Competition Between Ruminal Cellulolytic Bacteria for Adhesion to Cellulose

Competition for adhesion to cellulose among the three main ruminal cellulolytic bacterial species was studied using differential radiolabeling (¹⁴C/³H) of cells. When added simultaneously to cellulose, Ruminococcus flavefaciens FD1 and Fibrobacter succinogenes S85 showed some competition; however, both species were surpassed competitively by Ruminococcus albus 20. When R. flavefaciens FD1 and F. succinogenes S85 were already adherent, R. albus 20 adhesion occurred without inhibition but involved R. flavefaciens FD1 detachment. Received: 28 October 1996 / Accepted: 28 January 1997     

蕲蛇酶（Acutobin）对家兔及大鼠血小板超微结构的影响

目的 应用电镜研究蕲蛇酶对血小板超微结构的影响以探讨用药后血小板减少的原因。方法 （１）家停（Ｎ＝６）静脉注射蕲蛇酶前及注射蕲蛇酶（１ＡＵ／ｋｇ及３ＡＵ／ｋｇ）后４ｈ心脏采血,制备血以标本及超薄切片,同时摘取脾及肺按常规制作超汪切片,进行电镜观察;（２）大鼠（Ｎ＝４）腹腔注射蕲蛇酶３ＡＵ／ｋｇ后分别于２、８、１６及２４ｈ摘取脾、肺脏制作标本进行是观察。结果 蕲蛇酶１ＡＵ／ｋｇ静脉注射组的家兔,血小     

Effect of Milk Proteins on Adhesion of Bacteria to Stainless Steel Surfaces

Stainless steel coupons were treated with skim milk and subsequently challenged with individual bacterial suspensions of Staphylococcus aureus, Pseudomonas fragi, Escherichia coli, Listeria monocytogenes, and Serratia marcescens. The numbers of attached bacteria were determined by direct epifluorescence microscopy and compared with the attachment levels on clean stainless steel with two different surface finishes. Skim milk was found to reduce adhesion of S. aureus, L. monocytogenes, and S. marcescens. P. fragi and E. coli attached in very small numbers to the clear surfaces, making the effect of any adsorbed protein layer difficult to assess. Individual milk proteins α-casein, β-casein, κ-casein, and α-lactalbumin were also found to reduce the adhesion of S. aureus and L. monocytogenes. The adhesion of bacteria to samples treated with milk dilutions up to 0.001% was investigated. X-ray photoelectron spectroscopy was used to determine the proportion of nitrogen in the adsorbed films. Attached bacterial numbers were inversely related to the relative atomic percentage of nitrogen on the surface. A comparison of two types of stainless steel surface, a 2B and a no. 8 mirror finish, indicated that the difference in these levels of surface roughness did not greatly affect bacterial attachment, and reduction in adhesion to a milk-treated surface was still observed. Cross-linking of adsorbed proteins partially reversed the inhibition of bacterial attachment, indicating that protein chain mobility and steric exclusion may be important in this phenomenon.     

Specific Adhesion of Bacteria to Heterocysts of Anabaena spp. and Its Ecological Significance

Two bacterial isolates, Pseudomonas sp. SL10 and Zoogloea sp. SL20, attach to heterocysts of Anabaena spp. with a high degree of selectivity, and this attachment can be expressed quantitatively in terms of adsorption isotherms. Adhesion of Pseudomonas sp. SL10 was restricted to a monolayer and exhibited a type I (Langmuir) isotherm, whereas adhesion of Zoogloea sp. SL20 involved multilayer attachment and exhibited a type II isotherm. The degree of adhesion by the bacteria to heterocysts of different Anabaena species may reflect the distribution and abundance of binding sites on the surface of different heterocysts. Both Pseudomonas sp. SL10 and Zoogloea sp SL20 promoted higher rates of acetylene reduction by Anabaena spp. under oxygenated culture conditions when compared with a cyanobacterial control. At ambient oxygen levels, however, only Zoogloea sp. SL20 stimulated acetylene reduction by Anabaena spp.     

