Immune Responses to Plague Infection in Wild Rattus rattus, in Madagascar: A Role in Foci Persistence?

Plague is endemic within the central highlands of Madagascar, where its main reservoir is the black rat, Rattus rattus. Typically this species is considered susceptible to plague, rapidly dying after infection inducing the spread of infected fleas and, therefore, dissemination of the disease to humans. However, persistence of transmission foci in the same area from year to year, supposes mechanisms of maintenance among which rat immune responses could play a major role. Immunity against plague and subsequent rat survival could play an important role in the stabilization of the foci. In this study, we aimed to investigate serological responses to plague in wild black rats from endemic areas of Madagascar. In addition, we evaluate the use of a recently developed rapid serological diagnostic test to investigate the immune response of potential reservoir hosts in plague foci.We experimentally infected wild rats with Yersinia pestis to investigate short and long-term antibody responses. Anti-F1 IgM and IgG were detected to evaluate this antibody response. High levels of anti-F1 IgM and IgG were found in rats one and three weeks respectively after challenge, with responses greatly differing between villages. Plateau in anti-F1 IgM and IgG responses were reached for as few as 500 and 1500 colony forming units (cfu) inoculated respectively. More than 10% of rats were able to maintain anti-F1 responses for more than one year. This anti-F1 response was conveniently followed using dipsticks.Inoculation of very few bacteria is sufficient to induce high immune response in wild rats, allowing their survival after infection. A great heterogeneity of rat immune responses was found within and between villages which could heavily impact on plague epidemiology. In addition, results indicate that, in the field, anti-F1 dipsticks are efficient to investigate plague outbreaks several months after transmission

Relative contribution of three main virulence factors in Pseudomonas aeruginosa pneumonia:

Objective: The pathogenesis and the outcome of Pseudomonas aeruginosa ventilator-acquired pneumonia depend on the virulence factors displayed by the bacteria as well as the host response. Thus, quorum sensing, lipopolysaccharide, and type 3 secretion system have each individually been shown to be important virulence systems in laboratory reference strains. However, the relative contribution of these three factors to the in vivo pathogenicity of clinically relevant strains has never been studied. We analyzed the virulence of 56 nonclonal Pseudomonas aeruginosa strains isolated from critically ill patients with ventilator-acquired pneumonia. To avoid the variation of human immune response, we used a murine model of pneumonia. The aim was to determine which virulence factor was the most important.Setting: Research laboratory of a university. Subjects: Male adult BALB/c mice. Interventions: In vitro, the phenotype of each strain was established as to the expression of quorum sensing-regulated factors (elastase and pyocyanin), type 3 secretion system exotoxin secretion (Exotoxin U, S and/or T, or “nonsecreting”), and lipopolysaccharide O-antigen serotype. Strain pathogenicity was evaluated in vivo in a mouse model of acute pneumonia through lung injury assessment by measuring alveolar–capillary barrier permeability to proteins, lung wet/dry weight ratio, and bacterial dissemination. Associations were then sought between virulence system phenotypes and levels of lung injury. Measurements and Main Results: In univariate analysis, elastase production, O11 serotype, and type 3 secretion system exotoxin secretion were associated with increased lung injury and exotoxin U was linked to an increase risk of bacteremia. In multivariate analysis, we observed that type 3 secretion system exotoxin secretion and to a lesser degree elastase production were associated with increased lung injury. Conclusion: In a murine model of pneumonia, our data suggest that type 3 secretion system and elastase are the most important virulence factors in clinically relevant P. aeruginosa strains

Contributions of chaperone/usher systems to cell binding, biofilm formation and Yersinia pestis virulence

