometry, six of the proteins resolved by 2D analysis were identified as BtuB, EF-Tu, OmpA, OmpX, Slp, and TolC. While the first two proteins were unaffected by adhesion, the levels of the last four were moderately to strongly reduced. Based on the present results, it may be suggested that physical interactions between type 1 fimbriae and the surface are part of a surface-sensing mechanism in which protein turnover may contribute to the observed change in composition of outer membrane proteins. This change alters the surface characteristics of the cell envelope and may thus influence adhesion. PMID:11274103

  4. The Role of Host Factors and Bacterial Virulence Genes in the Development of Pyelonephritis Caused by Escherichia coli in Renal Transplant Recipients

    PubMed Central

    Siliano, Priscila Reina; Rocha, Lillian Andrade; Osmar Medina-Pestana, José

    2010-01-01

    Background and objectives: The aim of this study was to determine the role of host factors and bacterial virulence genes in the development of pyelonephritis caused by Escherichia coli in renal transplant (Tx) recipients. Design, setting, participants, & measurements: A total of 328 E. coli isolates from cases of cystitis (Cys; n = 239) or pyelonephritis (PN; n = 89), with 169 from renal Tx recipients, were subjected to molecular analyses to identify P-fimbria subunits (PapC, PapG II, and PapGIII), G- and M-fimbriae, and aerobactin. The presence of antibiotic resistance was also determined. Parameters such as gender, age, immunosuppression regimens, causes of ESRD, kidney donor, intraoperative anastomosis, use of double J stent, trimethoprim/sulfamethoxazole (TMP/SMZ) prophylaxis, and time after Tx were evaluated. Results: A multivariate analysis showed a significant association between PN and renal Tx. In renal Tx recipients, the risk of occurrence of PN was significantly higher among males and for those no longer receiving TMP/SMZ prophylaxis. E. coli strains isolated from PN presented a lower prevalence of papGIII and lower rates of resistance to pipemidic acid. Although papGII was more prevalent in PN than in Cys, it was not independently associated with PN. Conclusions: These findings suggested that renal Tx increases the risk for PN, and the male sex represented a host factor independently associated with risk, whereas the prophylaxis with TMP/SMZ was protective. The lack of papGIII and low resistance to first-generation quinolones were bacterial-independent risk factors for PN in Tx. PMID:20448070

  5. Effectiveness of F18+ Fimbrial Antigens Released by a Novel Autolyzed Salmonella Expression System as a Vaccine Candidate against Lethal F18+ STEC Infection

    PubMed Central

    Won, Gayeon; Lee, John H.

    2016-01-01

    Porcine edema disease (ED) caused by Shiga toxin 2e producing Escherichia coli expressing F18ab+ fimbriae (F18ab+STEC) frequently occurs in post-weaned piglets, resulting in a significant economic loss in swine industries worldwide. In the present study, we proposed an efficient prevention scheme against ED in which the attenuated Salmonella Typhimurium inactivated by the E-mediated cell lysis to deliver target antigens, FedF and FedA, which function in fimbrial-mediated adhesion and as a major subunit of F18ab+fimbriae, respectively. The co-expression of FedA and FedF protein with outer membrane protein A signal peptide was confirmed in the resultant strains JOL1460 and JOL1464 by immunoblot analysis. Immunization with the candidate strains in mice led to the significant generation of immunoglobulin (Ig) G, specific to both antigens and secretory IgA specific to FedF (P < 0.05). The titers of IgG isotypes, IgG1 and IgG2a, used as markers for T-helpers (Th)-2 and Th-1lymphocytes, respectively, also significantly increased in the immunized group (P < 0.05). The increase in CD3+CD4+ T lymphocyte subpopulation and in vitro proliferative activity was observed in in vivo stimulated splenocytes, which indicated the immunostimulatory effect of the candidate strains. Moreover, the immunized mice were completely protected from a lethal challenge against wild-type F18+STEC whereas 28% of mice died in the non-immunized group. This study demonstrated that the inactivated Salmonella system could efficiently release FedF and FedA and induce robust immune responses specific to the target antigens, which is sufficient to protect the mice from the lethal challenge. PMID:27920758

  6. Fallopian tube secretory cell expansion: a sensitive biomarker for ovarian serous carcinogenesis.

    PubMed

    Wang, Yiying; Li, Li; Wang, Yue; Tang, Sarah Ngocvi; Zheng, Wenxin

    2015-01-01

    Recent advances suggest that precancerous lesions of pelvic serous carcinoma originate from tubal secretory cells. The purpose of our study was to determine if an increased number of secretory cells varies with age or location in the fallopian tube and to examine its association with serous neoplasia. Three groups (benign control, high-risk, and pelvic serous carcinoma) of age-matched patients were studied. The age data were stratified into 10-year intervals ranging from 20-29 to older than 80. The number of secretory and ciliated cells from both tubal fimbria and ampulla segments was counted by microscopy and immunohistochemical staining methods. The data were analyzed by standard contingency table and Poisson distribution methods after age justification. We found that the absolute number of tubal secretory cells increased significantly with age in all three groups. But a more dramatic increase of secretory cells was observed in high-risk and pelvic serous carcinoma patients. Secretory cell expansion is more prevalent than secretory cell outgrowth in both fimbria and ampulla tubal segments and is significantly associated with serous neoplasia (P < 0.001). Furthermore, age remained a significant risk factor for serous neoplasia after age adjustment. These findings suggest that secretory cell expansion could serve as a potential sensitive biomarker for early serous carcinogenesis within the fallopian tube. The study also supports a relationship between serous neoplasia and increased secretory to ciliated cell ratios, and the relationship between frequency of secretory cell expansion within the fallopian tube and increasing age and-more significantly-presence of high-risk factors or co-existing serous cancers.

  7. Fallopian tube secretory cell expansion: a sensitive biomarker for ovarian serous carcinogenesis.

    PubMed

    Wang, Yiying; Li, Li; Wang, Yue; Tang, Sarah Ngocvi; Zheng, Wenxin

    2016-01-01

    Recent advances suggest that precancerous lesions of pelvic serous carcinoma originate from tubal secretory cells. The purpose of our study was to determine if an increased number of secretory cells vary with age or location in the fallopian tube and to examine its association with serous neoplasia. Three groups (benign control, high-risk, and pelvic serous carcinoma) of age-matched patients were studied. The age data were stratified into 10-year intervals ranging from 20-29 to older than 80. The number of secretory and ciliated cells from both tubal fimbria and ampulla segments was counted by microscopy and immunohistochemical staining methods. The data were analyzed by standard contingency table and Poisson distribution methods after age justification. We found that the absolute number of tubal secretory cells increased significantly with age in all three groups. But a more dramatic increase of secretory cells was observed in high-risk and pelvic serous carcinoma patients. Secretory cell expansion is more prevalent than secretory cell outgrowth in both fimbria and ampulla tubal segments and is significantly associated with serous neoplasia (p < 0.001). Furthermore, age remained a significant risk factor for serous neoplasia after age adjustment. These findings suggest that secretory cell expansion could serve as a potential sensitive biomarker for early serous carcinogenesis within the fallopian tube. The study also supports a relationship between serous neoplasia and increased secretory to ciliated cell ratios, and the relationship between frequency of secretory cell expansion within the fallopian tube and increasing age and-more significantly-presence of high-risk factors or co-existing serous cancers.

  8. RcsB contributes to the distinct stress fitness among Escherichia coli O157:H7 curli variants of the 1993 hamburger-associated outbreak strains.

    PubMed

    Carter, Michelle Q; Parker, Craig T; Louie, Jacqueline W; Huynh, Steven; Fagerquist, Clifton K; Mandrell, Robert E

    2012-11-01

    Curli are adhesive fimbriae of Enterobactericaeae and are involved in surface attachment, cell aggregation, and biofilm formation. We reported previously that curli-producing (C(+)) variants of E. coli O157:H7 (EcO157) were much more acid sensitive than their corresponding curli-deficient (C(-)) variants; however, this difference was not linked to the curli fimbriae per se. Here, we investigated the underlying molecular basis of this phenotypic divergence. We identified large deletions in the rcsB gene of C(+) variants isolated from the 1993 U.S. hamburger-associated outbreak strains. rcsB encodes the response regulator of the RcsCDB two-component signal transduction system, which regulates curli biogenesis negatively but acid resistance positively. Further comparison of stress fitness revealed that C(+) variants were also significantly more sensitive to heat shock but were resistant to osmotic stress and oxidative damage, similar to C(-) variants. Transcriptomics analysis uncovered a large number of differentially expressed genes between the curli variants, characterized by enhanced expression in C(+) variants of genes related to biofilm formation, virulence, catabolic activity, and nutrient uptake but marked decreases in transcription of genes related to various types of stress resistance. Supplying C(+) variants with a functional rcsB restored resistance to heat shock and acid challenge in cells but blocked curli production, confirming that inactivation of RcsB in C(+) variants was the basis of fitness segregation within the EcO157 population. This study provides an example of how genome instability of EcO157 promotes intrapopulation diversification, generating subpopulations carrying an array of distinct phenotypes that may confer the pathogen with survival advantages in diverse environments.

  9. Genetic exchange of fimbrial alleles exemplifies the adaptive virulence strategy of Porphyromonas gingivalis.

    PubMed

    Kerr, Jennifer E; Abramian, Jared R; Dao, Doan-Hieu V; Rigney, Todd W; Fritz, Jamie; Pham, Tan; Gay, Isabel; Parthasarathy, Kavitha; Wang, Bing-yan; Zhang, Wenjian; Tribble, Gena D

    2014-01-01

    Porphyromonas gingivalis is a gram-negative anaerobic bacterium, a member of the human oral microbiome, and a proposed "keystone" pathogen in the development of chronic periodontitis, an inflammatory disease of the gingiva. P. gingivalis is a genetically diverse species, and is able to exchange chromosomal DNA between strains by natural competence and conjugation. In this study, we investigate the role of horizontal DNA transfer as an adaptive process to modify behavior, using the major fimbriae as our model system, due to their critical role in mediating interactions with the host environment. We show that P. gingivalis is able to exchange fimbrial allele types I and IV into four distinct strain backgrounds via natural competence. In all recombinants, we detected a complete exchange of the entire fimA allele, and the rate of exchange varies between the different strain backgrounds. In addition, gene exchange within other regions of the fimbrial genetic locus was identified. To measure the biological implications of these allele swaps we compared three genotypes of fimA in an isogenic background, strain ATCC 33277. We demonstrate that exchange of fimbrial allele type results in profound phenotypic changes, including the quantity of fimbriae elaborated, membrane blebbing, auto-aggregation and other virulence-associated phenotypes. Replacement of the type I allele with either the type III or IV allele resulted in increased invasion of gingival fibroblast cells relative to the isogenic parent strain. While genetic variability is known to impact host-microbiome interactions, this is the first study to quantitatively assess the adaptive effect of exchanging genes within the pan genome cloud. This is significant as it presents a potential mechanism by which opportunistic pathogens may acquire the traits necessary to modify host-microbial interactions.

  10. Genetic Exchange of Fimbrial Alleles Exemplifies the Adaptive Virulence Strategy of Porphyromonas gingivalis

    PubMed Central

    Kerr, Jennifer E.; Abramian, Jared R.; Dao, Doan-Hieu V.; Rigney, Todd W.; Fritz, Jamie; Pham, Tan; Gay, Isabel; Parthasarathy, Kavitha; Wang, Bing-yan; Zhang, Wenjian; Tribble, Gena D.

    2014-01-01

    Porphyromonas gingivalis is a gram–negative anaerobic bacterium, a member of the human oral microbiome, and a proposed “keystone” pathogen in the development of chronic periodontitis, an inflammatory disease of the gingiva. P. gingivalis is a genetically diverse species, and is able to exchange chromosomal DNA between strains by natural competence and conjugation. In this study, we investigate the role of horizontal DNA transfer as an adaptive process to modify behavior, using the major fimbriae as our model system, due to their critical role in mediating interactions with the host environment. We show that P. gingivalis is able to exchange fimbrial allele types I and IV into four distinct strain backgrounds via natural competence. In all recombinants, we detected a complete exchange of the entire fimA allele, and the rate of exchange varies between the different strain backgrounds. In addition, gene exchange within other regions of the fimbrial genetic locus was identified. To measure the biological implications of these allele swaps we compared three genotypes of fimA in an isogenic background, strain ATCC 33277. We demonstrate that exchange of fimbrial allele type results in profound phenotypic changes, including the quantity of fimbriae elaborated, membrane blebbing, auto-aggregation and other virulence-associated phenotypes. Replacement of the type I allele with either the type III or IV allele resulted in increased invasion of gingival fibroblast cells relative to the isogenic parent strain. While genetic variability is known to impact host-microbiome interactions, this is the first study to quantitatively assess the adaptive effect of exchanging genes within the pan genome cloud. This is significant as it presents a potential mechanism by which opportunistic pathogens may acquire the traits necessary to modify host-microbial interactions. PMID:24626479

  11. Highly Differentiated Human Airway Epithelial Cells: a Model to Study Host cell-parasite Interactions in Pertussis

    PubMed Central

    Guevara, Claudia; Zhang, Chengxian; Gaddy, Jennifer A.; Iqbal, Junaid; Guerra, Julio; Greenberg, David P.; Decker, Michael D.; Carbonetti, Nicholas; Starner, Timothy D.; McCray, Paul B.; Mooi, Frits R.

    2017-01-01

    Background Bordetella pertussis colonizes the human respiratory mucosa. Most studies on B. pertussis adherence have relied on cultured mammalian cells that lack key features present in differentiated human airway cells or on animal models that are not natural hosts of B. pertussis. The objectives of this work are to evaluate B. pertussis infection on highly differentiated human airway cells in vitro and to show the role of B. pertussis fimbriae in cell adherence. Methods Primary human airway epithelial (PHAE) cells from human bronchi and a human bronchial epithelial (HBE) cell line were grown in vitro under air-liquid interface conditions. Results PHAE and HBE cells infected with B. pertussis wild type strain revealed bacterial adherence to cell’s apical surface and bacterial induced cytoskeleton changes and cell detachment. Mutations in the major fimbrial subunits Fim2/3 or in the minor fimbrial adhesin subunit FimD affected B. pertussis adherence to predominantly HBE cells. This cell model recapitulates the morphologic features of the human airway infected by B. pertussis and confirms the role of fimbriae in B. pertussis adherence. Furthemore, HBE cells show that fimbrial subunits, and specifically FimD adhesin, are critical in B. pertussis adherence to airway cells. Conclusions The relevance of this model to study host-parasite interaction in pertussis lies in the striking physiologic and morphologic similarity between the PHAE and HBE cells and the human airway ciliated and goblet cells in vivo. These cells can proliferate in vitro, differentiate, and express the same genetic profile as human respiratory cells in vivo. PMID:26492208

  12. Identification and Characterization of Heparin Binding Regions of the Fim2 Subunit of Bordetella pertussis

    PubMed Central

    Geuijen, Cecile A. W.; Willems, Rob J. L.; Hoogerhout, Peter; Puijk, Wouter C.; Meloen, Rob H.; Mooi, Frits R.

    1998-01-01

    Bordetella pertussis fimbriae bind to sulfated sugars such as heparin through the major subunit Fim2. The Fim2 subunit contains two regions, designated H1 and H2, which show sequence similarity with heparin binding regions of fibronectin, and the role of these regions in heparin binding was investigated with maltose binding protein (MBP)-Fim2 fusion proteins. Deletion derivatives of MBP-Fim2 showed that both regions are important for binding to heparin. The role of H2 in heparin binding was confirmed by site-directed mutagenesis in which basic amino acids were replaced by alanine. These studies revealed that Lys-186 and Lys-187 are important for heparin binding of MBP-Fim2, whereas Arg-179 is not required. Peptides derived from H1 and H2 (pepH1 and pepH2) also showed heparin binding activity. Using a series of peptides, in each of which a different basic amino acid was substituted for alanine, we demonstrated that the structural requirements for heparin binding differ significantly among pepH1 and pepH2 peptides. A Pepscan analysis of Fim2 revealed regions outside H1 and H2 which bind heparin and showed that not only basic amino acids but also tyrosines may be important for binding to sulfated sugars. A comparison of the heparin binding regions of Fim2 with homologous regions of Fim3 and FimX, two closely related but antigenically distinct fimbrial subunits, showed that basic amino acids and tyrosines are generally conserved. The major heparin binding regions identified in Fim2 are part of epitopes recognized by human antibodies, suggesting that the heparin binding regions are exposed at the fimbrial surface and are immunodominant. Since B. pertussis fimbriae show weak serological cross-reactivity, the differences in primary structure in the heparin binding regions of Fim2, Fim3, and FimX may affect antibody binding but not heparin binding, allowing the bacteria to evade antibody-mediated immunity by switching the fimbrial gene expressed. PMID:9573115

  13. Expression of Pseudomonas aeruginosa CupD Fimbrial Genes Is Antagonistically Controlled by RcsB and the EAL-Containing PvrR Response Regulators

    PubMed Central

    Mikkelsen, Helga; Ball, Geneviève; Giraud, Caroline; Filloux, Alain

    2009-01-01

    Pseudomonas aeruginosa is a gram-negative pathogenic bacterium with a high adaptive potential that allows proliferation in a broad range of hosts or niches. It is also the causative agent of both acute and chronic biofilm-related infections in humans. Three cup gene clusters (cupA-C), involved in the assembly of cell surface fimbriae, have been shown to be involved in biofilm formation by the P. aeruginosa strains PAO1 or PAK. In PA14 isolates, a fourth cluster, named cupD, was identified within a pathogenicity island, PAPI-I, and may contribute to the higher virulence of this strain. Expression of the cupA genes is controlled by the HNS-like protein MvaT, whereas the cupB and cupC genes are under the control of the RocS1A1R two-component system. In this study, we show that cupD gene expression is positively controlled by the response regulator RcsB. As a consequence, CupD fimbriae are assembled on the cell surface, which results in a number of phenotypes such as a small colony morphotype, increased biofilm formation and decreased motility. These behaviors are compatible with the sessile bacterial lifestyle. The balance between planktonic and sessile lifestyles is known to be linked to the intracellular levels of c-di-GMP with high levels favoring biofilm formation. We showed that the EAL domain-containing PvrR response regulator counteracts the activity of RcsB on cupD gene expression. The action of PvrR is likely to involve c-di-GMP degradation through phosphodiesterase activity, confirming the key role of this second messenger in the balance between bacterial lifestyles. The regulatory network between RcsB and PvrR remains to be elucidated, but it stands as a potential model system to study how the equilibrium between the two lifestyles could be influenced by therapeutic agents that favor the planktonic lifestyle. This would render the pathogen accessible for the immune system or conventional antibiotic treatment. PMID:19547710

  14. Depth related trends in proximate composition of demersal fishes in the eastern North Pacific

    NASA Astrophysics Data System (ADS)

    Drazen, J. C.

    2007-02-01

    The proximate chemistry of the white muscle and liver of 18 species of demersal fish from the eastern North Pacific was studied to determine trends with depth, locomotory mode and buoyancy mechanism, foraging strategy and to elucidate energetic strategies. Data for 24 species from shallow water were taken from the literature and included for analysis of muscle water content. Benthopelagic species, primarily gadiforms, have significantly larger lipid-rich livers than benthic species. The benthopelagic species may use this lipid to add buoyancy, but it is also used as energy storage. Buoyancy mechanism was directly related to proximate composition. Fishes using gasbladders had normal muscle composition. The two species of benthopelagic fishes without gasbladders have either very high muscle lipid content ( Anoplopoma fimbria) or gelatinous muscle ( Alepocephalus tenobrosus) to aid in achieving neutral buoyancy. The macrourid, Albatrossia pectoralis, has a very small gasbladder and also has gelatinous muscle. Both of these benthopelagic fishes with gelatinous muscle feed on pelagic organisms. Gelatinous muscle was also found in two flatfishes that inhabit the oxygen minimum zone. For these fishes, high water content may serve to lower metabolic costs while maintaining large body size. Scavengers such as Coryphaenoides armatus and Coryphaenoides acrolepis have lipid rich livers and others such as A. fimbria and Pachycara sp. have high and variable muscle lipid content. Thus foraging mode also acts to influence proximate composition. Several depth-related trends in proximate composition were found. White muscle water content increased significantly with depth, and all four gelatinous species occurred at bathyal depths. This adds evidence in support of the hypothesis that decreasing light levels shorten reactive distances and relax the selective pressure for high locomotory capacity. In addition significant declines in liver protein content were observed, suggesting that

  15. Surfaces Presenting α-Phenyl Mannoside Derivatives Enable Formation of Stable, High Coverage, Non-pathogenic Escherichia coli Biofilms against Pathogen Colonization.

    PubMed

    Zhu, Zhiling; Wang, Jun; Lopez, Analette I; Yu, Fei; Huang, Yongkai; Kumar, Amit; Li, Siheng; Zhang, Lijuan; Cai, Chengzhi

    2015-06-01

    Prevention of pathogenic colonization on medical devices over a long period of time remains a great challenge, especially in a high-nutrient environment that accelerates production of biomass leading to biofouling of the device. Since biofouling and the subsequent pathogen colonization is eventually inevitable, a new strategy using non-pathogenic bacteria as living guards against pathogenic colonization on medical devices has attracted increasing interest. Crucial to the success of this strategy is to pre-establish a high coverage and stable biofilm of benign bacteria on the surface. Silicone elastomers are one of the most widely used materials in biomedical devices. In this work, we modified silicone surfaces to promote formation of high coverage and stable biofilms by a non-pathogenic Escherichia coli strain 83972 with type 1 fimbriae (fim+) to interfere the colonization of an aggressive biofilm-forming, uropathogenic Enterococcus faecalis. Although it is well known that mannoside surfaces promote the initial adherence of fim+ E. coli through binding to the FimH receptor at the tip of the type 1 fimbriae, it is not clear whether the fast initial adherence could lead to a high coverage and stable protective biofilm. To explore the role of mannoside ligands, we synthesized a series of alkyl and aryl mannosides varied in structure and immobilized them on silicone surfaces pre-coated with poly(amidoamine) (PAMAM) dendrimer. We found that stable and densely packed benign E. coli biofilms were formed on the surfaces presenting biphenyl mannoside with the highest initial adherence of fim+ E. coli. These non-pathogenic biofilms prevented the colonization of E. faecalis for 11 days at a high concentration (10(8) CFU mL(-1), 100,000 times above the diagnostic threshold for urinary tract infection) in the nutrient-rich Lysogeny Broth (LB) media. The result shows a correlation among the initial adherence of fim+ E. coli 83972, the coverage and long-term stability of the

  16. Surfaces presenting α-phenyl mannoside derivatives enable formation of stable, high coverage, non-pathogenic Escherichia coli biofilms against pathogen colonization.

    PubMed

    Zhu, Zhiling; Wang, Jun; Lopez, Analette I; Yu, Fei; Huang, Yongkai; Kumar, Amit; Li, Siheng; Zhang, Lijuan; Cai, Chengzhi

    2015-06-01

    Prevention of pathogenic colonization on medical devices over a long period of time remains a great challenge, especially in a high-nutrient environment that accelerates the production of biomass leading to biofouling of the device. Since biofouling and the subsequent pathogen colonization is eventually inevitable, a new strategy using non-pathogenic bacteria as living guards against pathogenic colonization on medical devices has attracted increasing interest. Crucial to the success of this strategy is to pre-establish a high coverage and stable biofilm of benign bacteria on the surface. Silicone elastomers are one of the most widely used materials in biomedical devices. In this work, we modified silicone surfaces to promote formation of high coverage and stable biofilms by a non-pathogenic Escherichia coli strain 83972 with type 1 fimbriae (fim+) to interfere with the colonization of an aggressive biofilm-forming, uropathogenic Enterococcus faecalis. Although it is well known that mannoside surfaces promote the initial adherence of fim+ E. coli through binding to the FimH receptor at the tip of the type 1 fimbriae, it is not clear whether the fast initial adherence could lead to a high coverage and stable protective biofilm. To explore the role of mannoside ligands, we synthesized a series of alkyl and aryl mannosides varied in the structure and immobilized them on silicone surfaces pre-coated with a poly(amidoamine) (PAMAM) dendrimer. We found that stable and densely packed benign E. coli biofilms were formed on the surfaces presenting biphenyl mannoside with the highest initial adherence of fim+ E. coli. These non-pathogenic biofilms prevented the colonization of E. faecalis for 11 days at a high concentration (10(8) CFU mL(-1), 100,000 times above the diagnostic threshold for urinary tract infection) in the nutrient-rich Lysogeny Broth (LB) media. The result shows a correlation among the initial adherence of fim+ E. coli 83972, the coverage and long

  17. Preoperative automated fibre quantification predicts postoperative seizure outcome in temporal lobe epilepsy

    PubMed Central

    Glenn, G. Russell; Weber, Bernd; Kreilkamp, Barbara A. K.; Jensen, Jens H.; Helpern, Joseph A.; Wagner, Jan; Barker, Gareth J.; Richardson, Mark P.; Bonilha, Leonardo

    2017-01-01

    Approximately one in every two patients with pharmacoresistant temporal lobe epilepsy will not be rendered completely seizure-free after temporal lobe surgery. The reasons for this are unknown and are likely to be multifactorial. Quantitative volumetric magnetic resonance imaging techniques have provided limited insight into the causes of persistent postoperative seizures in patients with temporal lobe epilepsy. The relationship between postoperative outcome and preoperative pathology of white matter tracts, which constitute crucial components of epileptogenic networks, is unknown. We investigated regional tissue characteristics of preoperative temporal lobe white matter tracts known to be important in the generation and propagation of temporal lobe seizures in temporal lobe epilepsy, using diffusion tensor imaging and automated fibre quantification. We studied 43 patients with mesial temporal lobe epilepsy associated with hippocampal sclerosis and 44 healthy controls. Patients underwent preoperative imaging, amygdalohippocampectomy and postoperative assessment using the International League Against Epilepsy seizure outcome scale. From preoperative imaging, the fimbria-fornix, parahippocampal white matter bundle and uncinate fasciculus were reconstructed, and scalar diffusion metrics were calculated along the length of each tract. Altogether, 51.2% of patients were rendered completely seizure-free and 48.8% continued to experience postoperative seizure symptoms. Relative to controls, both patient groups exhibited strong and significant diffusion abnormalities along the length of the uncinate bilaterally, the ipsilateral parahippocampal white matter bundle, and the ipsilateral fimbria-fornix in regions located within the medial temporal lobe. However, only patients with persistent postoperative seizures showed evidence of significant pathology of tract sections located in the ipsilateral dorsal fornix and in the contralateral parahippocampal white matter bundle

  18. A Comparison of Diets Supplemented with a Feed Additive Containing Organic Acids, Cinnamaldehyde and a Permeabilizing Complex, or Zinc Oxide, on Post-Weaning Diarrhoea, Selected Bacterial Populations, Blood Measures and Performance in Weaned Pigs Experimentally Infected with Enterotoxigenic E. coli †

    PubMed Central

    Stensland, Ingunn; Kim, Jae Cheol; Bowring, Bethany; Collins, Alison M.; Mansfield, Josephine P.; Pluske, John R.

    2015-01-01

    Simple Summary This experiment was conducted to assess the effects of three diets on diarrhoea, performance (weight change, feed intake and feed conversion ratio), selected bacterial populations and blood measures of weaner pigs infected with enterotoxigenic E. coli. The three diets were: base diet (no antimicrobial compounds), base diet containing zinc oxide, and base diet containing a feed additive (blend of organic acids, cinnamaldehyde and permeabilizing complex). Only feeding zinc oxide decreased diarrhoea, with zinc oxide-fed pigs performing better than base diet-fed pigs. Zinc oxide-fed pigs performed similarly to pigs fed the organic acids, cinnamaldehyde and permeabilizing complex. Significant interactions between treatment and day after weaning were found for some bacterial populations, although the implications of such findings require further examination. Abstract The effects of feeding a diet supplemented with zinc oxide (ZnO) or a blend of organic acids, cinnamaldehyde and a permeabilizing complex (OACP) on post-weaning diarrhoea (PWD) and performance in pigs infected with enterotoxigenic E. coli (ETEC) were examined. Additionally, changes in selected bacterial populations and blood measures were assessed. A total of 72 pigs weaned at 22 d of age and weighing 7.2 ± 1.02 kg (mean ± SEM) was used. Treatments were: base diet (no antimicrobial compounds); base diet + 3 g ZnO/kg; base diet + 1.5 g OACP/kg. Dietary treatments started on the day of weaning and were fed ad libitum for 3 weeks. All pigs were infected with an F4 ETEC on d 4, 5 and 6 after weaning. The incidence of PWD was lower in pigs fed ZnO (p = 0.026). Overall, pigs fed ZnO grew faster (p = 0.013) and ate more (p = 0.004) than the base diet-fed pigs, with OACP-fed pigs performing the same (p > 0.05) as both the ZnO- and base diet-fed pigs. Feed conversion ratio was similar for all diets (p > 0.05). The percentage of E. coli with F4 fimbriae was affected a day by treatment interaction (p

  19. Prevalence and characteristics of the epidemic multiresistant Escherichia coli ST131 clonal group among extended-spectrum beta-lactamase-producing E. coli isolates in Copenhagen, Denmark.

    PubMed

    Olesen, Bente; Hansen, Dennis S; Nilsson, Frida; Frimodt-Møller, Jakob; Leihof, Rikke Fleron; Struve, Carsten; Scheutz, Flemming; Johnston, Brian; Krogfelt, Karen A; Johnson, James R

    2013-06-01

    We report the characteristics of 115 extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli clinical isolates, from 115 unique Danish patients, over a 1-year study interval (1 October 2008 to 30 September 2009). Forty-four (38%) of the ESBL isolates represented sequence type 131 (ST13)1, from phylogenetic group B2. The remaining 71 isolates were from phylogenetic groups D (27%), A (22%), B1 (10%), and B2 (3%). Serogroup O25 ST131 isolates (n = 42; 95% of ST131) comprised 7 different K antigens, whereas two ST131 isolates were O16:K100:H5. Compared to non-ST131 isolates, ST131 isolates were associated positively with CTX-M-15 and negatively with CTX-M-1 and CTX-M-14. They also were associated positively with 11 virulence genes, including afa and dra (Dr family adhesins), the F10 papA allele (P fimbria variant), fimH (type 1 fimbriae), fyuA (yersiniabactin receptor), iha (adhesin siderophore), iutA (aerobactin receptor), kpsM II (group 2 capsules), malX (pathogenicity island marker), ompT (outer membrane protease), sat (secreted autotransporter toxin), and usp (uropathogenicity-specific protein) and negatively with hra (heat-resistant agglutinin) and iroN (salmochelin receptor). The consensus virulence gene profile (>90% prevalence) of the ST131 isolates included fimH, fyuA, malX, and usp (100% each), ompT and the F10 papA allele (95% each), and kpsM II and iutA (93% each). ST131 isolates were also positively associated with community acquisition, extraintestinal pathogenic E. coli (ExPEC) status, and the O25, K100, and H4 antigens. Thus, among ESBL E. coli isolates in Copenhagen, ST131 was the most prevalent clonal group, was community associated, and exhibited distinctive and comparatively extensive virulence profiles, plus a greater variety of capsular antigens than reported previously.

  20. Histomorphometric evaluation of intestinal cellular immune responses in pigs immunized with live oral F4ac+ non-enterotoxigenic E. coli vaccine against postweaning colibacillosis

    PubMed Central

    Kovšca Janjatović, A.; Lacković, G.; Božić, F.; Kezić, D.; Popović, M.; Valpotić, H.; Harapin, I.; Pavižić, Ž.; Njari, B.; Valpotić, I.

