The Passage of Chaperonins to Extracellular Locations in Requires a Functional Dot/Icm System.

HtpB, the chaperonin of the bacterial pathogen , is found in extracellular locations, even the cytoplasm of host cells. Although chaperonins have an essential cytoplasmic function in protein folding, HtpB exits the cytoplasm to perform extracellular virulence-related functions that support 's lifestyle. The mechanism by which HtpB reaches extracellular locations is not currently understood.

The Functional Differences between the GroEL Chaperonin of and the HtpB Chaperonin of Can Be Mapped to Specific Amino Acid Residues.

Group I chaperonins are a highly conserved family of essential proteins that self-assemble into molecular nanoboxes that mediate the folding of cytoplasmic proteins in bacteria and organelles. GroEL, the chaperonin of , is the archetype of the family. Protein folding-independent functions have been described for numerous chaperonins, including HtpB, the chaperonin of the bacterial pathogen .

Subcellular Location of Heat Shock Protein 60 (Hsp60) Chaperone by Using Immunogold Labeling and Proteomic Analysis.

is the causative bacterial agent of piscirickettsiosis, a systemic fish disease that significantly impacts the Chilean salmon industry. This bacterium possesses a type IV secretion system (T4SS), several proteins of the type III secretion system (T3SS), and a single heat shock protein 60 (Hsp60/GroEL). It has been suggested that due to its high antigenicity, the Hsp60 could be surface-exposed, translocated across the membrane, and (or) secreted into the extracellular matrix.

Potential Ad Hoc Markers of Persistence and Virulence in Canadian Food and Clinical Isolates.

The gene encoding a surface virulence protein, was examined for the presence of premature stop codon (PMSC) mutations in 82 isolates obtained by the Canadian Food Inspection Agency (CFIA) from foods and food contact surfaces. These mutations were coanalyzed for the presence of stress survival islet 1 (SSI-1) and for the abilities of the isolates to invade Caco-2 intestinal epithelial cells and form biofilms on polystyrene. PMSC mutations were present in one-third of the isolates (predominantly those of serogroup 1/2a), and their presence was correlated with a noninvasive phenotype.

In vitro genomic and proteomic evidence of a type IV pili-like structure in the fish pathogen Piscirickettsia salmonis.

Piscirickettsia salmonis is an intracellular γ-proteobacteria and the etiological agent of piscirickettsiosis, which causes massive economic losses in the Chilean salmon industry. The type IV pili (T4P) play an important role in adherence to host cell surfaces and bacterial pathogenicity. T4P contains a variable number of components, as predicted in P.

Molecular and Biochemical Methods Useful for the Epigenetic Characterization of Chromatin-Associated Proteins in Bivalve Molluscs.

Bivalve molluscs constitute a ubiquitous taxonomic group playing key functions in virtually all ecosystems, and encompassing critical commercial relevance. Along with a sessile and filter-feeding lifestyle in most cases, these characteristics make bivalves model sentinel organisms routinely used for environmental monitoring studies in aquatic habitats. The study of epigenetic mechanisms linking environmental exposure and specific physiological responses (i.

Genome Sequence of Listeria monocytogenes Plasmid pLM-C-273 Carrying Genes Related to Stress Resistance.

Mobile genetic elements in bacteria, such as plasmids, act as important vectors for the transfer of antibiotic resistance, virulence, and metal resistance genes. Here, we report the genome sequence of a new plasmid pLM-C-273, identified in a Listeria monocytogenes strain isolated from a clinical sample in Ontario, Canada.

Identification of vacuoles containing extraintestinal differentiated forms of Legionella pneumophila in colonized Caenorhabditis elegans soil nematodes.

Legionella pneumophila, a causative agent of Legionnaires' disease, is a facultative intracellular parasite of freshwater protozoa. Legionella pneumophila features a unique developmental network that involves several developmental forms including the infectious cyst forms. Reservoirs of L.

The many forms of a pleomorphic bacterial pathogen-the developmental network of Legionella pneumophila.

Legionella pneumophila is a natural intracellular bacterial parasite of free-living freshwater protozoa and an accidental human pathogen that causes Legionnaires' disease. L. pneumophila differentiates, and does it in style.

Deletion of potD, encoding a putative spermidine-binding protein, results in a complex phenotype in Legionella pneumophila.

L. pneumophila is an intracellular pathogen that replicates in a membrane-bound compartment known as the Legionella-containing vacuole (LCV). We previously observed that the polyamine spermidine, produced by host cells or added exogenously, enhances the intracellular growth of L.

The progeny of Legionella pneumophila in human macrophages shows unique developmental traits.

The Gram-negative bacterium Legionella pneumophila is an intracellular parasite of amoebae and an accidental human pathogen that causes a noncommunicable atypical pneumonia known as Legionnaires' disease (LD). In some mammalian cells (e.g.

Stationary phase and mature infectious forms of Legionella pneumophila produce distinct viable but non-culturable cells.

Legionella pneumophila is an intracellular bacterial parasite of freshwater protozoa and an accidental waterborne human pathogen. L. pneumophila is highly pleomorphic showing several forms that differentiate within its developmental cycle.

The tetrahymena and acanthamoeba model systems.

