Peptidomic analysis of the host-defense peptides in skin secretions of the Amazon River frog Lithobates palmipes (Ranidae)

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Peptidomic Analysis of Skin Secretions of the Caribbean Frogs Leptodactylus insularum and Leptodactylus nesiotus (Leptodactylidae) Identifies an Ocellatin with Broad Spectrum Antimicrobial Activity

International audienceOcellatins are peptides produced in the skins of frogs belonging to the genus Leptodactylus that generally display weak antimicrobial activity against Gram-negative bacteria only. Peptidomic analysis of norepinephrine-stimulated skin secretions from Leptodactylus insularum Barbour 1906 and Leptodactylus nesiotus Heyer 1994, collected in the Icacos Peninsula, Trinidad, led to the purification and structural characterization of five ocellatin-related peptides from L. insularum (ocellatin-1I together with its (1-16) fragment, ocellatin-2I and its (1-16) fragment, and ocellatin-3I) and four ocellatins from L. nesiotus (ocellatin-1N,-2N,-3N, and-4N). While ocellatins-1I,-2I, and-1N showed a typically low antimicrobial potency against Gram-negative bacteria, ocellatin-3N (GIFDVLKNLAKGVITSLAS.NH 2) was active against an antibiotic-resistant strain of Klebsiella pneumoniae and reference strains of Escherichia coli, K. pneumoniae, Pseudomonas aeruginosa, and Salmonella typhimurium (minimum inhibitory concentrations (MICs) in the range 31.25-62.5 µM), and was the only peptide active against Gram-positive Staphylococcus aureus (MIC = 31.25 µM) and Enterococcus faecium (MIC = 62.5 µM). The therapeutic potential of ocellatin-3N is limited by its moderate hemolytic activity (LC 50 = 98 µM) against mouse erythrocytes. The peptide represents a template for the design of long-acting, non-toxic, and broad-spectrum antimicrobial agents for targeting multidrug-resistant pathogens

Transmission of Equine Influenza Virus to English Foxhounds

We retrospectively demonstrated that an outbreak of severe respiratory disease in a pack of English foxhounds in the United Kingdom in September 2002 was caused by an equine influenza A virus (H3N8). We also demonstrated that canine respiratory tissue possesses the relevant receptors for infection with equine influenza virus

The Salmonella effector SseJ disrupts microtubule dynamics when ectopically expressed in Normal Rat Kidney cells

Salmonella effector protein SseJ is secreted by Salmonella into the host cell cytoplasm where it can then modify host cell processes. Whilst host cell small GTPase RhoA has previously been shown to activate the acyl-transferase activity of SseJ we show here an un-described effect of SseJ protein production upon microtubule dynamism. SseJ prevents microtubule collapse and this is independent of SseJ's acyl-transferase activity. We speculate that the effects of SseJ on microtubules would be mediated via its known interactions with the small GTPases of the Rho family

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

Identification of Chromosomal Genes in Yersinia pestis that Influence Type III Secretion and Delivery of Yops into Target Cells

Pathogenic Yersinia species possess a type III secretion system, which is required for the delivery of effector Yop proteins into target cells during infection. Genes encoding the type III secretion machinery, its substrates, and several regulatory proteins all reside on a 70-Kb virulence plasmid. Genes encoded in the chromosome of yersiniae are thought to play important roles in bacterial perception of host environments and in the coordinated activation of the type III secretion pathway. Here, we investigate the contribution of chromosomal genes to the complex regulatory process controlling type III secretion in Yersinia pestis. Using transposon mutagenesis, we identified five chromosomal genes required for expression or secretion of Yops in laboratory media. Four out of the five chromosomal mutants were defective to various extents at injecting Yops into tissue culture cells. Interestingly, we found one mutant that was not able to secrete in vitro but was fully competent for injecting Yops into host cells, suggesting independent mechanisms for activation of the secretion apparatus. When tested in a mouse model of plague disease, three mutants were avirulent, whereas two strains were severely attenuated. Together these results demonstrate the importance of Y. pestis chromosomal genes in the proper function of type III secretion and in the pathogenesis of plague

Yersinia enterocolitica Serum Resistance Proteins YadA and Ail Bind the Complement Regulator C4b-Binding Protein

Many pathogens are equipped with factors providing resistance against the bactericidal action of complement. Yersinia enterocolitica, a Gram-negative enteric pathogen with invasive properties, efficiently resists the deleterious action of human complement. The major Y. enterocolitica serum resistance determinants include outer membrane proteins YadA and Ail. Lipopolysaccharide (LPS) O-antigen (O-ag) and outer core (OC) do not contribute directly to complement resistance. The aim of this study was to analyze a possible mechanism whereby Y. enterocolitica could inhibit the antibody-mediated classical pathway of complement activation. We show that Y. enterocolitica serotypes O:3, O:8, and O:9 bind C4b-binding protein (C4bp), an inhibitor of both the classical and lectin pathways of complement. To identify the C4bp receptors on Y. enterocolitica serotype O:3 surface, a set of mutants expressing YadA, Ail, O-ag, and OC in different combinations was tested for the ability to bind C4bp. The studies showed that both YadA and Ail acted as C4bp receptors. Ail-mediated C4bp binding, however, was blocked by the O-ag and OC, and could be observed only with mutants lacking these LPS structures. C4bp bound to Y. enterocolitica was functionally active and participated in the factor I-mediated degradation of C4b. These findings show that Y. enterocolitica uses two proteins, YadA and Ail, to bind C4bp. Binding of C4bp could help Y. enterocolitica to evade complement-mediated clearance in the human host