    2010-01-01

    Enterotoxigenic Escherichia coli (ETEC) infection is the most common type of porcine postweaning colibacillosis (PWC). Among fimbriae of porcine ETEC strains the best studied family of fimbriae are the members of F4 adhesins, existing in at least three variants: ab, ac, ad. Active immunization against porcine PWC is difficult due to: i) ETEC strains are only one of the essential predisposing factors, ii) the success of vaccinal antigen uptake depends on the presence of enterocyte receptors for F4 adhesins, iii) the intestinal immune system may react with tolerance or hypersensitivity to the same antigens depending on the dose and form of the vaccinal immunogen, and iv) kinetics of the specific immune responses may be different in the case of F4 (earlier) and the other ETEC adhesins, particularly F18 (later). The aim of this study was to test the effectiveness of a live attenuated F4ac+ non-ETEC vaccine against porcine PWC by analyzing quantitative differences in the small intestinal lymphoid and myeloid cell subsets of immunized (with or without levamisole given as an adjuvant) vs control non-immunized pigs. Four week-old pigs were intragastrically immunized with a vaccine candidate F4ac + non-ETEC strain 2407 at day 0, challenged 7 days later with a virulent F4ac+ strain ETEC 11-800/1/94, euthanatized at day 13 and sampled for immunohistology. Non-immunized pigs received saline at day 0 and were processed as the principals. Immunophenotypes of lymphoid and myeloid cell subsets were demonstrated within jejunal and ileal mucosa by immunohistochemical avidinbiotin complex method and corresponding morphometric data were analyzed using software program Lucia G for digital image analyses. Monoclonal antibodies reactive with surface molecules on porcine immune cells such as CD3, CD45RA, CD45RC, CD21 and SWC3 enabled clear insight into distribution patterns and amount of these cells within the gut-associated lymphoid tissues (GALT) examined. The numbers of jejunal and

  1. Wild birds as biological indicators of environmental pollution: antimicrobial resistance patterns of Escherichia coli and enterococci isolated from common buzzards (Buteo buteo).

    PubMed

    Radhouani, Hajer; Poeta, Patrícia; Gonçalves, Alexandre; Pacheco, Rui; Sargo, Roberto; Igrejas, Gilberto

    2012-06-01

    A total of 36 Escherichia coli and 31 enterococci isolates were recovered from 42 common buzzard faecal samples. The E. coli isolates showed high levels of resistance to streptomycin and tetracycline. The following resistance genes were detected: bla(TEM) (20 of 22 ampicillin-resistant isolates), tet(A) and/or tet(B) (16 of 27 tetracycline-resistant isolates), aadA1 (eight of 27 streptomycin-resistant isolates), cmlA (three of 15 chloramphenicol-resistant isolates), aac(3)-II with/without aac(3)-IV (all seven gentamicin-resistant isolates) and sul1 and/or sul2 and/or sul3 [all eight sulfamethoxazole/trimethoprim-resistant (SXT) isolates]. intI1 and intI2 genes were detected in four SXT-resistant isolates. The virulence-associated genes fimA (type 1 fimbriae), papC (P fimbriae) and aer (aerobactin) were detected in 61.1, 13.8 and 11.1% of the isolates, respectively. The isolates belonged to phylogroups A (47.2%), B1 (8.3%), B2 (13.9%) and D (30.5%). For the enterococci isolates, Enterococcus faecium was the most prevalent species (48.4%). High levels of tetracycline and erythromycin resistance were found among our isolates (87 and 81%, respectively). Most of the tetracycline-resistant strains carried the tet(M) and/or tet(L) genes. The erm(B) gene was detected in 80% of erythromycin-resistant isolates. The vat(D) and/or vat(E) genes were found in nine of the 17 quinupristin-dalfopristin-resistant isolates. The enterococcal isolates showing high-level resistance for kanamycin, gentamicin and streptomycin contained the aph(3')-IIIa, aac(6')-aph(2″) and ant(6)-Ia genes, respectively. This report reveals that common buzzards seem to represent an important reservoir, or at least a source, of multi-resistant E. coli and enterococci isolates, and consequently may represent a considerable hazard to human and animal health by transmission of these isolates to waterways and other environmental sources via their faecal deposits.

  2. Phenotypic and genotypic characterization of enterotoxigenic Escherichia coli serotype O8:KX105 and O8:K"2829" strains isolated from piglets with diarrhea.

    PubMed Central

    Broes, A; Fairbrother, J M; Mainil, J; Harel, J; Lariviere, S

    1988-01-01

    Twelve pathogenic and seven nonpathogenic enterotoxigenic Escherichia coli strains which were previously identified as belonging to serogroup O8:KX105 (A. Broes, J. M. Fairbrother, S. Larivière, M. Jacques, and W. M. Johnson, Infect. Immun. 56:241-246, 1988) were further examined for their phenotypic and genotypic properties. Only the 12 pathogenic strains were confirmed to possess the capsular antigen KX105. The seven nonpathogenic strains did not possess this antigen and thus were incorrectly assigned to have capsular antigen KX105. All seven nonpathogenic strains apparently possessed a previously unrecognized capsular antigen which has been designated K"2829". Studies with antisera prepared against F1 (type 1) fimbriae from three E. coli strains suggested that at least three antigenic subtypes of F1 fimbriae were represented among the O8:KX105 strains examined. By using serotyping, biotyping, and outer membrane protein profile analyses, the O8:KX105 strains were divided into at least two distinct clusters, whereas the O8:K"2829" strains were grouped into a single unique cluster. Most of the strains of the same cluster were further differentiated by testing for antibiotic resistance and colicin production and resistance and by analysis of plasmid content. With the exception of one strain which lost its enterotoxicity during storage, all of the O8:KX105 strains hybridized with the gene probes for the heat-labile (LT) and heat-stable (STb) enterotoxins. For each O8:KX105 strain, a single plasmid ranging in size from 61 to 77 megadaltons carried the LT and STb genes.All of the enterotoxigenic O8: KX105 strains fermented sorbose, whereas the nonenterotoxigenic strain did not. All of the O8:K "2829" strains hybridized with the STb probe only. For each O8:K "2829" strain, the STb genes were located on a single plasmid of 61 or 22 megadaltons. None of the strains demonstrated homology with the genes encoding the F4 (K88), F5 (K99), F6 (987P), and F41 fimbrial antigens

  3. Environmentally Endemic Pseudomonas aeruginosa Strains with Mutations in lasR Are Associated with Increased Disease Severity in Corneal Ulcers

    PubMed Central

    Hammond, John H.; Hebert, Wesley P.; Naimie, Amanda; Ray, Kathryn; Van Gelder, Rachel D.; DiGiandomenico, Antonio; Lalitha, Prajna; Srinivasan, Muthiah; Acharya, Nisha R.; Lietman, Thomas; Hogan, Deborah A.

    2016-01-01

    ABSTRACT The Steroids for Corneal Ulcers Trial (SCUT) was a multicenter, international study of bacterial keratitis in which 101 Pseudomonas aeruginosa infections were treated. Twenty-two of 101 P. aeruginosa isolates collected had a colony morphology characteristic of a loss-of-function mutation in lasR, the gene encoding a quorum-sensing master regulator. Ulcers caused by these 22 strains were associated with larger areas of corneal opacification, worse vision, and a lower rate of vision recovery in response to treatment than ulcers caused by the other isolates. The lasR sequences from these isolates each contained one of three nonsynonymous substitutions, and these strains were deficient in production of LasR-regulated protease and rhamnolipids. Replacement of lasR with either of the two most common lasR alleles from the SCUT isolates was sufficient to decrease protease and rhamnolipid production in PA14. Loss of LasR function is associated with increased production of CupA fimbriae, and the LasR-defective isolates exhibited higher production of CupA fimbriae than LasR-intact isolates. Strains with the same lasR mutation were of the same multilocus sequence type, suggesting that LasR-deficient, environmental P. aeruginosa strains were endemic to the area, and infections caused by these strains were associated with worse patient outcomes in the SCUT study. (This study has been registered at ClinicalTrials.gov under registration no. NCT00324168.) IMPORTANCE The LasR transcription factor is an important regulator of quorum sensing in P. aeruginosa and positively controls multiple virulence-associated pathways. The emergence of strains with lasR loss-of-function alleles in chronic disease is well described and is thought to represent a specific adaptation to the host environment. However, the prevalence and virulence of these strains in acute infections remain unclear. This report describes observations revealing that lasR mutants were common among isolates from

  4. Surface structures (peritrichous fibrils and tufts of fibrils) found on Streptococcus sanguis strains may be related to their ability to coaggregate with other oral genera.

    PubMed Central

    Handley, P S; Carter, P L; Wyatt, J E; Hesketh, L M

    1985-01-01

    We screened 36 strains of Streptococcus sanguis biotype I and 8 strains of S. sanguis biotype II for the presence of surface structures and for their ability to coaggregate with Actinomyces viscosus, Actinomyces naeslundii, and Fusobacterium nucleatum. Negative staining under an electron microscope revealed detectable surface structures on all S. sanguis strains. The majority of strains (38 of 44) carried peritrichous fibrils, which have an irregular profile and no distinct width. They usually appeared as a fringe with a constant width around the cell. Strains selected for measurement had a fringe with an average length of 72.4 +/- 8.5 nm on biotype I strains and 51.6 +/- 3.3 nm on biotype II strains. Some fibrillar biotype I strains carried an additional, longer (158.7 +/- 33.1 nm) type of fibril projecting through the shorter fibrils. Fibrillar density was characteristic for each strain, ranging from very dense on all cells in a population to very sparse on a few cells in a population. A small group of six strains carried tufts of fibrils in a lateral or polar position on the cell. Either one or two lengths of fibril were present in the tuft depending on the strain. One strain carried both peritrichous fibrils and fimbriae. Fimbriae are flexible structures with a constant width (4.5 to 5.0 nm) all along their length but very variable lengths (less than or equal to 0.7 micron) on each cell. S. sanguis I and II both included strains with peritrichous fibrils and tufts of fibrils, but the mixed morphotype strain was confined to biotype II. Fibrils were present on cells at all stages throughout the growth cycle for the strains tested. Freshly isolated fibrillar strains coaggregated consistently well with A. viscosus and A. naeslundii, although some fibrillar reference strains lacked the ability. In addition, all tufted strains could not coaggregate, but the strains with the mixed morphotype coaggregated well. Coaggregation with F. nucleatum was very strong for the

  5. A Dual Role for P2X7 Receptor during Porphyromonas gingivalis Infection

    PubMed Central

    Ramos-Junior, E.S.; Morandini, A.C.; Almeida-da-Silva, C.L.C.; Franco, E.J.; Potempa, J.; Nguyen, K.A.; Oliveira, A.C.; Zamboni, D.S.; Ojcius, D.M.; Scharfstein, J.

    2015-01-01

    Emerging evidence suggests a role for purinergic signaling in the activation of multiprotein intracellular complexes called inflammasomes, which control the release of potent inflammatory cytokines, such as interleukin (IL) -1β and -18. Porphyromonas gingivalis is intimately associated with periodontitis and is currently considered one of the pathogens that can subvert the immune system by limiting the activation of the NLRP3 inflammasome. We recently showed that P. gingivalis can dampen eATP-induced IL-1β secretion by means of its fimbriae in a purinergic P2X7 receptor–dependent manner. Here, we further explore the role of this purinergic receptor during eATP-induced IL-1β processing and secretion by P. gingivalis–infected macrophages. We found that NLRP3 was necessary for eATP-induced IL-1β secretion as well as for caspase 1 activation irrespective of P. gingivalis fimbriae. Additionally, although the secretion of IL-1β from P. gingivalis–infected macrophages was dependent on NLRP3, its adaptor protein ASC, or caspase 1, the cleavage of intracellular pro-IL-1β to the mature form was found to occur independently of NLRP3, its adaptor protein ASC, or caspase 1. Our in vitro findings revealed that P2X7 receptor has a dual role, being critical not only for eATP-induced IL-1β secretion but also for intracellular pro-IL-1β processing. These results were relevant in vivo since P2X7 receptor expression was upregulated in a P. gingivalis oral infection model, and reduced IFN-γ and IL-17 were detected in draining lymph node cells from P2rx7-/- mice. Furthermore, we demonstrated that P2X7 receptor and NLRP3 transcription were modulated in human chronic periodontitis. Overall, we conclude that the P2X7 receptor has a role in periodontal immunopathogenesis and suggest that targeting of the P2X7/NLRP3 pathway should be considered in future therapeutic interventions in periodontitis. PMID:26152185

  6. Regulation of neurotrophin and trkA, trkB and trkC tyrosine kinase receptor messenger RNA expression in kindling.

    PubMed

    Bengzon, J; Kokaia, Z; Ernfors, P; Kokaia, M; Leanza, G; Nilsson, O G; Persson, H; Lindvall, O

    1993-03-01

    Levels of messenger RNA for nerve growth factor, brain-derived neurotrophic factor, neurotrophin-3, and the tyrosine kinase receptors trkA, trkB and trkC have been studied using in situ hybridization in the rat brain 2 h and four weeks after kindling-induced seizures. Epileptiform activity evoked by hippocampal stimulation and exceeding 70 s lead to a concomitant and transient increase of brain- derived neurotrophic factor, nerve growth factor, trkB and trkC messenger RNA expression in dentate granule cells after both focal and generalized seizures. Brain-derived neurotrophic factor messenger RNA levels were also increased bilaterally in the CA1-CA3 regions, amygdala and the piriform, entorhinal, perirhinal, retrosplenial and temporal cortices after generalized seizures. The magnitude of the increases was similar throughout the development of kindling and in the fully kindled brain. No changes of trkA messenger RNA were observed. In amygdalar kindling, elevated brain-derived neurotrophic factor messenger RNA levels developed more rapidly in the amygdala-piriform cortex than after stimulation in the hippocampus but changes in the hippocampal formation were only seen in few animals. Intraventricular 6-hydroxydopamine or a bilateral fimbria-fornix lesion did not alter basal expression or seizure-evoked changes in messenger RNA levels for neurotrophins or trk receptors but increased the number of animals exhibiting elevated levels after the first stimulation, probably due to a prolongation of seizure activity. Both in sham-operated and fimbria-fornix-lesioned rats seizure activity caused a marked reduction of neurotrophin-3 messenger RNA levels in dentate granule cells. The results indicate that activation of the brain-derived neurotrophic factor gene, at least in dentate granule cells, is an "all-or-none" type of response and dependent on the duration but not the severity of seizures or the stage of kindling epileptogenesis. Changes in brain-derived neurotrophic

  7. Incidental serous tubal intraepithelial carcinoma and early invasive serous carcinoma in the nonprophylactic setting: analysis of a case series.

    PubMed

    Morrison, Jane C; Blanco, Luis Z; Vang, Russell; Ronnett, Brigitte M

    2015-04-01

    A precursor for invasive ovarian/pelvic high-grade serous carcinoma, termed serous tubal intraepithelial carcinoma (STIC), has been identified and characterized through careful analysis of the fallopian tubes in both prophylactic salpingo-oophorectomy specimens obtained from women with either a family history of breast and/or ovarian cancer or germline mutations of BRCA1 and BRCA2 and in cases of pelvic high-grade serous carcinoma. Data on incidental STICs and clinically occult microscopic invasive high-grade serous carcinomas are limited. We analyzed the clinicopathologic features of 22 cases, including 15 pure STICs and 7 STICs associated with microscopic invasive high-grade serous carcinomas, identified incidentally in fallopian tubes removed for nonprophylactic indications. Patient age ranged from 39 to 79 years (mean: 62.7; median: 61), with only 1 patient under the age of 50. No patients were known to carry BRCA1 or BRCA2 mutations. Of the 12 pure STICs for which the location in the fallopian tube could be established, 9 were in the fimbriated portion, 1 was at the junction of the fimbria and infundibulum, and 2 were in the nonfimbriated tube. Of the 7 STICs with associated invasive high-grade serous carcinoma, 3 were located in the fimbriated portion, 2 were at the junction of the fimbria and infundibulum, and 2 were in the nonfimbriated tube. The invasive components were in the fallopian tube in 6 cases, 4 in subepithelial stroma of tubal mucosa, and 2 as an intramucosal (exophytic) luminal lesion without invasion of underlying subepithelial stroma (size range: 1 to 4 mm). The remaining case had a microscopic focus of high-grade serous carcinoma within the ipsilateral ovary (1.3 mm cortical focus) identified only on deeper sections, without an associated invasive component in the fallopian tube. The preferential finding of atypical epithelium with the cytologic features of high-grade serous carcinoma, namely STIC, in the fallopian tubes rather than the

  8. Effective G-protein coupling of Y2 receptors along axonal fiber tracts and its relevance for epilepsy.

    PubMed

    Dum, Elisabeth; Fürtinger, Sabine; Gasser, Elisabeth; Bukovac, Anneliese; Drexel, Meinrad; Tasan, Ramon; Sperk, Günther

    2017-02-01

    Neuropeptide Y (NPY)-Y2 receptors are G-protein coupled receptors and, upon activation, induce opening of potassium channels or closing of calcium channels. They are generally presynaptically located. Depending on the neuron in which they are expressed they mediate inhibition of release of NPY and of the neuron's classical transmitter GABA, glutamate or noradrenaline, respectively. Here we provide evidence that Y2 receptor binding is inhibited dose-dependently by GTPγS along Schaffer collaterals, the stria terminalis and the fimbria indicating that Y2 receptors are functionally coupled to G-proteins along these fiber tracts. Double immune fluorescence revealed coexistence of Y2-immunoreactivity with β-tubulin, a marker for axons in the stria terminalis, but not with synaptophysin labeling presynaptic terminals, supporting the localization of Y2 receptors along axonal tracts. After kainic acid-induced seizures in rats, GTPγS-induced inhibition of Y2 receptor binding is facilitated in the Schaffer collaterals but not in the stria terminalis. Our data indicate that Y2 receptors are not only located at nerve terminals but also along fiber tracts and are there functionally coupled to G-proteins.

  9. Prevention of urinary tract infections with vaccinium products.

    PubMed

    Davidson, Elyad; Zimmermann, Benno F; Jungfer, Elvira; Chrubasik-Hausmann, Sigrun

    2014-03-01

    Cranberries exert a dose-dependent inhibition of the adherence of E. coli fimbriae to uroepithelial cells. This was demonstrated in vitro but also ex vivo in vitro with urine from cranberry consumers. The active principle has not been identified in detail but type-A proanthocyanidins (PAC) play an important role in the mechanism of action. Since the three species, American cranberry (Vaccinium macrocarpon), European cranberry (Vaccinium oxycoccus) and/or lingonberry (Vaccinium vitis-idaea), have different patterns of type-A PACs, results from one species cannot be transferred to the others. It seems likely that most of the studies with monopreparations from V. macrocarpon were underdosed. Whereas photometric PAC quantification may overestimate the true content on co-active compounds, reversed phase high-performance liquid chromatograpy may underestimate them. Recent studies with PAC doses in the upper range (DMAC method) or declared type-A PAC content in the daily dose reveal a dose-dependent trend of clinical effectiveness, however, with a possible ceiling effect. In order to clarify this, future three-arm studies should investigate Vaccinium preparations with higher type-A PAC doses than previously used. We analysed two popular European vitis-idaea products, a mother juice and a proprietary extract. Both preparations may be appropriate to confirm the Vaccinium urinary tract infection-preventive effect beyond doubt.

  10. Klebsiella pneumoniae yfiRNB operon affects biofilm formation, polysaccharide production and drug susceptibility.

    PubMed

    Huertas, Mónica G; Zárate, Lina; Acosta, Iván C; Posada, Leonardo; Cruz, Diana P; Lozano, Marcela; Zambrano, María M

    2014-12-01

    Klebsiella pneumoniae is an opportunistic pathogen important in hospital-acquired infections, which are complicated by the rise of drug-resistant strains and the capacity of cells to adhere to surfaces and form biofilms. In this work, we carried out an analysis of the genes in the K. pneumoniae yfiRNB operon, previously implicated in biofilm formation. The results indicated that in addition to the previously reported effect on type 3 fimbriae expression, this operon also affected biofilm formation due to changes in cellulose as part of the extracellular matrix. Deletion of yfiR resulted in enhanced biofilm formation and an altered colony phenotype indicative of cellulose overproduction when grown on solid indicator media. Extraction of polysaccharides and treatment with cellulase were consistent with the presence of cellulose in biofilms. The enhanced cellulose production did not, however, correlate with virulence as assessed using a Caenorhabditis elegans assay. In addition, cells bearing mutations in genes of the yfiRNB operon varied with respect to the WT control in terms of susceptibility to the antibiotics amikacin, ciprofloxacin, imipenem and meropenem. These results indicated that the yfiRNB operon is implicated in the production of exopolysaccharides that alter cell surface characteristics and the capacity to form biofilms--a phenotype that does not necessarily correlate with properties related with survival, such as resistance to antibiotics.

  11. Advanced glycation end products facilitate bacterial adherence in urinary tract infection in diabetic mice.

    PubMed

    Ozer, Ahmet; Altuntas, Cengiz Z; Izgi, Kenan; Bicer, Fuat; Hultgren, Scott J; Liu, Guiming; Daneshgari, Firouz

    2015-07-01

    Diabetic individuals have increased susceptibility to urinary tract infection (UTI), a common, painful condition. During diabetes mellitus, non-enzymatic reactions between reducing sugars and protein amine groups result in excessive production of advanced glycation end products (AGEs) that accumulate in tissues. Since bacteria adhere to cell surfaces by binding to carbohydrates, we hypothesized that adherence of bacteria to the bladder in diabetics may be enhanced by accumulation of AGEs on urothelial surface proteins. Using a murine model of UTI, we observed increased adherence of type 1 fimbriated uropathogenic Escherichia coli (UPEC) to the bladder in streptozotocin-induced diabetic female mice compared with age-matched controls, along with increased concentrations of two common AGEs in superficial urothelial cells from diabetic bladders. Several lectins with different specificities exhibited increased binding to urothelial homogenates from diabetic mice compared with controls, and two of those lectins also bound to AGEs. Furthermore, mannose-binding type 1 fimbriae isolated from UPEC bound to different AGEs, and UPEC adherence to the bladder in diabetic mice, were inhibited by pretreatment of mice with the AGE inhibitor pyridoxamine. These results strongly suggest a role for urothelial AGE accumulation in increased bacterial adherence during UTI in diabetes.

  12. Cloning, expression, and sequence analysis of the Haemophilus influenzae type b strain M43p+ pilin gene.

    PubMed Central

    Gilsdorf, J R; Marrs, C F; McCrea, K W; Forney, L J

    1990-01-01

    By using antiserum against Haemophilus influenzae type b (Hib) strain M43p+ denatured pilin, we screened a genomic library of Hib strain M43p+ and identified a clone that expressed pilin, but not assembled pili, on its surface. Southern blot analysis revealed the presence of one structural gene, which was also present in strain M42p-, a nonpiliated variant. Five exonuclease III deletion mutants, two of which had deletions that extended into the structural gene and failed to express pilin, were used to obtain the nucleotide sequence of the structural gene. The amino acid sequence of the open reading frame agrees with 38 of 40 amino acids from the published sequence of purified Hib M43p+ pilin. The pilin gene coded for a mature protein of 193 amino acids, with a calculated molecular mass of 21,101 daltons. Comparison of the Hib M43p+ pilin amino acid sequence with those of pilins of other bacteria revealed strong conservation of amino- and carboxy-terminal regions in M43p+ and Escherichia coli F17, type 1C, and several members of the P pili family, as well as Klebsiella pneumoniae type 3 MR/K, Bordetella pertussis serotype 2, and Serratia marcescens US46 fimbriae. Images PMID:1969389

  13. TolC promotes ExPEC biofilm formation and curli production in response to medium osmolarity.

    PubMed

    Hou, Bo; Meng, Xian-Rong; Zhang, Li-Yuan; Tan, Chen; Jin, Hui; Zhou, Rui; Gao, Jian-Feng; Wu, Bin; Li, Zi-Li; Liu, Mei; Chen, Huan-Chun; Bi, Ding-Ren; Li, Shao-Wen

    2014-01-01

    While a high osmolarity medium activates Cpx signaling and causes CpxR to repress csgD expression, and efflux protein TolC protein plays an important role in biofilm formation in Escherichia coli, whether TolC also responds to an osmolarity change to regulate biofilm formation in extraintestinal pathogenic E. coli (ExPEC) remains unknown. In this study, we constructed ΔtolC mutant and complement ExPEC strains to investigate the role of TolC in the retention of biofilm formation and curli production capability under different osmotic conditions. The ΔtolC mutant showed significantly decreased biofilm formation and lost the ability to produce curli fimbriae compared to its parent ExPEC strain PPECC42 when cultured in M9 medium or 1/2 M9 medium of increased osmolarity with NaCl or sucrose at 28°C. However, biofilm formation and curli production levels were restored to wild-type levels in the ΔtolC mutant in 1/2 M9 medium. We propose for the first time that TolC protein is able to form biofilm even under high osmotic stress. Our findings reveal an interplay between the role of TolC in ExPEC biofilm formation and the osmolarity of the surrounding environment, thus providing guidance for the development of a treatment for ExPEC biofilm formation.

  14. Effect of 2,4-Dichlorophenoxyacetic acid herbicide Escherichia coli growth, chemical, composition, and cellular envelope

    USGS Publications Warehouse

    Carr, R.S.; Biedenbach, J.M.; Hooten, R.L.

    2001-01-01

    2,4-Dichlorophenoxyacetic acid (2,4-D) is a herbicide widely used in the world and mainly excreted by the renal route in exposed humans and animals. Herbicides can affect other nontarget organisms, such as Escherichia coli. We observed that a single exposure to 1 mM 2,4-D diminished growth and total protein content in all E. coli strains tested in vitro. In addition, successive exposures to 0.01 mM 2,4-D had a toxic effect decreasing growth up to early stationary phase. Uropathogenic E. coli adhere to epithelial cells mediated by fimbriae, adhesins, and hydrophobic properties. 2,4-D exposure of uropathogenic E. coli demonstrated altered hydrophobicity and fimbriation. Hydrophobicity index values obtained by partition in p-xylene/water were 300-420% higher in exposed cells than in control ones. Furthermore, values of hemagglutination titer, protein contents in fimbrial crude extract, and electron microscopy demonstrated a significant diminution of fimbriation in treated cells. Other envelope alterations could be detected, such as lipoperoxidation, evidenced by decreased polyunsaturated fatty acids and increased lipid degradation products (malonaldehyde), and motility diminution. These alterations decreased cell adherence to erythrocytes, indicating a diminished pathogenic capacity of the 2,4-D-exposed E. coli. ?? 2001 by John Wiley & Sons, Inc.

  15. Neuropathological Sequelae of Long-Term Allogeneic and Syngeneic Neural Transplantation into the Hippocampus

    PubMed Central

    Patel, Sanjay N.; Kershaw, Tim R.; Williams, John; Gray, Jeffrey A.; Lantos, Peter L.; Sinden, John D.

    1994-01-01

    The long-term fate of multiple intrahippocampal allogeneic transplants of fetal basal forebrain tissue was studied in neonatally tolerised and immunised groups of rats with lesions of the fimbria-fornix. Despite the good survival of the allografts in all groups, unexpected transplant-associated host hippo-campal neuropathology was discovered 12 months after transplantation, which consisted of (i) CA1 cell degeneration and (ii) abnormal accumulations of phosphorylated neurofilaments in neuronal perikarya and axonal swellings only within the host hippocampal neuropil and not of the transplanted tissue. This neurofilament abnormality, identified by RT97 immunohistochemistry, was significantly greater in the transplanted rats compared to the non-grafted lesion-only and sham-lesioned rats (p<0.01). The same type of neurofilament abnormality was again observed in a second, separate experiment using unilateral and bilateral syngeneic and allogeneic transplants. The neuropathology was significantly (p<0.05) greater in the transplanted side of the unilateral transplanted rats compared to the non-transplanted lesion-only control side of the same animals, showing that transplantation per se was a major factor involved in the pathogenesis of this neuropathology, irrespective of the type of transplant (syngeneic or allogeneic). In addition, a small degree of neurofilament abnormality was also found within the transplants in the second experiment, but not in the first. The results show that, under certain conditions, specific local neuropathological damage to the surrounding host neural tissue can develop in long-surviving allografted and syngrafted animals. PMID:7578437

  16. IgG responses after booster vaccination with different pertussis vaccines in Dutch children 4 years of age: effect of vaccine antigen content.

    PubMed

    Hendrikx, Lotte H; Berbers, Guy A M; Veenhoven, Reinier H; Sanders, Elisabeth A M; Buisman, Anne-Marie

    2009-11-05

    Since whooping cough is reemerging in the Netherlands from 1996 onwards, several changes in the national immunization program have been implemented regarding the pertussis vaccinations. The aim of this study is to investigate IgG responses in whole cell (wP) and acellular (aP) pertussis vaccine primed children following revaccination with different pertussis booster vaccines at 4 years of age. IgG levels to pertussis toxin (Pt), filamentous heamagglutin (FHA), pertactin (Prn) and fimbriae type 2 and 3 (Fim2/3) and avidities of Pt and Prn antibodies were measured using a multiplex immunoassay. Before and after the booster we found significantly higher IgG levels to Pt, FHA and Prn in aP compared to wP primed children. In all children a booster vaccination with a pertussis vaccine containing a high antigen dose (Infanrix) induced higher IgG responses compared to a low antigen dose containing vaccine (Triaxis). Avidities of Pt- and Prn-antibodies before and after booster vaccination were significantly higher in aP than in wP primed children. This study shows that a booster vaccine with high pertussis antigen concentrations induces higher antibody levels than a low antigen containing vaccine. In children primed with the Dutch DTwP-IPV-Hib vaccine we suggest to administer a booster vaccine containing high pertussis antigens to optimize IgG responses. The pertussis vaccination history has to be taken into account in decisions on changes in pertussis vaccination policy.

  17. Identification of a capsular variant and characterization of capsular acetylation in Klebsiella pneumoniae PLA-associated type K57

    PubMed Central

    Hsu, Chun-Ru; Liao, Chun-Hsing; Lin, Tzu-Lung; Yang, Han-Ru; Yang, Feng-Ling; Hsieh, Pei-Fang; Wu, Shih-Hsiung; Wang, Jin-Town

    2016-01-01

    Klebsiella pneumoniae can cause community-acquired pyogenic liver abscess (PLA). Capsular polysaccharide (CPS) is important for its virulence. Among 79 capsular (K) types discovered thus far, K57 is often associated with PLA. Here, we report the identification of a K57 variant. Cps gene locus sequencing revealed differences between the K57 reference strain 4425/51 (Ref-K57) and a variant, the PLA isolate A1142. While Ref-K57 cps contained orf13 encoding a putative acetyltransferase, the insertion of a putative transposase-encoding gene at this position was detected in A1142. This variation was detected in other K57 clinical strains. Biochemical analyses indicated that A1142 was deficient in CPS acetylation. Genetic replacement and complementation verified that orf13 was responsible for CPS acetylation. Acetylation increased CPS immunoreactivity to antiserum and enhanced K. pneumoniae induction of pro-inflammatory cytokines through JNK and MAPK signaling. While acetylation diminished the serum resistance of bacteria, it promoted adhesion to intestinal epithelial cells possibly via increasing production of type I fimbriae. In conclusion, acetylation-mediated capsular variation in K57 was observed. Capsular acetylation contributed to the variety and antigenic diversity of CPS, influenced its biological activities, and was involved in K. pneumoniae-host interactions. These findings have implications for vaccine design and pathogenicity of K. pneumoniae. PMID:27550826

  18. Actinobacillus actinomycetemcomitans adheres to human gingival fibroblasts and modifies cytoskeletal organization.

    PubMed

    Gutiérrez-Venegas, Gloria; Kawasaki-Cárdenas, Perla; Garcés, Carla Portillo; Román-Alvárez, Patricia; Barajas-Torres, Carolina; Contreras-Marmolejo, Luis Arturo

    2007-09-01

    Adherence of Actinobacillus actinomycetemcomitans to human gingival fibroblast cells induces cytoskeletal reorganization. A. actinomycetemcomitans is considered a pathogenic bacteria involved in localized aggressive periodontitis. Studies with epithelial cells have shown an adherent capacity of bacteria that is increased under anaerobic conditions. For adherence to take place, there is a need for interaction between extracellular vesicles and bacterial fimbriae. However, molecular events associated with the adherence process are still unknown. The aim of this study was to investigate whether A. actinomycetemcomitans adherence to human gingival fibroblasts promotes cytoskeletal reorganization. Adherence was determined with light microscopy and scanning electron microscopy. For F-actin visualization, cells were treated with fluorescein-isothiocyanate-phalloidin and samples were examined with epifluorescence optics. Fluorescent was recorded on Kodak T-Max 400 film. We showed that A. actinomycetemcomitans adheres to human gingival fibroblast primary cultures, this property stimulating an increase in the intracellular calcium levels. In human gingival fibroblast primary cultures, we observed that maximal A. actinomycetemcomitans adherence took place 1.5h after culture infection occurred and remained for 6h. The adherence was associated with morphologic alterations and an increased in the intracellular calcium levels. These experiments suggest that A. actinomycetemcomitans adherence cause morphological alterations, induce actin stress fibers and recruitment of intracellular calcium levels.

