scholarly journals Quorum-Sensing and BvrR/BvrS Regulation, the Type IV Secretion System, Cyclic Glucans, and BacA in the Virulence of Brucella ovis: Similarities to and Differences from Smooth Brucellae

2012 ◽  
Vol 80 (5) ◽  
pp. 1783-1793 ◽  
Author(s):  
Ana I. Martín-Martín ◽  
Pilar Sancho ◽  
María Jesús de Miguel ◽  
Luis Fernández-Lago ◽  
Nieves Vizcaíno
Keyword(s):  
Quorum Sensing ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Multiplication Rate ◽  
Content Type ◽  
Murine Macrophages ◽  
Brucella Ovis ◽  
Type Iv

ABSTRACTBrucella ovisis a rough bacterium—lacking O-polysaccharide chains in the lipopolysaccharide—that is virulent in its natural host and whose virulence mechanisms remain almost unexplored. In a search for additional traits that distinguishB. ovisfrom smoothBrucella, which require O-polysaccharide chains for virulence, we have analyzed the significance inB. ovisof the main virulence factors described for smoothBrucella. Attempts to obtain strains of virulentB. ovisstrain PA that are mutated in the BvrR/BvrS two-component regulatory system were unsuccessful, suggesting the requirement of that system forin vitrosurvival, while the inactivation ofbacA—in contrast to the results seen with smoothBrucella—did not affect splenic colonization in mice or behavior in J774.A1 murine macrophages. Defects in the synthesis of cyclic ß-1,2 glucans reduced the uptake ofB. ovisPA in macrophages and, although the intracellular multiplication rate was unaffected, led to attenuation in mice. Growth of strains with mutations in the type IV secretion system (encoded by thevirBoperon) and the quorum-sensing-related regulator VjbR was severely attenuated in the mouse model, and although the mutant strains internalized like the parental strain in J774.A1 murine macrophages, they were impaired for intracellular replication. As described forB. melitensis, VjbR regulates the transcription of thevirBoperon positively, and theN-dodecanoyl-dl-homoserine lactone (C12-HSL) autoinducer abrogates this effect. In contrast, no apparent VjbR-mediated regulation of thefliFflagellar gene was observed inB. ovis, probably due to the two deletions detected upstream offliF. These results, together with others reported in the text, point to similarities between rough virulentB. ovisand smoothBrucellaspecies as regards virulence but also reveal distinctive traits that could be related to the particular pathogenicity and host tropism characteristics ofB. ovis.

scholarly journals Identification of ElpA, a Coxiella burnetii Pathotype-Specific Dot/Icm Type IV Secretion System Substrate

2015 ◽  
Vol 83 (3) ◽  
pp. 1190-1198 ◽  
Author(s):  
Joseph G. Graham ◽  
Caylin G. Winchell ◽  
Uma M. Sharma ◽  
Daniel E. Voth
Keyword(s):  
Coxiella Burnetii ◽  
Q Fever ◽  
Parasitophorous Vacuole ◽  
Secretion System ◽  
Terminal Region ◽  
Effector Proteins ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Content Type ◽  
Type Iv

Coxiella burnetiicauses human Q fever, a zoonotic disease that presents with acute flu-like symptoms and can result in chronic life-threatening endocarditis. In human alveolar macrophages,C. burnetiiuses a Dot/Icm type IV secretion system (T4SS) to generate a phagolysosome-like parasitophorous vacuole (PV) in which to replicate. The T4SS translocates effector proteins, or substrates, into the host cytosol, where they mediate critical cellular events, including interaction with autophagosomes, PV formation, and prevention of apoptosis. Over 100C. burnetiiDot/Icm substrates have been identified, but the function of most remains undefined. Here, we identified a novel Dot/Icm substrate-encoding open reading frame (CbuD1884) present in allC. burnetiiisolates except the Nine Mile reference isolate, where the gene is disrupted by a frameshift mutation, resulting in a pseudogene. The CbuD1884 protein contains two transmembrane helices (TMHs) and a coiled-coil domain predicted to mediate protein-protein interactions. The C-terminal region of the protein contains a predicted Dot/Icm translocation signal and was secreted by the T4SS, while the N-terminal portion of the protein was not secreted. When ectopically expressed in eukaryotic cells, the TMH-containing N-terminal region of the CbuD1884 protein trafficked to the endoplasmic reticulum (ER), with the C terminus dispersed nonspecifically in the host cytoplasm. This new Dot/Icm substrate is now termed ElpA (ER-localizingproteinA). Full-length ElpA triggered substantial disruption of ER structure and host cell secretory transport. These results suggest that ElpA is a pathotype-specific T4SS effector that influences ER function duringC. burnetiiinfection.