Yersinia pestis genome sequencing projects have revealed six intact uncharacterized chaperone/ usher systems with the potential to play roles in plague pathogenesis. We cloned each locus and expressed them in the Deltafim Escherichia coli strain AAEC185 to test the assembled Y. pestis surface structures for various activities. Expression of each chaperone/usher locus gave rise to specific novel fibrillar structures on the surface of E. coli. One locus, y0561-0563, was able to mediate attachment to human epithelial cells (HEp-2) and human macrophages (THP-1) but not mouse macrophages (RAW264.7), while several loci were able to facilitate E. coli biofilm formation. When each chaperone/usher locus was deleted in Y. pestis, only deletion of the previously described pH 6 antigen (Psa) chaperone/usher system resulted in decreased adhesion and biofilm formation. Quantitative RT-PCR (qRT-PCR) revealed low expression levels for each novel chaperone/usher system in vitro as well as in mouse tissues following intravenous infection. However, a Y. pestis mutant in the chaperone/usher locus y1858-1862 was attenuated for virulence in mice via the intravenous route of infection, suggesting that expression of this locus is, at some stage, sufficient to affect the outcome of a plague infection. qRT-PCR experiments also indicated that expression of the chaperone/usher-dependent capsule locus, caf1, was influenced by oxygen availability and that the well-described chaperone/usher-dependent pilus, Psa, was strongly induced in minimal medium even at 28 degrees C rather than 37 degrees C, a temperature previously believed to be required for Psa expression. These data indicate several potential roles for the novel chaperone/usher systems of Y. pestis in pathogenesis and infection-related functions such as cell adhesion and biofilm formation.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/91950/1/2011 Microbiology - Contributions of chaperone usher systems to cell binding biofilm formation and Yersinia pestis virulence.pd

Modified Needle-Tip PcrV Proteins Reveal Distinct Phenotypes Relevant to the Control of Type III Secretion and Intoxication by Pseudomonas aeruginosa

The type III secretion system (T3SS) is employed to deliver effector proteins to the cytosol of eukaryotic hosts by multiple species of Gram-negative bacteria, including Pseudomonas aeruginosa. Translocation of effectors is dependent on the proteins encoded by the pcrGVHpopBD operon. These proteins form a T3S translocator complex, composed of a needle-tip complex (PcrV), translocons (PopB and PopD), and chaperones (PcrG and PcrH). PcrV mediates the folding and insertion of PopB/PopD in host plasmic membranes, where assembled translocons form a translocation channel. Assembly of this complex and delivery of effectors through this machinery is tightly controlled by PcrV, yet the multifunctional aspects of this molecule have not been defined. In addition, PcrV is a protective antigen for P. aeruginosa infection as is the ortholog, LcrV, for Yersinia. We constructed PcrV derivatives containing in-frame linker insertions and site-specific mutations. The expression of these derivatives was regulated by a T3S-specific promoter in a pcrV-null mutant of PA103. Nine derivatives disrupted the regulation of effector secretion and constitutively released an effector protein into growth medium. Three of these regulatory mutants, in which the linker was inserted in the N-terminal globular domain, were competent for the translocation of a cytotoxin, ExoU, into eukaryotic host cells. We also isolated strains expressing a delayed-toxicity phenotype, which secrete translocators slowly despite the normal level of effector secretion. Most of the cytotoxic translocation-competent strains retained the protective epitope of PcrV derivatives, and Mab166 was able to protect erythrocytes during infection with these strains. The use of defined PcrV derivatives possessing distinct phenotypes may lead to a better understanding of the functional aspects of T3 needle-tip proteins and the development of therapeutic agents or vaccines targeting T3SS-mediated intoxication

Transformations mineralogiques et texturales de materiaux rocheux, mortiers et betons d'ouvrages varies. Approche de la cinetique des mecanismes et identification des facteurs responsables

SIGLEINIST T 75749 / INIST-CNRS - Institut de l'Information Scientifique et TechniqueFRFranc

Mécanismes de la sécrétion des toxines ExoS et ExoT par le système de sécrétion de type III de Pseudomonas aeruginosa (séquence d'adressage et rôle de la chaperonne SpcS)