  19. Proteus mirabilis and Urinary Tract Infections

    PubMed Central

    Schaffer, Jessica N.; Pearson, Melanie M.

    2015-01-01

    Proteus mirabilis is a Gram-negative bacterium which is well-known for its ability to robustly swarm across surfaces in a striking bulls’-eye pattern. Clinically, this organism is most frequently a pathogen of the urinary tract, particularly in patients undergoing long-term catheterization. This review covers P. mirabilis with a focus on urinary tract infections (UTI), including disease models, vaccine development efforts, and clinical perspectives. Flagella-mediated motility, both swimming and swarming, is a central facet of this organism. The regulation of this complex process and its contribution to virulence is discussed, along with the type VI-secretion system-dependent intra-strain competition which occurs during swarming. P. mirabilis uses a diverse set of virulence factors to access and colonize the host urinary tract, including urease and stone formation, fimbriae and other adhesins, iron and zinc acquisition, proteases and toxins, biofilm formation, and regulation of pathogenesis. While significant advances in this field have been made, challenges remain to combatting complicated UTI and deciphering P. mirabilis pathogenesis. PMID:26542036

  20. Porphyromonas gingivalis Fim-A genotype distribution among Colombians

    PubMed Central

    Jaramillo, Adriana; Parra, Beatriz; Botero, Javier Enrique; Contreras, Adolfo

    2015-01-01

    Introduction: Porphyromonas gingivalis is associated with periodontitis and exhibit a wide array of virulence factors, including fimbriae which is encoded by the FimA gene representing six known genotypes. Objetive: To identify FimA genotypes of P. gingivalis in subjects from Cali-Colombia, including the co-infection with Aggregatibacter actinomycetemcomitans, Treponema denticola, and Tannerella forsythia. Methods: Subgingival samples were collected from 151 people exhibiting diverse periodontal condition. The occurrence of P. gingivalis, FimA genotypes and other bacteria was determined by PCR. Results: P. gingivalis was positive in 85 patients. Genotype FimA II was more prevalent without reach significant differences among study groups (54.3%), FimA IV was also prevalent in gingivitis (13.0%). A high correlation (p= 0.000) was found among P. gingivalis, T. denticola, and T. forsythia co-infection. The FimA II genotype correlated with concomitant detection of T. denticola and T. forsythia. Conclusions: Porphyromonas gingivalis was high even in the healthy group at the study population. A trend toward a greater frequency of FimA II genotype in patients with moderate and severe periodontitis was determined. The FimA II genotype was also associated with increased pocket depth, greater loss of attachment level, and patients co-infected with T. denticola and T. forsythia. PMID:26600627

  1. Mesenchymal stromal-cell transplants induce oligodendrocyte progenitor migration and remyelination in a chronic demyelination model.

    PubMed

    Jaramillo-Merchán, J; Jones, J; Ivorra, J L; Pastor, D; Viso-León, M C; Armengól, J A; Moltó, M D; Geijo-Barrientos, E; Martínez, S

    2013-08-29

    Demyelinating disorders such as leukodystrophies and multiple sclerosis are neurodegenerative diseases characterized by the progressive loss of myelin that may lead toward a chronic demyelination of the brain's white matter, impairing normal axonal conduction velocity and ultimately causing neurodegeneration. Current treatments modifying the pathological mechanisms are capable of ameliorating the disease; however, frequently, these therapies are not sufficient to repress the progressive demyelination into a chronic condition and permanent loss of function. To this end, we analyzed the effect that bone marrow-derived mesenchymal stromal cell (BM-MSC) grafts exert in a chronically demyelinated mouse brain. As a result, oligodendrocyte progenitors were recruited surrounding the graft due to the expression of various trophic signals by the grafted MSCs. Although there was no significant reaction in the non-grafted side, in the grafted regions oligodendrocyte progenitors were detected. These progenitors were derived from the nearby tissue as well as from the neurogenic niches, including the subependymal zone and dentate gyrus. Once near the graft site, the cells matured to myelinating oligodendrocytes. Finally, electrophysiological studies demonstrated that axonal conduction velocity was significantly increased in the grafted side of the fimbria. In conclusion, we demonstrate here that in chronic demyelinated white matter, BM-MSC transplantation activates oligodendrocyte progenitors and induces remyelination in the tissue surrounding the stem cell graft.

  2. The majority of enteroaggregative Escherichia coli strains produce the E. coli common pilus when adhering to cultured epithelial cells.

    PubMed

    Avelino, Fabiola; Saldaña, Zeus; Islam, Sohidul; Monteiro-Neto, Valerio; Dall'Agnol, Monique; Eslava, Carlos A; Girón, Jorge A

    2010-11-01

    Enteroaggregative Escherichia coli (EAEC) have emerged as a significant worldwide cause of chronic diarrhea in the pediatric population and in HIV patients. The vast majority of EAEC strains do not produce the aggregative adherence fimbriae I-III (AAFs) so far reported and thus, what adherence factors are present in these strains remains unknown. Here, we investigated the prevalence of the chromosomal E. coli common pilus (ECP) genes and ECP production amongst 130 EAEC strains of diverse origin as well as the role of ECP in EAEC adherence. Through multiplex PCR analysis we found that 96% of EAEC strains contained the ecpA structural pilin gene whereas only 3.1% and 5.4% were positive for AAF fimbrial genes aggA or aafA, respectively. Among the ecpA(+) strains, 63% produced ECP when adhering to cultured epithelial cells. An ecpA mutant derived from prototypic strain 042 (AAF/II(+)) was not altered in adherence suggesting that the AAF/II, and not ECP, plays a major role in this strain. In contrast, strain 278-1 (AAF(-)) deleted of the ecpA gene was significantly reduced in adherence to cultured epithelial cells. In all, these data indicate a potential role of ECP in adherence for EAEC strains lacking the known AAFs and that in association with other adhesive determinants, ECP may contribute to their survival and persistence within the host and in the environment.

  3. Stenotrophomonas maltophilia strains from cystic fibrosis patients: genomic variability and molecular characterization of some virulence determinants.

    PubMed

    Nicoletti, Mauro; Iacobino, Angelo; Prosseda, Gianni; Fiscarelli, Ersilia; Zarrilli, Raffaele; De Carolis, Elena; Petrucca, Andrea; Nencioni, Lucia; Colonna, Bianca; Casalino, Mariassunta

    2011-01-01

    The genetic relatedness of 52 Stenotrophomonas maltophilia strains, collected from various environmental and clinical sources, including cystic fibrosis (CF) patients, as well as the presence and the expression of some virulence-associated genes were studied. Pulsed-field gel electrophoresis (PFGE) analysis identified 47 profiles and three clusters of isolates with an identical PFGE pattern considered to be indistinguishable strains. Restriction fragment length polymorphism of the gyrB gene grouped the 52 strains into nine different profiles. Most CF clinical isolates (29 out of 41) showed profile 1, while the analysis of the hypervariable regions of the 16S rRNA gene revealed five distinct allelic variations, with the majority of CF isolates (23 out of 41) belonging to sequence group 1. Furthermore, the strains were characterized for motility and expression of virulence-associated genes, including genes encoding type-1 fimbriae, proteases (StmPr1 and StmPr2) and esterase. All S. maltophilia strains exhibited a very broad range of swimming and twitching motility, while none showed swarming motility. A complete smf-1 gene was PCR-amplified only from clinically derived S. maltophilia strains. Finally, the virulence of representative S. maltophilia strains impaired in the expression of proteases and esterase activities was evaluated by infecting larvae of the wax moth Galleria mellonella. The results obtained strongly indicate that the major extracellular protease StmPr1 may be a relevant virulence factor of S. maltophilia.

  4. Complete genome sequence and comparative analysis of the wild-type commensal Escherichia coli strain SE11 isolated from a healthy adult.

    PubMed

    Oshima, Kenshiro; Toh, Hidehiro; Ogura, Yoshitoshi; Sasamoto, Hiroyuki; Morita, Hidetoshi; Park, Sang-Hee; Ooka, Tadasuke; Iyoda, Sunao; Taylor, Todd D; Hayashi, Tetsuya; Itoh, Kikuji; Hattori, Masahira

    2008-12-01

    We sequenced and analyzed the genome of a commensal Escherichia coli (E. coli) strain SE11 (O152:H28) recently isolated from feces of a healthy adult and classified into E. coli phylogenetic group B1. SE11 harbored a 4.8 Mb chromosome encoding 4679 protein-coding genes and six plasmids encoding 323 protein-coding genes. None of the SE11 genes had sequence similarity to known genes encoding phage- and plasmid-borne virulence factors found in pathogenic E. coli strains. The comparative genome analysis with the laboratory strain K-12 MG1655 identified 62 poorly conserved genes between these two non-pathogenic strains and 1186 genes absent in MG1655. These genes in SE11 were mostly encoded in large insertion regions on the chromosome or in the plasmids, and were notably abundant in genes of fimbriae and autotransporters, which are cell surface appendages that largely contribute to the adherence ability of bacteria to host cells and bacterial conjugation. These data suggest that SE11 may have evolved to acquire and accumulate the functions advantageous for stable colonization of intestinal cells, and that the adhesion-associated functions are important for the commensality of E. coli in human gut habitat.

  5. Complete Genome Sequence and Comparative Analysis of the Wild-type Commensal Escherichia coli Strain SE11 Isolated from a Healthy Adult

    PubMed Central

    Oshima, Kenshiro; Toh, Hidehiro; Ogura, Yoshitoshi; Sasamoto, Hiroyuki; Morita, Hidetoshi; Park, Sang-Hee; Ooka, Tadasuke; Iyoda, Sunao; Taylor, Todd D.; Hayashi, Tetsuya; Itoh, Kikuji; Hattori, Masahira

    2008-01-01

    We sequenced and analyzed the genome of a commensal Escherichia coli (E. coli) strain SE11 (O152:H28) recently isolated from feces of a healthy adult and classified into E. coli phylogenetic group B1. SE11 harbored a 4.8 Mb chromosome encoding 4679 protein-coding genes and six plasmids encoding 323 protein-coding genes. None of the SE11 genes had sequence similarity to known genes encoding phage- and plasmid-borne virulence factors found in pathogenic E. coli strains. The comparative genome analysis with the laboratory strain K-12 MG1655 identified 62 poorly conserved genes between these two non-pathogenic strains and 1186 genes absent in MG1655. These genes in SE11 were mostly encoded in large insertion regions on the chromosome or in the plasmids, and were notably abundant in genes of fimbriae and autotransporters, which are cell surface appendages that largely contribute to the adherence ability of bacteria to host cells and bacterial conjugation. These data suggest that SE11 may have evolved to acquire and accumulate the functions advantageous for stable colonization of intestinal cells, and that the adhesion-associated functions are important for the commensality of E. coli in human gut habitat. PMID:18931093

  6. Identification of bapA in Strains of Salmonella enterica subsp. enterica Isolated from Wild Animals Kept in Captivity in Sinaloa, Mexico

    PubMed Central

    López-Valenzuela, Martin; Cárcamo-Aréchiga, Nora; Cota-Guajardo, Silvia; López-Salazar, Mayra; Montiel-Vázquez, Edith

    2016-01-01

    bapA, previously named stm2689, encodes the BapA protein, which, along with cellulose and fimbriae, constitutes biofilms. Biofilms are communities of microorganisms that grow in a matrix of exopolysaccharides and may adhere to living tissues or inert surfaces. Biofilm formation is associated with the ability to persist in different environments, which contributes to the pathogenicity of several species. We analyzed the presence of bapA in 83 strains belonging to 17 serovars of Salmonella enterica subsp. enterica from wildlife in captivity at Culiacan's Zoo and Mazatlán's Aquarium. Each isolate amplified a product of 667 bp, which corresponds to the expected size of the bapA initiator, with no observed variation between different serovars analyzed. bapA gene was found to be highly conserved in Salmonella and can be targeted for the genus-specific detection of this organism from different sources. Since bapA expression improves bacterial proliferation outside of the host and facilitates resistance to disinfectants and desiccation, the survival of Salmonella in natural habitats may be favored. Thus, the risk of bacterial contamination from these animals is increased. PMID:27379195

  7. A food-grade fimbrial adhesin FaeG expression system in Lactococcus lactis and Lactobacillus casei.

    PubMed

    Lu, W W; Wang, T; Wang, Y; Xin, M; Kong, J

    2016-03-01

    Enterotoxigenic Escherichia coli (ETEC) infection is the major cause of diarrhea in neonatal piglets. The fimbriae as colonizing factor in the pathogenesis of ETEC constitute a primary target for vaccination against ETEC. Lactic acid bacteria (LAB) are attractive tools to deliver antigens at the mucosal level. With the safety of genetically modified LAB in mind, a food-grade secretion vector (pALRc or pALRb) was constructed with DNA entirely from LAB, including the replicon, promoter, signal peptide, and selection marker alanine racemase gene (alr). To evaluate the feasibility of the system, the nuclease gene (nuc) from Staphylococcus aureus was used as a reporter to be expressed in both Lactococcus lactis and Lactobacillus casei. Subsequently, the extracellular secretion of the fimbrial adhesin FaeG of ETEC was confirmed by Western blot analysis. These results showed that this food-grade expression system has potential as the delivery vehicle for the safe use of genetically modified LAB for the development of vaccines against ETEC infection.

  8. Biofunctionalization of silicone polymers using poly(amidoamine) dendrimers and a mannose derivative for prolonged interference against pathogen colonization

    PubMed Central

    Lopez, Analette I.; Kumar, Amit; Planas, Megan R.; Li, Yan; Nguyen, Thuy V.; Cai, Chengzhi

    2011-01-01

    Despite numerous preventive strategies on bacterial adhesion, pathogenic biofilm formation remained the major cause of medical device-related infections. Bacterial interference is a promising strategy that uses pre-established biofilms of benign bacteria to serve as live, protective coating against pathogen colonization. However, the application of this strategy to silicone urinary catheters was hampered by low adherence of benign bacteria onto silicone materials. In this work, we present a general method for biofunctionalization of silicone (PDMS) as one of the most widely used materials for biomedical devices. We used mild CO2 plasma to activate PDMS surface followed by simple attachment of generation 5 (G5) poly(amidoamine) (PAMAM) dendrimers to generate an amino-terminated surface that were maintained even after storage in PBS buffer for 36 days. We then covalently attach a carboxy-terminated mannose derivative to the modified PDMS to promote the adherence of benign Escherichia coli 83972 expressing mannose-binding type 1 fimbriae. We demonstrated that dense, stable biofilms of E. coli 83972 could be established within 48 h on the mannose-coated PDMS. Significantly, this benign biofilm reduced the adherence of the uropathogenic Enterococcus faecalis by 104-fold after 72 hours, while the benign bacteria on the unmodified substrate by only 5.5-fold. PMID:21435713

  9. Identification of Escherichia coli F4ac-binding proteins in porcine milk fat globule membrane.

    PubMed

    Novakovic, Predrag; Huang, Yanyun Y; Lockerbie, Betty; Shahriar, Farshid; Kelly, John; Gordon, John R; Middleton, Dorothy M; Loewen, Matthew E; Kidney, Beverly A; Simko, Elemir

    2015-04-01

    F4ac-positive enterotoxigenic Escherichia coli (ETEC) must attach to the intestinal mucosa to cause diarrhea in piglets. Prevention of bacterial attachment to the intestinal mucosa is the most effective defense against ETEC-induced diarrhea. Porcine milk fat globule membranes (MFGM) were shown to be able to inhibit attachment of ETEC to the intestinal brush border; however, the specific components of porcine MFGM that inhibited attachment of ETEC to enterocytes were not identified. Accordingly, the purpose of this study was to identify F4ac-binding MFGM proteins by overlay Western blot and affinity chromatography. The proteome of porcine MFGM was characterized and the following F4ac-binding proteins were detected by overlay Western blot and affinity chromatography: lactadherin, butyrophilin, adipophilin, acyl-CoA synthetase 3, and fatty acid-binding protein 3. The biological function of these proteins was not investigated but it is possible that their interaction with F4ac fimbria interferes with bacterial attachment and colonization.

  10. Characterization of a planctomycete associated with the marine dinoflagellate Prorocentrum micans Her.

    PubMed

    Lage, Olga Maria

    2013-10-01

    During attempts to obtain axenic the cultures of the marine dinoflagellate Prorocentrum micans, a microorganism with peculiar features was isolated. This contaminant resisted the physical and antibiotic treatments performed. Subsequent characterization showed that in agar plates this microorganism develops round granular pink colonies. It is a salt-dependent mesophilic and chemoheterotrophic Gram negative bacterium with a rod to ovoid shape, presenting cell motility in young cultures. Cell division occurs by cell budding. The bacterium forms aggregates with a variable number of cells that are stacked by fibrillar glycoproteic material, the holdfast. A tuft of numerous short glycoproteic fimbriae emerges from one pole of the cell. Preeminent granular inclusions, also of glycoproteic nature, are present in the cytoplasm. Several structural and compositional aspects of the cell envelope and cytoplasm are provided. The production of fibrillar material and the existence of the polar appendages suggest that this microorganism should occur in aquatic environments bound to substrates and could be associated with P. micans in natural marine habitats. Based on the characteristics displayed, this microorganism is a member of the Planctomycetes, order Planctomycetales.

  11. Uropathogenic virulence factor FimH facilitates binding of uteropathogenic Escherichia coli to canine endometrium.

    PubMed

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

    2012-09-01

    Pyometra is a potentially life-threatening condition in bitches and is often caused by Escherichia coli infection. Both pathogenic and non-pathogenic E. coli strains commonly carry the genes for type 1 fimbriae that mediate bacterial adhesion onto host epithelium. To investigate whether the type 1 fimbrial adhesin, FimH, facilitates the binding of uropathogenic E. coli to canine endometrium, the fimH gene was insertionally inactivated in a pathogenic E. coli strain. The ability of E. coli to bind to canine endometrial epithelial cells was determined in vitro using canine uterine biopsies. Binding of the fimH mutant was only 0.3% of that of the wild type. Complementation of the mutation restored the phenotype to that of the parent. This study has developed an in vitro model that allows quantitative and qualitative assessment of bacterial binding to canine endometrium and has demonstrated that the fimH gene plays a role in adherence of pathogenic E. coli to canine endometrium.

  12. Foraging behavior of fish-eating sperm whales in the Gulf of Alaska in the presence and absence of fishing vessels

    NASA Astrophysics Data System (ADS)

    Thode, Aaron; Straley, Jan; Folkert, Kendall; O'Connell, Victoria; Tiemann, Christopher

    2005-09-01

    Historical whaling records indicate that sperm whales off southeast Alaska incorporate fish into their diets, particularly black cod (Anoploploma fimbria). Since 1995 this fact has become relevant to fisheries' concerns in the form of increased depredation encounters between longline fishermen and over 40 sperm whales. Since 2002 the SE Alaska Sperm Whale Avoidance Project (SEASWAP) has been studying this phenomenon using fishermen reports, photo-ID, biopsy, and (since 2004) passive acoustics using both towed arrays and autonomous recorders placed on longline deployments. By using acoustic multipath the range and depths of foraging whales can be determined. Findings to date indicate that, under natural conditions, sperm whales are foraging at mid-depth in the water column (e.g., 250 m in 500-m-deep water), and that their dive cycle durations are similar to those reported in other oceans. This information is being compared with depth measurements of black cod at various stages of their life cycle. There is increasing evidence that distinctive acoustic cues made by longline vessels lead to changes in diving and acoustic behavior by the animals, when the animals are less than 10 nautical miles away. [Work supported by the North Pacific Research Board.

  13. The Rcs regulon in Proteus mirabilis: implications for motility, biofilm formation, and virulence.

    PubMed

    Howery, Kristen E; Clemmer, Katy M; Rather, Philip N

    2016-11-01

    The overall role of the Rcs phosphorelay in Proteus mirabilis is largely unknown. Previous work had demonstrated that the Rcs phosphorelay represses the flhDC operon and activates the minCDE cell division inhibition system. To identify additional cellular functions regulated by the Rcs phosphorelay, an analysis of RNA-seq data was undertaken. In this report, the results of the RNA-sequencing are discussed with an emphasis on the predicted roles of the Rcs phosphorelay in swarmer cell differentiation, motility, biofilm formation, and virulence. RcsB is shown to activate genes important for differentiation and fimbriae formation, while repressing the expression of genes important for motility and virulence. Additionally, to follow up on the RNA-Seq data, we demonstrate that an rcsB mutant is deficient in its ability to form biofilm and exhibits enhanced virulence in a Galleria mellonella waxworm model. Overall, these results indicate the Rcs regulon in P. mirabilis extends beyond flagellar genes to include those involved in biofilm formation and virulence. Furthermore, the information presented in this study may provide clues to additional roles of the Rcs phosphorelay in other members of the Enterobacteriaceae.

  14. Proteus mirabilis and Urinary Tract Infections.

    PubMed

    Schaffer, Jessica N; Pearson, Melanie M

    2015-10-01

    Proteus mirabilis is a Gram-negative bacterium and is well known for its ability to robustly swarm across surfaces in a striking bulls'-eye pattern. Clinically, this organism is most frequently a pathogen of the urinary tract, particularly in patients undergoing long-term catheterization. This review covers P. mirabilis with a focus on urinary tract infections (UTI), including disease models, vaccine development efforts, and clinical perspectives. Flagella-mediated motility, both swimming and swarming, is a central facet of this organism. The regulation of this complex process and its contribution to virulence is discussed, along with the type VI-secretion system-dependent intra-strain competition, which occurs during swarming. P. mirabilis uses a diverse set of virulence factors to access and colonize the host urinary tract, including urease and stone formation, fimbriae and other adhesins, iron and zinc acquisition, proteases and toxins, biofilm formation, and regulation of pathogenesis. While significant advances in this field have been made, challenges remain to combatting complicated UTI and deciphering P. mirabilis pathogenesis.

  15. Measuring Escherichia coli Gene Expression during Human Urinary Tract Infections

    PubMed Central

    Mobley, Harry L. T.

    2016-01-01

    Extraintestinal Escherichia coli (E. coli) evolved by acquisition of pathogenicity islands, phage, plasmids, and DNA segments by horizontal gene transfer. Strains are heterogeneous but virulent uropathogenic isolates more often have specific fimbriae, toxins, and iron receptors than commensal strains. One may ask whether it is the virulence factors alone that are required to establish infection. While these virulence factors clearly contribute strongly to pathogenesis, bacteria must survive by metabolizing nutrients available to them. By constructing mutants in all major metabolic pathways and co-challenging mice transurethrally with each mutant and the wild type strain, we identified which major metabolic pathways are required to infect the urinary tract. We must also ask what else is E. coli doing in vivo? To answer this question, we examined the transcriptome of E. coli CFT073 in the murine model of urinary tract infection (UTI) as well as for E. coli strains collected and analyzed directly from the urine of patients attending either a urology clinic or a university health clinic for symptoms of UTI. Using microarrays and RNA-seq, we measured in vivo gene expression for these uropathogenic E. coli strains, identifying genes upregulated during murine and human UTI. Our findings allow us to propose a new definition of bacterial virulence. PMID:26784237

  16. Immunoglobulin domains in Escherichia coli and other enterobacteria: from pathogenesis to applications in antibody technologies.

    PubMed

    Bodelón, Gustavo; Palomino, Carmen; Fernández, Luis Ángel

    2013-03-01

    The immunoglobulin (Ig) protein domain is widespread in nature having a well-recognized role in proteins of the immune system. In this review, we describe the proteins containing Ig-like domains in Escherichia coli and enterobacteria, reporting their structural and functional properties, protein folding, and diverse biological roles. In addition, we cover the expression of heterologous Ig domains in E. coli owing to its biotechnological application for expression and selection of antibody fragments and full-length IgG molecules. Ig-like domains in E. coli and enterobacteria are frequently found in cell surface proteins and fimbrial organelles playing important functions during host cell adhesion and invasion of pathogenic strains, being structural components of pilus and nonpilus fimbrial systems and members of the intimin/invasin family of outer membrane (OM) adhesins. Ig-like domains are also found in periplasmic chaperones and OM usher proteins assembling fimbriae, in oxidoreductases and hydrolytic enzymes, ATP-binding cassette transporters, sugar-binding and metal-resistance proteins. The folding of most E. coli Ig-like domains is assisted by periplasmic chaperones, peptidyl-prolyl cis/trans isomerases and disulfide bond catalysts that also participate in the folding of antibodies expressed in this bacterium. The technologies for expression and selection of recombinant antibodies in E. coli are described along with their biotechnological potential.

  17. Global assessment of molecularly identified Anisakis Dujardin, 1845 (Nematoda: Anisakidae) in their teleost intermediate hosts.

    PubMed

    Kuhn, Thomas; Hailer, Frank; Palm, Harry W; Klimpel, Sven

    2013-05-01

    Here, we present the ITS ribosomal DNA (rDNA) sequence data on 330 larvae of nematodes of the genus Anisakis Dujardin, 1845 collected from 26 different bony fish species from 21 sampling locations and different climatic zones. New host records are provided for Anisakis simplex (Rudolphi, 1809) sensu stricto (s.s.) and A. pegreffli Campana-Rouget et Biocca, 1955 from Anoplopoma fimbria (Pallas) (Santa Barbara, East Pacific), A. typica (Diesing, 1860) from Caesio cuning (Bloch), Lepturacanthus savala (Cuvier) and Katsuwonus pelamis (Linnaeus) (Indonesia, West Pacific), A. simplex s.s. from Cololabis saira (Brevoort) (Hawaii, Central Pacific), A. simplex C of Nascetti et al. (1986) from Sebastolobus alascanus Bean (Santa Barbara, East Pacific) and A. physeteris Baylis, 1923 from Synaphobranchus kaupii Johnson (Namibia, East Atlantic). Comparison with host records from 60 previous molecular studies of Anisakis species reveals the teleost host range so far recorded for the genus. Perciform (57 species) and gadiform (21) fishes were the most frequently infected orders, followed by pleuronectiforms (15) and scorpaeniforms (15). Most commonly infected fish families were Scombridae (12), Gadidae (10), Carangidae (8) and Clupeidae (7), with Merluccius merluccius (Linnaeus) alone harbouring eight Anisakis species. Different intermediate host compositions implicate differing life cycles for the so far molecularly identified Anisakis sibling species.

  18. Acoustic and diving behavior of sperm whales (Physeter macrocephalus) during natural and depredation foraging in the Gulf of Alaska.

    PubMed

    Mathias, Delphine; Thode, Aaron M; Straley, Jan; Calambokidis, John; Schorr, Gregory S; Folkert, Kendall

    2012-07-01

    Sperm whales have depredated black cod (Anoplopoma fimbria) from demersal longlines in the Gulf of Alaska for decades, but the behavior has recently spread in intensity and geographic coverage. Over a three-year period 11 bioacoustic tags were attached to adult sperm whales off Southeast Alaska during both natural and depredation foraging conditions. Measurements of the animals' dive profiles and their acoustic behavior under both behavioral modes were examined for statistically significant differences. Two rough categories of depredation are identified: "deep" and "shallow." "Deep depredating" whales consistently surface within 500 m of a hauling fishing vessel, have maximum dive depths greater than 200 m, and display significantly different acoustic behavior than naturally foraging whales, with shorter inter-click intervals, occasional bouts of high "creak" rates, and fewer dives without creaks. "Shallow depredating" whales conduct dives that are much shorter, shallower, and more acoustically active than both the natural and deep depredating behaviors, with median creak rates three times that of natural levels. These results suggest that depredation efforts might be measured remotely with passive acoustic monitoring at close ranges.

  19. Relationship between sperm whale (Physeter macrocephalus) click structure and size derived from videocamera images of a depredating whale (sperm whale prey acquisition).

    PubMed

    Mathias, Delphine; Thode, Aaron; Straley, Jan; Folkert, Kendall

    2009-05-01

    Sperm whales have learned to depredate black cod (Anoplopoma fimbria) from longline deployments in the Gulf of Alaska. On May 31, 2006, simultaneous acoustic and visual recordings were made of a depredation attempt by a sperm whale at 108 m depth. Because the whale was oriented perpendicularly to the camera as it contacted the longline at a known distance from the camera, the distance from the nose to the hinge of the jaw could be estimated. Allometric relationships obtained from whaling data and skeleton measurements could then be used to estimate both the spermaceti organ length and total length of the animal. An acoustic estimate of animal length was obtained by measuring the inter-pulse interval (IPI) of clicks detected from the animal and using empirical formulas to convert this interval into a length estimate. Two distinct IPIs were extracted from the clicks, one yielding a length estimate that matches the visually-derived length to within experimental error. However, acoustic estimates of spermaceti organ size, derived from standard sound production theories, are inconsistent with the visual estimates, and the derived size of the junk is smaller than that of the spermaceti organ, in contradiction with known anatomical relationships.

  20. Influence of Sublethal Antibiotic Concentrations on Bacterial Adherence to Saliva-Treated Hydroxyapatite

    PubMed Central

    Peros, W. J.; Gibbons, R. J.

    1982-01-01

    The influence of growth in the presence of sublethal concentrations of nine antibiotics on the ability of certain potentially odontopathic bacteria to attach to saliva-treated hydroxyapatite surfaces which mimic teeth was studied. Cells of Actinomyces viscosus LY7 and S2, Bacteroides gingivalis 381, Capnocytophaga ochraceus 6, and Actinobacillus actinomycetemcomitans N27 attached in lower numbers to saliva-treated hydroxyapatite when grown in the presence of 50% of the minimum inhibitory concentration of tetracycline. Electron microscopic observations of negatively stained preparations indicated that tetracycline-grown A. viscosus LY7 cells had fewer fimbriae than did untreated cells, which may account for the impaired ability of the treated cells to attach. However, cells of Actinomyces naeslundii L13 and S4 attached in higher numbers when grown in the presence of tetracycline, clindamycin, erythromycin, chloramphenicol, or neomycin. Streptococcus mutans strains H12 and JBP also exhibited increased adherence to saliva-treated hydroxyapatite when grown in the presence of 50 or 25% of the minimum inhibitory concentration of penicillin. Thus, growth in the presence of sublethal antibiotic concentrations could increase as well as decrease the adherence of bacteria to saliva-treated hydroxyapatite. Antibiotic-grown cells of the Actinomyces strains showed enhanced hemagglutination activity, but this did not correlate with their ability to attach to saliva-treated hydroxyapatite. Sublethal concentrations of antibiotics in the growth media also affected the coaggregation reactions of several organisms; the effects were specific for one member of the coaggregation pair. Images PMID:6274799

  1. The effects of upaB deletion and the double/triple deletion of upaB, aatA, and aatB genes on pathogenicity of avian pathogenic Escherichia coli.

