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PubMed2005

The fucose-binding lectin from Ralstonia solanacearum. A new type of beta-propeller architecture formed by oligomerization and interacting with fucoside, fucosyllactose, and plant xyloglucan.

Plant pathogens, like animal ones, use protein-carbohydrate interactions in their strategy for host recognition, attachment, and invasion. The bacterium Ralstonia solanacearum, which is distributed worldwide and causes lethal wilt in many agricultural crops, was shown to produce a potent L-fucose-binding lectin, R. solanacearum lectin, a small protein of 90 amino acids with a tandem repeat in its amino acid sequence. In the present study, surface plasmon resonance experiments conducted on a series of oligosaccharides show a preference for binding to alphaFuc1-2Gal and alphaFuc1-6Gal epitopes. Titration microcalorimetry demonstrates the presence of two binding sites per monomer and an unusually high affinity of the lectin for alphaFuc1-2Gal-containing oligosaccharides (KD = 2.5 x 10(-7) M for 2-fucosyllactose). R. solanacearum lectin has been crystallized with a methyl derivative of fucose and with the highest affinity ligand, 2-fucosyllactose. X-ray crystal structures, the one with alpha-methyl-fucoside being at ultrahigh resolution, reveal that each monomer consists of two small four-stranded anti-parallel beta-sheets. Trimerization through a 3-fold or pseudo-3-fold axis generates a six-bladed beta-propeller architecture, very similar to that previously described for the fungal lectin of Aleuria aurantia. This is the first report of a beta-propeller formed by oligomerization and not by sequential domains. Each monomer presents two fucose binding sites, resulting in six symmetrically arranged sugar binding sites for the beta-propeller. Crystals were also obtained for a mutated lectin complexed with a fragment of xyloglucan, a fucosylated polysaccharide from the primary cell wall of plants, which may be the biological target of the lectin.

Amino Acid Sequence↗

PubMed1994

Bordetella pertussis filamentous hemagglutinin interacts with a leukocyte signal transduction complex and stimulates bacterial adherence to monocyte CR3 (CD11b/CD18).

Bordetella pertussis, the causative agent of whooping cough, adheres to human monocytes/macrophages by means of a bacterial surface-associated protein, filamentous hemagglutinin (FHA) and the leukocyte integrin, complement receptor 3 (CR3, alpha M beta 2, CD11b/CD18). We show that an FHA Arg-Gly-Asp site induces enhanced B. pertussis binding to monocytes, and that this enhancement is blocked by antibodies directed against CR3. Enhancement requires a monocyte signal transduction complex, composed of leukocyte response integrin (alpha? beta 3) and integrin-associated protein (CD47). This complex is known to upregulate CR3 binding activity. Thus, a bacterial pathogen enhances its own attachment to host cells by coopting a host cell signaling pathway.

Adhesins, Bacterial↗

PubMed1995

Adherence of pilus- Opa+ gonococci to epithelial cells in vitro involves heparan sulfate.

Neisseria gonorrhoeae attaches to host epithelial cells via pili and opacity-associated (Opa) outer membrane proteins. Pilus- gonococci (Gc) of strain MS11 adhere to both human and nonhuman cells, but only when particular Opa proteins are expressed; OpaA+ variants adhere best, OpaC+ variants are next best, and the seven other Opa+ variants adhere poorly or not at all. The adherence of OpaA+ Gc to Chinese hamster ovary (CHO) cells is inhibited by heparin or heparan sulfate (HS), but not by chondroitin sulfate. OpaA+ Gc do not adhere to CHO cells devoid of HS proteoglycans; low concentrations of heparin restore OpaA+ Gc adherence to these HS-deficient CHO cells and high concentrations inhibit it. 3H-heparin binding to whole Gc parallels their adherence abilities (OpaA+ > OpaC+ > OpaH+ >> Opas B, D, E, F, G, I = Opa- = 0). Opa proteins separated by SDS-PAGE also bind 3H-heparin. These data suggest that adherence of pilus-, Opa+ Gc involves HS-proteoglycan of eukaryotic cells.

Animals↗

PubMed2002

Quantitative ecology of ticks as a basis for transmission models of tick-borne pathogens.

