Lsa63, a newly identified surface protein of Leptospira interrogans binds laminin and collagen IV

Leptospira interrogans is the etiological agent of leptospirosis, a zoonotic disease that affects populations worldwide. We have identified in proteomic studies a protein that is encoded by the gene LIC10314 and expressed in virulent strain of L. interrogans serovar Pomona. This protein was predicted to be surface exposed by PSORT program and contains a p83/100 domain identified by BLAST analysis that is conserved in protein antigens of several strains of Borrelia and Treponema spp. The proteins containing this domain have been claimed antigen candidates for serodiagnosis of Lyme borreliosis. Thus, we have cloned the LIC10314 and expressed the protein in Escherichia coli BL21-SI strain by using the expression vector pAE. The recombinant protein tagged with N-terminal hexahistidine was purified by metal-charged chromatography and characterized by circular dichroism spectroscopy. This protein is conserved among several species of pathogenic Leptospira and absent in the saprophytic strain L. biflexa. We confirm by liquid-phase immunofluorescence assays with living organisms that this protein is most likely a new surface leptospiral protein. The ability of the protein to mediate attachment to ECM components was evaluated by binding assays. The leptospiral protein encoded by LIC10314, named Lsa63 (Leptospiral surface adhesin of 63 kDa), binds strongly to laminin and collagen IV in a dose-dependent and saturable fashion. In addition, Lsa63 is probably expressed during infection since it was recognized by antibodies of serum samples of confirmed-leptospirosis patients in convalescent phase of the disease. Altogether, the data suggests that this novel identified surface protein may be involved in leptospiral pathogenesis. (C) 2009 The British Infection Society. Published by Elsevier Ltd. All rights reserved.

Lp95, a novel leptospiral protein that binds extracellular matrix components and activates e-selectin on endothelial cells

Objectives: The study of a predicted outer membrane leptospiral protein encoded by the gene LIC12690 in mediating the adhesion process. Methods: The gene was cloned and expressed in Escherichia coli BL21 (SI) strain by using the expression vector pAE. The recombinant protein tagged with N-terminal hexahistidine was purified by metal-charged chromatography and used to assess its ability to activate human umbilical vein endothelial cells (HUVECs). Results: The recombinant leptospiral protein of 95 kDa, named Lp95, activated E-selectin in a dose-dependent fashion but not the intercellular adhesion molecule 1 (ICAM-1). In addition, we show that pathogenic and non-pathogenic Leptospira are both capable to stimulate endothelium E-selectin and ICAM-1, but the pathogenic L. interrogans serovar Copenhageni strain promotes a statistically significant higher activation than the non-pathogenic L. biflexa serovar Patoc (P < 0.01). The Lp95 was identified in vivo in the renal tubules of animal during experimental infection with L. interrogans. The whole Lp95 as well as its fragments, the C-terminal containing the domain of unknown function (DUF), the N-terminal and the central overlap regions bind laminin and fibronectin ECM molecules, being the binding stronger with the DUF containing fragment. Conclusion: This is the first leptospiral protein capable to mediate the adhesion to ECM components and the activation of HUVECS, thus suggesting its participation in the pathogenesis of Leptospira. (C) 2009 The British Infection Society. Published by Elsevier Ltd. All rights reserved.

A newly identified protein of Leptospira interrogans mediates binding to laminin

Pathogenic Leptospira is the aetiological agent of leptospirosis, a life-threatening disease that affects populations worldwide. The search for novel antigens that could be relevant in host-pathogen interactions is being pursued. These antigens have the potential to elicit several activities, including adhesion. This study focused on a hypothetical predicted lipoprotein of Leptospira, encoded by the gene LIC12895, thought to mediate attachment to extracellular matrix (ECM) components. The gene was cloned and expressed in Escherichia coli BL21 Star (DE3)pLys by using the expression vector pAE. The recombinant protein tagged with N-terminal hexahistidine was purified by metal-charged chromatography and characterized by circular dichroism spectroscopy. The capacity of the protein to mediate attachment to ECM components was evaluated by binding assays. The leptospiral protein encoded by LIC12895, named Lsa27 (leptospiral surface adhesin, 27 kDa), bound strongly to laminin in a dose-dependent and saturable fashion. Moreover, Lsa27 was recognized by antibodies from serum samples of confirmed leptospirosis specimens in both the initial and the convalescent phases of the disease. Lsa27 is most likely a surface protein of Leptospira as revealed in liquid-phase immunofluorescence assays with living organisms. Taken together, these data indicate that this newly identified membrane protein is expressed during natural infection and may play a role in mediating adhesion of L. interrogans to its host.