    PubMed

    Zhu-Ge, Xiang-Kai; Pan, Zi-Hao; Tang, Fang; Mao, Xiang; Hu, Lin; Wang, Shao-Hui; Xu, Bin; Lu, Cheng-Ping; Fan, Hong-Jie; Dai, Jian-Jun

    2015-12-01

    Autotransporters (ATs) are associated with pathogenesis of Avian Pathogenic Escherichia coli (APEC). The molecular characterization of APEC ATs can provide insights about their relevance to APEC pathogenesis. Here, we characterized a conventional autotransporter UpaB in APEC DE205B genome. The upaB existed in 41.9 % of 236 APEC isolates and was predominantly associated with ECOR B2 and D. Our studies showed that UpaB mediates the DE205B adhesion in DF-1 cells, and enhances autoaggregation and biofilm formation of fimbria-negative E. coli AAEC189 (MG1655Δfim) in vitro. Deletion of upaB of DE205B attenuates the virulence in duck model and early colonization in the duck lungs during APEC systemic infection. Furthermore, double and triple deletion of upaB, aatA, and aatB genes cumulatively attenuated DE205B adhesion in DF-1 cells, accompanying with decreased 50 % lethal dose (LD50) in duck model and the early colonization in the duck lungs. However, DE205BΔupaB/ΔaatA/ΔaatB might "compensate" the influence of gene deletion by upregulating the expression of fimbrial adhesin genes yqiL, yadN, and vacuolating autotransporter vat during early colonization of APEC. Finally, we demonstrated that vaccination with recombinant UpaB, AatA, and AatB proteins conferred protection against colisepticemia caused by DE205B infection in duck model.

  2. Subnanometric Roughness Affects the Deposition and Mobile Adhesion of Escherichia coli on Silanized Glass Surfaces.

    PubMed

    Sharma, Sumedha; Jaimes-Lizcano, Yuly Andrea; McLay, Ryan B; Cirino, Patrick C; Conrad, Jacinta C

    2016-05-31

    We investigate the deposition and transient adhesion of Escherichia coli on alkyl and fluoroalkyl silanized glass surfaces of different carbon chain lengths. The rate at which bacteria deposit onto these surfaces decreases as the shear stress is increased from 3 to 67 mPa, but trends in the deposition rate across all surfaces cannot be predicted from extended DLVO calculations of the interaction potential. As the surface root-mean-square (rms) roughness increases, the deposition rate increases and the percentage of motile tethered cells decreases. Furthermore, on surfaces of root-mean-square roughness of less than 0.2 nm, bacteria exhibit mobile adhesion, for which surface-associated cells linearly translate distances greater than approximately 1.5 times their average body length along the flow direction. E. coli bacteria with and without flagella exhibit mobile adhesion, indicating that this behavior is not driven by these appendages. Cells that express fimbriae do not exhibit mobile adhesion. These results suggest that even subnanoscale roughness can influence the deposition and transient adhesion of bacteria and imply that strategies to reduce frictional interactions by making cells or surfaces smoother may help to control the initial fouling of surfaces by E. coli bacteria.

  3. Afa/Dr diffusely adhering Escherichia coli strain C1845 induces neutrophil extracellular traps that kill bacteria and damage human enterocyte-like cells.

    PubMed

    Marin-Esteban, Viviana; Turbica, Isabelle; Dufour, Guillaume; Semiramoth, Nicolas; Gleizes, Aude; Gorges, Roseline; Beau, Isabelle; Servin, Alain L; Lievin-Le Moal, Vanessa; Sandré, Catherine; Chollet-Martin, Sylvie

    2012-05-01

    We recently documented the neutrophil response to enterovirulent diffusely adherent Escherichia coli expressing Afa/Dr fimbriae (Afa/Dr DAEC), using the human myeloid cell line PLB-985 differentiated into fully mature neutrophils. Upon activation, particularly during infections, neutrophils release neutrophil extracellular traps (NETs), composed of a nuclear DNA backbone associated with antimicrobial peptides, histones, and proteases, which entrap and kill pathogens. Here, using fluorescence microscopy and field emission scanning electron microscopy, we observed NET production by PLB-985 cells infected with the Afa/Dr wild-type (WT) E. coli strain C1845. We found that these NETs were able to capture, immobilize, and kill WT C1845 bacteria. We also developed a coculture model of human enterocyte-like Caco-2/TC7 cells and PLB-985 cells previously treated with WT C1845 and found, for the first time, that the F-actin cytoskeleton of enterocyte-like cells is damaged in the presence of bacterium-induced NETs and that this deleterious effect is prevented by inhibition of protease release. These findings provide new insights into the neutrophil response to bacterial infection via the production of bactericidal NETs and suggest that NETs may damage the intestinal epithelium, particularly in situations such as inflammatory bowel diseases.

  4. Ultrastructural and chemotaxonomic analysis of a xylanolytic strain of Cryptococcus adeliensis isolated from sheep droppings in Spain.

    PubMed

    Velázquez, Encarna; del Villar, María; Grondona, Isabel; Monte, Enrique; González-Villa, Tomás

    2006-09-01

    Cryptococcus adeliensis was initially described as a psycrophilic species containing a single strain CBS 8351(T) isolated from decayed algae in Terre Adelie (Antartida). Later, a second strain of this species was isolated from an immunosuppressed patient affected by leukaemia in Germany and recently several strains from this species have been found in human patients and pigeon droppings of the same country. In this study, we isolated from sheep droppings in Spain a xylanolytic strain named LEVX01 that was phenotypically related to the strain CBS 8351(T) and showed a 100% similarity in the D1/D2 domain and 5.8S-ITS region sequences with respect to the remaining described strains of C. adeliensis. These findings suggest that this species has a wide geographical distribution and that the animal faeces are a common habitat for C. adeliensis. The chemotaxonomic analyses showed the absence of detectable amounts of xylose in the cell walls of the strains LEVX01 and CBS8351(T) in contrast to other Cryptococcus species. Interestingly, the ultrastructural study showed the presence of fimbriae in these two strains that could be involved in the attachment to the host cells and, as occurs in Candida albicans, they could also be a pathogenicity factor for the man.

  5. Identification of Enterohemorrhagic Escherichia coli O26:H− Genes Required for Intestinal Colonization in Calves

    PubMed Central

    van Diemen, Pauline M.; Dziva, Francis; Stevens, Mark P.; Wallis, Timothy S.

    2005-01-01

    Enterohemorrhagic Escherichia coli (EHEC) infections in humans are an important public health problem and are commonly acquired via contact with ruminant feces. The serogroups that are predominantly associated with human infection in the United States and Europe are O157 and O26. Serotypes O157:H7 and O26:H− differ in their virulence and tissue tropism in calves and therefore may colonize calves by distinct mechanisms. The mechanisms underlying EHEC intestinal colonization and pathogenesis are poorly understood. Signature-tagged mutagenesis was used to identify 59 genes of EHEC O26:H− that are required for the intestinal colonization of calves. Our results indicate important roles for locus of enterocyte effacement (LEE)-encoded type III secreted proteins in intestinal colonization. In addition, colonization is facilitated by cytotoxins, putative type III secreted proteins unlinked to the LEE, a putative fimbrial operon, and numerous genes involved in central metabolism and transport and genes of unknown function. Our data also imply that the elaboration of type I fimbriae by EHEC O26:H− is disadvantageous for persistence within the bovine intestines. These observations have important implications for the design of vaccines to control these important zoonotic pathogens. PMID:15731074

  6. Double dissociation of fornix and caudate nucleus lesions on acquisition of two water maze tasks: further evidence for multiple memory systems.

    PubMed

    Packard, M G; McGaugh, J L

    1992-06-01

    The present study examined the effect of lesions of the caudate nucleus or fimbria-fornix on the acquisition of two water maze tasks. In both tasks, two rubber balls with different visual patterns were used as platforms (i.e., cues). The "correct" cue was attached to a submerged rectangular platform and could be mounted by an animal to escape the water. The "incorrect" cue was attached to a thin round pedestal and could not be mounted. In a spatial version of the task, the correct cue was located in the same quadrant of the maze on all trials, whereas the visual pattern on the cue was varied from trial to trial. Lesions of the fornix, but not the caudate nucleus, impaired acquisition of this spatial task in relation to control animals. In a simultaneous visual discrimination version of the task, the correct cue on all trials was one with a specific visual pattern, and the spatial location of the correct cue was varied from trial to trial. Lesions of the caudate nucleus, but not the fornix, impaired acquisition of this visual discrimination task in relation to control animals. The double dissociation observed supports the hypothesis that the hippocampus and caudate nucleus are parts of systems that differ in the type of memory they mediate.

  7. MOLECULAR CHARACTERIZATION OF VIRULENCE AND ANTIMICROBIAL SUSCEPTIBILITY PROFILES OF UROPATHOGENIC ESCHERICHIA COLI FROM PATIENTS IN A TERTIARY HOSPITAL, SOUTHERN THAILAND.

    PubMed

    Themphachanal, Monchanok; Kongpheng, Suttiporn; Rattanachuay, Pattamarat; Khianngam, Saowapar; Singkhamanan, Kamonnut; Sukhumungoon, Pharanai

    2015-11-01

    Among uropathogens, uropathogenic Escherichia coli (UPEC) is the most common cause of urinary tract infection (UTI) worldwide, but clinical aspects due to this bacterial species is not fully understood in southern Thailand. Two hundred fifty-four UPEC isolates from patients admitted to Maharaj Nakhon Si Thammarat Hospital, southern Thailand were examined for crucial virulence genes, showing that 33.5% contained at least one of the virulence, genes tested. Genes encoding P fimbria, cytotoxic necrotizing factor-1 and α-hemolysin constituted the majority (15.8%) carried by UPEC isolates. Phylogenetic group classification revealed that 57.5% of UPEC belonged to group D. Antimicrobial susceptibility tests showed that 70.5% and 65.1% of the isolates were resistant to ciprofloxacin and norfloxacin, respectively. Moreover, 50.0% of UPEC were capable of producing extended spectrum beta-lactamases. These findings should be of benefit for more appropriate treatment of UTI patients in this region of Thailand. Keywords: uropathogenic Escherichia coli, antibiotics resistance, cnfl, hlyA, pap, Thailand

  8. A vaccine candidate for post-weaning diarrhea in swine constructed with a live attenuated Salmonella delivering Escherichia coli K88ab, K88ac, FedA, and FedF fimbrial antigens and its immune responses in a murine model.

    PubMed

    Hur, Jin; Stein, Barry D; Lee, John Hwa

    2012-07-01

    In order to construct a novel vaccine candidate for preventing post-weaning diarrhea in swine, the individual genes for Escherichia coli K88ab, K88ac, FedA, and FedF fimbriae were inserted into a secretion plasmid pBP244 containing asd, lepB, secA, and secB. These were transformed into Salmonella Typhimurium Δlon ΔcpxR Δasd. Secretion of the individual recombinant fimbrial antigens was confirmed by immunoblot analysis. Groups 1 and 2 mice received a single oral dose of the vaccine mixture and S. Typhimurium carrying pBP244 only as a control, respectively. In groups 3 and 4, mice were primed and boosted with the vaccine mixture and S. Typhimurium carrying pBP244 only as a control, respectively. In general, all immunized mice had significantly increased serum immunoglobulin (Ig)G (P < 0.05) and intestinal secretory IgA against the individual fimbrial antigens compared with those mice in the control group. In the IgG2a and IgG1 titer assay, only IgG2a titer was increased in group 1, while both IgG2a and IgG1 titers were increased in group 3. Furthermore, the vaccine strains were not detected in the excreted feces of any immunized mice. Thus, the vaccine candidate can be highly immunogenic and be safe to the environment.

  9. Chronic cocaine administration causes extensive white matter damage in brain: diffusion tensor imaging and immunohistochemistry studies.

    PubMed

    Narayana, Ponnada A; Herrera, Juan J; Bockhorst, Kurt H; Esparza-Coss, Emilio; Xia, Ying; Steinberg, Joel L; Moeller, F Gerard

    2014-03-30

    The effect of chronic cocaine exposure on multiple white matter structures in rodent brain was examined using diffusion tensor imaging (DTI), locomotor behavior, and end point histology. The animals received either cocaine at a dose of 100mg/kg (N=19), or saline (N=17) for 28 days through an implanted osmotic minipump. The animals underwent serial DTI scans, locomotor assessment, and end point histology for determining the expressions of myelin basic protein (MBP), neurofilament-heavy protein (NF-H), proteolipid protein (PLP), Nogo-A, aquaporin-4 (AQP-4), and growth associated protein-43 (GAP-43). Differences in the DTI measures were observed in the splenium (scc) and genu (gcc) of the corpus callosum (cc), fimbria (fi), and the internal capsule (ic). A significant increase in the activity in the fine motor movements and a significant decrease in the number of rearing events were observed in the cocaine-treated animals. Reduced MBP and Nogo-A and increased GAP-43 expressions were most consistently observed in these structures. A decrease in the NF-H expression was observed in fi and ic. The reduced expression of Nogo-A and the increased expression of GAP-43 may suggest destabilization of axonal connectivity and increased neurite growth with aberrant connections. Increased GAP-43 suggests drug-induced plasticity or a possible repair mechanism response. The findings indicated that multiple white matter tracts are affected following chronic cocaine exposure.

  10. Autolysis and autoaggregation in Pseudomonas aeruginosa colony morphology mutants.

    PubMed

    D'Argenio, David A; Calfee, M Worth; Rainey, Paul B; Pesci, Everett C

    2002-12-01

    Two distinctive colony morphologies were noted in a collection of Pseudomonas aeruginosa transposon insertion mutants. One set of mutants formed wrinkled colonies of autoaggregating cells. Suppressor analysis of a subset of these mutants showed that this was due to the action of the regulator WspR and linked this regulator (and the chemosensory pathway to which it belongs) to genes that encode a putative fimbrial adhesin required for biofilm formation. WspR homologs, related in part by a shared GGDEF domain, regulate cell surface factors, including aggregative fimbriae and exopolysaccharides, in diverse bacteria. The second set of distinctive insertion mutants formed colonies that lysed at their center. Strains with the most pronounced lysis overproduced the Pseudomonas quinolone signal (PQS), an extracellular signal that interacts with quorum sensing. Autolysis was suppressed by mutation of genes required for PQS biosynthesis, and in one suppressed mutant, autolysis was restored by addition of synthetic PQS. The mechanism of autolysis may involve activation of the endogenous prophage and phage-related pyocins in the genome of strain PAO1. The fact that PQS levels correlated with autolysis suggests a fine balance in natural populations of P. aeruginosa between survival of the many and persistence of the few.

  11. Detection of neuritic plaques in Alzheimer’s disease by magnetic resonance microscopy

    PubMed Central

    Benveniste, Helene; Einstein, Gillian; Kim, Katie R.; Hulette, Christine; Johnson, G. Allan

    1999-01-01

    Magnetic resonance microscopy (MRM) theoretically provides the spatial resolution and signal-to-noise ratio needed to resolve neuritic plaques, the neuropathological hallmark of Alzheimer’s disease (AD). Two previously unexplored MR contrast parameters, T2* and diffusion, are tested for plaque-specific contrast to noise. Autopsy specimens from nondemented controls (n = 3) and patients with AD (n = 5) were used. Three-dimensional T2* and diffusion MR images with voxel sizes ranging from 3 × 10−3 mm3 to 5.9 × 10−5 mm3 were acquired. After imaging, specimens were cut and stained with a microwave king silver stain to demonstrate neuritic plaques. From controls, the alveus, fimbria, pyramidal cell layer, hippocampal sulcus, and granule cell layer were detected by either T2* or diffusion contrast. These structures were used as landmarks when correlating MRMs with histological sections. At a voxel resolution of 5.9 × 10−5 mm3, neuritic plaques could be detected by T2*. The neuritic plaques emerged as black, spherical elements on T2* MRMs and could be distinguished from vessels only in cross-section when presented in three dimension. Here we provide MR images of neuritic plaques in vitro. The MRM results reported provide a new direction for applying this technology in vivo. Clearly, the ability to detect and follow the early progression of amyloid-positive brain lesions will greatly aid and simplify the many possibilities to intervene pharmacologically in AD. PMID:10570201

  12. Fungal lectin of Peltigera canina induces chemotropism of compatible Nostoc cells by constriction-relaxation pulses of cyanobiont cytoskeleton

    PubMed Central

    Díaz, Eva Maria; Vicente-Manzanares, Miguel; Sacristan, Mara; Legaz, Maria-Estrella

    2011-01-01

    A glycosylated arginase acting as a fungal lectin from Peltigera canina is able to produce recruitment of cyanobiont Nostoc cells and their adhesion to the hyphal surface. This implies that the cyanobiont would develop organelles to motility toward the chemoattractant. However when visualized by transmission electron microscopy, Nostoc cells recently isolated from P. canina thallus do not reveal any motile, superficial organelles, although their surface was covered by small spindles and serrated layer related to gliding. The use of S-(3,4-dichlorobenzyl)isothiourea, blebbistatin, phalloidin and latrunculin A provide circumstantial evidence that actin microfilaments rather than MreB, the actin-like protein from prokaryota, and probably, an ATPase which develops contractile function similar to that of myosin II, are involved in cell motility. These experimental facts, the absence of superficial elements (fimbriae, pili or flagellum) related to cell movement, and the appearance of sunken cells during of after movement verified by scanning electron microscopy, support the hypothesis that the motility of lichen cyanobionts could be achieved by contraction-relaxation episodes of the cytoskeleton induced by fungal lectin act as a chemoattractant. PMID:21897128

  13. Fungal lectin of Peltigera canina induces chemotropism of compatible Nostoc cells by constriction-relaxation pulses of cyanobiont cytoskeleton.

    PubMed

    Díaz, Eva Maria; Vicente-Manzanares, Miguel; Sacristan, Mara; Vicente, Carlos; Legaz, Maria-Estrella

    2011-10-01

    A glycosylated arginase acting as a fungal lectin from Peltigera canina is able to produce recruitment of cyanobiont Nostoc cells and their adhesion to the hyphal surface. This implies that the cyanobiont would develop organelles to motility towards the chemoattractant. However when visualized by transmission electron microscopy, Nostoc cells recently isolated from P. canina thallus do not reveal any motile, superficial organelles, although their surface was covered by small spindles and serrated layer related to gliding. The use of S-(3,4-dichlorobenzyl)isothiourea, blebbistatin, phalloidin and latrunculin A provide circumstantial evidence that actin microfilaments rather than MreB, the actin-like protein from prokaryota, and, probably, an ATPase which develops contractile function similar to that of myosin II, are involved in cell motility. These experimental facts, the absence of superficial elements (fimbriae, pili or flagellum) related to cell movement, and the appearance of sunken cells during of after movement verified by scanning electron microscopy, support the hypothesis that the motility of lichen cyanobionts could be achieved by contraction-relaxation episodes of the cytoskeleton induced by fungal lectin act as a chemoattractant.

  14. Alkaloids: an overview of their antibacterial, antibiotic-enhancing and antivirulence activities.

    PubMed

    Cushnie, T P Tim; Cushnie, Benjamart; Lamb, Andrew J

    2014-11-01

    With reports of pandrug-resistant bacteria causing untreatable infections, the need for new antibacterial therapies is more pressing than ever. Alkaloids are a large and structurally diverse group of compounds that have served as scaffolds for important antibacterial drugs such as metronidazole and the quinolones. In this review, we highlight other alkaloids with development potential. Natural, semisynthetic and synthetic alkaloids of all classes are considered, looking first at those with direct antibacterial activity and those with antibiotic-enhancing activity. Potent examples include CJ-13,136, a novel actinomycete-derived quinolone alkaloid with a minimum inhibitory concentration of 0.1 ng/mL against Helicobacter pylori, and squalamine, a polyamine alkaloid from the dogfish shark that renders Gram-negative pathogens 16- to >32-fold more susceptible to ciprofloxacin. Where available, information on toxicity, structure-activity relationships, mechanisms of action and in vivo activity is presented. The effects of alkaloids on virulence gene regulatory systems such as quorum sensing and virulence factors such as sortases, adhesins and secretion systems are also described. The synthetic isoquinoline alkaloid virstatin, for example, inhibits the transcriptional regulator ToxT in Vibrio cholerae, preventing expression of cholera toxin and fimbriae and conferring in vivo protection against intestinal colonisation. The review concludes with implications and limitations of the described research and directions for future research.

  15. The role of H4 flagella in Escherichia coli ST131 virulence.

    PubMed

    Kakkanat, Asha; Totsika, Makrina; Schaale, Kolja; Duell, Benjamin L; Lo, Alvin W; Phan, Minh-Duy; Moriel, Danilo G; Beatson, Scott A; Sweet, Matthew J; Ulett, Glen C; Schembri, Mark A

    2015-11-09

    Escherichia coli sequence type 131 (ST131) is a globally dominant multidrug resistant clone associated with urinary tract and bloodstream infections. Most ST131 strains exhibit resistance to multiple antibiotics and cause infections associated with limited treatment options. The largest sub-clonal ST131 lineage is resistant to fluoroquinolones, contains the type 1 fimbriae fimH30 allele and expresses an H4 flagella antigen. Flagella are motility organelles that contribute to UPEC colonisation of the upper urinary tract. In this study, we examined the specific role of H4 flagella in ST131 motility and interaction with host epithelial and immune cells. We show that the majority of H4-positive ST131 strains are motile and are enriched for flagella expression during static pellicle growth. We also tested the role of H4 flagella in ST131 through the construction of specific mutants, over-expression strains and isogenic mutants that expressed alternative H1 and H7 flagellar subtypes. Overall, our results revealed that H4, H1 and H7 flagella possess conserved phenotypes with regards to motility, epithelial cell adhesion, invasion and uptake by macrophages. In contrast, H4 flagella trigger enhanced induction of the anti-inflammatory cytokine IL-10 compared to H1 and H7 flagella, a property that may contribute to ST131 fitness in the urinary tract.

  16. Flagella interact with ionic plant lipids to mediate adherence of pathogenic Escherichia coli to fresh produce plants.

    PubMed

    Rossez, Yannick; Holmes, Ashleigh; Wolfson, Eliza B; Gally, David L; Mahajan, Arvind; Pedersen, Henriette L; Willats, William G T; Toth, Ian K; Holden, Nicola J

    2014-07-01

    Bacterial attachment to plant and animal surfaces is generally thought to constitute the initial step in colonization, requiring adherence factors such as flagella and fimbriae. We describe the molecular mechanism underpinning flagella-mediated adherence to plant tissue for the foodborne pathogen, enterohaemorrhagic Escherichia coli. Escherichia coli H7 flagella interacted with a sulphated carbohydrate (carrageenan) on a glycan array, which occurred in a dose-dependent manner. Adherence of E. coli O157 : H-expressing flagella of serotype H7, H6 or H48 to plants associated with outbreaks from fresh produce and to Arabidopsis thaliana, was dependent on flagella interactions with phospholipids and sulpholipids in plasma membranes. Adherence of purified H7 and H48 flagella to carrageenan was reduced at higher concentrations of KH2 PO4 or KCl, showing an ionic basis to the interactions. Purified H7 flagella were observed to physically interact with plasma membranes in spinach plants and in A.thaliana. The results show a specific interaction between E. coli H7, H6 and H48 flagella and ionic lipids in plant plasma membranes. The work extends our understanding of the molecular mechanisms underpinning E.coli flagella targeting of plant hosts and suggests a generic mechanism of recognition common in eukaryotic hosts belonging to different biological kingdoms.

  17. The profile of Porphyromonas gingivalis kgp biotype and fimA genotype mosaic in subgingival plaque samples.

    PubMed

    Nadkarni, Mangala A; Chhour, Kim-Ly; Chapple, Cheryl C; Nguyen, Ky-Anh; Hunter, Neil

    2014-12-01

    Combined analysis of allelic variation of the virulence-associated, strain-specific lys-gingipain gene (kgp) and major fimbrial gene (fimA) of Porphyromonas gingivalis was undertaken in 116 subgingival plaque samples to understand the kgp biotype and fimA genotype profile in a subject-specific manner. Allelic variation in the polyadhesin domain of kgp from P. gingivalis strains 381 (ATCC 33277), HG66 and W83 generated four isoforms corresponding to four biotypes of P. gingivalis. Similarly, variation in the fimA subunit of the fimA gene cluster of P. gingivalis resulted in six fimA genotypes. Strain-specific differential PCR was performed for kgp and fimA using DNA isolated from subgingival plaque samples. Our findings demonstrate that all of the P. gingivalis kgp biotypes detected in this study were predominantly associated with the fimA II genotype. Dominance of kgp biotypes 381 or HG66 combined with fimA II fimbriae could imply an adaptive strategy by P. gingivalis to generate the fittest strains for survival in the host environment.

  18. In vitro adherence patterns of Shigella serogroups to bovine recto-anal junction squamous epithelial (RSE) cells are similar to those of Escherichia coli O157.

    PubMed

    Kudva, Indira T

    2012-04-01

    The aims of this study were to determine whether Shigella species, which are human gastrointestinal pathogens, can adhere to cattle recto-anal junction squamous epithelial (RSE) cells using a recently standardized in vitro adherence assay, and to compare their adherence patterns with that of Escherichia coli O157. Shigella dysenteriae (serogroup A), S. flexneri (serogroup B), S. boydii (serogroup C), and S. sonnei (serogroup D) were tested in adherence assays using both RSE and HEp-2 cells, in the presence or absence of D+mannose. Escherichia coli O157, which adheres to RSE cells in a Type I fimbriae-independent manner, was used as a positive control. Shigella serogroups A, B, D, but not C adhered to RSE cells with distinct adherence patterns in the presence of D+mannose. No such distinction could be made between the four Shigella serogroups based on the HEp-2 cell adherence patterns. Thus, this study provides evidence that certain Shigella serogroups adhere to RSE cells in a manner that is similar to the adherence pattern of E. coli O157. These unexpected observations of in vitro binding of these foodborne human pathogens to cells of the bovine gastrointestinal tract warrant evaluation of Shigella carriage by cattle using both experimental and observational studies, especially for serogroups B and D. Such studies are currently underway.

  19. General N-and O-Linked Glycosylation of Lipoproteins in Mycoplasmas and Role of Exogenous Oligosaccharide

    PubMed Central

    Daubenspeck, James M.; Jordan, David S.; Simmons, Warren; Renfrow, Matthew B.; Dybvig, Kevin

    2015-01-01

    The lack of a cell wall, flagella, fimbria, and other extracellular appendages and the possession of only a single membrane render the mycoplasmas structurally simplistic and ideal model organisms for the study of glycoconjugates. Most species have genomes of about 800 kb and code for few proteins predicted to have a role in glycobiology. The murine pathogens Mycoplasma arthritidis and Mycoplasma pulmonis have only a single gene annotated as coding for a glycosyltransferase but synthesize glycolipid, polysaccharide and glycoproteins. Previously, it was shown that M. arthritidis glycosylated surface lipoproteins through O-linkage. In the current study, O-linked glycoproteins were similarly found in M. pulmonis and both species of mycoplasma were found to also possess N-linked glycans at residues of asparagine and glutamine. Protein glycosylation occurred at numerous sites on surface-exposed lipoproteins with no apparent amino acid sequence specificity. The lipoproteins of Mycoplasma pneumoniae also are glycosylated. Glycosylation was dependent on the glycosidic linkages from host oligosaccharides. As far as we are aware, N-linked glycoproteins have not been previously described in Gram-positive bacteria, the organisms to which the mycoplasmas are phylogenetically related. The findings indicate that the mycoplasma cell surface is heavily glycosylated with implications for the modulation of mycoplasma-host interactions. PMID:26599081

  20. Epithelialization of mouse ovarian tumor cells originating in the fallopian tube stroma

    PubMed Central

    Hua, Yuanyuan; Choi, Pui-Wah; Trachtenberg, Alexander J.; Ng, Allen C.; Kuo, Winston P.; Ng, Shu-Kay; Dinulescu, Daniela M.; Matzuk, Martin M.; Berkowitz, Ross S.; Ng, Shu-Wing

    2016-01-01

    Epithelial ovarian carcinoma accounts for 90% of all ovarian cancer and is the most deadly gynecologic malignancy. Recent studies have suggested that fallopian tube fimbriae can be the origin of cells for high-grade serous subtype of epithelial ovarian carcinoma (HGSOC). A mouse HGSOC model with conditional Dicer-Pten double knockout (Dicer-Pten DKO) developed primary tumors, intriguingly, from the fallopian tube stroma. We examined the growth and epithelial phenotypes of the Dicer-Pten DKO mouse tumor cells contributable by each gene knockout. Unlike human ovarian epithelial cancer cells that expressed full-length E-cadherin, the Dicer-Pten DKO stromal tumor cells expressed cleaved E-cadherin fragments and metalloproteinase 2, a mixture of epithelial and mesenchymal markers. Although the Dicer-Pten DKO tumor cells lost the expression of mature microRNAs as expected, they showed high levels of tRNA fragment expression and enhanced AKT activation due to the loss of PTEN function. Introduction of a Dicer1-expressing construct into the DKO mouse tumor cells significantly reduced DNA synthesis and the cell growth rate, with concurrent diminished adhesion and ZO1 epithelial staining. Hence, it is likely that the loss of Dicer promoted mesenchymal-epithelial transition in fallopian tube stromal cells, and in conjunction with Pten loss, further promoted cell proliferation and epithelial-like tumorigenesis. PMID:27602775

  1. Comparative Genome Analysis of Enterobacter cloacae

    PubMed Central

    Liu, Wing-Yee; Wong, Chi-Fat; Chung, Karl Ming-Kar; Jiang, Jing-Wei; Leung, Frederick Chi-Ching

    2013-01-01

    The Enterobacter cloacae species includes an extremely diverse group of bacteria that are associated with plants, soil and humans. Publication of the complete genome sequence of the plant growth-promoting endophytic E. cloacae subsp. cloacae ENHKU01 provided an opportunity to perform the first comparative genome analysis between strains of this dynamic species. Examination of the pan-genome of E. cloacae showed that the conserved core genome retains the general physiological and survival genes of the species, while genomic factors in plasmids and variable regions determine the virulence of the human pathogenic E. cloacae strain; additionally, the diversity of fimbriae contributes to variation in colonization and host determination of different E. cloacae strains. Comparative genome analysis further illustrated that E. cloacae strains possess multiple mechanisms for antagonistic action against other microorganisms, which involve the production of siderophores and various antimicrobial compounds, such as bacteriocins, chitinases and antibiotic resistance proteins. The presence of Type VI secretion systems is expected to provide further fitness advantages for E. cloacae in microbial competition, thus allowing it to survive in different environments. Competition assays were performed to support our observations in genomic analysis, where E. cloacae subsp. cloacae ENHKU01 demonstrated antagonistic activities against a wide range of plant pathogenic fungal and bacterial species. PMID:24069314

  2. A proteoliposome formulation derived from Bordetella pertussis induces protection in two murine challenge models.

    PubMed

    Fernández, Sonsire; Fajardo, Esther M; Mandiarote, Aleida; Año, Gemma; Padrón, Maria A; Acosta, Michel; Cabrera, Rubén A; Riverón, Luis A; Álvarez, Maydelis; Blaín, Kirenia; Fariñas, Mildrey; Cardoso, Daniel; García, Luis G; Campa, Concepción; Pérez, José L

    2013-01-01

    Whooping cough remains a health problem despite high vaccination coverage. It has been recommended that development of new strategies provide long-lasting immunity. The aim of this work was to evaluate the potential of proteoliposomes (PL) extracted from Bordetella pertussis as a vaccine candidate against whooping cough. The size of the B. pertussis PL was estimated to be 96.7 ± 50.9 nm by Scanning Correlation Spectroscopy and the polydispersity index was 0.268. Western blots using monoclonal antibodies revealed the presence of pertussis toxin, pertactin, and fimbriae 3. The Limulus Amebocyte Lisate (LAL) assay showed endotoxin levels lower than those reported for whole cell pertussis licensed vaccines, while the Pyrogen Test indicated 75 ng/mL/Kg. The PL showed high protection capacity in mouse challenge models. There was 89.7% survival in the intracerebral challenge and total reduction of the number of CFU in the intranasal challenge. No significant differences (p > 0.05) were observed between mice immunized with B. pertussis PL and the Cuban DTwP vaccine, whichever challenge model used. These results encouraged us to continue the development of the B. pertussis PL as a component of a new combined vaccine formulated with tetanus and diphtheria toxoids or as a booster dose for adolescents and adults.