Tick population ecology is the basis of spatiotemporal variation in the risk of infection by tick-borne pathogens. The continental distribution of tick-borne encephalitis (TBE) in Europe is statistically associated with a specific pattern of the seasonal dynamics of Ixodes ricinus, and a particular characteristic of the seasonal land surface temperature profile. A tick population model would allow us to move from this statistical analysis to a biological, process-based model for TBE virus and the many other pathogens vectored by Ixodes spp. ticks in Europe and the United States. Long-term field data on I. ricinus are analyzed to provide empirical estimates of rates of the major demographic processes, development, questing, attachment to hosts, and mortality.

Animals↗

PubMedDate not supplied

In vivo response of polylactic acid-alginate scaffolds and bone marrow-derived cells for cartilage tissue engineering.

Successful application of tissue-engineering techniques to damaged biological structures is determined by functional performance in vivo. This study evaluated the in vivo response of a tissue-engineered construct composed of a polylactic acid-alginate amalgam seeded with bone marrow-derived mesenchymal stem cells and stimulated in vitro with transforming growth factor beta for cartilage tissue engineering. Constructs were placed in cylindrical osteochondral defects in the canine femoral condyle and examined 6 weeks postoperatively by gross, histological, immunohistochemical, and biomechanical analyses. In the course of 6 weeks in vivo, the defects filled with a cartilaginous tissue regardless of whether cell-seeded (experimental) or cell-free (control) constructs were implanted; however, the quality of the tissue differed between the experimental and control defects. Cell-seeded experimental defects showed more cartilage-like matrix quality, cell distribution, and proteoglycan staining. Biomechanically, experimental and control specimens exhibited similar behavior; however, both tissues were still immature compared with normal cartilage. The evidence accumulated in this study showed a modest acceleration of the in vivo healing of cell-seeded constructs but also demonstrated a reparative response of cell-free constructs. This finding suggests that the constructs prepared from the PLA-alginate amalgam may serve as a means for host cell attachment.

Alginates↗

PubMed2006

STD NMR spectroscopy and molecular modeling investigation of the binding of N-acetylneuraminic acid derivatives to rhesus rotavirus VP8* core.

The VP8* subunit of rotavirus spike protein VP4 contains a sialic acid (Sia)-binding domain important for host cell attachment and infection. In this study, the binding epitope of the N-acetylneuraminic acid (Neu5Ac) derivatives has been characterized by saturation transfer difference (STD) nuclear magnetic resonance (NMR) spectroscopy. From this STD NMR data, it is proposed that the VP8* core recognizes an identical binding epitope in both methyl alpha-D-N-acetylneuraminide (Neu5Acalpha2Me) and the disaccharide methyl S-(alpha-D-N-acetylneuraminosyl)-(2-->6)-6-thio-beta-D-galactopyranoside (Neu5Ac-alpha(2,6)-S-Galbeta1Me). In the VP8*-disaccharide complex, the Neu5Ac moiety contributes to the majority of interaction with the protein, whereas the galactose moiety is solvent-exposed. Molecular dynamics calculations of the VP8*-disaccharide complex indicated that the galactose moiety is unable to adopt a conformation that is in close proximity to the protein surface. STD NMR experiments with methyl 9-O-acetyl-alpha-D-N-acetylneuraminide (Neu5,9Ac(2)alpha2Me) in complex with rhesus rotavirus (RRV) VP8* revealed that both the N-acetamide and 9-O-acetate moieties are in close proximity to the Sia-binding domain, with the N-acetamide's methyl group being saturated to a larger extent, indicating a closer association with the protein. RRV VP8* does not appear to significantly recognize the unsaturated Neu5Ac derivative [2-deoxy-2,3-didehydro-D-N-acetylneuraminic acid (Neu5Ac2en)]. Molecular modeling of the protein-Neu5Ac2en complex indicates that key interactions between the protein and the unsaturated Neu5Ac derivative when compared with Neu5Acalpha2Me would not be sustained. Neu5Acalpha2Me, Neu5Ac-alpha(2,6)-S-Galbeta1Me, Neu5,9Ac(2)alpha2Me, and Neu5Ac2en inhibited rotavirus infection of MA104 cells by 61%, 35%, 30%, and 0%, respectively, at 10 mM concentration. NMR spectroscopic, molecular modeling, and infectivity inhibition results are in excellent agreement and provide valuable information for the design of inhibitors of rotavirus infection.