Hydrophobicity, Adhesion, and Surface-Exposed Proteins of Gliding Bacteria

The cell surface hydrophobicities of a variety of aquatic and terrestrial gliding bacteria were measured by an assay of bacterial adherence to hydrocarbons (BATH), hydrophobic interaction chromatography, and the salt aggregation test. The bacteria demonstrated a broad range of hydrophobicities. Results among the three hydrophobicity assays performed on very hydrophilic strains were quite consistent. Bacterial adhesion to glass did not correlate with any particular measure of surface hydrophobicity. Several adhesion-defective mutants of Cytophaga sp. strain U67 were found to be more hydrophilic than the wild type, particularly by the BATH assay and hydrophobic interaction chromatography. The very limited adhesion of these mutants correlated well with hydrophilicity as determined by the BATH assay. The hydrophobicities of several adhesion-competent revertants ranged between those of the wild type and the mutants. As measured by the BATH assay, starvation increased hydrophobicity of both the wild type and an adhesion-defective mutant. During filament fragmentation of Flexibacter sp. strain FS-1, marked changes in hydrophobicity and adhesion were accompanied by changes in the arrays of surface-exposed proteins as detected by an immobilized radioiodination procedure.     

In Vitro Adhesion of N2-Fixing Enteric Bacteria to Roots of Grasses and Cereals

Nitrogen-fixing Klebsiella and Enterobacter strains isolated from several plants were assayed for fimbriae and for adhesion to plant roots in vitro. All eight Klebsiella strains formed type 3 fimbriae, and five strains also formed type 1 fimbriae; all 21 Enterobacter strains had type 1 fimbriae. Three strains of Klebsiella carrying either type 1, type 3, or no fimbriae were used as model organisms in developing an in vitro adhesion test. Adhesion was assayed with bacterial cells labeled with [³H]leucine. Fifteen N₂-fixing strains and the three model strains were compared for adhesion to the roots of seven grasses and five cereals. Type 3-fimbriated Klebsiella strains adhered better than the other strains, and type 3 fimbriae appeared to be major adhesins for the Klebsiella strains. Although variations between plants were observed, no host specificity for bacterial adhesion was found.     

Adhesion of Dissimilatory Fe(III)-Reducing Bacteria to Fe(III) Minerals

The metabolism of dissimilatory iron-reducing bacteria (DIRB) may provide a means of remediating contaminated subsurface soils. The factors controlling the rate and extent of bacterial F(III) mineral reduction are poorly understood. Recent research suggests that molecular-scale interactions between DIRB cells and Fe(III) mineral particles play an important role in this process. One of these interactions, cell adhesion to Fe(III) mineral particles, appears to be a complex process that is, at least in part, mediated by a variety of surface proteins. This study examined the hypothesis that the flagellum serves as an adhesin to different Fe(III) minerals that range in their surface area and degree of crystallinity. Deflagellated cells of the DIRB Shewanella algae BrY showed a reduced ability to adhere to hydrous ferric oxide (HFO) relative to flagellated cells. Flagellated cells were also more hydrophobic than deflagellated cells. This was significant because hydrophobic interactions have been previously shown to dominate S. algae cell adhesion to Fe(III) minerals. Pre-incubating HFO, goethite, or hematite with purified flagella inhibited the adhesion of S. algae BrY cells to these minerals. Transposon mutagenesis was used to generate a flagellum-deficient mutant designated S. algae strain NF. There was a significant difference in the rate and extent of S. algae NF adhesion to HFO, goethite, and hematite relative to that of S. algae BrY. Amiloride, a specific inhibitor of Na + -driven flagellar motors, inhibited S. algae BrY motility but did not affect the adhesion of S. algae BrY to HFO. S.algae NF reduced HFO at the same rate as S. algae BrY. Collectively, the results of this study support the hypothesis that the flagellum of S. algae functions as a specific Fe(III) mineral adhesin. However, these results suggest that flagellum-mediated adhesion is not requisite for Fe(III) mineral reduction.     

激光扫描共聚焦显微镜术检测单个血小板钙波动方法的探讨

目的 探讨血小板贴壁的简易方法,同时,观察单个血小板激活前后细胞内钙波动及形态变化的规律。方法 采用多种粘附剂固定血小板,利用钙荧光探针（Fluo-3/AM)（终浓度10－20μmol)进行染色,在激光扫描共聚焦显同镜检测下加入二磷酸腺苷,观察和分析血小板内Ca^2 浓度及形态变化。结果 多聚赖氨酸促进血小板贴壁固定的效果最佳;单个血小板活后细胞内Ca^2 浓度为激活前的128％,形态圆形或椭形转为不规则形,有空泡,突起形成。结论 多聚赖氨酸是血小板贴壁的理想粘附剂,本方法能简易,快捷地监测血小板激活过程中胞浆内钙离子动态变化及形态改变,有助于血小板功能的深入研究。     