scholarly journals Cytotoxicity in Macrophages Infected with Rough Brucella Mutants Is Type IV Secretion System Dependent

2007 ◽  
Vol 76 (1) ◽  
pp. 30-37 ◽  
Author(s):  
Jianwu Pei ◽  
Qingmin Wu ◽  
Melissa Kahl-McDonagh ◽  
Thomas A. Ficht
Keyword(s):  
Cell Death ◽  
Brucella Melitensis ◽  
Regulatory Element ◽  
Random Mutagenesis ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Type Iv ◽  
Mutant Bank

ABSTRACT Smooth Brucella spp. inhibit macrophage apoptosis, whereas rough Brucella mutants induce macrophage oncotic and necrotic cell death. However, the mechanisms and genes responsible for Brucella cytotoxicity have not been identified. In the current study, a random mutagenesis approach was used to create a mutant bank consisting of 11,354 mutants by mariner transposon mutagenesis using Brucella melitensis rough mutant 16MΔmanBA as the parental strain. Subsequent screening identified 56 mutants (0.49% of the mutant bank) that failed to cause macrophage cell death (release of 10% or less of the lactate dehydrogenase). The absence of cytotoxicity during infection with these mutants was independent of demonstrable defects in in vitro bacterial growth or uptake and survival in macrophages. Interrupted genes in 51 mutants were identified by DNA sequence analysis, and the mutations included interruptions in virB encoding the type IV secretion system (T4SS) (n = 36) and in vjbR encoding a LuxR-like regulatory element previously shown to be required for virB expression (n = 3), as well as additional mutations (n = 12), one of which also has predicted roles in virB expression. These results suggest that the T4SS is associated with Brucella cytotoxicity in macrophages. To verify this, deletion mutants were constructed in B. melitensis 16M by removing genes encoding phosphomannomutase/phosphomannoisomerase (ΔmanBA) and the T4SS (ΔvirB). As predicted, deletion of virB from 16MΔmanBA and 16M resulted in a complete loss of cytotoxicity in rough strains, as well as the low level cytotoxicity observed with smooth strains at extreme multiplicities of infection (>1,000). Taken together, these results demonstrate that Brucella cytotoxicity in macrophages is T4SS dependent.

scholarly journals Membrane and Core Periplasmic Agrobacterium tumefaciens Virulence Type IV Secretion System Components Localize to Multiple Sites around the Bacterial Perimeter during Lateral Attachment to Plant Cells

mBio ◽  
2011 ◽  
Vol 2 (6) ◽  
Author(s):  
Julieta Aguilar ◽  
Todd A. Cameron ◽  
John Zupan ◽  
Patricia Zambryski
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Plant Cells ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Host Cells ◽  
Type Iv Secretion System ◽  
Secretion Systems ◽  
Content Type ◽  
Protein Substrates ◽  
Type Iv

ABSTRACTType IV secretion systems (T4SS) transfer DNA and/or proteins into recipient cells. Here we performed immunofluorescence deconvolution microscopy to localize the assembled T4SS by detection of its native components VirB1, VirB2, VirB4, VirB5, VirB7, VirB8, VirB9, VirB10, and VirB11 in the C58 nopaline strain ofAgrobacterium tumefaciens, following induction of virulence (vir) gene expression. These different proteins represent T4SS components spanning the inner membrane, periplasm, or outer membrane. Native VirB2, VirB5, VirB7, and VirB8 were also localized in theA. tumefaciensoctopine strain A348. Quantitative analyses of the localization of all the above Vir proteins in nopaline and octopine strains revealed multiple foci in single optical sections in over 80% and 70% of the bacterial cells, respectively. Green fluorescent protein (GFP)-VirB8 expression followingvirinduction was used to monitor bacterial binding to live host plant cells; bacteria bind predominantly along their lengths, with few bacteria binding via their poles or subpoles.vir-induced attachment-defective bacteria or bacteria without the Ti plasmid do not bind to plant cells. These data support a model where multiplevir-T4SS around the perimeter of the bacterium maximize effective contact with the host to facilitate efficient transfer of DNA and protein substrates.IMPORTANCETransfer of DNA and/or proteins to host cells through multiprotein type IV secretion system (T4SS) complexes that span the bacterial cell envelope is critical to bacterial pathogenesis. Early reports suggested that T4SS components localized at the cell poles. Now, higher-resolution deconvolution fluorescence microscopy reveals that all structural components of theAgrobacterium tumefaciens vir-T4SS, as well as its transported protein substrates, localize to multiple foci around the cell perimeter. These results lead to a new model ofA. tumefaciensattachment to a plant cell, whereA. tumefacienstakes advantage of the multiplevir-T4SS along its length to make intimate lateral contact with plant cells and thereby effectively transfer DNA and/or proteins through thevir-T4SS. The T4SS ofA. tumefaciensis among the best-studied T4SS, and the majority of its components are highly conserved in different pathogenic bacterial species. Thus, the results presented can be applied to a broad range of pathogens that utilize T4SS.

