scholarly journals Role of Extracellular Transaldolase from Bifidobacterium bifidum in Mucin Adhesion and Aggregation

2012 ◽  
Vol 78 (11) ◽  
pp. 3992-3998 ◽  
Author(s):  
Irene González-Rodríguez ◽  
Borja Sánchez ◽  
Lorena Ruiz ◽  
Francesca Turroni ◽  
Marco Ventura ◽  
...  
Keyword(s):  
Bacterial Species ◽  
Bifidobacterium Bifidum ◽  
Surface Proteins ◽  
Proteinase K ◽  
Intestinal Cell ◽  
Immunogold Electron Microscopy ◽  
Content Type ◽  
Colonization Factor ◽  
Intestinal Cell Line

ABSTRACTThe ability of bifidobacteria to establish in the intestine of mammals is among the main factors considered to be important for achieving probiotic effects. The role of surface molecules fromBifidobacterium bifidumtaxon in mucin adhesion capability and the aggregation phenotype of this bacterial species was analyzed. Adhesion to the human intestinal cell line HT29 was determined for a collection of 12B. bifidumstrains. In four of them—B. bifidumLMG13195, DSM20456, DSM20239, and A8—the involvement of surface-exposed macromolecules in the aggregation phenomenon was determined. The aggregation ofB. bifidumA8 and DSM20456 was abolished after treatment with proteinase K, this effect being more pronounced for the strain A8. Furthermore, a mucin binding assay ofB. bifidumA8 surface proteins showed a high adhesive capability for its transaldolase (Tal). The localization of this enzyme on the surface ofB. bifidumA8 was unequivocally demonstrated by immunogold electron microscopy experiments. The gene encoding Tal fromB. bifidumA8 was expressed inLactococcus lactis, and the protein was purified to homogeneity. The pure protein was able to restore the autoaggregation phenotype of proteinase K-treatedB. bifidumA8 cells. A recombinantL. lactisstrain, engineered to secrete Tal, displayed a mucin- binding level more than three times higher than the strain not producing the transaldolase. These findings suggest that Tal, when exposed on the cell surface ofB. bifidum, could act as an important colonization factor favoring its establishment in the gut.

scholarly journals BopA Does Not Have a Major Role in the Adhesion of Bifidobacterium bifidum to Intestinal Epithelial Cells, Extracellular Matrix Proteins, and Mucus

2013 ◽  
Vol 79 (22) ◽  
pp. 6989-6997 ◽  
Author(s):  
Veera Kainulainen ◽  
Justus Reunanen ◽  
Kaisa Hiippala ◽  
Simone Guglielmetti ◽  
Satu Vesterlund ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
Cell Lines ◽  
Bifidobacterium Bifidum ◽  
Intestinal Cell ◽  
Content Type ◽  
Colonic Mucin ◽  
Epithelial Cell Lines ◽  
High Concentrations

ABSTRACTThe ability of bifidobacteria to adhere to the intestine of the human host is considered to be important for efficient colonization and achieving probiotic effects.Bifidobacterium bifidumstrains DSM20456 and MIMBb75 adhere well to the human intestinal cell lines Caco-2 and HT-29. The surface lipoprotein BopA was previously described to be involved in mediating adherence ofB. bifidumto epithelial cells, but thioacylated, purified BopA inhibited the adhesion ofB. bifidumto epithelial cells in competitive adhesion assays only at very high concentrations, indicating an unspecific effect. In this study, the role of BopA in the adhesion ofB. bifidumwas readdressed. The gene encoding BopA was cloned and expressed without its lipobox and hydrophobic signal peptide inEscherichia coli, and an antiserum against the recombinant BopA was produced. The antiserum was used to demonstrate the abundant localization of BopA on the cell surface ofB. bifidum. However, blocking ofB. bifidumBopA with specific antiserum did not reduce adhesion of bacteria to epithelial cell lines, arguing that BopA is not an adhesin. Also, adhesion ofB. bifidumto human colonic mucin and fibronectin was found to be BopA independent. The recombinant BopA bound only moderately to human epithelial cells and colonic mucus, and it failed to bind to fibronectin. Thus, our results contrast the earlier findings on the major role of BopA in adhesion, indicating that the strong adhesion ofB. bifidumto epithelial cell lines is BopA independent.

scholarly journals hfqPlays Important Roles in Virulence and Stress Adaptation in Cronobacter sakazakii ATCC 29544