Although the study of protozoology has been active for centuries, very few current academic curricula incorporate requirements or even options for coursework on the study of protists; yet, protozoa are becoming widely recognized by investigators as organisms that play a significant role in the evolution, pathogenicity, protection and amplification of human pathogens in the environment. This is particularly true for the study of Legionella, as this accidental human pathogen has naturally evolved to infect protozoa in fresh water environments. Researchers have made great progress in the study of pathogenicity, evolution, and ecology of Legionella and its protozoan hosts, which include amoebae and ciliated protozoa.

Electron microscopy of legionella and legionella-infected cells.

Those investigators who study the morphology of Legionella and Legionella-infected cells have greatly benefited from the superior resolution afforded by electron microscopy (EM). It can also be said with confidence that EM will continue to reveal as yet to be discovered features of this fascinating intracellular pathogen. In this chapter we detail our practical experience in the application of three transmission electron microscopy (TEM) techniques to the study of Legionella: conventional ultrastructural analysis, immuno-gold labeling, and negative staining.

Mechanism of invasion of lung epithelial cells by filamentous Legionella pneumophila.

Legionella, the aetiological agent responsible for Legionellosis, is an opportunistic pathogen that infects humans upon the inhalation of contaminated aerosolized water droplets. Legionella is pleomorphic and its different morphotypes exhibit varying degrees of virulence. While the filamentous forms of Legionella pneumophila (Lp) have been reported in patient samples since the first description of legionellosis, their role in disease has not been studied.

Passage through Tetrahymena tropicalis enhances the resistance to stress and the infectivity of Legionella pneumophila.

Legionella pneumophila is a gram-negative bacterium prevalent in fresh water which accidentally infects humans and is responsible for the disease called legionellosis. Intracellular growth of L. pneumophila in Tetrahymena is inconsistent; in the species Tetrahymena tropicalis stationary-phase forms (SPFs) of L.

The htpAB operon of Legionella pneumophila cannot be deleted in the presence of the groE chaperonin operon of Escherichia coli.

HtpB, the chaperonin of the intracellular bacterial pathogen Legionella pneumophila , displays several virulence-related functions in vitro. To confirm HtpB's role in vivo, host infections with an htpB deletion mutant would be required. However, we previously reported that the htpAB operon (encoding co-chaperonin and chaperonin) is essential.

Legionella pneumophila requires polyamines for optimal intracellular growth.

The Gram-negative intracellular pathogen Legionella pneumophila replicates in a membrane-bound compartment known as the Legionella-containing vacuole (LCV), into which it abundantly releases its chaperonin, HtpB. To determine whether HtpB remains within the LCV or reaches the host cell cytoplasm, we infected U937 human macrophages and CHO cells with L. pneumophila expressing a translocation reporter consisting of the Bordetella pertussisa denylate cyclase fused to HtpB.

The Legionella pneumophila Chaperonin - An Unusual Multifunctional Protein in Unusual Locations.

The Legionella pneumophila chaperonin, high temperature protein B (HtpB), was discovered as a highly immunogenic antigen, only a few years after the identification of L. pneumophila as the causative agent of Legionnaires' disease. As its counterparts in other bacterial pathogens, HtpB did not initially receive further attention, particularly because research was focused on a few model chaperonins that were used to demonstrate that chaperonins are essential stress proteins, present in all cellular forms of life and involved in helping other proteins to fold.

DsbA2 (27 kDa Com1-like protein) of Legionella pneumophila catalyses extracytoplasmic disulphide-bond formation in proteins including the Dot/Icm type IV secretion system.

In Gram-negative bacteria, thiol oxidoreductases catalyse the formation of disulphide bonds (DSB) in extracytoplasmic proteins. In this study, we sought to identify DSB-forming proteins required for assembly of macromolecular structures in Legionella pneumophila. Here we describe two DSB-forming proteins, one annotated as dsbA1 and the other annotated as a 27 kDa outer membrane protein similar to Com1 of Coxiella burnetii, which we designate as dsbA2.

The purified and recombinant Legionella pneumophila chaperonin alters mitochondrial trafficking and microfilament organization.

A portion of the total cellular pool of the Legionella pneumophila chaperonin, HtpB, is found on the bacterial cell surface, where it can mediate invasion of nonphagocytic cells. HtpB continues to be abundantly produced and released by internalized L. pneumophila and may thus have postinvasion functions.

Application of an immunoaffinity-based preconcentration method for mass spectrometric analysis of the O-chain polysaccharide of Aeromonas salmonicida from in vitro- and in vivo-grown cells.

In this study, application of magnetic beads (Dynabeads) coated with Aeromonas salmonicida lipopolysaccharide-specific polyclonal antisera to MS-based characterization of bacterial lipopolysaccharides has been evaluated. The results showed that the affinity-based preconcentration strategy resulted in at least a 100-fold increase in the detection of sensitivity, affording direct capillary electrophoresis (CE)-MS analysis of A. salmonicida lipopolysaccharide O-chain polysaccharide from in vitro-cultured cells.

Reciprocal expression of integration host factor and HU in the developmental cycle and infectivity of Legionella pneumophila.

Legionella pneumophila is an intracellular parasite of protozoa that differentiates late in infection into metabolically dormant cysts that are highly infectious. Regulation of this process is poorly understood. Here we report that the small DNA binding regulatory proteins integration host factor (IHF) and HU are reciprocally expressed over the developmental cycle, with HU expressed during exponential phase and IHF expressed postexponentially.