  3. [Development studies of lyophilized standard diphtheria-tetanus-pertussis vaccines].

    PubMed

    Ozcengiz, Erkan; Unver, Derya; Cayan, H Hüseyin; Atakan Ablay, Pinar; Kanik, Esin

    2007-04-01

    In this study, diphtheria, tetanus and pertussis vaccine components were prepared as the formulations of diphtheria-tetanus-pertussis (DTP), diphtheria-tetanus (DT) for children, diphtheria-tetanus (Td) for adults, and tetanus toxoid (TT), respectively. Alhydrogel-adsorbed vaccines prepared to contain the stabilizing substances were lyophilized and the immunogenicity tests were carried out both in vivo and in vitro. The potencies of the tetanus component of the vaccines were obtained by the lethal challenge test in mice. The values were found as 144.86 IU/ml for lyophilized adsorbed (LA)-DTP, 116.5 IU/ml for LA-DT, 98.25 IU/ml for LA-Td and 96.2 IU/ml for LA-TT. Anti-tetanus IgG and anti-diphteria IgG levels determined by ELISA method were found high in the sera taken from the mice immunized with the above-mentioned vaccines. Anti-B.pertussis fimbria IgG antibody levels were also high by both ELISA and microagglutination tests. The test preparations were then compared to adsorbed liquid vaccines and it was shown that the components were quite stable in the lyophilized formulations. It was concluded that the formulations prepared in this study can be used as standard vaccines after being calibrated against World Health Organization standards.

  4. Effect of heat inactivation of serum on Bordetella pertussis antibody determination by enzyme-linked immunosorbent assay.

    PubMed

    Lopez, A L; Pineda, E; Garakian, A; Cherry, J D

    1998-01-01

    The effect of heat inactivation on Bordetella pertussis antibodies determined by enzyme-linked immunosorbent assay (ELISA) was studied. Sera were heated at increasing temperatures (from 30 to 50 degrees C at 5 degrees C increments and from 52 to 70 degrees C at 2 degrees C increments). Between 30 and 50 degrees C, no significant differences were observed in immunoglobin G (IgG) antibodies to pertussis toxin (PT). From 50 to 56 degrees C the antibody values were twofold higher than those of uninactivated sera; at 64 degrees C the values were 3.6- to 9.1-fold higher. The increase in PT IgG antibody values was more pronounced in sera with low antibody values. ELISA antibody values of sera from a vaccine trial were determined in unheated and heat inactivated sera. The geometric mean value (GMV) of the heat inactivated samples was 3.2 times the geometric mean value of the uninactivated sera. ELISA IgG antibodies to filamentous hemagglutinin, fimbriae-2, and pertactin were studied, and the values of heat inactivated sera did not differ significantly from the values of the uninactivated sera. Our findings indicate that heat inactivation of sera leads to higher, variable, and false-positive PT and IgG values.

  5. Leukoencephalopathy Associated with Parvovirus Infection in Cretan Hound Puppies▿

    PubMed Central

    Schaudien, D.; Polizopoulou, Z.; Koutinas, A.; Schwab, S.; Porombka, D.; Baumgärtner, W.; Herden, C.

    2010-01-01

    Leukoencephalopathies in dogs encompass presumably inherited conditions such as leukodystrophies, hypomyelination or spongiform degeneration, but other causes, such as virus infections and toxic or nutritional factors, might also play a contributory role. In this report, we provide evidence of parvovirus infection and replication in the brains of five 6-week-old Cretan hound puppies suffering from a puppy shaker syndrome and leukoencephalopathy. Although these puppies belonged to two different litters, they were closely related, tracing back two generations to the same sire. Histologically, a mild to moderate lymphohistiocytic meningitis, with focal lymphohistiocytic leukoencephalitis in two animals, and a mild to moderate vacuolation with myelin loss, mainly in the white matter of the cerebellum was detected. Vacuolation was also found in the corpus callosum, fimbria hippocampi, mesencephalon, capsula interna, basal ganglia, and hypothalamus. By immunohistology and in situ hybridization, either parvoviral antigen, DNA, mRNA, or replicative intermediate DNA were detected in the cerebellum, hippocampus, periventricular areas, corpus callosum, cerebral cortex, medulla oblongata, and spinal cord. Parvovirus antigen, DNA, and mRNA were present in cells of the outer granular layer of the cerebellum and in periventricular cells, most likely representing spongioblasts, glial cells, neurons, endothelial cells, occasional macrophages, and ependymal cells. Sequencing revealed canine parvovirus type 2 stretches. Thus, an association of parvovirus infection with the leukoencephalopathy seems likely, possibly facilitated by a genetic predisposition due to the mode of inbreeding in this particular dog breed. PMID:20592142

  6. Characterization of five ECF sigma factors in the genome of Pseudomonas syringae pv. syringae B728a.

    PubMed

    Thakur, Poulami Basu; Vaughn-Diaz, Vanessa L; Greenwald, Jessica W; Gross, Dennis C

    2013-01-01

    Pseudomonas syringae pv. syringae B728a, a bacterial pathogen of bean, utilizes large surface populations and extracellular signaling to initiate a fundamental change from an epiphytic to a pathogenic lifestyle. Extracytoplasmic function (ECF) sigma (σ) factors serve as important regulatory factors in responding to various environmental signals. Bioinformatic analysis of the B728a genome revealed 10 ECF sigma factors. This study analyzed deletion mutants of five previously uncharacterized ECF sigma factor genes in B728a, including three FecI-type ECF sigma factors (ECF5, ECF6, and ECF7) and two ECF sigma factors placed in groups ECF11 and ECF18. Transcriptional profiling by qRT-PCR analysis of ECF sigma factor mutants was used to measure expression of their associated anti-sigma and outer membrane receptor proteins, and expression of genes associated with production of extracellular polysaccharides, fimbriae, glycine betaine and syringomycin. Notably, the B728aΔecf7 mutant displayed reduced swarming and had decreased expression of CupC fimbrial genes. Growth and pathogenicity assays, using a susceptible bean host, revealed that none of the tested sigma factor genes are required for in planta growth and lesion formation.

  7. Genome-Wide Screening of Genes Required for Swarming Motility in Escherichia coli K-12▿ †

    PubMed Central

    Inoue, Tetsuyoshi; Shingaki, Ryuji; Hirose, Shotaro; Waki, Kaori; Mori, Hirotada; Fukui, Kazuhiro

    2007-01-01

    Escherichia coli K-12 has the ability to migrate on semisolid media by means of swarming motility. A systematic and comprehensive collection of gene-disrupted E. coli K-12 mutants (the Keio collection) was used to identify the genes involved in the swarming motility of this bacterium. Of the 3,985 nonessential gene mutants, 294 were found to exhibit a strongly repressed-swarming phenotype. Further, 216 of the 294 mutants displayed no significant defects in swimming motility; therefore, the 216 genes were considered to be specifically associated with the swarming phenotype. The swarming-associated genes were classified into various functional categories, indicating that swarming is a specialized form of motility that requires a wide variety of cellular activities. These genes include genes for tricarboxylic acid cycle and glucose metabolism, iron acquisition, chaperones and protein-folding catalysts, signal transduction, and biosynthesis of cell surface components, such as lipopolysaccharide, the enterobacterial common antigen, and type 1 fimbriae. Lipopolysaccharide and the enterobacterial common antigen may be important surface-acting components that contribute to the reduction of surface tension, thereby facilitating the swarm migration in the E. coli K-12 strain. PMID:17122336

  8. Defining the human hippocampus in cerebral magnetic resonance images—An overview of current segmentation protocols

    PubMed Central

    Konrad, C.; Ukas, T.; Nebel, C.; Arolt, V.; Toga, A.W.; Narr, K.L.

    2011-01-01

    Due to its crucial role for memory processes and its relevance in neurological and psychiatric disorders, the hippocampus has been the focus of neuroimaging research for several decades. In vivo measurement of human hippocampal volume and shape with magnetic resonance imaging has become an important element of neuroimaging research. Nevertheless, volumetric findings are still inconsistent and controversial for many psychiatric conditions including affective disorders. Here we review the wealth of anatomical protocols for the delineation of the hippocampus in MR images, taking into consideration 71 different published protocols from the neuroimaging literature, with an emphasis on studies of affective disorders. We identified large variations between protocols in five major areas. 1) The inclusion/exclusion of hippocampal white matter (alveus and fimbria), 2) the definition of the anterior hippocampal–amygdala border, 3) the definition of the posterior border and the extent to which the hippocampal tail is included, 4) the definition of the inferior medial border of the hippocampus, and 5) the use of varying arbitrary lines. These are major sources of variance between different protocols. In contrast, the definitions of the lateral, superior, and inferior borders are less disputed. Directing resources to replication studies that incorporate characteristics of the segmentation protocols presented herein may help resolve seemingly contradictory volumetric results between prior neuroimaging studies and facilitate the appropriate selection of protocols for manual or automated delineation of the hippocampus for future research purposes. PMID:19447182

  9. Whooping Cough in Adults: A Series of Severe Cases.

    PubMed

    Zycinska, Katarzyna; Cieplak, M; Chmielewska, M; Nitsch-Osuch, A; Klaczkow, A; Hadzik-Blaszczyk, M; Kur, Z; Wardyn, K A

    2016-12-31

    Bordetella pertussis is a gram-negative aerobic coccobacillus causing contagious respiratory tract disease called whooping cough. The virulence factors consist of pertussis toxin, filamentous hemagglutinin, fimbriae, lipooligosaccharide, and adenylate cyclase toxin. The disease causes a worldwide threat to public health despite a high vaccination coverage. The course of whooping cough in adults is frequently atypical, causing difficulty in diagnosis. In this report we present five patients hospitalized with Bordetella pertussis infection manifesting atypical and severe symptoms. The diagnosis was based on serological tests: serum concentration of specific antibodies against pertussis toxin and sputum cultures. We observed a wide spectrum of symptoms, from benign (sinus pain - 80 %, headaches - 20 %), through moderate (hemoptysis - 40 %; chest pain 60 %) to severe symptoms (cardiac arrhythmia - 40 %; syncope - 60 %). Bordetella pertussis infection can cause life-threatening complications and exacerbation of concomitant chronic diseases. Most vaccination programs cover only the first few months of life. Booster doses should be considered in adults, especially those immunocompromised or with pulmonary complications, but also in healthcare workers who are exposed to the contagion and also may spread the infection.

  10. Dancing to Another Tune—Adhesive Moonlighting Proteins in Bacteria

    PubMed Central

    Kainulainen, Veera; Korhonen, Timo K.

    2014-01-01

    Biological moonlighting refers to proteins which express more than one function. Moonlighting proteins occur in pathogenic and commensal as well as in Gram-positive and Gram-negative bacteria. The canonical functions of moonlighting proteins are in essential cellular processes, i.e., glycolysis, protein synthesis, chaperone activity, and nucleic acid stability, and their moonlighting functions include binding to host epithelial and phagocytic cells, subepithelia, cytoskeleton as well as to mucins and circulating proteins of the immune and hemostatic systems. Sequences of the moonlighting proteins do not contain known motifs for surface export or anchoring, and it has remained open whether bacterial moonlighting proteins are actively secreted to the cell wall or whether they are released from traumatized cells and then rebind onto the bacteria. In lactobacilli, ionic interactions with lipoteichoic acids and with cell division sites are important for surface localization of the proteins. Moonlighting proteins represent an abundant class of bacterial adhesins that are part of bacterial interactions with the environment and in responses to environmental changes. Multifunctionality in bacterial surface proteins appears common: the canonical adhesion proteins fimbriae express also nonadhesive functions, whereas the mobility organelles flagella as well as surface proteases express adhesive functions. PMID:24833341

  11. Temperature control of molecular circuit switch responsible for virulent phenotype expression in uropathogenic Escherichia coli

    NASA Astrophysics Data System (ADS)

    Samoilov, Michael

    2010-03-01

    The behavior and fate of biological organisms are to a large extent dictated by their environment, which can be often viewed as a collection of features and constraints governed by physics laws. Since biological systems comprise networks of molecular interactions, one such key physical property is temperature, whose variations directly affect the rates of biochemical reactions involved. For instance, temperature is known to control many gene regulatory circuits responsible for pathogenicity in bacteria. One such example is type 1 fimbriae (T1F) -- the foremost virulence factor in uropathogenic E. coli (UPEC), which accounts for 80-90% of all community-acquired urinary tract infections (UTIs). The expression of T1F is randomly `phase variable', i.e. individual cells switch between virulent/fimbriate and avirulent/afimbriate phenotypes, with rates regulated by temperature. Our computational investigation of this process, which is based on FimB/FimE recombinase-mediated inversion of fimS DNA element, offers new insights into its discrete-stochastic kinetics. In particular, it elucidates the logic of T1F control optimization to the host temperature and contributes further understanding toward the development of novel therapeutic approaches to UPEC-caused UTIs.

  12. Identification of a CO2 responsive regulon in Bordetella.

    PubMed

    Hester, Sara E; Lui, Minghsun; Nicholson, Tracy; Nowacki, Daryl; Harvill, Eric T

    2012-01-01

    Sensing the environment allows pathogenic bacteria to coordinately regulate gene expression to maximize survival within or outside of a host. Here we show that Bordetella species regulate virulence factor expression in response to carbon dioxide levels that mimic in vivo conditions within the respiratory tract. We found strains of Bordetella bronchiseptica that did not produce adenylate cyclase toxin (ACT) when grown in liquid or solid media with ambient air aeration, but produced ACT and additional antigens when grown in air supplemented to 5% CO(2). Transcriptome analysis and quantitative real time-PCR analysis revealed that strain 761, as well as strain RB50, increased transcription of genes encoding ACT, filamentous hemagglutinin (FHA), pertactin, fimbriae and the type III secretion system in 5% CO(2) conditions, relative to ambient air. Furthermore, transcription of cyaA and fhaB in response to 5% CO(2) was increased even in the absence of BvgS. In vitro analysis also revealed increases in cytotoxicity and adherence when strains were grown in 5% CO(2). The human pathogens B. pertussis and B. parapertussis also increased transcription of several virulence factors when grown in 5% CO(2), indicating that this response is conserved among the classical bordetellae. Together, our data indicate that Bordetella species can sense and respond to physiologically relevant changes in CO(2) concentrations by regulating virulence factors important for colonization, persistence and evasion of the host immune response.

  13. Identification of a sustained neurogenic zone at the dorsal surface of the adult mouse hippocampus and its regulation by the chemokine SDF-1.

    PubMed

    Belmadani, Abdelhak; Ren, Dongjun; Bhattacharyya, Bula J; Rothwangl, Katharina B; Hope, Thomas J; Perlman, Harris; Miller, Richard J

    2015-11-01

    We identified a previously unknown neurogenic region at the dorsal surface of the hippocampus; (the "subhippocampal zone," SHZ) in the adult brain. Using a reporter mouse in which SHZ cells and their progeny could be traced through the expression of EGFP under the control of the CXCR4 chemokine receptor promoter we observed the presence of a pool of EGFP expressing cells migrating in direction of the dentate gyrus (DG), which is maintained throughout adulthood. This population appeared to originate from the SHZ where cells entered a caudal migratory stream (aCMS) that included the fimbria, the meninges and the DG. Deletion of CXCR4 from neural stem cells (NSCs) or neuroinflammation resulted in the appearance of neurons in the DG, which were the result of migration of NSCs from the SHZ. Some of these neurons were ectopically placed. Our observations indicate that the SHZ is a neurogenic zone in the adult brain through migration of NSCs in the aCMS. Regulation of CXCR4 signaling in these cells may be involved in repair of the DG and may also give rise to ectopic granule cells in the DG in the context of neuropathology.

  14. Label-free identification of the hippocampus and surrounding structures by multiphoton microscopy

    NASA Astrophysics Data System (ADS)

    Wang, Shu; Jiang, Liwei; Du, Huiping; Wang, Xingfu; Zheng, Liqin; Li, Lianhuang; Zhuo, Shuangmu; Zhu, Xiaoqin; Chen, Jianxin

    2016-05-01

    The hippocampus is one of the essential neuroanatomical substrates and plays an important role in different neurological illnesses. In this work, multiphoton microscopy (MPM) based on intrinsic nonlinear optical processes two-photon excited fluorescence (TPEF) and second harmonic generation (SHG), was applied to label-freely detect the entire hippocampus and surrounding structures in high-magnification imaging, as well as acquire large-scale MPM images at subcellular resolution. It was found that MPM has the capability to identify cornu ammonis, dentate gyrus (DG), alveus, and fimbria of the entire hippocampus, choroid plexus in lateral ventricles, and white matter tracts. MPM also can be used to quantitatively describe the differences of the cellular nucleus in the cornu ammonis and the DG, further identify the morphological features of hippocampal subfields. In addition, the surrounding structures of the hippocampus including the lateral ventricles and white matter serve as useful information to determine the position of the hippocampus. Our results suggest that with the development of the clinical feasibility of two-photon fiberscopes and microendoscope probes, MPM has the potential for in vivo intraoperative identification and monitoring of hippocampus-related lesions without the need for tissue labelling or fluorescent markers.

  15. Role of enteroaggregative Escherichia coli virulence factors in uropathogenesis.

    PubMed

    Boll, Erik J; Struve, Carsten; Boisen, Nadia; Olesen, Bente; Stahlhut, Steen G; Krogfelt, Karen A

    2013-04-01

    A multiresistant clonal Escherichia coli O78:H10 strain qualifying molecularly as enteroaggregative Escherichia coli (EAEC) was recently shown to be the cause of a community-acquired outbreak of urinary tract infection (UTI) in greater Copenhagen, Denmark, in 1991. This marks the first time EAEC has been associated with an extraintestinal disease outbreak. Importantly, the outbreak isolates were recovered from the urine of patients with symptomatic UTI, strongly implying urovirulence. Here, we sought to determine the uropathogenic properties of the Copenhagen outbreak strain and whether these properties are conferred by the EAEC-specific virulence factors. We demonstrated that through expression of aggregative adherence fimbriae, the principal adhesins of EAEC, the outbreak strain exhibited pronouncedly increased adherence to human bladder epithelial cells compared to prototype uropathogenic strains. Moreover, the strain was able to produce distinct biofilms on abiotic surfaces, including urethral catheters. These findings suggest that EAEC-specific virulence factors increase uropathogenicity and may have played a significant role in the ability of the strain to cause a community-acquired outbreak of UTI. Thus, inclusion of EAEC-specific virulence factors is warranted in future detection and characterization of uropathogenic E. coli.

  16. Ginkgolic acids and Ginkgo biloba extract inhibit Escherichia coli O157:H7 and Staphylococcus aureus biofilm formation.

    PubMed

    Lee, Jin-Hyung; Kim, Yong-Guy; Ryu, Shi Yong; Cho, Moo Hwan; Lee, Jintae

    2014-03-17

    Infection by enterohemorrhagic Escherichia coli O157:H7 (EHEC) is a worldwide problem, and there is no effective therapy. Biofilm formation is closely related to EHEC infection and is also a mechanism of antimicrobial resistance. Antibiofilm screening of 560 purified phytochemicals against EHEC showed that ginkgolic acids C15:1 and C17:1 at 5μg/ml and Ginkgo biloba extract at 100μg/ml significantly inhibited EHEC biofilm formation on the surfaces of polystyrene and glass, and on nylon membranes. Importantly, at their working concentrations, ginkgolic acids and G. biloba extract did not affect bacterial growth. Transcriptional analyses showed that ginkgolic acid C15:1 repressed curli genes and prophage genes in EHEC, and these findings were in-line with reduced fimbriae production and biofilm reductions. Interestingly, ginkgolic acids and G. biloba extract did not inhibit the biofilm formation of a commensal E. coli K-12 strain. In addition, ginkgolic acids and G. biloba extract inhibited the biofilm formation of three Staphylococcus aureus strains. The findings of this study suggest that plant secondary metabolites represent an important resource for biofilm inhibitors.

  17. Natural rpoS mutations contribute to population heterogeneity in Escherichia coli O157:H7 strains linked to the 2006 US spinach-associated outbreak.

    PubMed

    Carter, Michelle Qiu; Louie, Jacqueline W; Huynh, Steven; Parker, Craig T

    2014-12-01

    We previously reported significantly different acid resistance between curli variants derived from the same Escherichia coli O157:H7 strain, although the curli fimbriae were not associated with this phenotypic divergence. Here we investigated the underlying molecular mechanism by examining the genes encoding the common transcriptional regulators of curli biogenesis and acid resistance. rpoS null mutations were detected in all curli-expressing variants of the 2006 spinach-associated outbreak strains, whereas a wild-type rpoS was present in all curli-deficient variants. Consequently curli-expressing variants were much more sensitive to various stress challenges than curli-deficient variants. This loss of general stress fitness appeared solely to be the result of rpoS mutation since the stress resistances could be restored in curli-expressing variants by a functional rpoS. Comparative transcriptomic analyses between the curli variants revealed a large number of differentially expressed genes, characterized by the enhanced expression of metabolic genes in curli-expressing variants, but a marked decrease in transcription of genes related to stress resistances. Unlike the curli-expressing variants of the 1993 US hamburger-associated outbreak strains (Applied Environmental Microbiology 78: 7706-7719), all curli-expressing variants of the 2006 spinach-associated outbreak strains carry a functional rcsB gene, suggesting an alternative mechanism governing intra-strain phenotypic divergence in E. coli O157:H7.

  18. Cultivation of 'Candidatus Liberibacter asiaticus', 'Ca. L. africanus', and 'Ca. L. americanus' associated with huanglongbing.

    PubMed

    Sechler, A; Schuenzel, E L; Cooke, P; Donnua, S; Thaveechai, N; Postnikova, E; Stone, A L; Schneider, W L; Damsteegt, V D; Schaad, N W

    2009-05-01

    A new medium designated Liber A has been designed and used to successfully cultivate all three 'Candidatus Liberibacter spp.,' the suspect causative agents of huanglongbing (HLB) in citrus. The medium containing citrus vein extract and a growth factor sustained growth of 'Ca. Liberibacter spp.' for four or five single-colony transfers before viability declined. Colonies, positive for 'Ca. L. asiaticus' by a 16s-based rDNA real-time polymerase chain reaction (RT-PCR) assay and sequencing, were irregular-shaped, convex, and 0.1 to 0.3 mm after 3 to 4 days. Suspect 'Ca. L. asiaticus' and 'Ca. L. americanus' cells were observed in infected tissue and on agar culture by scanning electron microscopy. The cells were ovoid to rod shaped, 0.3 to 0.4 by 0.5 to 2.0 microm, often with fimbriae-like appendages. Two strains of 'Ca. L. asiaticus' and one of 'Ca. L. americanus' grown on Liber A medium were pathogenic on citrus and could be isolated from noninoculated tissues of inoculated trees and seedlings 9 and 2 months later, respectively. The identity was confirmed by RT-PCR and 16s rDNA sequencing. This is the first report of the cultivation and pathogenicity of 'Ca. L. asiaticus' and 'Ca. L. americanus' associated with symptoms of HLB.

  19. The role of H4 flagella in Escherichia coli ST131 virulence

    PubMed Central

    Kakkanat, Asha; Totsika, Makrina; Schaale, Kolja; Duell, Benjamin L.; Lo, Alvin W.; Phan, Minh-Duy; Moriel, Danilo G.; Beatson, Scott A.; Sweet, Matthew J.; Ulett, Glen C.; Schembri, Mark A.

    2015-01-01

    Escherichia coli sequence type 131 (ST131) is a globally dominant multidrug resistant clone associated with urinary tract and bloodstream infections. Most ST131 strains exhibit resistance to multiple antibiotics and cause infections associated with limited treatment options. The largest sub-clonal ST131 lineage is resistant to fluoroquinolones, contains the type 1 fimbriae fimH30 allele and expresses an H4 flagella antigen. Flagella are motility organelles that contribute to UPEC colonisation of the upper urinary tract. In this study, we examined the specific role of H4 flagella in ST131 motility and interaction with host epithelial and immune cells. We show that the majority of H4-positive ST131 strains are motile and are enriched for flagella expression during static pellicle growth. We also tested the role of H4 flagella in ST131 through the construction of specific mutants, over-expression strains and isogenic mutants that expressed alternative H1 and H7 flagellar subtypes. Overall, our results revealed that H4, H1 and H7 flagella possess conserved phenotypes with regards to motility, epithelial cell adhesion, invasion and uptake by macrophages. In contrast, H4 flagella trigger enhanced induction of the anti-inflammatory cytokine IL-10 compared to H1 and H7 flagella, a property that may contribute to ST131 fitness in the urinary tract. PMID:26548325

  20. S-layers at second glance? Altiarchaeal grappling hooks (hami) resemble archaeal S-layer proteins in structure and sequence

    PubMed Central

    Perras, Alexandra K.; Daum, Bertram; Ziegler, Christine; Takahashi, Lynelle K.; Ahmed, Musahid; Wanner, Gerhard; Klingl, Andreas; Leitinger, Gerd; Kolb-Lenz, Dagmar; Gribaldo, Simonetta; Auerbach, Anna; Mora, Maximilian; Probst, Alexander J.; Bellack, Annett; Moissl-Eichinger, Christine

    2015-01-01

    The uncultivated “Candidatus Altiarchaeum hamiconexum” (formerly known as SM1 Euryarchaeon) carries highly specialized nano-grappling hooks (“hami”) on its cell surface. Until now little is known about the major protein forming these structured fibrous cell surface appendages, the genes involved or membrane anchoring of these filaments. These aspects were analyzed in depth in this study using environmental transcriptomics combined with imaging methods. Since a laboratory culture of this archaeon is not yet available, natural biofilm samples with high Ca. A. hamiconexum abundance were used for the entire analyses. The filamentous surface appendages spanned both membranes of the cell, which are composed of glycosyl-archaeol. The hami consisted of multiple copies of the same protein, the corresponding gene of which was identified via metagenome-mapped transcriptome analysis. The hamus subunit proteins, which are likely to self-assemble due to their predicted beta sheet topology, revealed no similiarity to known microbial flagella-, archaella-, fimbriae- or pili-proteins, but a high similarity to known S-layer proteins of the archaeal domain at their N-terminal region (44–47% identity). Our results provide new insights into the structure of the unique hami and their major protein and indicate their divergent evolution with S-layer proteins. PMID:26106369

  1. Transcriptional analysis of the MrpJ network: modulation of diverse virulence-associated genes and direct regulation of mrp fimbrial and flhDC flagellar operons in Proteus mirabilis.

    PubMed

    Bode, Nadine J; Debnath, Irina; Kuan, Lisa; Schulfer, Anjelique; Ty, Maureen; Pearson, Melanie M

    2015-06-01

    The enteric bacterium Proteus mirabilis is associated with a significant number of catheter-associated urinary tract infections (UTIs). Strict regulation of the antagonistic processes of adhesion and motility, mediated by fimbriae and flagella, respectively, is essential for disease progression. Previously, the transcriptional regulator MrpJ, which is encoded by the mrp fimbrial operon, has been shown to repress both swimming and swarming motility. Here we show that MrpJ affects an array of cellular processes beyond adherence and motility. Microarray analysis found that expression of mrpJ mimicking levels observed during UTIs leads to differential expression of 217 genes related to, among other functions, bacterial virulence, type VI secretion, and metabolism. We probed the molecular mechanism of transcriptional regulation by MrpJ using transcriptional reporters and chromatin immunoprecipitation (ChIP). Binding of MrpJ to two virulence-associated target gene promoters, the promoters of the flagellar master regulator flhDC and mrp itself, appears to be affected by the condensation state of the native chromosome, although both targets share a direct MrpJ binding site proximal to the transcriptional start. Furthermore, an mrpJ deletion mutant colonized the bladders of mice at significantly lower levels in a transurethral model of infection. Additionally, we observed that mrpJ is widely conserved in a collection of recent clinical isolates. Altogether, these findings support a role of MrpJ as a global regulator of P. mirabilis virulence.

  2. Resource subsidies from multi-trophic aquaculture affect isotopic niche width in wild blue mussels (Mytilus edulis)

    NASA Astrophysics Data System (ADS)

    Weldrick, C. K.; Jelinski, D. E.

    2016-05-01

    Blue mussels (Mytilus edulis) are boreo-temperate, filter-feeding bivalves common to intertidal areas. As filter-feeders they have been employed in open-water, multi-tropic aquaculture systems to reduce organic benthic loading though the exploitation of suspended particulate organic materials. We compared δ13C and δ15N signatures and the isotopic niches of mussels growing in, and adjacent to, an integrated multi-trophic aquaculture (IMTA) farm in British Columbia, Canada, and using this information evaluated the contribution of aquaculture-derived effluent to their diet. Farm-sampled mussels had the least intraspecific isotopic variation compared to mussels sampled at the reference site. The interaction between time (i.e. sampling dates) and site did not significantly affect the isotopic composition of mussels; however significant variation was detected in δ15N values as a function of sampling date and particulate organic matter. A two-source isotopic mixing model indicated that marine particulate organic matter and IMTA farm effluent were approximately equal in importance (~ 46 % and ~ 54 %, respectively) to the diet of IMTA-retrieved mussels. Uptake of IMTA farm waste by M. edulis supports their use as economic extractives while also mitigating farmed sablefish (Anoplopoma fimbria) nutrient loading to the aquatic environment.