2015 ◽  
Vol 83 (5) ◽  
pp. 2089-2098 ◽  
Author(s):  
Seongok Kim ◽  
Hyelyeon Hwang ◽  
Kwang-Pyo Kim ◽  
Hyunjin Yoon ◽  
Dong-Hyun Kang ◽  
...  
Keyword(s):  
Pathogenic Bacteria ◽  
Bacterial Species ◽  
Soft Agar ◽  
Host Cells ◽  
Neonatal Meningitis ◽  
Animal Cells ◽  
Content Type ◽  
Flagellar Biosynthesis

Cronobacterspp. are opportunistic pathogens that cause neonatal meningitis and sepsis with high mortality in neonates. Despite the peril associated withCronobacterinfection, the mechanisms of pathogenesis are still being unraveled. Hfq, which is known as an RNA chaperone, participates in the interaction with bacterial small RNAs (sRNAs) to regulate posttranscriptionally the expression of various genes. Recent studies have demonstrated that Hfq contributes to the pathogenesis of numerous species of bacteria, and its roles are varied between bacterial species. Here, we tried to elucidate the role of Hfq inC. sakazakiivirulence. In the absence ofhfq,C. sakazakiiwas highly attenuated in disseminationin vivo, showed defects in invasion (3-fold) into animal cells and survival (103-fold) within host cells, and exhibited low resistance to hydrogen peroxide (102-fold). Remarkably, the loss ofhfqled to hypermotility on soft agar, which is contrary to what has been observed in other pathogenic bacteria. The hyperflagellated bacteria were likely to be attributable to the increased transcription of genes associated with flagellar biosynthesis in a strain lackinghfq. Together, these data strongly suggest thathfqplays important roles in the virulence ofC. sakazakiiby participating in the regulation of multiple genes.

scholarly journals Effect of Helicobacter pylori on Polymorphonuclear Leukocyte Migration across Polarized T84 Epithelial Cell Monolayers: Role of Vacuolating Toxin VacA and cagPathogenicity Island

2000 ◽  
Vol 68 (9) ◽  
pp. 5225-5233 ◽  
Author(s):  
Véronique Hofman ◽  
Vittorio Ricci ◽  
Antoine Galmiche ◽  
Patrick Brest ◽  
Patrick Auberger ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Polymorphonuclear Leukocyte ◽  
Broth Culture ◽  
Intestinal Cell ◽  
T84 Cells ◽  
Vacuolating Cytotoxin ◽  
Intestinal Cell Line ◽  
H Pylori

ABSTRACT Helicobacter pylori infection can induce polymorphonuclear leukocyte (PMNL) infiltration of the gastric mucosa, which characterizes acute chronic gastritis. The mechanisms underlying this process are poorly documented. The lack of an in vitro model has considerably impaired the study of transepithelial migration of PMNL induced by H. pylori. In the present work, we used confluent polarized monolayers of the human intestinal cell line T84 grown on permeable filters to analyze the epithelial PMNL response induced by broth culture filtrates (BCFs) and bacterial suspensions from different strains of H. pylori. We have evaluated the role of the vacuolating cytotoxin VacA and of the cagpathogenicity island (PAI) of H. pylori in PMNL migration via their effects on T84 epithelial cells. We noted no difference in the rates of PMNL transepithelial migration after epithelial preincubation with bacterial suspensions or with BCFs of VacA-negative or VacA-positive H. pylori strains. In contrast, PMNL transepithelial migration was induced after incubation of the T84 cells with cag PAI-positive and cagE-positiveH. pylori strains. Finally, PMNL migration was correlated with a basolateral secretion of interleukin-8 by T84 cells, thus creating a subepithelial chemotactic gradient for PMNL. These data provide evidence that the vacuolating cytotoxin VacA is not involved in PMNL transepithelial migration and that the cag PAI, with a pivotal role for the cagE gene, provokes a transcellular signal across T84 monolayers, inducing a subepithelial PMNL response.

scholarly journals Role of Neuraminidase-Producing Bacteria in Exposing Cryptic Carbohydrate Receptors forStreptococcus gordoniiAdherence