Animals↗

PubMed1991

Experimental acquisition of the Lyme disease spirochete, Borrelia burgdorferi, by larval Ixodes dammini (Acari: Ixodidae) during partial blood meals.

The duration of feeding by larval Ixodes dammini was directly correlated with the proportion of larvae acquiring infection with the Lyme disease spirochete. The infection rate of ticks attached to hosts for 8 h (1.4%) was much lower than ticks feeding to repletion (87.5%). Infection was also detected in partially fed larvae dropping off dead animals. These partially fed larvae successfully reattached to rodents, fed to repletion, and molted to nymphs. Partially fed larvae did not transmit Borrelia burgdorferi when refeeding as larvae, but the resultant nymphs were capable of transmitting spirochetes. Infected host-seeking larvae collected in nature cannot be judged a priori to have acquired B. burgdorferi transovarially.

Animals↗

PubMed1983

Neutrophil function and HLA-B27.

The pathogenesis of HLA-B27 linked diseases, including reactive enteroarthritides, is not known. Differences in immune response, cross-reaction between microbial antigens and host cell structures, attachment of microbial components on the host cell surfaces with subsequent autoimmune reaction, and inflammatory responsiveness of the host may be involved. In this contribution we present evidence to show that HLA-B27 positive subjects may be prone to an exaggerated inflammatory response, as compared with HLA-B27 negative subjects.

Arthritis, Infectious↗

PubMed1999

Isolation of respiratory bacterial pathogens from the throats of healthy infants fed by different methods.

Most bacterial infections are caused by organisms that have already colonized the host. Bacterial attachment to pharyngeal cells and proliferation may be necessary to infect the lower respiratory tract or middle ear. We investigated the incidence of pathogenic bacteria isolated from the throat of healthy infants with different feeding methods. The protecting role of breastmilk is also discussed. The incidence of respiratory bacterial pathogens isolated from the oropharynx of 113 normal infants with different feeding methods was investigated. Group A beta haemolytic Streptococcus, Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis were selected as respiratory bacterial pathogens. No respiratory bacterial pathogens were detected in breastfed and mixed-fed infants. Haemophilus influenzae and Moraxella catarrhalis were isolated from the oropharynx of formula-fed infants. The incidence of respiratory bacterial pathogens did differ among infants with different feeding methods. These results suggest that breastmilk may inhibit the colonization by respiratory bacterial pathogens of the throat of infants, by enhancing mucosal immunity against respiratory tract infection.

Bacteria↗

PubMed1996

Lyme disease.

In the United States, there was an increase in the number of reported cases of Lyme disease in 1994 compared with previous years. The majority of cases occurred in the Northeast and Midwest, which are areas with established foci of Borrelia burgdorferi. Advances in the understanding of the epidemiology and ecology of ixodes ticks, the established vector for transmission of Lyme disease, provided strategies for reducing tick populations and the risk of acquiring disease. Dissemination of B. burgdorferi from the site of a tick bite likely occurs, in part, via attachment to host plasmin and subsequent degradation of glycoproteins. Analysis of T cell immune responses in mice contributed to our understanding of differences in disease susceptibility. Vaccination of patients with previous Lyme disease has been found to be safe, and clinical trials of vaccine efficacy are currently under way.

Humans↗

PubMed2002

Mining the Plasmodium genome database to define organellar function: what does the apicoplast do?

Apicomplexan species constitute a diverse group of parasitic protozoa, which are responsible for a wide range of diseases in many organisms. Despite differences in the diseases they cause, these parasites share an underlying biology, from the genetic controls used to differentiate through the complex parasite life cycle, to the basic biochemical pathways employed for intracellular survival, to the distinctive cell biology necessary for host cell attachment and invasion. Different parasites lend themselves to the study of different aspects of parasite biology: Eimeria for biochemical studies, Toxoplasma for molecular genetic and cell biological investigation, etc. The Plasmodium falciparum Genome Project contributes the first large-scale genomic sequence for an apicomplexan parasite. The Plasmodium Genome Database (http://PlasmoDB.org) has been designed to permit individual investigators to ask their own questions, even prior to formal release of the reference P. falciparum genome sequence. As a case in point, PlasmoDB has been exploited to identify metabolic pathways associated with the apicomplexan plastid, or 'apicoplast' - an essential organelle derived by secondary endosymbiosis of an alga, and retention of the algal plastid.