Phosphorus kinetics in calves experimentally submitted to a trickle infection with Cooperia punctata

Ten male Holstein calves (74.3 +/- 3.2 kg LW) were used for a trial with trickle infection with Cooperia punctata to evaluate phosphorus (P) kinetics. Five calves were inoculated with 10,000 L(3) stage larvae per week during 35 days, while the other group of five calves was kept as a control. On the 29th day each calf was intravenously injected with 29.6 MBq of a (32)p solution. Blood samples were taken at 24 h periods for 7 days, after which all calves were slaughtered and worms burdens. Faeces, urine and tissue samples were taken for analysis using isotopic dilution and modeling techniques. The number of eggs per gram of faeces (EPG)was 1920 +/- 168 on 28th day and the total number of worms burdens was 11,131 +/- 1500. Infected calves showed lower feed intake and live weight gain, as well as lower P intake, absorption and retention than control calves. The P flows between body compartments were lower for blood to gastrointestinal tract (TGI), TGI to blood, blood to soft tissues, bone balance and soft tissue balance in infected calves when compared to the control. The trickle infection of C punctata affected P metabolism due to the decrease in P retained and live weight due to fall in feed intake. (C) 2009 Elsevier B.V. All rights reserved.

Two Atypical Enteropathogenic Escherichia coli Strains Induce the Production of Secreted and Membrane-Bound Mucins To Benefit Their Own Growth at the Apical Surface of Human Mucin-Secreting Intestinal HT29-MTX Cells

In rabbit ligated ileal loops, two atypical enteropathogenic Escherichia coli (aEPEC) strains, 3991-1 and 0421-1, intimately associated with the cell membrane, forming the characteristic EPEC attachment and effacement lesion of the brush border, induced a mucous hypersecretion, whereas typical EPEC (tEPEC) strain E2348/69 did not. Using cultured human mucin-secreting intestinal HT29-MTX cells, we demonstrate that apically aEPEC infection is followed by increased production of secreted MUC2 and MUC5AC mucins and membrane-bound MUC3 and MUC4 mucins. The transcription of the MUC5AC and MUC4 genes was transiently upregulated after aEPEC infection. We provide evidence that the apically adhering aEPEC cells exploit the mucins` increased production since they grew in the presence of membrane-bound mucins, whereas tEPEC did not. The data described herein report a putative new virulence phenomenon in aEPEC.

The essential role of toll like receptor-4 in the control of Aggregatibacter actinomycetemcomitans infection in mice

Objective: Aggregatibacter actinomycetemcomitans is an oral Gram-negative bacterium that contributes to periodontitis progression. Isolated antigens from A. actinomycetemcomitans could be activating innate immune cells through Toll-like receptors (TLRs). In this study, we evaluated the role of TLR4 in the control of A. actinomycetemcomitans infection. Material and Methods: We examined the mechanisms that modulate the outcome of A. actinomycetemcomitans-induced periodontal disease in TLR4(-/-) mice. The production of cytokines was evaluated by ELISA. The bacterial load was determined by counting the number of colony-forming units per gram of tissue. Results: The results showed that TLR4-deficient mice developed less severe periodontitis after A. actinomycetemcomitans infection, characterized by significantly lower bone loss and inflammatory cell migration to periodontal tissues. However, the absence of TLR4 facilitated the A. actinomycetemcomitans dissemination. Myeloperoxidase activity was diminished in the periodontal tissue of TLR4(-/-) mice. We observed a significant reduction in the production of tumour necrosis factor-alpha (TNF-alpha) and interleukin (IL)-1 beta in the periodontal tissue of TLR4(-/-) mice. Conclusion: The results of this study highlighted the role of TLR4 in controlling A. actinomycetemcomitans infection.

The Role of Toll-Like Receptor 2 in the Recognition of Aggregatibacter actinomycetemcomitans