  3. Mesenchymal stromal-cell transplants induce oligodendrocyte progenitor migration and remyelination in a chronic demyelination model

    PubMed Central

    Jaramillo-Merchán, J; Jones, J; Ivorra, J L; Pastor, D; Viso-León, M C; Armengól, J A; Moltó, M D; Geijo-Barrientos, E; Martínez, S

    2013-01-01

    Demyelinating disorders such as leukodystrophies and multiple sclerosis are neurodegenerative diseases characterized by the progressive loss of myelin that may lead toward a chronic demyelination of the brain's white matter, impairing normal axonal conduction velocity and ultimately causing neurodegeneration. Current treatments modifying the pathological mechanisms are capable of ameliorating the disease; however, frequently, these therapies are not sufficient to repress the progressive demyelination into a chronic condition and permanent loss of function. To this end, we analyzed the effect that bone marrow-derived mesenchymal stromal cell (BM-MSC) grafts exert in a chronically demyelinated mouse brain. As a result, oligodendrocyte progenitors were recruited surrounding the graft due to the expression of various trophic signals by the grafted MSCs. Although there was no significant reaction in the non-grafted side, in the grafted regions oligodendrocyte progenitors were detected. These progenitors were derived from the nearby tissue as well as from the neurogenic niches, including the subependymal zone and dentate gyrus. Once near the graft site, the cells matured to myelinating oligodendrocytes. Finally, electrophysiological studies demonstrated that axonal conduction velocity was significantly increased in the grafted side of the fimbria. In conclusion, we demonstrate here that in chronic demyelinated white matter, BM-MSC transplantation activates oligodendrocyte progenitors and induces remyelination in the tissue surrounding the stem cell graft. PMID:23990019

  4. An in silico model for identification of small RNAs in whole bacterial genomes: characterization of antisense RNAs in pathogenic Escherichia coli and Streptococcus agalactiae strains

    PubMed Central

    Pichon, Christophe; du Merle, Laurence; Caliot, Marie Elise; Trieu-Cuot, Patrick; Le Bouguénec, Chantal

    2012-01-01

    Characterization of small non-coding ribonucleic acids (sRNA) among the large volume of data generated by high-throughput RNA-seq or tiling microarray analyses remains a challenge. Thus, there is still a need for accurate in silico prediction methods to identify sRNAs within a given bacterial species. After years of effort, dedicated software were developed based on comparative genomic analyses or mathematical/statistical models. Although these genomic analyses enabled sRNAs in intergenic regions to be efficiently identified, they all failed to predict antisense sRNA genes (asRNA), i.e. RNA genes located on the DNA strand complementary to that which encodes the protein. The statistical models enabled any genomic region to be analyzed theorically but not efficiently. We present a new model for in silico identification of sRNA and asRNA candidates within an entire bacterial genome. This model was successfully used to analyze the Gram-negative Escherichia coli and Gram-positive Streptococcus agalactiae. In both bacteria, numerous asRNAs are transcribed from the complementary strand of genes located in pathogenicity islands, strongly suggesting that these asRNAs are regulators of the virulence expression. In particular, we characterized an asRNA that acted as an enhancer-like regulator of the type 1 fimbriae production involved in the virulence of extra-intestinal pathogenic E. coli. PMID:22139924

  5. Transcriptomic analysis of the highly efficient oil-degrading bacterium Acinetobacter venetianus RAG-1 reveals genes important in dodecane uptake and utilization

    PubMed Central

    Kothari, Ankita; Charrier, Marimikel; Wu, Yu-Wei; Malfatti, Stephanie; Zhou, Carol E.; Singer, Steven W.; Dugan, Larry; Mukhopadhyay, Aindrila

    2016-01-01

    The hydrocarbonoclastic bacterium Acinetobacter venetianus RAG-1 has attracted substantial attention due to its powerful oil-degrading capabilities and its potential to play an important ecological role in the cleanup of alkanes. In this study, we compare the transcriptome of the strain RAG-1 grown in dodecane, the corresponding alkanol (dodecanol), and sodium acetate for the characterization of genes involved in dodecane uptake and utilization. Comparison of the transcriptional responses of RAG-1 grown on dodecane led to the identification of 1074 genes that were differentially expressed relative to sodium acetate. Of these, 622 genes were upregulated when grown in dodecane. The highly upregulated genes were involved in alkane catabolism, along with stress response. Our data suggest AlkMb to be primarily involved in dodecane oxidation. Transcriptional response of RAG-1 grown on dodecane relative to dodecanol also led to the identification of permease, outer membrane protein and thin fimbriae coding genes potentially involved in dodecane uptake. This study provides the first model for key genes involved in alkane uptake and metabolism in A. venetianus RAG-1. PMID:27664055

  6. Biofilm formation by enteric pathogens and its role in plant colonization and persistence

    PubMed Central

    Yaron, Sima; Römling, Ute

    2014-01-01

    The significant increase in foodborne outbreaks caused by contaminated fresh produce, such as alfalfa sprouts, lettuce, melons, tomatoes and spinach, during the last 30 years stimulated investigation of the mechanisms of persistence of human pathogens on plants. Emerging evidence suggests that Salmonella enterica and Escherichia coli, which cause the vast majority of fresh produce outbreaks, are able to adhere to and to form biofilms on plants leading to persistence and resistance to disinfection treatments, which subsequently can cause human infections and major outbreaks. In this review, we present the current knowledge about host, bacterial and environmental factors that affect the attachment to plant tissue and the process of biofilm formation by S. enterica and E. coli, and discuss how biofilm formation assists in persistence of pathogens on the plants. Mechanisms used by S. enterica and E. coli to adhere and persist on abiotic surfaces and mammalian cells are partially similar and also used by plant pathogens and symbionts. For example, amyloid curli fimbriae, part of the extracellular matrix of biofilms, frequently contribute to adherence and are upregulated upon adherence and colonization of plant material. Also the major exopolysaccharide of the biofilm matrix, cellulose, is an adherence factor not only of S. enterica and E. coli, but also of plant symbionts and pathogens. Plants, on the other hand, respond to colonization by enteric pathogens with a variety of defence mechanisms, some of which can effectively inhibit biofilm formation. Consequently, plant compounds might be investigated for promising novel antibiofilm strategies. PMID:25351039

  7. Selective adherence of non-typeable Haemophilus influenzae (NTHi) to mucus or epithelial cells in the chinchilla eustachian tube and middle ear.

    PubMed

    Miyamoto, N; Bakaletz, L O

    1996-11-01

    Frozen sections of chinchilla Eustachian tube (ET) and middle ear mucosa were incubated with either FITC-labeled non-typeable Haemophilus influenzae (NTHi) or Bordetella pertussis. The number of bacteria adherent to "roof" vs "floor" regions was compared for each of three anatomic portions of the ET and for middle ear epithelium noting whether bacteria adhered to mucus or to epithelial cells. NTHi strains adhered significantly greater to mucus in the ET lumen whereas B. pertussis preferentially adhered to epithelial cells lining the ET (P < or = 0.05). A non-fimbriated isogenic mutant of NTHi adhered significantly less to mucus than the parental isolate at all sites of the ET floor (P < or = 0.05). Isolated fimbrin protein adhered to ET mucus and blocked adherence of whole organisms. Treatment with the mucolytic agent N-acetyl-L-cysteine resulted in significantly reduced adherence of NTHi to mucus (P < or = 0.001) and eliminated the ability to detect binding of isolated fimbrin protein. N-acetyl-L-cysteine treatment did not affect adherence of either B. pertussis or NTHi to epithelial cells. These data indicated that NTHi may mediate ascension of the ET from the nasopharynx primarily via adherence to and growth in mucus overlying the floor region of the tubal lumen. The OMP P5-homologous fimbriae were shown to contribute to this binding.

  8. Limited genetic diversity and gene expression differences between egg- and non-egg-related Salmonella Enteritidis strains.

    PubMed

    Botteldoorn, N; Van Coillie, E; Goris, J; Werbrouck, H; Piessens, V; Godard, C; Scheldeman, P; Herman, L; Heyndrickx, M

    2010-08-01

    Salmonella Enteritidis strains of egg- and non-egg-related origin were characterized molecularly to find markers correlated with the egg-contaminating property of Salmonella Enteritidis. Isolates were examined by random amplified polymorphic DNA (RAPD), plasmid profiling and phage typing. Furthermore, the presence of 30 virulence genes was tested by PCR. In genetic fingerprinting and gene content, only small differences between the strains were found and no correlation was observed with the origin (egg-related versus non-egg-related). A major RADP group was present in both egg- and non-egg-related strains, but other smaller RAPD groups were present as well in both categories of strains. Phage types PT4 and PT21 were predominant. Differential mRNA expression levels of fimA and agfA under conditions of growth simulating the conditions during egg formation were determined by real-time RT-PCR. Although differences in fimA and agfA expression levels were observed between the strains, these could not be correlated with the origin of the strains (egg-related versus non-egg-related). The highest expression levels of agfA and fimA were only found in two non-egg-related strains, which seemed to be correlated with the presence of a 93 kb plasmid instead of the 60 kb virulence plasmid. Our results seem to indicate only a limited role for at least type I fimbriae (encoded by fim operon) in egg contamination by Salmonella Enteritidis.

  9. Fiber optic laser light scattering measurement of ciliary function of the fallopian tube

    NASA Astrophysics Data System (ADS)

    Halbert, Sheridan A.; Lim, Kap; Lee, Wylie I.

    1990-07-01

    A fiber-optic laser light-scattering system (FLS) for measuring ciliary function was evaluated by means of three sets of in vitro experiments. First, FLS performance was compared to that of a previously proven benchtop laser system (BLS). Using tissue excised from rabbit fallopian tubes, ciliary beat frequency (CBF) of each sample was measured with FLS and BLS. Paired CBF measurements showed excellent correlation between the two systems (r =0.93). Second, the FLS was used to evaluate the dependency of CBF on temperature (T) by using tissue sampies of rabbit oviductal fimbna. Regression analysis of CBF vs T showed a linear relationship over the range of 18-37°C for both individual samples (r =0.98) and pooled data from all experiments (r = 0.84). Fmally, the relalionship between CBF and ciliary ovum transport rate (TR) was tested by using T to modulate CBF of rabbit fimbria, in vitro. The relationship was linear over the range of CBF from 10 to 30 Hz (r2 = 0.83). At 37°C, CBF = 31+/-1 Hz, and TR = O.12+/-.02 mm/sec. equal to ovum transport rate in situ. The FLS is a valuable tool for characterizing ciliary activity and thus ovum transport function. Owing to the fact that ciliary dyskinesia resulting from disease of the fallopian tube is associated with infeitility, the FLS may be useful to acquire data important to the clinical evaluation of fallopian tube function and female infertility.

  10. Characterization of a CTX-M-15 Producing Klebsiella Pneumoniae Outbreak Strain Assigned to a Novel Sequence Type (1427)

    PubMed Central

    Zhou, Kai; Lokate, Mariëtte; Deurenberg, Ruud H.; Arends, Jan; Lo-Ten Foe, Jerome; Grundmann, Hajo; Rossen, John W. A.; Friedrich, Alexander W.

    2015-01-01

    Extended-spectrum -lactamase producing Klebsiella pneumoniae have emerged as one of the major nosocomial pathogens. Between July and September 2012, a CTX-M-15 producing K. pneumoniae caused an outbreak in a university hospital in the Netherlands. The outbreak isolates were characterized and assigned to a novel sequence type (ST1427). An epidemiological link between affected patients was supported by patient contact tracing and whole-genome phylogenetic analysis. Intra-strain polymorphism was detected among multiple isolates obtained from different body sites of the index patient, which may relate to antibiotic treatment and/or host adaptation. Environmental contamination caused by the outbreak clone was found in the patient rooms even on medical equipment. The novel clone was not closely related to any known endemic/epidemic clone, but carried a set of a plasmid-borne resistance genes [blaCTX−M−15, blaTEM−1, blaOXA−1, aac(6′)-Ib-cr, qnrB1, tetA(A), aac(3)-II]. Analysis of its virulence factors revealed a previously uncharacterized capsular biosynthesis region and two uncharacterized fimbriae gene clusters, and suggested that the new clone was not hypervirulent. To our knowledge, this is the first outbreak report of K. pneumoniae ST1427, and our study could be of help to understand the features of this newly emerging clone. PMID:26617589

  11. Radiation-Induced Growth Retardation and Microstructural and Metabolite Abnormalities in the Hippocampus

    PubMed Central

    Zawaski, Janice A.; Sahnoune, Iman

    2016-01-01

    Cranial radiotherapy (CRT) increases survival in pediatric brain-tumor patients but can cause deleterious effects. This study evaluates the acute and long-term impact of CRT delivered during childhood/adolescence on the brain and body using a rodent model. Rats received CRT, either 4 Gy fractions × 5 d (fractionated) or a cumulative dose of 20 Gy (single dose) at 28 d of age. Animals were euthanized 1 d, 5 d, or 3.5 mo after CRT. The 3.5 mo group was imaged prior to euthanasia. At 3.5 mo, we observed significant growth retardation in irradiated animals, versus controls, and the effects of single dose on brain and body weights were more severe than fractionated. Acutely single dose significantly reduced body weight but increased brain weight, whereas fractionation significantly reduced brain but not body weights, versus controls. CRT suppressed cell proliferation in the hippocampal subgranular zone acutely. Fractional anisotropy (FA) in the fimbria was significantly lower in the single dose versus controls. Hippocampal metabolite levels were significantly altered in the single dose animals, reflecting a heightened state of inflammation that was absent in the fractionated. Our findings indicate that despite the differences in severity between the doses they both demonstrated an effect on cell proliferation and growth retardation, important factors in pediatric CRT. PMID:27242931

  12. Isolation and characterization of cytotoxic, aggregative Citrobacter freundii.

    PubMed

    Bai, Li; Xia, Shengli; Lan, Ruiting; Liu, Liyun; Ye, Changyun; Wang, Yiting; Jin, Dong; Cui, Zhigang; Jing, Huaiqi; Xiong, Yanwen; Bai, Xuemei; Sun, Hui; Zhang, Jin; Wang, Lei; Xu, Jianguo

    2012-01-01

    Citrobacter freundii is an infrequent but established cause of diarrhea in humans. However, little is known of its genetic diversity and potential for virulence. We analyzed 26 isolates, including 12 from human diarrheal patients, 2 from human fecal samples of unknown diarrheal status, and 12 from animals, insects, and other sources. Pulsed field gel electrophoresis using XbaI allowed us to divide the 26 isolates into 20 pulse types, while multi-locus sequence typing using 7 housekeeping genes allowed us to divide the 26 isolates into 6 sequence types (STs) with the majority belonging to 4 STs. We analyzed adhesion and cytotoxicity to HEp-2 cells in these 26 strains. All were found to adhere to HEp-2 cells. One strain, CF74, which had been isolated from a goat, showed the strongest aggregative adhesion pattern. Lactate dehydrogenase (LDH) released from HEp-2 cells was evaluated as a measure of cytotoxicity, averaging 7.46%. Strain CF74 induced the highest level of LDH, 24.3%, and caused >50% cell rounding, detachment, and death. We named strain CF74 "cytotoxic and aggregative C. freundii." Genome sequencing of CF74 revealed that it had acquired 7 genomic islands, including 2 fimbriae islands and a type VI secretion system island, all of which are potential virulence factors. Our results show that aggregative adherence and cytotoxicity play an important role in the pathogenesis of C. freundii.

  13. Klebsiella: a long way to go towards understanding this enigmatic jet-setter

    PubMed Central

    Broberg, Christopher A.; Palacios, Michelle

    2014-01-01

    Klebsiella pneumoniae is the causative agent of a variety of diseases, including pneumonia, urinary tract infections, septicemia, and the recently recognized pyogenic liver abscesses (PLA). Renewed efforts to identify and understand the bacterial determinants required to cause disease have come about because of the worldwide increase in the isolation of strains resistant to a broad spectrum of antibiotics. The recent increased isolation of carbapenem-resistant strains further reduces the available treatment options. The rapid geographic spread of the resistant isolates and the spread to other pathogens are of particular concern. For many years, the best characterized virulence determinants were capsule, lipopolysaccharide, siderophores, and types 1 and 3 fimbriae. Recent efforts to expand this list include in vivo screens and whole-genome sequencing. However, we still know little about how this bacterium is able to cause disease. Some recent clonal analyses of K. pneumoniae strains indicate that there are distinct clonal groups, some of which may be associated with specific disease syndromes. However, what makes one clonal group more virulent and what changes the disease pattern are not yet clear and remain important questions for the future. PMID:25165563

  14. [Fusion expression of SLT-IIeB gene and FedF gene of Ee in Escherichia coli and its immunogenicity].

    PubMed

    Liu, Guo-Ping; Wu, Bin; Lin, Yi-Yuan; Liu, Meng-Yuan; Chen, Huan-Chun

    2007-12-01

    The DNA fragment encoding the truncated SLT-IIeB and FedF of Ee strain were fused to the downstream of glutathione S-transferase (GST) of pGEX-KG expression vector, resulting in the fusion expression plasmid pKSF. After transformed into E. coli BL21 (DE3) and induced by IPTG, the results of SDS-PAGE showed that the GST-SF fusion protein was expressed in high level. Western blot was performed to confirm that the expressed fusion protein could specifically react with mouse anti-SLT-IIeB antiserum, mouse anti-FedF antiserum and moue anti-GST monoclonal antibody respectively. The fusion protein was further purified and used as an antigen for preparation of immune serum. The anti-sera of GST-SF were able to restrain the toxicity of SLT-IIe to Vero-E6 cells and inhibit the adhesin of F18 fimbriae to brush borders of swine in vitro. Groups of SPF KM mice were vaccinated subcutaneously at 0 week with 25 micrograms and at 2 weeks with 25 micrograms of purified GST-SF, GST-B, GST-F and challenged intraperitoneally with volume of 5 LD50 Ee strain. The results show the fusion protein GST-SF had more shrong immunogenicity and better protection against Ee strain.

  15. Nerve growth factor promotes survival of septal cholinergic neurons after fimbrial transections.

    PubMed

    Hefti, F

    1986-08-01

    Several findings obtained in recent years suggest that NGF, aside from its well-established function as a neurotrophic factor for peripheral sympathetic and sensory neurons, also has trophic influence on the cholinergic neurons of the basal forebrain. The present study assessed whether NGF was able to affect survival of central cholinergic neurons after axonal transections in adult rats. The septo-hippocampal pathway was transected unilaterally by cutting the fimbria, and animals were implanted with a cannula through which NGF or control solutions were injected intraventricularly over 4 weeks. The lesions reduced the number of large cell bodies, as visualized by Nissl staining in the medial septal nucleus and in the vertical limb of the diagonal band of Broca. Furthermore, in the same nuclei, they reduced the number of cell bodies positively stained for AChE after pretreatment with diisopropylfluorophosphate (a method known to result in reliable identification of cholinergic neurons in the septal area). On lesioned sides, the number of cholinergic cells in medial septal nucleus and the vertical limb of the diagonal band was reduced by 50 +/- 4%, as compared to the number on contralateral sides. On lesioned sides of animals chronically treated with NGF, the number of AChE-positive cells in these areas was reduced only by 12 +/- 6%, as compared to control levels. These findings suggest that fimbrial transections resulted in retrograde degeneration of cholinergic septo-hippocampal neurons and that NGF treatment strongly attenuated this lesion-induced degeneration.(ABSTRACT TRUNCATED AT 250 WORDS)

  16. Short communication: Distribution of Porphyromonas gulae fimA genotypes in oral specimens from dogs with mitral regurgitation.

    PubMed

    Shirai, Mitsuyuki; Nomura, Ryota; Kato, Yukio; Murakami, Masaru; Kondo, Chihiro; Takahashi, Soraaki; Yamasaki, Yoshie; Matsumoto-Nakano, Michiyo; Arai, Nobuaki; Yasuda, Hidemi; Nakano, Kazuhiko; Asai, Fumitoshi

    2015-10-01

    Porphyromonas gulae, a suspected pathogen for periodontal disease in dogs, possesses approximately 41-kDa fimbriae (FimA) that are encoded by the fimA gene. In the present study, the association of fimA genotypes with mitral regurgitation (MR) was investigated. Twenty-five dogs diagnosed with MR (age range 6-13 years old, average 10.8 years) and 32 healthy dogs (8-15 years old, average 10.8 years) were selected at the participating clinics in a consecutive manner during the same time period. Oral swab specimens were collected from the dogs and bacterial DNA was extracted, then polymerase chain reaction analysis was performed using primers specific for each fimA genotype, with the dominant genotype determined. The rate for genotype C dominant specimens was 48.0% in the MR group, which was significantly higher than that in the control group (18.8%) (P <0.05). These results suggest that P. gulae fimA genotype C is associated with MR.

  17. Plasticity of fimbrial genotype and serotype within populations of Bordetella pertussis: analysis by paired flow cytometry and genome sequencing.

    PubMed

    Vaughan, Thomas E; Pratt, Catherine B; Sealey, Katie; Preston, Andrew; Fry, Norman K; Gorringe, Andrew R

    2014-09-01

    The fimbriae of Bordetella pertussis are required for colonization of the human respiratory tract. Two serologically distinct fimbrial subunits, Fim2 and Fim3, considered important vaccine components for many years, are included in the Sanofi Pasteur 5-component acellular pertussis vaccine, and the World Health Organization recommends the inclusion of strains expressing both fimbrial serotypes in whole-cell pertussis vaccines. Each of the fimbrial major subunit genes, fim2, fim3, and fimX, has a promoter poly(C) tract upstream of its -10 box. Such monotonic DNA elements are susceptible to changes in length via slipped-strand mispairing in vitro and in vivo, which potentially causes on/off switching of genes at every cell division. Here, we have described intra-culture variability in poly(C) tract lengths and the resulting fimbrial phenotypes in 22 recent UK B. pertussis isolates. Owing to the highly plastic nature of fimbrial promoters, we used the same cultures for both genome sequencing and flow cytometry. Individual cultures of B. pertussis contained multiple fimbrial serotypes and multiple different fimbrial promoter poly(C) tract lengths, which supports earlier serological evidence that B. pertussis expresses both serotypes during infection.

  18. In vivo magnetic resonance imaging of the human limbic white matter

    PubMed Central

    Mori, Susumu; Aggarwal, Manisha

    2014-01-01

    The limbic system mediates memory, behavior, and emotional output in the human brain, and is implicated in the pathology of Alzheimer’s disease and a wide spectrum of related neurological disorders. In vivo magnetic resonance imaging (MRI) of structural components comprising the limbic system and their interconnections via white matter pathways in the human brain has helped define current understanding of the limbic model based on the classical circuit proposed by Papez. MRI techniques, including diffusion MR imaging, provide a non-invasive method to characterize white matter tracts of the limbic system, and investigate pathological changes that affect these pathways in clinical settings. This review focuses on delineation of the anatomy of major limbic tracts in the human brain, namely, the cingulum, the fornix and fimbria, and the stria terminalis, based on in vivo MRI contrasts. The detailed morphology and intricate trajectories of these pathways that can be identified using relaxometry-based and diffusion-weighted MRI provide an important anatomical reference for evaluation of clinical disorders commonly associated with limbic pathology. PMID:25505883

  19. Bordetella pertussis: new concepts in pathogenesis and treatment

    PubMed Central

    Carbonetti, Nicholas H.

    2016-01-01

    Purpose of review The purpose of this review is to summarize and discuss recent findings and selected topics of interest in Bordetella pertussis virulence and pathogenesis and treatment of pertussis. It is not intended to cover issues on immune responses to B. pertussis infection or problems with currently used pertussis vaccines. Recent findings Studies on the activities of various B. pertussis virulence factors include the immunomodulatory activities of filamentous hemagglutinin, fimbriae, and adenylate cyclase toxin. Recently emerging B. pertussis strains show evidence of genetic selection for vaccine escape mutants, with changes in vaccine antigen-expressing genes, some of which may have increased the virulence of this pathogen. Severe and fatal pertussis in young infants continues to be a problem, with several studies highlighting predictors of fatality, including the extreme leukocytosis associated with this infection. Treatments for pertussis are extremely limited, though early antibiotic intervention may be beneficial. Neutralizing pertussis toxin activity may be an effective strategy, as well as targeting two host proteins, pendrin and sphingosine-1-phosphate receptors, as novel potential therapeutic interventions. Summary Pertussis is reemerging as a major public health problem and continued basic research is revealing information on bacterial virulence and disease pathogenesis, as well as potential novel strategies for vaccination and targets for therapeutic intervention. PMID:26906206

  20. Intestinal anastomotic injury alters spatially defined microbiome composition and function

    PubMed Central

    2014-01-01

    Background When diseased intestine (i.e., from colon cancer, diverticulitis) requires resection, its reconnection (termed anastomosis) can be complicated by non-healing of the newly joined intestine resulting in spillage of intestinal contents into the abdominal cavity (termed anastomotic leakage). While it is suspected that the intestinal microbiota have the capacity to both accelerate and complicate anastomotic healing, the associated genotypes and functions have not been characterized. Results Using 16S rRNA amplicon sequencing of samples collected on the day of surgery (postoperative day 0 (POD0)) and the 6th day following surgery (postoperative day 0 (POD6)), we analyzed the changes in luminal versus tissue-associated microbiota at anastomotic sites created in the colon of rats. Results indicated that anastomotic injury induced significant changes in the anastomotic tissue-associated microbiota with minimal differences in the luminal microbiota. The most striking difference was a 500-fold and 200-fold increase in the relative abundance of Enterococcus and Escherichia/Shigella, respectively. Functional profiling predicted the predominance of bacterial virulence-associated pathways in post-anastomotic tissues, including production of hemolysin, cytolethal toxins, fimbriae, invasins, cytotoxic necrotizing factors, and coccolysin. Conclusion Taken together, our results suggest that compositional and functional changes accompany anastomotic tissues and may potentially accelerate or complicate anastomotic healing. PMID:25250176

  1. The primary transcriptome of the Escherichia coli O104:H4 pAA plasmid and novel insights into its virulence gene expression and regulation

    PubMed Central

    Berger, Petya; Knödler, Michael; Förstner, Konrad U.; Berger, Michael; Bertling, Christian; Sharma, Cynthia M.; Vogel, Jörg; Karch, Helge; Dobrindt, Ulrich; Mellmann, Alexander

    2016-01-01

    Escherichia coli O104:H4 (E. coli O104:H4), which caused a massive outbreak of acute gastroenteritis and hemolytic uremic syndrome in 2011, carries an aggregative adherence fimbriae I (AAF/I) encoding virulence plasmid, pAA. The importance of pAA in host-pathogen interaction and disease severity has been demonstrated, however, not much is known about its transcriptional organization and gene regulation. Here, we analyzed the pAA primary transcriptome using differential RNA sequencing, which allows for the high-throughput mapping of transcription start site (TSS) and non-coding RNA candidates. We identified 248 TSS candidates in the 74-kb pAA and only 21% of them could be assigned as TSS of annotated genes. We detected TSS for the majority of pAA-encoded virulence factors. Interestingly, we mapped TSS, which could allow for the transcriptional uncoupling of the AAF/I operon, and potentially regulatory antisense RNA candidates against the genes encoding dispersin and the serine protease SepA. Moreover, a computational search for transcription factor binding sites suggested for AggR-mediated activation of SepA expression, which was additionally experimentally validated. This work advances our understanding of the molecular basis of E. coli O104:H4 pathogenicity and provides a valuable resource for further characterization of pAA virulence gene regulation. PMID:27748404

  2. A novel Pseudomonas aeruginosa bacteriophage, Ab31, a chimera formed from temperate phage PAJU2 and P. putida lytic phage AF: characteristics and mechanism of bacterial resistance.

    PubMed

    Latino, Libera; Essoh, Christiane; Blouin, Yann; Vu Thien, Hoang; Pourcel, Christine

    2014-01-01

    A novel temperate bacteriophage of Pseudomonas aeruginosa, phage vB_PaeP_Tr60_Ab31 (alias Ab31) is described. Its genome is composed of structural genes related to those of lytic P. putida phage AF, and regulatory genes similar to those of temperate phage PAJU2. The virion structure resembles that of phage AF and other lytic Podoviridae (S. enterica Epsilon 15 and E. coli phiv10) with similar tail spikes. Ab31 was able to infect P. aeruginosa strain PA14 and two genetically related strains called Tr60 and Tr162, out of 35 diverse strains from cystic fibrosis patients. Analysis of resistant host variants revealed different phenotypes, including induction of pigment and alginate overproduction. Whole genome sequencing of resistant variants highlighted the existence of a large deletion of 234 kbp in two strains, encompassing a cluster of genes required for the production of CupA fimbriae. Stable lysogens formed by Ab31 in strain Tr60, permitted the identification of the insertion site. During colonization of the lung in cystic fibrosis patients, P. aeruginosa adapts by modifying its genome. We suggest that bacteriophages such as Ab31 may play an important role in this adaptation by selecting for bacterial characteristics that favor persistence of bacteria in the lung.

  3. The pilus usher controls protein interactions via domain masking and is functional as an oligomer

    DOE PAGES

    Werneburg, Glenn T.; Li, Huilin; Henderson, Nadine S.; ...

    2015-06-08

    The chaperone/usher (CU) pathway is responsible for biogenesis of organelles termed pili or fimbriae in Gram-negative bacteria. Type 1 pili expressed by uropathogenic Escherichia coli are prototypical structures assembled by the CU pathway. Assembly and secretion of pili by the CU pathway requires a dedicated periplasmic chaperone and a multidomain outer membrane protein termed the usher (FimD). We show that the FimD C-terminal domains provide the high-affinity substrate binding site, but that these domains are masked in the resting usher. Domain masking requires the FimD plug domain, which served as a central switch controlling usher activation. In addition, we demonstratemore » that usher molecules can act in trans for pilus biogenesis, providing conclusive evidence for a functional usher oligomer. These results reveal mechanisms by which molecular machines such as the usher regulate and harness protein-protein interactions, and suggest that ushers may interact in a cooperative manner during pilus assembly in bacteria.« less

  4. Subinhibitory Concentrations of Allicin Decrease Uropathogenic Escherichia coli (UPEC) Biofilm Formation, Adhesion Ability, and Swimming Motility.

    PubMed

    Yang, Xiaolong; Sha, Kaihui; Xu, Guangya; Tian, Hanwen; Wang, Xiaoying; Chen, Shanze; Wang, Yi; Li, Jingyu; Chen, Junli; Huang, Ning

    2016-06-29

    Uropathogenic Escherichia coli (UPEC) biofilm formation enables the organism to avoid the host immune system, resist antibiotics, and provide a reservoir for persistent infection. Once the biofilm is established, eradication of the infection becomes difficult. Therefore, strategies against UPEC biofilm are urgently required. In this study, we investigated the effect of allicin, isolated from garlic essential oil, on UPEC CFT073 and J96 biofilm formation and dispersal, along with its effect on UPEC adhesion ability and swimming motility. Sub-inhibitory concentrations (sub-MICs) of allicin decreased UPEC biofilm formation and affected its architecture. Allicin was also capable of dispersing biofilm. Furthermore, allicin decreased the bacterial adhesion ability and swimming motility, which are important for biofilm formation. Real-time quantitative polymerase chain reaction (RT-qPCR) revealed that allicin decreased the expression of UPEC type 1 fimbriae adhesin gene fimH. Docking studies suggested that allicin was located within the binding pocket of heptyl α-d-mannopyrannoside in FimH and formed hydrogen bonds with Phe1 and Asn135. In addition, allicin decreased the expression of the two-component regulatory systems (TCSs) cognate response regulator gene uvrY and increased the expression of the RNA binding global regulatory protein gene csrA of UPEC CFT073, which is associated with UPEC biofilm. The findings suggest that sub-MICs of allicin are capable of affecting UPEC biofilm formation and dispersal, and decreasing UPEC adhesion ability and swimming motility.

  5. Inhibition of enteroaggregative Escherichia coli cell adhesion in-vitro by designed peptides.

    PubMed

    Gupta, Deepika; Sarkar, Subendu; Sharma, Monica; Thapa, B R; Chakraborti, Anuradha

    2016-09-01

    Enteroaggregative Escherichia coli (EAEC) bears remarkable capacity to adhere the host intestinal mucosal surface and results in acute or persistent childhood diarrhea worldwide. In this study, an attempt has been made to inhibit EAEC cell adherence in-vitro using synthetic peptides. E. coli isolates (n = 54) were isolated from the stool samples of clinically diagnosed pediatric diarrheal patients. 92.8% isolates showed different types of aggregative adherence patterns with HEp-2 cells. AAF-II (Aggregative Adherence Fimbriae-II) EAEC exhibited the maximum ability to form biofilm and intracellular survival. Peptides were designed against the high antigenic epitopic regions of AAF-II adhesin of EAEC O42 using prediction algorithms like BcePred and ProPred software to block the EAEC cell adhesion in-vitro. Peptides P2 (DITITPATNRDVNV) and P3 (MRIKAWGEANHGQL) demonstrated higher inhibition of EAEC cell adhesion than P1 (GMQGSITPAIPLRPG). Interestingly, increasing the pre-incubation time of the peptides with HEp-2 cells from 1 h to 2 h showed the maximum inhibition. The data suggested the potential role of P2 and P3 peptides in successfully blocking the binding of AAF-II EAEC with HEp-2 cell receptors. Hence, the peptides may be efficacious in designing new chemotherapeutic for the management of EAEC mediated diarrhea.