2018 ◽  
Vol 86 (7) ◽  
pp. e00068-18 ◽  
Author(s):  
Alex Wong ◽  
Margaret A. Grau ◽  
Anirudh K. Singh ◽  
Shireen A. Woodiga ◽  
Samantha J. King
Keyword(s):  
Sialic Acid ◽  
Oral Cavity ◽  
Bacterial Species ◽  
Dependent Manner ◽  
Neuraminidase Treatment ◽  
Content Type ◽  
Oral Epithelial Cells ◽  
Oral Environment ◽  
Oral Epithelial

ABSTRACTStreptococcus gordoniiis an early colonizer of the oral cavity. Although a variety ofS. gordoniiadherence mechanisms have been described, current dogma is that the major receptor forS. gordoniiis sialic acid. However, as many bacterial species in the oral cavity produce neuraminidase that can cleave terminal sialic acid, it is unclear whetherS. gordoniirelies on sialic acid for adherence to oral surfaces or if this species has developed alternative binding strategies. Previous studies have examined adherence to immobilized glycoconjugates and identified binding to additional glycans, but no prior studies have defined the contribution of these different glycan structures in adherence to oral epithelial cells. We determined that the majority ofS. gordoniistrains tested did not rely on sialic acid for efficient adherence. In fact, adherence of some strains was significantly increased following neuraminidase treatment. Further investigation of representative strains that do not rely on sialic acid for adherence revealed binding not only to sialic acid via the serine-rich repeat protein GspB but also to β-1,4-linked galactose. Adherence to this carbohydrate occurs via an unknown adhesin distinct from those utilized byStreptococcus oralisandStreptococcus pneumoniae. Demonstrating the potential biological relevance of binding to this cryptic receptor, we established thatS. oralisincreasesS. gordoniiadherence in a neuraminidase-dependent manner. These data suggest thatS. gordoniihas evolved to simultaneously utilize both terminal and cryptic receptors in response to the production of neuraminidase by other species in the oral environment.

Protection against ethanol injury by prostaglandin in a human intestinal cell line: role of microtubules

1998 ◽  
Vol 274 (1) ◽  
pp. G111-G121 ◽  
Author(s):  
A. Banan ◽  
G. S. Smith ◽  
C. L. Rieckenberg ◽  
E. R. Kokoska ◽  
T. A. Miller
Keyword(s):  
Cell Line ◽  
Protective Action ◽  
Microtubule Assembly ◽  
Intestinal Cell ◽  
Major Protein ◽  
Cellular Viability ◽  
Intestinal Cell Line ◽  
Cytoskeletal Component ◽  
Human Intestinal Cell Line

Prostaglandins have been shown to protect the gastrointestinal (GI) epithelium from injury induced by various luminal insults independent of their known acid-inhibitory effects, a process termed “cytoprotection.” The mechanism of this protective action remains unknown. The present investigation determined the role of microtubules (a major cytoskeletal component) in GI injury induced by ethanol (EtOH) and its prevention by 16,16-dimethylprostaglandin E2(dmPGE2) using cells from a human colonic cell line known as Caco-2 cells. These cells were preincubated in Eagle’s minimum essential medium with and without dmPGE2 (2.6 μM) for 15 min and subsequently incubated in media containing 1, 2.5, 5, 7.5, and 10% EtOH. The effects on cell viability and tubulin (the major protein backbone of microtubules) were then determined. EtOH concentrations ≥2.5% extensively disrupted the microtubules as demonstrated by fragmentation, kinking, and perturbation of the microtubule organizer center. EtOH treatment also led to a significant decrease in the S2 (polymerized) fraction and an increase in the S1 (monomeric) pool of tubulin. Concomitant with these effects were marked decreases in cellular viability. DmPGE2pretreatment abolished the disruption of microtubules, significantly increased the S2 fraction of tubulin, and increased cellular viability in cultures exposed to EtOH. Furthermore, pretreatment with colchicine, an inhibitor of microtubule assembly, prevented the cytoprotective action of dmPGE2. Taxol, a microtubule stabilizing agent, mimicked the effects of dmPGE2 by also enhancing microtubule integrity and increasing cellular viability in cells exposed to EtOH. Our data indicate that organization and stabilization of microtubules may play an essential role in the mechanism of prostaglandin-induced protection.

scholarly journals Antimicrobial effect of probiotics on bacterial species from dental plaque