The virB-encoded type IV secretion system is critical for establishment of infection and persistence of Brucella ovis infection in mice

2012 ◽  
Vol 159 (1-2) ◽  
pp. 130-140 ◽  
Author(s):  
Joicy C. Sá ◽  
Teane M.A. Silva ◽  
Érica A. Costa ◽  
Ana P.C. Silva ◽  
Renée M. Tsolis ◽  
...  
Keyword(s):  
Secretion System ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Brucella Ovis ◽  
Type Iv

scholarly journals Putative Quorum-Sensing Regulator BlxR of Brucella melitensis Regulates Virulence Factors Including the Type IV Secretion System and Flagella

2008 ◽  
Vol 190 (9) ◽  
pp. 3274-3282 ◽  
Author(s):  
Amy A. Rambow-Larsen ◽  
Gireesh Rajashekara ◽  
Erik Petersen ◽  
Gary Splitter
Keyword(s):  
Quorum Sensing ◽  
Virulence Factors ◽  
Brucella Melitensis ◽  
Regulatory Protein ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Deletion Strain ◽  
Type Iv Secretion System ◽  
Type Iv ◽  
Systemic Spread

ABSTRACT Brucella melitensis is an intracellular pathogen that establishes a replicative niche within macrophages. While the intracellular lifestyle of Brucella is poorly understood and few virulence factors have been identified, components of a quorum-sensing pathway in Brucella have recently been identified. The LuxR-type regulatory protein, VjbR, and an N-acylhomoserine lactone signaling molecule are both involved in regulating expression of the virB-encoded type IV secretion system. We have identified a second LuxR-type regulatory protein (BlxR) in Brucella. Microarray analysis of a blxR mutant suggests that BlxR regulates the expression of a number of genes, including those encoding the type IV secretion system and flagella. Confirming these results, deletion of blxR in B. melitensis reduced the transcriptional activities of promoters for the virB operon, flagellar genes, and another putative virulence factor gene, bopA. Furthermore, our data suggested that both BlxR and VjbR are positively autoregulated and cross-regulate the expression of each other. The blxR deletion strain exhibited reduced growth in macrophages, similar to that observed for a vjbR deletion strain. However, unlike the vjbR deletion, the blxR deletion did not fully attenuate virulence in mice. More strikingly, bioluminescent imaging revealed that dissemination of the blxR mutant was similar to that of wild-type B. melitensis, while the vjbR mutant was defective for systemic spread in IRF-1−/− mice, suggesting that these regulators are not functionally redundant but that they converge in a common pathway regulating bacterial processes.

scholarly journals Type IV Secretion System Is Not Involved in Infection Process in Citrus

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Tiago Rinaldi Jacob ◽  
Marcelo Luiz de Laia ◽  
Leandro Marcio Moreira ◽  
Janaína Fernandes Gonçalves ◽  
Flavia Maria de Souza Carvalho ◽  
...  
Keyword(s):  
Cell Envelope ◽  
Positive Control ◽  
Secretion System ◽  
Infection Process ◽  
Type Iv Secretion ◽  
Target Cells ◽  
Type Iv Secretion System ◽  
In Planta ◽  
Type Iv

The type IV secretion system (T4SS) is used by Gram-negative bacteria to translocate protein and DNA substrates across the cell envelope and into target cells.Xanthomonas citrisubsp.citricontains two copies of the T4SS, one in the chromosome and the other is plasmid-encoded. To understand the conditions that induce expression of the T4SS inXcc, we analyzed,in vitroandin planta, the expression of 18 ORFs from the T4SS and 7 hypothetical flanking genes by RT-qPCR. As a positive control, we also evaluated the expression of 29 ORFs from the type III secretion system (T3SS), since these genes are known to be expressed during plant infection condition, but not necessarily in standard culture medium. From the 29 T3SS genes analyzed by qPCR, onlyhrpAwas downregulated at 72 h after inoculation. All genes associated with the T4SS were downregulated onCitrusleaves 72 h after inoculation. Our results showed that unlike the T3SS, the T4SS is not induced during the infection process.