  6. Cell Adhesion Modification of Streptococcus viridians in the Presence of Xylitol

    NASA Astrophysics Data System (ADS)

    Esmacher, Jason; Vidakovich, Blair; Giangrande, Michael; Hoffmann, Peter

    2012-10-01

    There is scientific documentation that those who chew gum sweetened by the sugar alcohol xylitol report a dramatically lower incident of both dental caries and otitis media compared to those who chew conventional gum sweetened by sucrose. An explanation contends that xylitol interferes with the ability of Streptococcus viridian (SV) to form biofilms which is a necessary precursor to the bacteria's ability to damage human tissues. We have used atomic force microscopy to study the cell wall/fimbria properties at the nanonewton level in both the presence and absence of xylitol. The first set of measurements used varying concentrations of xylitol incorporated within the incubation medium. The second used non-xylitol grown bacteria, the xylitol was added externally at various concentrations. Our study suggests that growing SV with xylitol reduces their ability to adhere together. Additionally, externally added xylitol showed grouping of cell adhesion to a relatively narrow nanonewton spread that is concentration dependent. Measurement of the adhesion properties of the bacterial cell wall have found that there is a dramatic increase in the cell wall's firmness which simultaneously accompanied a decrease in its ability to support adhesion, even at very low concentrations of xylitol.

  7. Epidemiology and Clinical Manifestations of Enteroaggregative Escherichia coli

    PubMed Central

    Hebbelstrup Jensen, Betina; Olsen, Katharina E. P.; Struve, Carsten; Petersen, Andreas Munk

    2014-01-01

    SUMMARY Enteroaggregative Escherichia coli (EAEC) represents a heterogeneous group of E. coli strains. The pathogenicity and clinical relevance of these bacteria are still controversial. In this review, we describe the clinical significance of EAEC regarding patterns of infection in humans, transmission, reservoirs, and symptoms. Manifestations associated with EAEC infection include watery diarrhea, mucoid diarrhea, low-grade fever, nausea, tenesmus, and borborygmi. In early studies, EAEC was considered to be an opportunistic pathogen associated with diarrhea in HIV patients and in malnourished children in developing countries. In recent studies, associations with traveler's diarrhea, the occurrence of diarrhea cases in industrialized countries, and outbreaks of diarrhea in Europe and Asia have been reported. In the spring of 2011, a large outbreak of hemolytic-uremic syndrome (HUS) and hemorrhagic colitis occurred in Germany due to an EAEC O104:H4 strain, causing 54 deaths and 855 cases of HUS. This strain produces the potent Shiga toxin along with the aggregative fimbriae. An outbreak of urinary tract infection associated with EAEC in Copenhagen, Denmark, occurred in 1991; this involved extensive production of biofilm, an important characteristic of the pathogenicity of EAEC. However, the heterogeneity of EAEC continues to complicate diagnostics and also our understanding of pathogenicity. PMID:24982324

  8. Identification of bapA in Strains of Salmonella enterica subsp. enterica Isolated from Wild Animals Kept in Captivity in Sinaloa, Mexico.

    PubMed

    Silva-Hidalgo, Gabriela; López-Valenzuela, Martin; Cárcamo-Aréchiga, Nora; Cota-Guajardo, Silvia; López-Salazar, Mayra; Montiel-Vázquez, Edith

    2016-01-01

    bapA, previously named stm2689, encodes the BapA protein, which, along with cellulose and fimbriae, constitutes biofilms. Biofilms are communities of microorganisms that grow in a matrix of exopolysaccharides and may adhere to living tissues or inert surfaces. Biofilm formation is associated with the ability to persist in different environments, which contributes to the pathogenicity of several species. We analyzed the presence of bapA in 83 strains belonging to 17 serovars of Salmonella enterica subsp. enterica from wildlife in captivity at Culiacan's Zoo and Mazatlán's Aquarium. Each isolate amplified a product of 667 bp, which corresponds to the expected size of the bapA initiator, with no observed variation between different serovars analyzed. bapA gene was found to be highly conserved in Salmonella and can be targeted for the genus-specific detection of this organism from different sources. Since bapA expression improves bacterial proliferation outside of the host and facilitates resistance to disinfectants and desiccation, the survival of Salmonella in natural habitats may be favored. Thus, the risk of bacterial contamination from these animals is increased.

  9. Galanin inhibits acetylcholine release in the ventral hippocampus of the rat: histochemical, autoradiographic, in vivo, and in vitro studies

    SciTech Connect

    Fisone, G.; Wu, C.F.; Consolo, S.; Nordstroem, O.; Brynne, N.; Bartfai, T.; Melander, T.; Hoekfelt, T.

    1987-10-01

    A high density of galanin binding sites was found by using /sup 125/I-labeled galanin, iodinated by chloramine-T, followed by autoradiography in the ventral, but not in the dorsal, hippocampus of the rat. Lesions of the fimbria and of the septum caused disappearance of a major population of these binding sites, suggesting that a large proportion of them is localized on cholinergic nerve terminals of septal afferents. As a functional correlate to these putative galanin receptor sites, it was shown, both in vivo and in vitro, that galanin, in a concentration-dependent manner, inhibited the evoked release of acetylcholine in the ventral, but not in the dorsal, hippocampus. Intracerebroventricularly applied galanin fully inhibited the scopolamine stimulated release of acetylcholine in the ventral, but not in the dorsal, hippocampus, as measured by the microdialysis technique. In vitro, galanin inhibited the 25 mM K/sup +/-evoked release of (/sup 3/H)acetylcholine from slices of the ventral hippocampus, with an IC/sub 50/ value of approx. = 50 nM. These results are discussed with respect to the colocalization of galanin- and choline acetyltransferase-like immunoreactivity in septal somata projecting to the hippocampus.

  10. Cytochemical Labeling for Fungal and Host Components in Plant Tissues Inoculated with Fungal Wilt Pathogens

    NASA Astrophysics Data System (ADS)

    Ouellette, G. B.; Baayen, R. P.; Chamberland, H.; Simard, M.; Rioux, D.; Charest, P. M.

    2004-08-01

    Antibodies to detect pectin in present investigations attached to distinct fibrils in vessel lumina. In carnation infected with an isolate of Fusarium oxysporum f.sp., labeling of pathogen cells also occurred; in a resistant cultivar (cv.), it was coincident with proximate pectin fibrils and linked to altered fungal walls, which was the opposite in the susceptible cv., indicating that hindrance of pathogen ability to degrade pectin may be related to resistance. Labeling of the fungus in culture was nil, except in media containing pectin, showing that pectin is not native to the pathogen. Labeling of fungal walls for cellulose in elm (inoculated with Ophiostoma novo-ulmi) and carnation also occurred, linked to adsorbed host wall components. The chitin probe often attached to dispersed matter, in vessel lumina, traceable to irregularly labeled fungal cells and host wall degradation products. With an anti-horseradish peroxidase probe, host and fungal walls were equally labeled, and with a glucosidase, differences of labeling between these walls were observed, depending on pH of the test solution. Fungal extracellular matter and filamentous structures, present in fungal walls, predominantly in another elm isolate (Phaeotheca dimorphospora), did not label with any of the probes used. However, in cultures of this fungus, extracellular material labeled, even at a distance from the colony margin, with an anti-fimbriae probe.

  11. Stress induced by hooking, net towing, elevated sea water temperature and air in sablefish: Lack of concordance between mortality and physiological measures of stress

    USGS Publications Warehouse

    Davis, M.W.; Olla, B.L.; Schreck, C.B.

    2001-01-01

    In a series of laboratory studies designed to simulate bycatch processes, sablefish Anoplopoma fimbria were either hooked for up to 24 h or towed in a net for 4 h and then subjected to an abrupt transfer to elevated sea water temperature and air. Mortality did not result from hooking or net towing followed by exposure to air, but increased for both capture methods as fish were exposed to elevated temperatures, reflecting the magnifying effect of elevated temperature on mortality. Hooking and exposure to air resulted in increased plasma cortisol and lactate concentrations, while the combination of hooking and exposure to elevated temperature and air resulted in increased lactate and potassium concentrations. In fish that were towed in a net and exposed to air, cortisol, lactate, potassium and sodium concentrations increased, but when subjected to elevated temperature and air, no further increases occurred above the concentrations induced by net towing and air, suggesting a possible maximum of the physiological stress response. The results suggest that caution should be exercised when using physiological measures to quantify stress induced by capture and exposure to elevated temperature and air, that ultimately result in mortality, since the connections between physiological stress and mortality in bycatch processes remain to be fully understood.

  12. ExPEC-typical virulence-associated genes correlate with successful colonization by intestinal E. coli in a small piglet group.

    PubMed

    Schierack, Peter; Walk, Nicole; Ewers, Christa; Wilking, Hendrik; Steinrück, Hartmut; Filter, Matthias; Wieler, Lothar H

    2008-07-01

    Upon studying the transmission of Escherichia coli from a sow to five of her piglets, we observed domination of the coliform flora in piglets by a single E. coli clone, especially after weaning. This haemolytic cloneH1 did not harbour any virulence determinants typical for intestinal pathogenic E. coli isolates from swine but had a virulence gene profile very similar to extraintestinal E. coli (ExPEC), including genes coding for P fimbriae and several iron acquisition systems, besides having an affiliation to the phylogenetic B2 group. Overall, we show that the presence of higher numbers of ExPEC-typical virulence-associated genes (VAGs) in clones correlate with their successful colonization ability in piglets. We conclude that VAGs typical for ExPEC also support intestinal colonization in healthy pigs. Faeces of healthy domestic pigs can harbour high numbers of ExPEC-similar E. coli and are suggested to be a potential risk for the transmission of such bacteria to other hosts.

  13. Environmental scanning electron microscopy analysis of Proteus mirabilis biofilms grown on chitin and stainless steel.

    PubMed

    Fernández-Delgado, Milagro; Duque, Zoilabet; Rojas, Héctor; Suárez, Paula; Contreras, Monica; García-Amado, María A; Alciaturi, Carlos

    Proteus mirabilis is a human pathogen able to form biofilms on the surface of urinary catheters. Little is known about P. mirabilis biofilms on natural or industrial surfaces and the potential consequences for these settings. The main aim of this work was to assess and compare the adhesion and biofilm formation of P. mirabilis strains from different origins on chitin and stainless steel surfaces within 4 to 96 h. Using environmental scanning electron microscopy, the biofilms of a clinical strain grown on chitin at 4 h showed greater adhesion, aggregation, thickness, and extracellular matrix production than those grown on stainless steel, whereas biofilms of an environmental strain had less aggregation on both surfaces. Biofilms of both P. mirabilis strains developed different structures on chitin, such as pillars, mushrooms, channels, and crystalline-like precipitates between 24 and 96 h, in contrast with flat-layer biofilms produced on stainless steel. Significant differences (p < 0.05) were found in the frequency of pillars and channels. Images of transmission electron microscopy demonstrated abundant fimbriae in 100 % of cells from both strains, which could be related to surface adherence and biofilm formation. This represents the first study of P. mirabilis showing adhesion, biofilm formation, and development of different structures on surfaces found outside the human host.

  14. Towards a better understanding of Lactobacillus rhamnosus GG - host interactions

    PubMed Central

    2014-01-01

    Lactobacillus rhamnosus GG (LGG) is one of the most widely used probiotic strains. Various health effects are well documented including the prevention and treatment of gastro-intestinal infections and diarrhea, and stimulation of immune responses that promote vaccination or even prevent certain allergic symptoms. However, not all intervention studies could show a clinical benefit and even for the same conditions, the results are not univocal. Clearly, the host phenotype governed by age, genetics and environmental factors such as the endogenous microbiota, plays a role in whether individuals are responders or non-responders. However, we believe that a detailed knowledge of the bacterial physiology and the LGG molecules that play a key role in its host-interaction capacity is crucial for a better understanding of its potential health benefits. Molecules that were yet identified as important factors governing host interactions include its adhesive pili or fimbriae, its lipoteichoic acid molecules, its major secreted proteins and its galactose-rich exopolysaccharides, as well as specific DNA motifs. Nevertheless, future studies are needed to correlate specific health effects to these molecular effectors in LGG, and also in other probiotic strains. PMID:25186587

  15. Comparable Genital Tract Infection, Pathology, and Immunity in Rhesus Macaques Inoculated with Wild-Type or Plasmid-Deficient Chlamydia trachomatis Serovar D

    PubMed Central

    Qu, Yanyan; Frazer, Lauren C.; O'Connell, Catherine M.; Tarantal, Alice F.; Andrews, Charles W.; O'Connor, Shelby L.; Russell, Ali N.; Sullivan, Jeanne E.; Poston, Taylor B.; Vallejo, Abbe N.

    2015-01-01

    Rhesus macaques were studied to directly address the potential for plasmid-deficient Chlamydia trachomatis to serve as a live attenuated vaccine in the genital tract. Five repeated cervical inoculations of rhesus macaques with wild-type serovar D strain D/UW-3/Cx or a plasmid-deficient derivative of this strain, CTD153, resulted in infections with similar kinetics and induced comparable levels of protective immunity. After all animals received five challenges with D/UW-3/Cx, levels of inflammation observed grossly and histologically were similar between the groups. Animals in both groups developed evidence of oviduct dilatation; however, reduced oviduct dilatation was observed for “controllers,” i.e., animals without detectable chlamydial DNA in the fimbriae at weeks 5 and 12. Grouping animals into “ascenders” and “controllers” revealed that elevated early T cell responses were associated with protection, whereas higher antibody responses were associated with ascension. Protected animals shared common major histocompatibility complex (MHC) alleles. Overall, genetic differences of individual animals, rather than the presence or absence of the chlamydial plasmid in the primary infecting strain, appeared to play a role in determining the outcome of infection. PMID:26216426

  16. Comparable Genital Tract Infection, Pathology, and Immunity in Rhesus Macaques Inoculated with Wild-Type or Plasmid-Deficient Chlamydia trachomatis Serovar D.

    PubMed

    Qu, Yanyan; Frazer, Lauren C; O'Connell, Catherine M; Tarantal, Alice F; Andrews, Charles W; O'Connor, Shelby L; Russell, Ali N; Sullivan, Jeanne E; Poston, Taylor B; Vallejo, Abbe N; Darville, Toni

    2015-10-01

    Rhesus macaques were studied to directly address the potential for plasmid-deficient Chlamydia trachomatis to serve as a live attenuated vaccine in the genital tract. Five repeated cervical inoculations of rhesus macaques with wild-type serovar D strain D/UW-3/Cx or a plasmid-deficient derivative of this strain, CTD153, resulted in infections with similar kinetics and induced comparable levels of protective immunity. After all animals received five challenges with D/UW-3/Cx, levels of inflammation observed grossly and histologically were similar between the groups. Animals in both groups developed evidence of oviduct dilatation; however, reduced oviduct dilatation was observed for "controllers," i.e., animals without detectable chlamydial DNA in the fimbriae at weeks 5 and 12. Grouping animals into "ascenders" and "controllers" revealed that elevated early T cell responses were associated with protection, whereas higher antibody responses were associated with ascension. Protected animals shared common major histocompatibility complex (MHC) alleles. Overall, genetic differences of individual animals, rather than the presence or absence of the chlamydial plasmid in the primary infecting strain, appeared to play a role in determining the outcome of infection.

  17. Ex vivo diffusion tensor imaging and neuropathological correlation in a murine model of hypoxia-ischemia-induced thrombotic stroke.

    PubMed

    Shereen, Ahmed; Nemkul, Niza; Yang, Dianer; Adhami, Faisal; Dunn, R Scott; Hazen, Missy L; Nakafuku, Masato; Ning, Gang; Lindquist, Diana M; Kuan, Chia-Yi

    2011-04-01

    Diffusion tensor imaging (DTI) is a powerful method to visualize white matter, but its use in patients with acute stroke remains limited because of the lack of corresponding histologic information. In this study, we addressed this issue using a hypoxia-ischemia (HI)-induced thrombotic model of stroke in adult mice. At 6, 15, and 24  hours after injury, animals were divided into three groups for (1) in vivo T2- and diffusion-weighted magnetic resonance imaging, followed by histochemistry, (2) ex vivo DTI and electron microscopy, and (3) additional biochemical or immunochemical assays. The temporal changes of diffusion anisotropy and histopathology were compared in the fimbria, internal capsule, and external capsule. We found that HI caused a rapid reduction of axial and radial diffusivities in all three axonal bundles. A large decrease in fractional anisotropy, but not in axial diffusivity per se, was associated with structural breakdown of axons. Furthermore, the decrease in radial diffusivity correlated with swelling of myelin sheaths and compression of the axoplasma. The gray matter of the hippocampus also exhibited a high level of diffusion anisotropy, and its reduction signified dendritic degeneration. Taken together, these results suggest that cross-evaluation of multiple DTI parameters may provide a fuller picture of axonal and dendritic injury in acute ischemic stroke.

  18. Epigenetic Control of Virulence Gene Expression in Pseudomonas aeruginosa by a LysR-Type Transcription Regulator

    PubMed Central

    Turner, Keith H.; Vallet-Gely, Isabelle; Dove, Simon L.

    2009-01-01

    Phenotypic variation within an isogenic bacterial population is thought to ensure the survival of a subset of cells in adverse conditions. The opportunistic pathogen Pseudomonas aeruginosa variably expresses several phenotypes, including antibiotic resistance, biofilm formation, and the production of CupA fimbriae. Here we describe a previously unidentified bistable switch in P. aeruginosa. This switch controls the expression of a diverse set of genes, including aprA, which encodes the secreted virulence factor alkaline protease. We present evidence that bistable expression of PA2432, herein named bexR (bistable expression regulator), which encodes a LysR-type transcription regulator, controls this switch. In particular, using DNA microarrays, quantitative RT–PCR analysis, chromatin immunoprecipitation, and reporter gene fusions, we identify genes directly under the control of BexR and show that these genes are bistably expressed. Furthermore, we show that bexR is itself bistably expressed and positively autoregulated. Finally, using single-cell analyses of a GFP reporter fusion, we present evidence that positive autoregulation of bexR is necessary for bistable expression of the BexR regulon. Our findings suggest that a positive feedback loop involving a LysR-type transcription regulator serves as the basis for an epigenetic switch that controls virulence gene expression in P. aeruginosa. PMID:20041030

  19. Androgen uptake in the female reproductive system of a nonhuman primate

    SciTech Connect

    Weaker, F.J.; Sheridan, P.J.

    1981-07-15

    The nuclear uptake and retention of /sup 3/H-dehydrotestosterone (/sup 3/H-DHT) were studied in the reproductive organs of the female rhesus monkey by autoradiography. Ovariectomized and adrenalectomized adult female monkeys were injected with 1 microgram of /sup 3/H-DHT (107 Ci/mmole) and were exsanguinated 1 hour later. The uterus, cervix, vagina, and fimbriae of the fallopian tubes were removed and processed for autoradiography. Localization of the androgen was observed in the nuclei of fibroblasts, but not in the nuclei of smooth muscle and tissues of the blood vessels, of all four organs. The uptake of androgen by the epithelium varied for each organ. The simple columnar epithelium of the fallopian tube demonstrated nuclear labeling, but the epithelium of the uterus did not. The cells of the germinative layer of the stratified squamous epithelium of both the cervix and vagina exhibited nuclear concentration of label; however, the simple columnar epithelium of the cervix contiguous with the uterus was free of label. The results suggest that androgens may have a physiologic role in the female reproductive organs, and that the therapeutic effect of drugs such as danazol may be mediated via an androgen receptor.

  20. Delayed voluntary exercise does not enhance cognitive performance after hippocampal injury: an investigation of differentially distributed exercise protocols

    PubMed Central

    Wogensen, Elise; Gram, Marie Gajhede; Sommer, Jens Bak; Vilsen, Christina Rytter; Mogensen, Jesper; Malá, Hana

    2016-01-01

    Voluntary exercise has previously been shown to enhance cognitive recovery after acquired brain injury (ABI). The present study evaluated effects of two differentially distributed protocols of delayed, voluntary exercise on cognitive recovery using an allocentric place learning task in an 8-arm radial maze. Fifty-four Wistar rats were subjected to either bilateral transection of the fimbria-fornix (FF) or to sham surgery. Twenty-one days postinjury, the animals started exercising in running wheels either for 14 consecutive days (FF/exercise daily [ExD], sham/ExD) or every other day for 14 days (FF/exercise every second day [ExS], sham/ExS). Additional groups were given no exercise treatment (FF/not exercise [NE], sham/NE). Regardless of how exercise was distributed, we found no cognitively enhancing effects of exercise in the brain injured animals. Design and protocol factors possibly affecting the efficacy of post-ABI exercise are discussed. PMID:27807517

  1. Delphi definition of the EADC-ADNI Harmonized Protocol for hippocampal segmentation on magnetic resonance

    PubMed Central

    Boccardi, Marina; Bocchetta, Martina; Apostolova, Liana G.; Barnes, Josephine; Bartzokis, George; Corbetta, Gabriele; DeCarli, Charles; deToledo-Morrell, Leyla; Firbank, Michael; Ganzola, Rossana; Gerritsen, Lotte; Henneman, Wouter; Killiany, Ronald J.; Malykhin, Nikolai; Pasqualetti, Patrizio; Pruessner, Jens C.; Redolfi, Alberto; Robitaille, Nicolas; Soininen, Hilkka; Tolomeo, Daniele; Wang, Lei; Watson, Craig; Wolf, Henrike; Duvernoy, Henri; Duchesne, Simon; Jack, Clifford R.; Frisoni, Giovanni B.

    2015-01-01

    Background This study aimed to have international experts converge on a harmonized definition of whole hippocampus boundaries and segmentation procedures, to define standard operating procedures for magnetic resonance (MR)-based manual hippocampal segmentation. Methods The panel received a questionnaire regarding whole hippocampus boundaries and segmentation procedures. Quantitative information was supplied to allow evidence-based answers. A recursive and anonymous Delphi procedure was used to achieve convergence. Significance of agreement among panelists was assessed by exact probability on Fisher’s and binomial tests. Results Agreement was significant on the inclusion of alveus/fimbria (P =.021), whole hippocampal tail (P =.013), medial border of the body according to visible morphology (P =.0006), and on this combined set of features (P =.001). This definition captures 100% of hippocampal tissue, 100% of Alzheimer’s disease-related atrophy, and demonstrated good reliability on preliminary intrarater (0.98) and inter-rater (0.94) estimates. Discussion Consensus was achieved among international experts with respect to hippocampal segmentation using MR resulting in a harmonized segmentation protocol. PMID:25130658

  2. Chronic Cocaine Administration Causes Extensive White Matter Damage in Brain: Diffusion Tensor Imaging and Immunohistochemistry Studies

    PubMed Central

    Narayana, Ponnada A.; Herrera, Juan J.; Bockhorst, Kurt H; Esparza-Coss, Emilio; Xia, Ying; Steinberg, Joel L.; Moeller, F. Gerard

    2014-01-01

    The effect of chronic cocaine exposure on multiple white matter structures in rodent brain was examined using diffusion tensor imaging (DTI), locomotor behavior, and end point histology. The animals received either cocaine at a dose of 100 mg/kg (N=19), or saline (N=17) for 28 days through an implanted osmotic minipump. The animals underwent serial DTI scans, locomotor assessment, and end point histology for determining the expressions of myelin basic protein (MBP), neurofilament-heavy protein (NF-H), proteolipid protein (PLP), Nogo-A, aquaporin-4 (AQP-4), and growth associated protein – 43 (GAP-43). Differences in the DTI measures were observed in the splenium (scc) and genu (gcc) of the corpus callosum (cc), fimbria (fi), and the internal capsule (ic). Significant increase in the activity in the fine motor movements and decrease in the number of rearing events were observed in the cocaine treated animals. Reduced MBP and Nogo-A, and increased GAP-43 expressions were most consistently observed in these structures. A decrease in the NF-H expression was observed in fi and ic. The reduced expression of Nogo-A and increased GAP-43 may suggest destabilization of axonal connectivity and increased neurite growth with aberrant connections. Increased GAP-43 suggests drug induced plasticity or a possible repair mechanism response. The findings indicated that multiple white matter tracts are affected following chronic cocaine exposure. PMID:24507117

  3. Mutant p53 regulates ovarian cancer transformed phenotypes through autocrine matrix deposition

    PubMed Central

    Iwanicki, Marcin P.; Chen, Hsing-Yu; Iavarone, Claudia; Zervantonakis, Ioannis K.; Muranen, Taru; Novak, Marián; Ince, Tan A.; Brugge, Joan S.

    2016-01-01

    High-grade serous ovarian carcinoma (HGS-OvCa) harbors p53 mutations and can originate from the epithelial cell compartment of the fallopian tube fimbriae. From this site, neoplastic cells detach, survive in the peritoneal cavity, and form cellular clusters that intercalate into the mesothelium to form ovarian and peritoneal masses. To examine the contribution of mutant p53 to phenotypic alterations associated with HGS-OvCA, we developed live-cell microscopy assays that recapitulate these early events in cultured fallopian tube nonciliated epithelial (FNE) cells. Expression of stabilizing mutant variants of p53, but not depletion of endogenous wild-type p53, in FNE cells promoted survival and cell-cell aggregation under conditions of cell detachment, leading to the formation of cell clusters with mesothelium-intercalation capacity. Mutant p53R175H-induced phenotypes were dependent on fibronectin production, α5β1 fibronectin receptor engagement, and TWIST1 expression. These results indicate that FNE cells expressing stabilizing p53 mutants acquire anchorage independence and subsequent mesothelial intercalation capacity through a mechanism involving mesenchymal transition and matrix production. These findings provide important new insights into activities of mutant p53 in the cells of origin of HGS-OvCa. PMID:27482544

  4. Subinhibitory Concentrations of Allicin Decrease Uropathogenic Escherichia coli (UPEC) Biofilm Formation, Adhesion Ability, and Swimming Motility

    PubMed Central

    Yang, Xiaolong; Sha, Kaihui; Xu, Guangya; Tian, Hanwen; Wang, Xiaoying; Chen, Shanze; Wang, Yi; Li, Jingyu; Chen, Junli; Huang, Ning

    2016-01-01

    Uropathogenic Escherichia coli (UPEC) biofilm formation enables the organism to avoid the host immune system, resist antibiotics, and provide a reservoir for persistent infection. Once the biofilm is established, eradication of the infection becomes difficult. Therefore, strategies against UPEC biofilm are urgently required. In this study, we investigated the effect of allicin, isolated from garlic essential oil, on UPEC CFT073 and J96 biofilm formation and dispersal, along with its effect on UPEC adhesion ability and swimming motility. Sub-inhibitory concentrations (sub-MICs) of allicin decreased UPEC biofilm formation and affected its architecture. Allicin was also capable of dispersing biofilm. Furthermore, allicin decreased the bacterial adhesion ability and swimming motility, which are important for biofilm formation. Real-time quantitative polymerase chain reaction (RT-qPCR) revealed that allicin decreased the expression of UPEC type 1 fimbriae adhesin gene fimH. Docking studies suggested that allicin was located within the binding pocket of heptyl α-d-mannopyrannoside in FimH and formed hydrogen bonds with Phe1 and Asn135. In addition, allicin decreased the expression of the two-component regulatory systems (TCSs) cognate response regulator gene uvrY and increased the expression of the RNA binding global regulatory protein gene csrA of UPEC CFT073, which is associated with UPEC biofilm. The findings suggest that sub-MICs of allicin are capable of affecting UPEC biofilm formation and dispersal, and decreasing UPEC adhesion ability and swimming motility. PMID:27367677

  5. Effect of Temperature on Fimbrial Gene Expression and Adherence of Enteroaggregative Escherichia coli.

    PubMed

    Hinthong, Woranich; Indrawattana, Nitaya; Pitaksajjakul, Pannamthip; Pipattanaboon, Chonlatip; Kongngoen, Thida; Tharnpoophasiam, Prapin; Worakhunpiset, Suwalee

    2015-07-23

    The influence of temperature on bacterial virulence has been studied worldwide from the viewpoint of climate change and global warming. The bacterium enteroaggregative Escherichia coli (EAEC) is the causative agent of watery diarrhea and shows an increasing incidence worldwide. Its pathogenicity is associated with the virulence factors aggregative adherence fimbria type I and II (AAFI and AAFII), encoded by aggA and aafA in EAEC strains 17-2 and 042, respectively. This study focused on the effect of temperature increases from 29 °C to 40 °C on fimbrial gene expression using real-time PCR, and on its virulence using an aggregative adherence assay and biofilm formation assay. Incubation at 32 °C caused an up-regulation in both EAEC strains 17-2 and strain 042 virulence gene expression. EAEC strain 042 cultured at temperature above 32 °C showed down-regulation of aafA expression except at 38 °C. Interestingly, EAEC cultured at a high temperature showed a reduced adherence to cells and an uneven biofilm formation. These results provide evidence that increases in temperature potentially affect the virulence of pathogenic EAEC, although the response varies in each strain.

  6. Prophylactic salpingectomy and prophylactic salpingoophorectomy for adnexal high-grade serous epithelial carcinoma: A reappraisal.

    PubMed

    Oliver Perez, M Reyes; Magriñá, Javier; García, Alvaro Tejerizo; Jiménez Lopez, Jesus Salvador

    2015-12-01

    At present, there is no effective screening of ovarian cancer. Primary prevention may be the only strategy to decrease the mortality from ovarian cancer, not only in women at high risk but also at low risk. Several recent studies have identified the distal fimbriae end of the fallopian tubes as primary precursor of High-grade serous carcinoma. Serous tubal intraepithelial carcinomas and occult invasive serous carcinomas have been identified in 2-17% of the fallopian tubes of BRCA1/2 positive women undergoing risk-reducing salpingo-oophorectomy. Removal of the fallopian tubes with ovarian preservation has been suggested as a reasonable strategy that could reduce the risk of developing ovarian carcinoma in both low and high-risk women. It has been proposed after childbearing in women at high risk to be followed by bilateral oophorectomy at a later date. Bilateral salpingectomy is also suggested for low risk women, at the time of other benign gynaecologic surgery as a primary preventive strategy. Some studies have shown a risk reduction of ovarian cancer in women with bilateral prophylactic salpingectomy. Current research regarding bilateral salpingoophorectomy as primary prevention approach of ovarian cancer is reviewed here. In addition, the potential use of bilateral salpingectomy as prevention approach of ovarian cancer is discussed.

  7. Role of the PhoP-PhoQ gene regulatory system in adaptation of Yersinia pestis to environmental stress in the flea digestive tract.

    PubMed

    Vadyvaloo, Viveka; Viall, Austin K; Jarrett, Clayton O; Hinz, Angela K; Sturdevant, Daniel E; Joseph Hinnebusch, B

    2015-06-01

    The Yersinia pestis PhoPQ gene regulatory system is induced during infection of the flea digestive tract and is required to produce adherent biofilm in the foregut, which greatly enhances bacterial transmission during a flea bite. To understand the in vivo context of PhoPQ induction and to determine PhoP-regulated targets in the flea, we undertook whole-genome comparative transcriptional profiling of Y. pestis WT and ΔphoP strains isolated from infected fleas and from temperature-matched in vitro planktonic and flow-cell biofilm cultures. In the absence of PhoP regulation, the gene expression program indicated that the bacteria experienced diverse physiological stresses and were in a metabolically less active state. Multiple stress response genes, including several toxin-antitoxin loci and YhcN family genes responsible for increased acid tolerance, were upregulated in the phoP mutant during flea infection. The data implied that PhoPQ was induced by low pH in the flea gut, and that PhoP modulated physiological adaptation to acid and other stresses encountered during infection of the flea. This adaptive response, together with PhoP-dependent modification of the bacterial outer surface that includes repression of pH 6 antigen fimbriae, supports stable biofilm development in the flea foregut.