Animals↗

PubMed2002

Group A streptococcal RofA-type global regulators exhibit a strain-specific genomic presence and regulation pattern.

RofA-like protein (RALP) type regulators have been shown to exist in different forms in group A streptococci (GAS) and to regulate the expression of important bacterial adhesins. This study shows that the vast majority of strains from different GAS M serotypes carried a rofA virulence regulator gene in their genome and that this gene could be detected in combination with other RALP genes and RALP-dependent adhesin genes in a strain-specific manner. The gene encoding the Nra regulator was predominantly found in opacity factor (OF)-negative serotypes. When analysing a rofA mutant in a serotype M2 strain, the strain specificity was also found in the positive and negative regulatory functions of RALP genes as well as in the type and number of virulence genes and functions controlled by the RALP genes. Of 17 virulence-associated genes tested, only one, the putative streptolysin S gene, was observed to be derepressed in RALP mutants of three different GAS serotype strains. This strain-specific variability of RALP regulon sizes is associated with different patterns of host cell attachment and internalization. In addition, RofA2 was shown to control expression of the ribosomal protein gene rpsL. As a consequence, it was demonstrated for the first time in streptococci that aminoglycoside resistance mediated by rpsL expression is apparently controlled by a virulence gene regulator.

Bacterial Adhesion↗

PubMed2004

Significant differences in type IV pilin allele distribution among Pseudomonas aeruginosa isolates from cystic fibrosis (CF) versus non-CF patients.

Type IV pili (TFP) are important colonization factors of the opportunistic pathogen Pseudomonas aeruginosa, involved in biofilm formation and attachment to host cells. This study undertook a comprehensive analysis of TFP alleles in more than 290 environmental, clinical, rectal and cystic fibrosis (CF) isolates of P. aeruginosa. Based on the results, a new system of nomenclature is proposed, in which P. aeruginosa TFP are divided into five distinct phylogenetic groups. Each pilin allele is stringently associated with characteristic, distinct accessory genes that allow the identification of the allele by specific PCR. The invariant association of the pilin and accessory genes implies horizontal transfer of the entire locus. Analysis of pilin allele distribution among isolates from various sources revealed a striking bias in the prevalence of isolates with group I pilin genes from CF compared with non-CF human sources (P<0.0001), suggesting this particular pilin type, which can be post-translationally modified by glycosylation via the action of TfpO (PilO), may confer a colonization or persistence advantage in the CF host. This allele was also predominant in paediatric CF isolates (29 of 43; 67.4 %), showing that this bias is apparent early in colonization. Group I pilins were also the most common type found in environmental isolates tested. To the authors' knowledge, this is the first example of a P. aeruginosa virulence factor allele that is strongly associated with CF isolates.

Adolescent↗

PubMed1992

Adhesion Pad Formation and the Involvement of Cutinase and Esterases in the Attachment of Uredospores to the Host Cuticle.

We have investigated the basis of adhesion of uredospores of the obligately parasitic rust fungus Uromyces viciae-fabae to leaves of its broad bean host. Upon contact with an aqueous environment, spores form a structure that we have termed an adhesion pad. The adhesion pad is formed by both living and autoclaved spores, but only adhesion pads formed by living spores adhered to the cuticle of leaves of the host plant. Treatment of living spores with the serine-esterase inhibitor diisopropyl fluorophosphate prevented the adhesion of the pad to the leaf surface, suggesting a functional role for esterase or cutinase in the process of adhesion. A cutinase and two nonspecific serine-esterases were found to be localized on the surface of spores. These enzymes were released rapidly from the spore surface upon contact with an aqueous environment. The addition of the cutinase and the nonspecific esterases to autoclaved spores restored their ability to adhere to the host cuticle. Thus, whereas pad formation appears to be a passive response to the aqueous environment, the actual adhesion of pads to the host cuticle appears to depend on the cutinase and esterases associated with the spore surface. These results suggest a new role for cutinases and serine-esterases in the fungal infection process.