Background: Aggregatibacter actinomycetemcomitans (previously Actinobacillus actinomycetemcomitans) is a Gram-negative bacterium present in the oral cavity and is usually associated with localized aggressive periodontitis. Isolated antigens from A. actinomycetemcomitans can activate innate immune cells through Toll-like receptors (TLRs), which are molecules that recognize structural components conserved among microorganisms. In this study, we evaluate the role of TLR2 in the recognition of A. actinomycetemcomitans. Methods: Macrophages and neutrophils from knockout mice with targeted disruption of TLR2 (TLR2(-/-) mice) and wild-type mice were collected and used for the subsequent assays. The production of cytokines and chemokines was evaluated by enzyme-linked immunosorbent assay (ELISA), and the presence of apoptotic cells was determined by flow cytometry. In addition, the mechanisms that modulate the outcome of A. actinomycetemcomitans-induced periodontal disease in TLR2(-/-) mice were examined. Results: The results show that TLR2-deficient mice developed more severe periodontitis after A. actinomycetemcomitans infection, characterized by significantly higher bone loss and inflammatory cell migration to periodontal tissues. The inflammatory cell influx into the peritoneal cavities of TLR2(-/-) mice was three-fold lower than that observed for the littermate controls. A significantly diminished production of the cytokines tumor necrosis factor-alpha and interleukin-1 beta as well as the chemokine CC-ligand-5 in the peritoneal cavities of TLR2(-/-) mice was observed. In addition, a high frequency of apoptotic cells in the inflammatory exudates from TLR2(-/-) mice was observed. Phagocytosis and nitric oxide production was diminished in cells from TLR2(-/-) mice, facilitating the dissemination of the pathogen to the spleen. Conclusion: The results of this study highlight the involvement of TLR2 in recognizing A. actinomycetemcomitans and its essential role in controlling A. actinomycetemcomitans infection. J Periodontot 2009,80:2070-2019.

Differential inclusions and robust control theory

Uncontrolled systems (x) over dot is an element of Ax, where A is a non-empty compact set of matrices, and controlled systems (x) over dot is an element of Ax + Bu are considered. Higher-order systems 0 is an element of Px - Du, where and are sets of differential polynomials, are also studied. It is shown that, under natural conditions commonly occurring in robust control theory, with some mild additional restrictions, asymptotic stability of differential inclusions is guaranteed. The main results are variants of small-gain theorems and the principal technique used is the Krasnosel'skii-Pokrovskii principle of absence of bounded solutions.

Human susceptibility to Schistosoma japonicum in China correlates with antibody isotypes to native antigens

Antibody isotypic responses (IgE, IgA, IgG1, IgG2, IgG3 and IgG4) to Schistosoma japonicum antigens-adult worm (AWA), soluble egg (SEA) and the recombinant proteins TEG (22.6-kDa tegumental antigen, Sj22) and PMY (paramyosin, Sj97)-were measured (in 1998) in a cohort of 179 Chinese subjects 2 years post-treatment. Subjects in the highest intensity re-infection group (> 100 eggs per gram faeces) had significantly higher levels of IgG1 and IgG4 against AWA. Analysis of IgG4/IgE ratios for AWA and SEA linked IgG4 excess to re-infection and IgE excess to non-re-infection. Two years after chemotherapeutic cure, 29 subjects, who were re-infected or never infected but highly water-exposed, were classified as epidemiologically susceptible (n = 15) or epidemiologically insusceptible to infection (n = 14). IgG4 levels against native antigens (AWA and SEA) were higher in susceptibles and IgE levels were higher in insusceptibles but antibody responses to the recombinant proteins (PMY and TEG) showed no clear pattern or difference between susceptibility groups. These and earlier findings provide evidence that immunity develops against schistosomiasis japonica in China and that susceptibility/resistance correlates with antibody isotypes against native schistosome antigens.

Efficacy of praziquantel against Schistosoma japonicum: field evaluation in an area with repeated chemotherapy compared with a newly identified endemic focus in Hunan, China

This study conducted in 1999/2000 was designed to evaluate the efficacy of praziquantel against Schistosoma japonicum in an area with repeated chemotherapy (Area A) compared with a newly identified endemic focus (Area B) in Hunan Province, China. The population size was 2015 and 2180 in Areas A and B, respectively, of which 1129 and 1298 subjects received stool examination. A total of 230 subjects were identified by the Kato-Katz technique (4 smears per person) as being infected with S. japonicum, 124 in Area A (prevalence 11 %) and 106 in Area B (prevalence 8.2%). They were treated with a single oral dose of praziquantel (40 mg/kg) in the non-transmission season. A follow-up stool examination was made 50 days after treatment. Among the 220 cases followed, 22 were found stool-egg-positive, with an overall cure rate of 90 %, and 99 % reduction of infection intensity (eggs per gram stool). No significant difference was found in cure rates between the 2 areas (89.7% vs 90.3%). The efficacy of the drug in the area with repeated chemotherapy was not significantly different from that in the newly identified endemic focus. This study, therefore, suggests that the efficacy of praziquantel against S. japonicum has not changed in the Dongting Lake region after more than 14 years of mass chemotherapy, and there is no evidence of tolerance or resistance of S. japonicum against praziquantel.