  8. RosE represses Std fimbrial expression in Salmonella enterica serotype Typhimurium

    PubMed Central

    Chessa, Daniela; Winter, Maria G; Nuccio, Sean-Paul; Tükel, Çagla; Bäumler, Andreas J

    2008-01-01

    The Salmonella enterica serotype Typhimurium (S. typhimurium) genome contains a large repertoire of putative fimbrial operons that remain poorly characterized because they are not expressed in vitro. In this study, insertions that induced expression of the putative stdABCD fimbrial operon were identified from a random bank of transposon mutants by screening with immuno-magnetic particles for ligand expression (SIMPLE). Transposon insertions upstream of csgC and lrhA or within dam, setB and STM4463 (renamed rosE) resulted in expression of StdA and its assembly into fimbrial filaments on the cell surface. RosE is a novel negative regulator of Std fimbrial expression as indicated by its repression of a std::lacZ reporter construct and by binding of the purified protein to a DNA region upstream of the stdA start codon. Expression of Std fimbriae in the rosE mutant resulted in increased attachment of S. typhimurium to human colonic epithelial cell lines (T-84 and CaCo-2). A rosE mutant exhibited a reduced ability to compete with virulent S. typhimurium for colonization of murine organs, while no defect was observed when both competing strains carried a stdAB deletion. These data suggest that a tight control of Std fimbrial expression mediated by RosE is required during host pathogen interaction. PMID:18331470

  9. Cranberry Xyloglucan Structure and Inhibition of Escherichia coli Adhesion to Epithelial Cells.

    PubMed

    Hotchkiss, Arland T; Nuñez, Alberto; Strahan, Gary D; Chau, Hoa K; White, André K; Marais, Jannie P J; Hom, Kellie; Vakkalanka, Malathi S; Di, Rong; Yam, Kit L; Khoo, Christina

    2015-06-17

    Cranberry juice has been recognized as a treatment for urinary tract infections on the basis of scientific reports of proanthocyanidin anti-adhesion activity against Escherichia coli as well as from folklore. Xyloglucan oligosaccharides were detected in cranberry juice and the residue remaining following commercial juice extraction that included pectinase maceration of the pulp. A novel xyloglucan was detected through tandem mass spectrometry analysis of an ion at m/z 1055 that was determined to be a branched, three hexose, four pentose oligosaccharide consistent with an arabino-xyloglucan structure. Two-dimensional nuclear magnetic resonance spectroscopy analysis provided through-bond correlations for the α-L-Araf (1→2) α-D-Xylp (1→6) β-D-Glcp sequence, proving the S-type cranberry xyloglucan structure. Cranberry xyloglucan-rich fractions inhibited the adhesion of E. coli CFT073 and UTI89 strains to T24 human bladder epithelial cells and that of E. coli O157:H7 to HT29 human colonic epithelial cells. SSGG xyloglucan oligosaccharides represent a new cranberry bioactive component with E. coli anti-adhesion activity and high affinity for type 1 fimbriae.

  10. Anchoring of LPXTG-Like Proteins to the Gram-Positive Cell Wall Envelope.

    PubMed

    Siegel, Sara D; Reardon, Melissa E; Ton-That, Hung

    2016-04-21

    In Gram-positive bacteria, protein precursors with a signal peptide and a cell wall sorting signal (CWSS)-which begins with an LPXTG motif, followed by a hydrophobic domain and a tail of positively charged residues-are targeted to the cell envelope by a transpeptidase enzyme call sortase. Evolution and selective pressure gave rise to six classes of sortase, i.e., SrtA-F. Only class C sortases are capable of polymerizing substrates harboring the pilin motif and CWSS into protein polymers known as pili or fimbriae, whereas the others perform cell wall anchoring functions. Regardless of the products generated from these sortases, the basic principle of sortase-catalyzed transpeptidation is the same. It begins with the cleavage of the LPXTG motif, followed by the cross-linking of this cleaved product at the threonine residue to a nucleophile, i.e., an active amino group of the peptidoglycan stem peptide or the lysine residue of the pilin motif. This chapter will summarize the efforts to identify and characterize sortases and their associated pathways with emphasis on the cell wall anchoring function.

  11. Polymorphism of Bordetella pertussis Isolates Circulating for the Last 10 Years in France, Where a Single Effective Whole-Cell Vaccine Has Been Used for More than 30 Years

    PubMed Central

    Weber, Christian; Boursaux-Eude, Caroline; Coralie, Gilberte; Caro, Valérie; Guiso, Nicole

    2001-01-01

    We compared Bordetella pertussis isolates collected in France over the last 10 years, the vaccine strains used for more than 30 years, and isolates collected before the introduction of generalized vaccination. The analysis included serotyping, pulsed-field gel electrophoresis of chromosomal DNA after digestion with XbaI and SpeI, and sequencing of the pt S1 gene, encoding the S1 subunit of pertussis toxin, and the prn gene, encoding the adhesin pertactin. We found that the incidence of infection increases every 3 years. Ninety-five per cent of the isolates analyzed express type 3 fimbriae. Most of the isolates circulating since 1991, unlike the vaccinal strains, express a type A pertussis toxin and a type 2 pertactin. The isolates could be classified into five major groups by pulsed-field gel electrophoresis. Most of these groups correlated with the pertactin type expressed by the isolates. Pulsed-field gel electrophoresis is more discriminative than sequencing particular genes since it could differentiate isolates expressing type 2 pertactin into two subgroups: those circulating in 1993 to 1997 and those circulating in 1997 to 2001. This observation suggests that there has been continuous evolution of the B. pertussis population. PMID:11724851

  12. Immunogenicity and safety of a monovalent, multicomponent acellular pertussis vaccine in 15 month-6-year-old German children. Monovalent Acellular Pertussis Vaccine Study Group.

    PubMed

    Stehr, K; Heininger, U; Uhlenbusch, R; Angersbach, P; Hackell, J; Eckhardt, T

    1995-03-01

    Immunization against pertussis has been re-recommended for healthy children in Germany in 1991. In addition the former restriction of immunizing only in the first 2 years of life was abolished. In children born before 1991 immunization rates against pertussis were 15% or less. With the new recommendations physicians are now faced with an increasing demand of parents for catch-up vaccinations in these children. Since they were immunized against diphtheria and tetanus previously monovalent pertussis vaccines are needed for this indication. Therefore a monovalent, multicomponent acellular pertussis vaccine was studied in 249 German children 15 months to 6 years of age. Three doses were administered at 6-10 week intervals. Reactogenicity and antibody responses against the vaccine antigens pertussis toxin (PT), filamentous haemagglutinin (FHA), 69-kd antigen (pertactin) and fimbriae-2 (agglutinogen) were investigated. Local and systemic reactions were minimal in frequency and severity. Antibody responses against all vaccine antigens were pronounced with 93%-100% of vaccinees demonstrating at least four fold titre rises above pre-immunization after the third dose. These findings indicate that this monovalent, multicomponent acellular pertussis vaccine with excellent immunogenicity and low reactogenicity is an appropriate candidate for closing immunization gaps in older children in countries with previously low vaccination rates against pertussis. Based on the results of this study the monovalent acellular pertussis vaccine was licensed in Germany in January 1994.

  13. The pilus usher controls protein interactions via domain masking and is functional as an oligomer

    SciTech Connect

    Werneburg, Glenn T.; Li, Huilin; Henderson, Nadine S.; Portnoy, Erica B.; Sarowar, Samema; Hultgren, Scott J.; Thanassi, David G.

    2015-06-08

    The chaperone/usher (CU) pathway is responsible for biogenesis of organelles termed pili or fimbriae in Gram-negative bacteria. Type 1 pili expressed by uropathogenic Escherichia coli are prototypical structures assembled by the CU pathway. Assembly and secretion of pili by the CU pathway requires a dedicated periplasmic chaperone and a multidomain outer membrane protein termed the usher (FimD). We show that the FimD C-terminal domains provide the high-affinity substrate binding site, but that these domains are masked in the resting usher. Domain masking requires the FimD plug domain, which served as a central switch controlling usher activation. In addition, we demonstrate that usher molecules can act in trans for pilus biogenesis, providing conclusive evidence for a functional usher oligomer. These results reveal mechanisms by which molecular machines such as the usher regulate and harness protein-protein interactions, and suggest that ushers may interact in a cooperative manner during pilus assembly in bacteria.

  14. Milk-Based Nutraceutical for Treating Autoimmune Arthritis via the Stimulation of IL-10- and TGF-β-producing CD39+ Regulatory T Cells

    PubMed Central

    Maddaloni, Massimo; Kochetkova, Irina; Jun, SangMu; Callis, Gayle; Thornburg, Theresa; Pascual, David W.

    2015-01-01

    Autoimmune diseases arise from the loss of tolerance to self, and because the etiologies of such diseases are largely unknown, symptomatic treatments rely on anti-inflammatory and analgesic agents. Tolerogenic treatments that can reverse disease are preferred, but again, often thwarted by not knowing the responsible auto-antigens (auto-Ags). Hence, a viable alternative to stimulating regulatory T cells (Tregs) is to induce bystander tolerance. Colonization factor antigen I (CFA/I) has been shown to evoke bystander immunity and to hasten Ag-specific Treg development independent of auto-Ag. To translate in treating human autoimmune diseases, the food-based Lactococcus was engineered to express CFA/I fimbriae, and Lactococcus-CFA/I fermented milk fed to arthritic mice proved highly efficacious. Protection occurred via CD39+ Tregs producing TGF-β and IL-10 to potently suppress TNF-α production and neutrophil influx into the joints. Thus, these data demonstrate the feasibility of oral nutraceuticals for treating arthritis, and potency of protection against arthritis was improved relative to that obtained with Salmonella-CFA/I. PMID:25629976

  15. Identification of a Sustained Neurogenic Zone at the Dorsal Surface of the Adult Mouse Hippocampus and Its Regulation by the Chemokine SDF-1

    PubMed Central

    Belmadani, Abdelhak; Ren, Dongjun; Bhattacharyya, Bula J.; Rothwangl, Katharina B.; Hope, Thomas J.; Perlman, Harris; Miller, Richard J.

    2015-01-01

    We identified a previously unknown neurogenic region at the dorsal surface of the hippocampus; (the “subhippocampal zone,” SHZ) in the adult brain. Using a reporter mouse in which SHZ cells and their progeny could be traced through the expression of EGFP under the control of the CXCR4 chemokine receptor promoter we observed the presence of a pool of EGFP expressing cells migrating in direction of the dentate gyrus (DG), which is maintained throughout adulthood. This population appeared to originate from the SHZ where cells entered a caudal migratory stream (aCMS) that included the fimbria, the meninges and the DG. Deletion of CXCR4 from neural stem cells (NSCs) or neuroinflammation resulted in the appearance of neurons in the DG, which were the result of migration of NSCs from the SHZ. Some of these neurons were ectopically placed. Our observations indicate that the SHZ is a neurogenic zone in the adult brain through migration of NSCs in the aCMS. Regulation of CXCR4 signaling in these cells may be involved in repair of the DG and may also give rise to ectopic granule cells in the DG in the context of neuropathology. PMID:25656357

  16. Expression of verocytotoxic Escherichia coli antigens in tobacco seeds and evaluation of gut immunity after oral administration in mouse model.

    PubMed

    Rossi, Luciana; Di Giancamillo, Alessia; Reggi, Serena; Domeneghini, Cinzia; Baldi, Antonella; Sala, Vittorio; Dell'Orto, Vittorio; Coddens, Annelies; Cox, Eric; Fogher, Corrado

    2013-01-01

    Verocytotoxic Escherichia (E.) coli strains are responsible for swine oedema disease, which is an enterotoxaemia that causes economic losses in the pig industry. The production of a vaccine for oral administration in transgenic seeds could be an efficient system to stimulate local immunity. This study was conducted to transform tobacco plants for the seed-specific expression of antigenic proteins from a porcine verocytotoxic E. coli strain. Parameters related to an immunological response and possible adverse effects on the oral administration of obtained tobacco seeds were evaluated in a mouse model. Tobacco was transformed via Agrobacteium tumefaciens with chimeric constructs containing structural parts of the major subunit FedA of the F18 adhesive fimbriae and VT2e B-subunit genes under control of a seed specific GLOB promoter. We showed that the foreign Vt2e-B and F18 genes were stably accumulated in storage tissue by the immunostaining method. In addition, Balb-C mice receiving transgenic tobacco seeds via the oral route showed a significant increase in IgA-positive plasma cell presence in tunica propria when compared to the control group with no observed adverse effects. Our findings encourage future studies focusing on swine for evaluation of the protective effects of transformed tobacco seeds against E. coli infection.

  17. Structure, Function, and Assembly of Adhesive Organelles by Uropathogenic Bacteria

    PubMed Central

    Chahales, Peter; Thanassi, David G.

    2015-01-01

    Bacteria assemble a wide range of adhesive proteins, termed adhesins, to mediate binding to receptors and colonization of surfaces. For pathogenic bacteria, adhesins are critical for early stages of infection, allowing the bacteria to initiate contact with host cells, colonize different tissues, and establish a foothold within the host. The adhesins expressed by a pathogen are also critical for bacterial-bacterial interactions and the formation of bacterial communities such as biofilms. The ability to adhere to host tissues is particularly important for bacteria that colonize sites such as the urinary tract, where the flow of urine functions to maintain sterility by washing away non-adherent pathogens. Adhesins vary from monomeric proteins that are directly anchored to the bacterial surface to polymeric, hairlike fibers that extend out from the cell surface. These latter fibers are termed pili or fimbriae, and were among the first identified virulence factors of uropathogenic Escherichia coli. Studies since then have identified a range of both pilus and non-pilus adhesins that contribute to bacterial colonization of the urinary tract, and have revealed molecular details of the structures, assembly pathways, and functions of these adhesive organelles. In this review, we describe the different types of adhesins expressed by both Gram-negative and Gram-positive uropathogens, what is known about their structures, how they are assembled on the bacterial surface, and the functions of specific adhesins in the pathogenesis of urinary tract infections. PMID:26542038

  18. N-Acetylglucosamine Functions in Cell Signaling

    PubMed Central

    Konopka, James B.

    2012-01-01

    The amino sugar N-acetylglucosamine (GlcNAc) is well known for the important structural roles that it plays at the cell surface. It is a key component of bacterial cell wall peptidoglycan, fungal cell wall chitin, and the extracellular matrix of animal cells. Interestingly, recent studies have also identified new roles for GlcNAc in cell signaling. For example, GlcNAc stimulates the human fungal pathogen Candida albicans to undergo changes in morphogenesis and expression of virulence genes. Pathogenic E. coli responds to GlcNAc by altering the expression of fimbriae and CURLI fibers that promote biofilm formation and GlcNAc stimulates soil bacteria to undergo changes in morphogenesis and production of antibiotics. Studies with animal cells have revealed that GlcNAc influences cell signaling through the posttranslational modification of proteins by glycosylation. O-linked attachment of GlcNAc to Ser and Thr residues regulates a variety of intracellular proteins, including transcription factors such as NFκB, c-myc, and p53. In addition, the specificity of Notch family receptors for different ligands is altered by GlcNAc attachment to fucose residues in the extracellular domain. GlcNAc also impacts signal transduction by altering the degree of branching of N-linked glycans, which influences cell surface signaling proteins. These emerging roles of GlcNAc as an activator and mediator of cellular signaling in fungi, animals, and bacteria will be the focus of this paper. PMID:23350039

  19. Serous tubal intraepithelial carcinoma localizes to the tubal-peritoneal junction: a pivotal clue to the site of origin of extrauterine high-grade serous carcinoma (ovarian cancer).

    PubMed

    Seidman, Jeffrey D

    2015-03-01

    Recent data suggest that intraepithelial carcinoma of the fallopian tube [serous tubal intraepithelial carcinoma (STIC)] is the precursor of high-grade extrauterine serous carcinoma. A more specific location for the origin of this lesion is suggested by the recently described junction between the fallopian tubal epithelium and the peritoneum [tubal-peritoneal junction (TPJ)]. Fallopian tubes from 202 patients with advanced-stage high-grade extrauterine serous carcinoma or carcinosarcoma were evaluated histologically as were 124 prophylactic salpingo-oophorectomy specimens. These included 54 patients with BRCA or other high-risk mutation or a family history of BRCA mutation and 70 with a personal or family history of breast carcinoma. STIC was found in 81 of 202 patients with serous carcinoma (40.1%). STIC was present in 73 of 141 (52%) cases in which the fimbriae were present and in 62 of 100 (62%) cases in which the TPJ was present (P not significant). In comparison with these groups, when fimbriae and TPJ were absent, STIC was found in 8 of 61 (13%) cases (P<0.0001). None of the prophylactic specimens contained STIC. The mean size of STIC was 1.7 mm. In 32 cases (39.5%), the lesion was flat and in 49 (60.5%), papillary. The mean size of flat STICs was 0.8 mm as compared with 2.3 mm for papillary STICs (P=0.00005). STIC was identified in the same tissue fragment as the junction in 48 cases. The mean distance of STIC to the junction was 1.8 mm. In 11 cases, STIC was flanked by peritoneal mesothelium on one side and tubal epithelium on the opposite side. In 51 patients, the mean distance of invasive carcinoma from the TPJ was 1.8 mm. This distance was 1.9 mm when STIC was present (37 cases) in comparison with 1.5 mm when STIC was absent (14 cases) (P not significant). In 27 of 42 cases (64%), STIC was contiguous with invasive carcinoma. Lamina propria invasion was present in 71% of cases in which STIC was present as compared with 26% of cases in which STIC was

  20. Bacterial Surface Appendages Strongly Impact Nanomechanical and Electrokinetic Properties of Escherichia coli Cells Subjected to Osmotic Stress

    PubMed Central

    Francius, Grégory; Polyakov, Pavel; Merlin, Jenny; Abe, Yumiko; Ghigo, Jean-Marc; Merlin, Christophe; Beloin, Christophe; Duval, Jérôme F. L.

    2011-01-01

    The physicochemical properties and dynamics of bacterial envelope, play a major role in bacterial activity. In this study, the morphological, nanomechanical and electrohydrodynamic properties of Escherichia coli K-12 mutant cells were thoroughly investigated as a function of bulk medium ionic strength using atomic force microscopy (AFM) and electrokinetics (electrophoresis). Bacteria were differing according to genetic alterations controlling the production of different surface appendages (short and rigid Ag43 adhesins, longer and more flexible type 1 fimbriae and F pilus). From the analysis of the spatially resolved force curves, it is shown that cells elasticity and turgor pressure are not only depending on bulk salt concentration but also on the presence/absence and nature of surface appendage. In 1 mM KNO3, cells without appendages or cells surrounded by Ag43 exhibit large Young moduli and turgor pressures (∼700–900 kPa and ∼100–300 kPa respectively). Under similar ionic strength condition, a dramatic ∼50% to ∼70% decrease of these nanomechanical parameters was evidenced for cells with appendages. Qualitatively, such dependence of nanomechanical behavior on surface organization remains when increasing medium salt content to 100 mM, even though, quantitatively, differences are marked to a much smaller extent. Additionally, for a given surface appendage, the magnitude of the nanomechanical parameters decreases significantly when increasing bulk salt concentration. This effect is ascribed to a bacterial exoosmotic water loss resulting in a combined contraction of bacterial cytoplasm together with an electrostatically-driven shrinkage of the surface appendages. The former process is demonstrated upon AFM analysis, while the latter, inaccessible upon AFM imaging, is inferred from electrophoretic data interpreted according to advanced soft particle electrokinetic theory. Altogether, AFM and electrokinetic results clearly demonstrate the intimate

  1. Two consecutive randomized controlled pertussis booster trials in children initially vaccinated in infancy with an acellular vaccine: The first with a five-component Tdap vaccine to 5-year olds and the second with five- or monocomponent Tdap vaccines at age 14-15 years.

    PubMed

    Carlsson, R M; Gustafsson, L; Hallander, H O; Ljungman, M; Olin, P; Gothefors, L; Nilsson, L; Netterlid, E

    2015-07-17

    Prior study children from a DTaP efficacy trial were recruited at ages 5 and 15 years to randomized booster trials addressing immunogenicity and reactogenicity; 475 preschool children received mixed or separate injections of a reduced antigen vaccine (Tdap5, Sanofi Pasteur MSD) and an inactivated polio vaccine, and 230 adolescents received the same or another booster vaccine (Tdap1, SSI, Denmark). Pre-vaccination antibody concentrations against pertussis antigens were significantly higher at 15 than 5 years of age, probably due to natural boosting between the studies. Tdap5 induced comparable anti-PT concentrations at both ages, but antibody responses were significantly higher to filamentous haemagglutinin, pertactin and fimbriae 2/3 in adolescents. As expected, a higher amount of PT (Tdap1, 20μg) induced a stronger anti-PT response than a lower amount (Tdap5, 2.5μg). The frequency of adverse events was low and there were no serious adverse reactions. All local reactions had an early onset and a short duration. A large swelling or redness of more than half of the upper arm circumference was reported in 8/475 5-year-olds and in 6/230 15-year-olds. Children vaccinated with Tdap5 reported more moderate pain in adolescence than at preschool age, whereas itching was only reported in preschool children. Sweden introduced DTaP vaccines in 1996 after a 17-year hiatus with no general pertussis vaccination and pertussis was still endemic at the time of the studies. The frequency of adverse events was nevertheless low in both preschool children and adolescents and antibody responses were adequate. These studies document immunogenicity and reactogenicity in a trial cohort consecutively vaccinated with acellular pertussis vaccines from infancy to adolescence. The adolescent study was registered at ClinicalTrials.gov on 26 March 2009 (NCT00870350).

  2. Post-breeding season distribution of black-footed and Laysan albatrosses satellite-tagged in Alaska: Inter-specific differences in spatial overlap with North Pacific fisheries

    USGS Publications Warehouse

    Fischer, K.N.; Suryan, R.M.; Roby, D.D.; Balogh, G.R.

    2009-01-01

    We integrated satellite-tracking data from black-footed albatrosses (Phoebastria nigripes; n = 7) and Laysan albatrosses captured in Alaska (Phoebastria immutabilis; n = 18) with data on fishing effort and distribution from commercial fisheries in the North Pacific in order to assess potential risk from bycatch. Albatrosses were satellite-tagged at-sea in the Central Aleutian Islands, Alaska, and tracked during the post-breeding season, July-October 2005 and 2006. In Alaskan waters, fishing effort occurred almost exclusively within continental shelf and slope waters. Potential fishery interaction for black-footed albatrosses, which most often frequented shelf-slope waters, was greatest with sablefish (Anoplopoma fimbria) longline and pot fisheries and with the Pacific halibut (Hippoglossus stenolepsis) longline fishery. In contrast, Laysan albatrosses spent as much time over oceanic waters beyond the continental shelf and slope, thereby overlapping less with fisheries in Alaska than black-footed albatrosses. Regionally, Laysan albatrosses had the greatest potential fishery interaction with the Atka mackerel (Pleurogrammus monopterygius) trawl fishery in the Western Aleutian Islands and the sablefish pot fishery in the Central Aleutian Islands. Black-footed albatrosses ranged further beyond Alaskan waters than Laysan albatrosses, overlapping west coast Canada fisheries and pelagic longline fisheries in the subarctic transition domain; Laysan albatrosses remained north of these pelagic fisheries. Due to inter-specific differences in oceanic distribution and habitat use, the overlap of fisheries with the post-breeding distribution of black-footed albatrosses is greater than that for Laysan albatrosses, highlighting inter-specific differences in potential vulnerability to bycatch and risk of population-level impacts from fisheries. ?? 2008 Elsevier Ltd.

  3. Plasmid-Encoded Tetracycline Efflux Pump Protein Alters Bacterial Stress Responses and Ecological Fitness of Acinetobacter oleivorans

    PubMed Central

    Hong, Hyerim; Jung, Jaejoon; Park, Woojun

    2014-01-01

    Acquisition of the extracellular tetracycline (TC) resistance plasmid pAST2 affected host gene expression and phenotype in the oil-degrading soil bacterium, Acinetobacter oleivorans DR1. Whole-transcriptome profiling of DR1 cells harboring pAST2 revealed that all the plasmid genes were highly expressed under TC conditions, and the expression levels of many host chromosomal genes were modulated by the presence of pAST2. The host energy burden imposed by replication of pAST2 led to (i) lowered ATP concentrations, (ii) downregulated expression of many genes involved in cellular growth, and (iii) reduced growth rate. Interestingly, some phenotypes were restored by deleting the plasmid-encoded efflux pump gene tetH, suggesting that the membrane integrity changes resulting from the incorporation of efflux pump proteins also resulted in altered host response under the tested conditions. Alteration of membrane integrity by tetH deletion was shown by measuring permeability of fluorescent probe and membrane hydrophobicity. The presence of the plasmid conferred peroxide and superoxide resistance to cells, but only peroxide resistance was diminished by tetH gene deletion, suggesting that the plasmid-encoded membrane-bound efflux pump protein provided peroxide resistance. The downregulation of fimbriae-related genes presumably led to reduced swimming motility, but this phenotype was recovered by tetH gene deletion. Our data suggest that not only the plasmid replication burden, but also its encoded efflux pump protein altered host chromosomal gene expression and phenotype, which also alters the ecological fitness of the host in the environment. PMID:25229538

  4. Cranberry extract inhibits in vitro adhesion of F4 and F18(+)Escherichia coli to pig intestinal epithelium and reduces in vivo excretion of pigs orally challenged with F18(+) verotoxigenic E. coli.

    PubMed

    Coddens, Annelies; Loos, Michaela; Vanrompay, Daisy; Remon, Jean Paul; Cox, Eric

    2017-01-20

    F4(+)E. coli and F18(+)E. coli infections are an important threat for pig industry worldwide. Antibiotics are commonly used to treat infected piglets, but the emerging development of resistance against antibiotics raises major concerns. Hence, alternative therapies to prevent pigs from F4(+)E. coli and F18(+)E. coli infections need to be developed. Since cranberry previously showed anti-adhesive activity against uropathogenic E. coli, we aimed to investigate whether cranberry extract could also inhibit binding of F4(+)E. coli and F18(+)E. coli to pig intestinal epithelium. Using the in vitro villus adhesion assay, we found that low concentrations of cranberry extract (20μg or 100μg/ml) have strong inhibitory activity on F4(+)E. coli (75.3%, S.D.=9.31 or 95.8%, S.D.=2.56, respectively) and F18(+)E. coli adherence (100% inhibition). This effect was not due to antimicrobial activity. Moreover, cranberry extract (10mg or 100mg) could also abolish in vivo binding of F4 and F18 fimbriae to the pig intestinal epithelium in ligated loop experiments. Finally, two challenge experiments with F18(+)E. coli were performed to address the efficacy of in-feed or water supplemented cranberry extract. No effect could be observed in piglets that received cranberry extract only in feed (1g/kg or 10g/kg). However, supplementation of feed (10g/kg) and drinking water (1g/L) significantly decreased excretion and diarrhea. The decreased infection resulted in a decreased serum antibody response indicating reduced exposure to F18(+)E. coli.

  5. Maternal exposure to hexachlorophene targets intermediate-stage progenitor cells in the hippocampal neurogenesis involving myelin vacuolation of cholinergic and glutamatergic inputs in mice.

    PubMed

    Kato, Mizuho; Abe, Hajime; Itahashi, Megu; Kikuchihara, Yoh; Kimura, Masayuki; Mizukami, Sayaka; Yoshida, Toshinori; Shibutani, Makoto

    2016-02-01

    Hexachlorophene (HCP) has been shown to induce myelin vacuolation due to intramyelinic edema of the nerve fibers in animal neural tissue. We investigated the maternal exposure effect of HCP on hippocampal neurogenesis in the offspring of pregnant mice supplemented with 0 (control), 33 or 100 ppm HCP in diet from gestational day 6 to day 21 after delivery. On postnatal day (PND) 21, offspring as examined in males exhibited decreased granule cell lineage populations expressing paired box 6, sex-determining region Y-box 2 and eomesodermin in the hippocampal subgranular zone (SGZ) accompanied by myelin vacuolation involving white matter tracts of the hippocampal fimbria at ≥ 33 ppm. However, SGZ cellular populations expressing brain lipid binding protein and doublecortin were unchanged at any dose. Transcript expression of cholinergic receptor genes, Chrna4 and Chrnb2, and glutamate receptor genes, Grm1 and Grin2d, examined at 100 ppm, decreased in the dentate gyrus. HCP exposure did not alter the number of proliferating or apoptotic cells in the SGZ, or reelin- or calcium-binding protein-expressing γ-aminobutyric acid (GABA)ergic interneurons in the dentate hilus, on PND 21 and PND 77. All neurogenesis-related changes observed in HCP-exposed offspring on PND 21 disappeared on PND 77, suggesting that maternal HCP exposure at ≥ 33 ppm reversibly decreased type 2 intermediate-stage progenitor cells in the hippocampal neurogenesis. Myelin vacuolation might be responsible for changes in neurogenesis possibly by reducing nerve conduction velocity of cholinergic inputs from the septal-hippocampal pathway to granule cell lineages and/or GABAergic interneurons, and of glutamatergic inputs to granule cell lineages.

  6. The isotopic biosignatures of photo- vs. thiotrophic bivalves: are they preserved in fossil shells?

    PubMed

    Dreier, A; Loh, W; Blumenberg, M; Thiel, V; Hause-Reitner, D; Hoppert, M

    2014-09-01

    Symbiont-bearing and non-symbiotic marine bivalves were used as model organisms to establish biosignatures for the detection of distinctive symbioses in ancient bivalves. For this purpose, the isotopic composition of lipids (δ13C) and bulk organic shell matrix (δ13C, δ34S, δ15N) from shells of several thiotrophic, phototrophic, or non-symbiotic bivalves were compared (phototrophic: Fragum fragum, Fragum unedo, Tridacna maxima; thiotrophic: Codakia tigerina, Fimbria fimbriata, Anodontia sp.; non-symbiotic: Tapes dorsatus, Vasticardium vertebratum, Scutarcopagia sp.). ∆13C values of bulk organic shell matrices, most likely representing mainly original shell protein/chitin biomass, were depleted in thio- and phototrophic bivalves compared to non-symbiotic bivalves. As the bulk organic shell matrix also showed a major depletion of δ15N (down to -2.2 ‰) for thiotrophic bivalves, combined δ13C and δ15N values are useful to differentiate between thio-, phototrophic, and non-symbiotic lifestyles. However, the use of these isotopic signatures for the study of ancient bivalves is limited by the preservation of the bulk organic shell matrix in fossils. Substantial alteration was clearly shown by detailed microscopic analyses of fossil (late Pleistocene) T. maxima and Trachycardium lacunosum shell, demonstrating a severe loss of quantity and quality of bulk organic shell matrix with time. Likewise, the composition and δ13C-values of lipids from empty shells indicated that a large part of these compounds derived from prokaryotic decomposers. The use of lipids from ancient shells for the reconstruction of the bivalve's life style therefore appears to be restricted.

  7. Molecular Requirements for Transformation of Fallopian Tube Epithelial Cells into Serous Carcinoma12

    PubMed Central

    Jazaeri, Amir A; Bryant, Jennifer L; Park, Hong; Li, Hui; Dahiya, Neetu; Stoler, Mark H; Ferriss, James Stuart; Dutta, Anindya

    2011-01-01

    Although controversial, recent studies suggest that serous ovarian carcinomas may arise from fallopian tube fimbria rather than ovarian surface epithelium. We developed an in vitro model for serous carcinogenesis in which primary human fallopian tube epithelial cells (FTECs) were exposed to potentially oncogenic molecular alterations delivered by retroviral vectors. To more closely mirror in vivo conditions, transformation of FTECs was driven by the positive selection of growth-promoting alterations rather antibiotic selection. Injection of the transformed FTEC lines in SCID mice resulted in xenografts with histologic and immunohistochemical features indistinguishable from poorly differentiated serous carcinomas. Transcriptional profiling revealed high similarity among the transformed and control FTEC lines and patient-derived serous ovarian carcinoma cells and was used to define a malignancy-related transcriptional signature. Oncogene-treated FTEC lines were serially analyzed using quantitative reverse transcription-polymerase chain reaction and immunoblot analysis to identify oncogenes whose expression was subject to positive selection. The combination of p53 and Rb inactivation (mediated by SV40 T antigen), hTERT expression, and oncogenic C-MYC and HRAS accumulation showed positive selection during transformation. Knockdown of each of these selected components resulted in significant growth inhibition of the transformed cell lines that correlated with p27 accumulation. The combination of SV40 T antigen and hTERT expression resulted in immortalized cells that were nontumorigenic in mice, whereas forced expression of a dominant-negative p53 isoform (p53DD) and hTERT resulted in