Journal Article↗

PubMed2005

Crystallization and preliminary X-ray diffraction analysis of the sialic acid-binding domain (VP8*) of porcine rotavirus strain CRW-8.

Rotavirus recognition and attachment to host cells involves interaction with the spike protein VP4 that projects outwards from the surface of the virus particle. An integral component of these spikes is the VP8* domain, which is implicated in the direct recognition and binding of sialic acid-containing cell-surface carbohydrates and facilitates subsequent invasion by the virus. The expression, purification, crystallization and preliminary X-ray diffraction analysis of VP8* from porcine CRW-8 rotavirus is reported. Diffraction data have been collected to 2.3 A resolution, enabling the determination of the VP8* structure by molecular replacement.

Animals↗

PubMed2005

Crystallization of the C-terminal globular domain of avian reovirus fibre.

Avian reovirus fibre, a homotrimer of the sigmaC protein, is responsible for primary host-cell attachment. Using the protease trypsin, a C-terminal sigmaC fragment containing amino acids 156-326 has been generated which was subsequently purified and crystallized. Two different crystal forms were obtained, one grown in the absence of divalent cations and belonging to space group P6(3)22 (unit-cell parameters a = 75.6, c = 243.1 A) and one grown in the presence of either zinc or cadmium sulfate and belonging to space group P321 (unit-cell parameters a = 74.7, c = 74.5 A and a = 73.1, c = 69.9 A for the Zn(II)- and Cd(II)-grown crystals, respectively). The first crystal form diffracted synchrotron radiation to 3.0 A resolution and the second form to 2.2-2.3 A. Its closest related structure, the C-terminal fragment of mammalian reovirus fibre, has only 18% sequence identity and molecular-replacement attempts were unsuccessful. Therefore, a search is under way for suitable heavy-atom derivatives and attempts are being made to grow protein crystals containing selenomethionine instead of methionine.

Cadmium Compounds↗

PubMed2006

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

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 A resolution for both native and derivatized crystals and showed the symmetry of space group P6(2)22, with unit-cell parameters a = b = 142.9, c = 231.9 A. 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.

Bacterial Adhesion↗

PubMed1983

Penetration of Rickettsia tsutsugamushi into cultured mouse fibroblasts (L cells): an electron microscopic observation.

The mechanism of penetration of purified Rickettsia tsutsugamushi (Gilliam strain) into cultured mouse fibroblasts (L cells) was examined by electron microscopy. After 10-40 min of infection, rickettsiae in the process of being phagocytized were often seen on the cell surface. These were restricted to the rickettsiae which seemed to be intact in morphology, while heavy plasmolyzed ones were never phagocytized. Additionally, rickettsiae were taken up individually into a phagosome, and phagocytosis of several rickettsiae together was rarely observed, except in the case of heat-inactivated microorganisms. In the cells, phagosomes whose membranes enclosed rickettsiae either tightly or loosely were seen. Rickettsiae in the loose phagosomes often showed signs of plasmolysis and were rarely released into the cell cytoplasm. Partial disintegration of phagosomal membranes and the escape of rickettsiae from the phagosomes were seen only in tight phagosomes. Large phagosomes containing a clump of several rickettsiae were observed occasionally, in which case the microorganisms were deformed and seemed to be denatured. From the above observations and the frequency of appearance of these different penetration stages in the specimens 10, 20, and 40 min after infection, it was concluded that the rickettsiae enter initially into a tight phagosome by phagocytosis and are then released into the cell cytoplasm by disruption of the phagosomal membrane. No other mechanisms of penetration were found. On the other hand, rickettsiae inactivated by trypsin did not attach to host cells. Inactivation by heat or UV irradiation resulted in reduction of phagocytosis, and rickettsiae treated with rifamycin could penetrate into the host cell cytoplasm to the same extent as in the case of infection with intact rickettsiae.

Animals↗