2016 ◽  
Vol 10 (03) ◽  
pp. 214-221 ◽  
Author(s):  
Csilla Zambori ◽  
Attila Alexandru Morvay ◽  
Claudia Sala ◽  
Monica Licker ◽  
Camelia Gurban ◽  
...  
Keyword(s):  
Dental Plaque ◽  
Bacterial Species ◽  
Antimicrobial Effect ◽  
Bactericidal Effect ◽  
Bifidobacterium Bifidum ◽  
Bacteriostatic Effect ◽  
Zoonotic Infections ◽  
Probiotic Lactobacillus

Introduction: The antimicrobial role of probiotic Lactobacillus casei subspecies casei DG (L. casei DG) and of the mix culture of probiotic Lactobacillus acidophilus LA-5 and Bifidobacterium BB-12 was tested on species of Staphylococcus, Streptococcus, Pasteurella, and Neisseria genera from supragingival sites from dogs with dental disease of different breed, age, sex, weight, and diet. The research was conducted on these four genera because of their importance in zoonotic infections after dog bites. Methodology: Species from Staphylococcus, Streptococcus, Pasteurella, and Neisseria genera were isolated and identified. To test the antimicrobial efficacy of L. casei DG and the mixed culture of probiotic L. acidophilus LA-5 and Bifidobacterium bifidum BB-12 on the pathogenic species, the agar overlay method was used. Results: L. casei DG had a bactericidal effect on all analyzed species isolated from Staphylococcus, Streptococcus, Pasteurella, and Neisseria genera after 24 hours of incubation. The mixed probiotic culture made up of L. acidophilus LA-5 and Bifidobacterium BB-12 species had no bactericidal effect on the species of Staphylococcus and Streptococcus genera, which were resistant. However, it had a bacteriostatic effect on several species of Pasteurella and Neisseria genera. Conclusions: This work highlights the antimicrobial potential of probiotics in vitro, demonstrating that the probiotic L. casei DG has a bactericidal effect on all analyzed species isolated from dental plaque and that the mix culture of probiotic L. acidophilus LA-5 and Bifidobacterium BB-12 has only a bacteriostatic effect.

scholarly journals Structural basis for the role of serine-rich repeat proteins from Lactobacillus reuteri in gut microbe–host interactions

2018 ◽  
Vol 115 (12) ◽  
pp. E2706-E2715 ◽  
Author(s):  
Saannya Sequeira ◽  
Devon Kavanaugh ◽  
Donald A. MacKenzie ◽  
Tanja Šuligoj ◽  
Samuel Walpole ◽  
...  
Keyword(s):  
Lactobacillus Reuteri ◽  
Structural Information ◽  
Bacterial Species ◽  
Surface Proteins ◽  
Structural Basis ◽  
Gene Encoding ◽  
Rhamnogalacturonan I ◽  
Host Microbe Interactions ◽  
Dynamics Simulations

Lactobacillus reuteri, a Gram-positive bacterial species inhabiting the gastrointestinal tract of vertebrates, displays remarkable host adaptation. Previous mutational analyses of rodent strain L. reuteri 100-23C identified a gene encoding a predicted surface-exposed serine-rich repeat protein (SRRP100-23) that was vital for L. reuteri biofilm formation in mice. SRRPs have emerged as an important group of surface proteins on many pathogens, but no structural information is available in commensal bacteria. Here we report the 2.00-Å and 1.92-Å crystal structures of the binding regions (BRs) of SRRP100-23 and SRRP53608 from L. reuteri ATCC 53608, revealing a unique β-solenoid fold in this important adhesin family. SRRP53608-BR bound to host epithelial cells and DNA at neutral pH and recognized polygalacturonic acid (PGA), rhamnogalacturonan I, or chondroitin sulfate A at acidic pH. Mutagenesis confirmed the role of the BR putative binding site in the interaction of SRRP53608-BR with PGA. Long molecular dynamics simulations showed that SRRP53608-BR undergoes a pH-dependent conformational change. Together, these findings provide mechanistic insights into the role of SRRPs in host–microbe interactions and open avenues of research into the use of biofilm-forming probiotics against clinically important pathogens.

scholarly journals Functional Characterization of the Pseudomonas aeruginosa Ribosome Hibernation-Promoting Factor

2020 ◽  
Vol 202 (19) ◽  
Author(s):  
Michael J. Franklin ◽  
Elizabeth Sandvik ◽  
Sila Yanardag ◽  
Kerry S. Williamson
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Bacterial Species ◽  
Point Mutations ◽  
Single Amino Acid ◽  
Physiological Changes ◽  
Alpha Helices ◽  
Content Type ◽  
Charged Residues ◽  
Bacterial Starvation