scholarly journals Novel Plasmid and Its Variant Harboring both ablaNDM-1Gene and Type IV Secretion System in Clinical Isolates of Acinetobacter lwoffii

2012 ◽  
Vol 56 (4) ◽  
pp. 1698-1702 ◽  
Author(s):  
Hongyan Hu ◽  
Yongfei Hu ◽  
Yuanlong Pan ◽  
Hui Liang ◽  
Haiyan Wang ◽  
...  
Keyword(s):  
Secretion System ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Content Type ◽  
Type Iv ◽  
Insertion Elements ◽  
Complete Sequences ◽  
Large Plasmids ◽  
P Type ◽  
T4ss Gene

ABSTRACTThe spread of theblaNDM-1gene is gaining worldwide attentions. This gene is usually carried by large plasmids and has been discovered in diverse bacteria since it was originally found inKlebsiella pneumoniae. Here we report the complete sequences of ablaNDM-1-bearing plasmid, pNDM-BJ01, and its variant, pNDM-BJ02, isolated from clinicalAcinetobacter lwoffiistrains. The plasmid pNDM-BJ01 is 47.3 kb in size and cannot be classified into any known plasmid incompatibility group, thus representing a novel plasmid with an unknown maintenance mechanism. This plasmid contains both ablaNDM-1gene and a type IV secretion system (T4SS) gene cluster. The T4SS is assigned to the P-type T4SS group, which usually encode a short, rigid pilus, and theblaNDM-1gene is located within a composite transposon flanked by two insertion elements of ISAba125. Plasmid pNDM-BJ02 is nearly identical to pNDM-BJ01 except that one copy of the ISAba125element is missing, and it is therefore regarded as a variant of pNDM-BJ01. Sequence alignment indicated that thisblaNDM-1-containing composite transposon, which can also be captured by other mobile elements, was probably a product of multiple recombination events and can move as a whole by transposition.

scholarly journals A quorum-sensing regulator controls expression of both the type IV secretion system and the flagellar apparatus of Brucella melitensis

2005 ◽  
Vol 7 (8) ◽  
pp. 1151-1161 ◽  
Author(s):  
Rose-May Delrue ◽  
Chantal Deschamps ◽  
Sandrine Leonard ◽  
Caroline Nijskens ◽  
Isabelle Danese ◽  
...  
Keyword(s):  
Quorum Sensing ◽  
Brucella Melitensis ◽  
Flagellar Apparatus ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Type Iv

scholarly journals Brucella melitensis Invades Murine Erythrocytes during Infection

2014 ◽  
Vol 82 (9) ◽  
pp. 3927-3938 ◽  
Author(s):  
Marie-Alice Vitry ◽  
Delphine Hanot Mambres ◽  
Michaël Deghelt ◽  
Katrin Hack ◽  
Arnaud Machelart ◽  
...  
Keyword(s):  
Brucella Melitensis ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Bacterial Cells ◽  
Content Type ◽  
Type Iv ◽  
Transmission Electron ◽  
Time Point

ABSTRACTBrucellaspp. are facultative intracellular Gram-negative coccobacilli responsible for brucellosis, a worldwide zoonosis. We observed thatBrucella melitensisis able to persist for several weeks in the blood of intraperitoneally infected mice and that transferred blood at any time point tested is able to induce infection in naive recipient mice. Bacterial persistence in the blood is dramatically impaired by specific antibodies induced followingBrucellavaccination. In contrast toBartonella, the type IV secretion system and flagellar expression are not critically required for the persistence ofBrucellain blood. ImageStream analysis of blood cells showed that following a brief extracellular phase,Brucellais associated mainly with the erythrocytes. Examination by confocal microscopy and transmission electron microscopy formally demonstrated thatB. melitensisis able to invade erythrocytesin vivo. The bacteria do not seem to multiply in erythrocytes and are found free in the cytoplasm. Our results open up new areas for investigation and should serve in the development of novel strategies for the treatment or prophylaxis of brucellosis. Invasion of erythrocytes could potentially protect the bacterial cells from the host's immune response and hamper antibiotic treatment and suggests possibleBrucellatransmission by bloodsucking insects in nature.

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