X-ray fluorescence with synchrotron radiation to elemental analysis of lead and calcium content of primary teeth

Primary teeth were analyzed by micro-SRXRF. The aim of this study was to determine the elemental distribution of lead and calcium in different regions of primary incisor of children living in a notoriously contaminated area (Santo Amaro da Purificacao, Bahia State, Brazil). The measurements were performed in standard geometry of 45 incidence, exciting with a white beam and using a conventional system collimation (orthogonal slits) in the XRF beamline at the Synchrotron Light National Laboratory (Campinas, Brazil). (C) 2009 Elsevier Ltd. All rights reserved.

An in vitro study of non-axial forces upon the retention of an O-ring attachment

Objective The purpose of this study was to evaluate the retention force of an O-ring attachment system in different inclinations to the ideal path of insertion, using devices to compensate angulations. Material and methods Two implants were inserted into an aluminum base, and ball attachments were screwed to implants. Cylinders with O-rings were placed on ball attachments and connected to the test device using positioners to compensate implant angulations (0 degrees, 7 degrees, and 14 degrees). Plexiglass bases were used to simulate implant angulations. The base and the test device were positioned in a testing apparatus, which simulated insertion/removal of an overdenture. A total of 2900 cycles, simulating 2 years of overdenture use, were performed and 36 O-rings were tested. The force required for each cycle was recorded with computer software. Longitudinal sections of ball attachment-positioner-cylinder with O-rings of each angulation were obtained to analyze the relationship among them, and O-ring sections tested in each angulation were compared with an unused counterpart. A mixed linear model was used to analyze the data, and the comparison was performed by orthogonal contrasts (alpha=0.05). Results At 0 degrees, the retention force decreased significantly over time, and the retention force was significantly different in all comparisons, except from 12 to 18 months. When the implants were positioned at 7 degrees, the retention force was statistically different at 0 and 24 months. At 14 degrees, significant differences were found from 6 and 12 to 24 months. Conclusions Within the limitations of this study, it was concluded that O-rings for implant/attachments perpendicular to the occlusal plane were adequately retentive over the first year and that the retentive capacity of O-ring was affected by implant inclinations despite the proposed positioners. To cite this article:Rodrigues RCS, Faria ACL, Macedo AP, Sartori IAM, de Mattos MGC, Ribeiro RF. An in vitro study of non-axial forces upon the retention of an O-ring attachment.Clin. Oral Impl. Res. 20, 2009; 1314-1319.doi: 10.1111/j.1600-0501.2009.01742.x.

Assessing alternative models of individualism and collectivism: A confirmatory factor analysis

Six alternative structural models of individualism-collectivism are reviewed and empirically compared in a confirmatory factor analysis of questionnaire data from an Australian student sample (N=340). Central to the debate about the structure of this broad social attitude are the issues of (I) polarity (are individualism and collectivism bipolar opposites, or orthogonal factors?) and (2) dimensionality (are individualism and collectivism themselves higher-order constructs subsuming several more specific factors and, if so, what are they?). The data from this Australian sample support a model that represents individualism and collectivism as a higher-order bipolar factor hierarchically subsuming several bipolar reference-group-specific individualisms and collectivisms. Copyright (C) 2001 John Wiley & Sons, Ltd.

Appraisal ratings, halo, and selection: A study using sales staff

Supervisor ratings are useful criteria for the validation of selection instruments but maybe limited because of the presence of rating errors, such as halo. This study set out to show that supervisor ratings which are high in halo remain successful criteria in selection. Following a thorough job analysis, a customer service questionnaire was designed to assess the potential of retail sales staff on three orthogonal subscales labelled Dealing with people, Emotions and energy, and Solitary style. These subscales were uncorrelated with supervisor ratings made about 8 weeks later. However, the supervisor ratings were correlated with an overall scale derived from the three scales of the customer service questionnaire. These results support the view that supervisor ratings generally consist of global impressions and suggest that these global impressions are useful measures of overall performances. This field study confirms laboratory results that halo does not necessarily reduce rating accuracy.

Stochastic gauges in quantum dynamics for many-body simulations

Quantum dynamics simulations can be improved using novel quasiprobability distributions based on non-orthogonal Hermitian kernel operators. This introduces arbitrary functions (gauges) into the stochastic equations. which can be used to tailor them for improved calculations. A possible application to full quantum dynamic simulations of BEC's is presented. (C) 2001 Elsevier Science B.V. All rights reserved.