P-Selectin-Mediated Adhesion between Platelets and Tumor Cells Promotes Intestinal Tumorigenesis in ApcMin/+ Mice

Studies have indicated that platelets play an important role in tumorigenesis, and an abundance of platelets accumulate in the ovarian tumor microenvironment outside the vasculature. However, whether cancer cells recruit platelets within intestinal tumors and how they signal adherent platelets to enter intestinal tumor tissues remain unknown. Here, we unexpectedly found that large numbers of platelets were deposited within human colorectal tumor specimens using immunohistochemical staining, and these platelets were fully associated with tumor development. We further report the robust adhesion of platelet aggregates to tumor cells within intestinal tumors, which occurs via a mechanism that is dependent on P-selectin (CD62P), a cell adhesion molecule that is abundantly expressed on activated platelets. Using spontaneous intestinal tumor mouse models, we determined that the genetic deletion of P-selectin suppressed intestinal tumor growth, which was rescued by the infusion of wild-type platelets but not P-selectin^-/- platelets. Mechanistically, platelet adhesion to tumor cells induced the secretion of vascular endothelial growth factor (VEGF) to promote angiogenesis and accelerate intestinal tumor cell proliferation. Our results indicate that the adherence of platelets to tumor cells could promote tumor growth and metastasis. By targeting this platelet-tumor cell interaction, recombinant soluble P-selectin may have therapeutic value for the treatment of intestinal tumors.     

Effects of Lipopolysaccharides from Gram-Negative Bacteria on the Level of Thiols in Blood Platelets

Lipopolysaccharide (endotoxin, LPS) activates blood platelets and stimulates generation of free radicals in these cells. The mechanism of platelet activation induced by LPS is not known. The aim of the present study was to examine how glutathione (GSH) and other thiol-containing compounds are involved in the oxidative stress in blood platelets caused by LPS. The HPLC technique has been used on the analysis of non-protein thiols from human blood platelets treated with lipopolysaccharides of different Gram-negative bacteria (Proteus mirabilis, Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa). Our results show that LPSs caused an increase (about 10%) of the level of reduced glutathione (GSH) and other nonprotein thiols such as cysteine (CSH) and cysteinylglycine (CGSH), whereas the total pool of these compounds was almost unchanged. LPS may react directly with thiols, since after incubation of LPSs with glutathione alone (in reduced form) we observed a distinct decrease of the level of platelet GSH.     

Biosurfactants as Agents to Reduce Adhesion of Pathogenic Bacteria to Polystyrene Surfaces: Effect of Temperature and Hydrophobicity

Polystyrene surfaces were conditioned with surfactin and rhamnolipid biosurfactants and then assessed regarding the attachment of Staphylococcus aureus, Listeria monocytogenes, and Micrococcus luteus. The effect of different temperatures (35, 25, and 4°C) on the anti-adhesive activity was also studied. Microbial adhesion to solvents and contact angle measurements were performed to characterize bacteria and material surfaces. The results showed that surfactin was able to inhibit bacterial adhesion in all the conditions analyzed, giving a 63–66% adhesion reduction in the bacterial strains at 4°C. Rhamnolipid promoted a slight decrease in the attachment of S. aureus. The anti-adhesive activity of surfactin increased with the decrease in temperature, showing that this is an important parameter to be considered in surface conditioning tests. Surfactin showed good potential as an anti-adhesive compound that can be explored to protect surfaces from microbial contamination.     

Relationship between Rapid, Firm Adhesion and Long-Term Colonization of Roots by Bacteria

For rhizobacteria to exert physiological effects on plant growth, the bacteria must first effectively colonize the root surface. To examine the relationship between long-term colonization of root systems and adherence to roots in the short term, a binding assay was developed. Adherence was determined by incubating roots of intact radish seedlings with bacteria, washing and homogenizing the roots, and dilution plating the resulting homogenate. Irreversible binding of bacteria was rapid, reaching half-maximum by 5 min. All of the rhizosphere bacteria tested showed similar, concentration-dependent binding (ranging from 10⁴ to 10⁸ CFU/ml), as well as long-term colonization of radish roots under sterile conditions. Escherichia coli, which is not a root colonizer, showed about 10-fold less binding, but still demonstrated concentration-dependent binding and rapid kinetics of adherence at high concentrations (10⁶ to 10⁸ CFU/ml). The bacteria tested were very different with respect to source or habitat and plant response, yet they showed similar concentration-dependent binding. There was no correlation between the relative hydrophobicities of the cell surfaces of strains and the adherence of the strains to roots. Binding of Pseudomonas fluorescens E6-22 was promoted by divalent cations (Ca²⁺ and Mg²⁺) at concentrations of 5 to 10 mM, whereas monovalent cations (Na⁺ and K⁺) had little effect; electrostatic phenomena may partially explain adherence in the short term, an important prelude to long-term colonization of root surfaces.     