Pseudomonas aeruginosa, pathogène opportuniste, est responsable d'infections nosocomiales et d'infections pulmonaires chez les patients atteints de mucoviscidose. L'un des facteurs déterminants dans la virulence de cette bactérie est son système de sécrétion de type III (SSTT). Le SSTT permet aux bactéries d'injecter des toxines directement dans le cytosol d'ne cellule eucaryote cible. ce facteur de virulence peut être aussi considéré comme un système biologique de transfert de protéines. Nous avons donc défini les modalités d'utilisation de la souche CHA de P.aeruginosa comme outil de vectorialisation. L'invalidation des gènes exoS et exoT a permis de créer une souche peu toxique pour les cellules eucaryotes, qui présente une capacité d'injection accrue pour les protéines d'intérêt. Des protéines de fusion ont été réalisées avec différents segments N-terminaux de ExoS et des protéines rapporteuses. La capacité de ces protéines à être produites et injectées par le SSTT de la souche CHA, a permis de définir la séquence d'adressage permettant l'injection optimale dans une cellule cible. Les 54 premiers acides aminés de ExoS sont le meilleur compromis entre un domaine minimum de la toxine native et une information suffisante pour un adressage efficace vers le SSTT. le gène orf1 code pour une chaperonne spécifique : SpcS, qui interagit avec les toxines ExoS et ExoT dans le cytosol de la souche CHA et favorise leur sécrétion, sans pour autant être indispensable à un mécanisme ou à la toxicité bactérienne. Avec le complexe ExoS/Spcs nous avons copurifié la protéine CysB. Cette protéine est un nouvel inhibiteur du SSTT, impliqué dans une voie de régulation encore indéterminée.GRENOBLE1-BU Sciences (384212103) / SudocSudocFranceF

Les facteurs prédictifs de gravité de la leptospirose en Polynésie française

AIX-MARSEILLE2-BU Méd/Odontol. (130552103) / SudocPARIS-BIUM (751062103) / SudocSudocFranceF

Considerations for Infectious Disease Research Studies Using Animals

Animal models are vital in understanding the transmission and pathogenesis of infectious organisms and the host immune response to infection. In addition, animal models are essential in vaccine and therapeutic drug development and testing. Prior to selecting an animal model to use when studying an infectious agent, the scientific team must determine that sufficient in vitro and ex vivo data are available to justify performing research in an animal model, that ethical considerations are addressed, and that the data generated from animal work will add useful information to the body of scientific knowledge. Once it is established that an animal should be used, the questions become 'Which animal model is most suitable?' and 'Which experimental design issues should be considered?' The answers to these questions take into account numerous factors, including scientific, practical, welfare, and regulatory considerations, which are the focus of this article

Polymorphisms in the lcrV Gene of Yersinia enterocolitica and Their Effect on Plague Protective Immunity

Current efforts to develop plague vaccines focus on LcrV, a polypeptide that resides at the tip of type III secretion needles. LcrV-specific antibodies block Yersinia pestis type III injection of Yop effectors into host immune cells, thereby enabling phagocytes to kill the invading pathogen. Earlier work reported that antibodies against Y. pestis LcrV cannot block type III injection by Yersinia enterocolitica strains and suggested that lcrV polymorphisms may provide for escape from LcrV-mediated plague immunity. We show here that polyclonal or monoclonal antibodies raised against Y. pestis KIM D27 LcrV (LcrVD27) bind LcrV from Y. enterocolitica O:9 strain W22703 (LcrVW22703) or O:8 strain WA-314 (LcrVWA-314) but are otherwise unable to block type III injection by Y. enterocolitica strains. Replacing the lcrV gene on the pCD1 virulence plasmid of Y. pestis KIM D27 with either lcrVW22703 or lcrVWA-314 does not affect the ability of plague bacteria to secrete proteins via the type III pathway, to inject Yops into macrophages, or to cause lethal plague infections in mice. LcrVD27-specific antibodies blocked type III injection by Y. pestis expressing lcrVW22703 or lcrVWA-314 and protected mice against intravenous lethal plague challenge with these strains. Thus, although antibodies raised against LcrVD27 are unable to block the type III injection of Y. enterocolitica strains, expression of lcrVW22703 or lcrVWA-314 in Y. pestis did not allow these strains to escape LcrV-mediated plague protective immunity in the intravenous challenge model