ABSTRACT Hibernation-promoting factor (HPF) is a ribosomal accessory protein that inactivates ribosomes during bacterial starvation. In Pseudomonas aeruginosa, HPF protects ribosome integrity while the cells are dormant. The sequence of HPF has diverged among bacteria but contains conserved charged amino acids in its two alpha helices that interact with the rRNA. Here, we characterized the function of HPF in P. aeruginosa by performing mutagenesis of the conserved residues and then assaying mutant HPF alleles for their ability to protect ribosome integrity of starved P. aeruginosa cells. The results show that HPF functionally tolerates point mutations in charged residues and in the conserved Y71 residue as well as a C-terminal truncation. Double and triple mutations of charged residues in helix 1 in combination with a Y71F substitution reduce HPF activity. Screening for single point mutations that caused impaired HPF activity identified additional substitutions in the two HPF alpha helices. However, alanine substitutions in equivalent positions restored HPF activity, indicating that HPF is tolerant to mutations that do not disrupt the protein structure. Surprisingly, heterologous HPFs from Gram-positive bacteria that have long C-terminal domains functionally complement the P. aeruginosa Δhpf mutant, suggesting that HPF may play a similar role in ribosome protection in other bacterial species. Collectively, the results show that HPF has diverged among bacteria and is tolerant to most single amino acid substitutions. The Y71 residue in combination with helix 1 is important for the functional role of HPF in ribosome protection during bacterial starvation and resuscitation of the bacteria from dormancy. IMPORTANCE In most environments, bacteria experience conditions where nutrients may be readily abundant or where nutrients are limited. Under nutrient limitation conditions, even non-spore-forming bacteria may enter a dormant state. Dormancy is accompanied by a variety of cellular physiological changes that are required for the cells to remain viable during dormancy and to resuscitate when nutrients become available. Among the physiological changes that occur in dormant bacteria is the inactivation and preservation of ribosomes by the dormancy protein, hibernation-promoting factor (HPF). In this study, we characterized the activity of HPF of Pseudomonas aeruginosa, an opportunistic pathogen that causes persistent infections, and analyzed the role of HPF in ribosome protection and bacterial survival during dormancy.

scholarly journals The Identification and Functional Characterization of WxL Proteins from Enterococcus faecium Reveal Surface Proteins Involved in Extracellular Matrix Interactions

2014 ◽  
Vol 197 (5) ◽  
pp. 882-892 ◽  
Author(s):  
Jessica R. Galloway-Peña ◽  
Xiaowen Liang ◽  
Kavindra V. Singh ◽  
Puja Yadav ◽  
Chungyu Chang ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Cell Surface ◽  
Bile Salt ◽  
Enterococcus Faecium ◽  
Bacterial Species ◽  
Functional Characterization ◽  
Surface Proteins ◽  
Type I ◽  
Gram Positive ◽  
Content Type

The WxL domain recently has been identified as a novel cell wall binding domain found in numerous predicted proteins within multiple Gram-positive bacterial species. However, little is known about the function of proteins containing this novel domain. Here, we identify and characterize 6Enterococcus faeciumproteins containing the WxL domain which, by reverse transcription-PCR (RT-PCR) and genomic analyses, are located in three similarly organized operons, deemed WxL loci A, B, and C. Western blotting, electron microscopy, and enzyme-linked immunosorbent assays (ELISAs) determined that genes of WxL loci A and C encode antigenic, cell surface proteins exposed at higher levels in clinical isolates than in commensal isolates. Secondary structural analyses of locus A recombinant WxL domain-containing proteins found they are rich in β-sheet structure and disordered segments. Using Biacore analyses, we discovered that recombinant WxL proteins from locus A bind human extracellular matrix proteins, specifically type I collagen and fibronectin. Proteins encoded by locus A also were found to bind to each other, suggesting a novel cell surface complex. Furthermore, bile salt survival assays and animal models using a mutant from which all three WxL loci were deleted revealed the involvement of WxL operons in bile salt stress and endocarditis pathogenesis. In summary, these studies extend our understanding of proteins containing the WxL domain and their potential impact on colonization and virulence inE. faeciumand possibly other Gram-positive bacterial species.

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