Effects of Hydrogel/Silver Coatings on In Vitro Adhesion to Catheters of Bacteria Associated with Urinary Tract Infections

Sections of sterile all-silicone-, hydrogel/silver-all-silicone-, and hydrogel/silver-latex-Foley urinary catheters were exposed to suspensions of bacteria and Candida albicans associated with urinary tract infections. The adhesion of these microorganisms to the catheters was determined with a radiolabel–cell procedure and scanning electron microscopy. Anomalous data with the radiolabel procedure were produced with the hydrogel/silver-latex catheters for certain species. These aberrant data were related to adhesion on the untreated cut ends of the latex catheter. Radiolabel-cell-adhesion procedures that involve sections of coated materials may need to be supplemented with additional procedures such as scanning electron microscopy for valid interpretations of the data. Adhesion to the hydrogel/silver catheters by both Gram-positive- and Gram-negative bacteria most commonly associated with nosocomial urinary tract infections, including a strain of Pseudomonas aeruginosa noted for its superior adhesion capacity, was significantly lower than the adhesion to the control all-silicone catheter. Received: 21 January 2000 / Accepted: 26 February 2000     

Quantitative Approach in the Study of Adhesion of Lactic Acid Bacteria to Intestinal Cells and Their Competition with Enterobacteria

To describe the phenomena of bacterial adhesion to intestinal cells and the competition for adhesion between bacteria, mathematical equations based on a simple dissociation process involving a finite number of bacterial receptors on intestinal cell surface were developed. The equations allow the estimation of the maximum number of Lactobacillus sp. and Escherichia coli cells that can adhere to Caco-2 cells and intestinal mucus; they also characterize the affinity of the bacteria to Caco-2 cells and intestinal and fecal mucus and the theoretical adhesion ratio of two bacteria present in a mixed suspension. The competition for adhesion between Lactobacillus rhamnosus GG and E. coli TG1 appeared to follow the proposed kinetics, whereas the competition between Lactobacillus casei Shirota and E. coli TG1 may involve multiple adhesion sites or a soluble factor in the culture medium of the former. The displacement of the adhered Lactobacillus by E. coli TG1 seemed to be a rapid process, whereas the displacement of E. coli TG1 by the Lactobacillus took more than an hour.     

A Small Fibronectin-mimicking Protein from Bacteria Induces Cell Spreading and Focal Adhesion Formation

Fibronectin, a 250-kDa eukaryotic extracellular matrix protein containing an RGD motif plays crucial roles in cell-cell communication, development, tissue homeostasis, and disease development. The highly complex fibrillar fibronectin meshwork orchestrates the functions of other extracellular matrix proteins, promoting cell adhesion, migration, and intracellular signaling. Here, we demonstrate that CagL, a 26-kDa protein of the gastric pathogen and type I carcinogen Helicobacter pylori, mimics fibronectin in various cellular functions. Like fibronectin, CagL contains a RGD motif and is located on the surface of the bacterial type IV secretion pili as previously shown. CagL binds to the integrin receptor α₅β₁ and mediates the injection of virulence factors into host target cells. We show that purified CagL alone can directly trigger intracellular signaling pathways upon contact with mammalian cells and can complement the spreading defect of fibronectin^−/− knock-out cells in vitro. During interaction with various human and mouse cell lines, CagL mimics fibronectin in triggering cell spreading, focal adhesion formation, and activation of several tyrosine kinases in an RGD-dependent manner. Among the activated factors are the nonreceptor tyrosine kinases focal adhesion kinase and Src but also the epidermal growth factor receptor and epidermal growth factor receptor family member Her3/ErbB3. Interestingly, fibronectin activates a similar range of tyrosine kinases but not Her3/ErbB3. These findings suggest that the bacterial protein CagL not only exhibits functional mimicry with fibronectin but is also capable of activating fibronectin-independent signaling events. We thus postulate that CagL may contribute directly to H. pylori pathogenesis by promoting aberrant signaling cross-talk within host cells.