scholarly journals Transcriptional Regulation of the virR Operon of the Intracellular Pathogen Rhodococcus equi

2007 ◽  
Vol 189 (14) ◽  
pp. 5082-5089 ◽  
Author(s):  
Gavin A. Byrne ◽  
Dean A. Russell ◽  
Xiaoxiao Chen ◽  
Wim G. Meijer
Keyword(s):  
Response Regulator ◽  
Sequence Similarity ◽  
High Growth ◽  
Rhodococcus Equi ◽  
Low Ph ◽  
Initiation Codon ◽  
Virulence Plasmid ◽  
Intracellular Pathogen ◽  
Share Sequence Similarity ◽  
A Site

ABSTRACT The virR operon, located on the virulence plasmid of the intracellular pathogen Rhodococcus equi, contains five genes, two of which (virR and orf8) encode transcriptional regulators. The first gene of the operon (virR), encoding a LysR-type transcriptional regulator, is transcribed at a constitutive low level, whereas the four downstream genes are induced by low pH and high growth temperature. Differential regulation of the virR operon genes could not be explained by differential mRNA stability, as there were no major differences in mRNA half-lives of the transcripts representing each of the five genes within the virR operon. Transcription of virR is driven by the P virR promoter, with a transcription start site 53 bp upstream of the virR initiation codon. The four genes downstream of virR are transcribed from P virR and from a second promoter, P orf5 , located 585 bp downstream of the virR initiation codon. VirR binds to a site overlapping the initiation codon of virR, resulting in negative autoregulation of the virR gene, explaining its low constitutive transcription level. The P orf5 promoter is induced by high temperature and low pH, thus explaining the observed differential gene expression of the virR operon. VirR has a positive effect on P orf5 activity, whereas the response regulator encoded by orf8 is not involved in regulating transcription of the virR operon. The P virR promoter is strikingly similar to those recognized by the principal sigma factors of Streptomyces and Mycobacterium, whereas the P orf5 promoter does not share sequence similarity with P virR . This suggests that P orf5 is recognized by an alternative sigma factor.

scholarly journals The LysR-Type Transcriptional Regulator VirR Is Required for Expression of the Virulence Gene vapA of Rhodococcus equi ATCC 33701

2004 ◽  
Vol 186 (17) ◽  
pp. 5576-5584 ◽  
Author(s):  
Dean A. Russell ◽  
Gavin A. Byrne ◽  
Enda P. O'Connell ◽  
Clara A. Boland ◽  
Wim G. Meijer
Keyword(s):  
Response Regulator ◽  
Transcriptional Start Site ◽  
Virulence Gene ◽  
Transcriptional Regulator ◽  
Rhodococcus Equi ◽  
Virulence Plasmid ◽  
Binding Studies ◽  
Nickel Affinity Chromatography ◽  
Virulence Protein ◽  
Dna Binding Studies

ABSTRACT The virulence of the intracellular pathogen Rhodococcus equi in foals is dependent on the presence of an 81-kb virulence plasmid encoding the virulence protein VapA. Expression of this protein is induced by exposure to oxidative stress, high temperatures, and low pHs, which reflect the conditions encountered by R. equi when it enters the host environment. The aim of this study was to determine whether the LysR-type transcriptional regulator VirR, which is encoded by the virulence plasmid, is required for the expression of vapA. It was shown that the virR gene is cotranscribed with four downstream genes, one of which encodes a two-component response regulator. The expression of VapA, as monitored by Western blotting, was completely dependent on the presence of virR. Maximal expression was observed when vapA was present together with the complete virR operon, suggesting that at least one of the virR operon genes, in addition to virR, is required for the expression of vapA to wild-type levels. The transcriptional start site of vapA was determined to be a cytidine located 226 bp upstream from the vapA initiation codon. His-tagged VirR protein was expressed in Escherichia coli and purified by nickel affinity chromatography. DNA binding studies showed that purified VirR binds to a DNA fragment containing the vapA promoter. We therefore conclude that VirR is required for the activation of vapA transcription.

Activation of hilA expression at low pH requires the signal sensor CpxA, but not the cognate response regulator CpxR, in Salmonella enterica serovar Typhimurium

Microbiology ◽  
2003 ◽  
Vol 149 (10) ◽  
pp. 2809-2817 ◽  
Author(s):  
Shu-ichi Nakayama ◽  
Akira Kushiro ◽  
Takashi Asahara ◽  
Ryu-ichiro Tanaka ◽  
Lan Hu ◽  
...  
Keyword(s):  
Mutant Strain ◽  
Cell Invasion ◽  
Salmonella Enterica ◽  
Response Regulator ◽  
Low Ph ◽  
Apparent Difference ◽  
Virulence Determinants ◽  
Serovar Typhimurium ◽  
Cognate Response Regulator ◽  
Invasion Capacity

A two-component regulatory system, cpxR–cpxA, plays an important role in the pH-dependent regulation of virF, a global activator for virulence determinants including invasion genes, in Shigella sonnei. The authors examined whether the cpxR–cpxA homologues have some function in the expression of Salmonella enterica serovar Typhimurium invasion genes via the regulation of hilA, an activator for these genes. In a Salmonella cpxA mutant, the hilA expression level was reduced to less than 10 % of that in the parent strain at pH 6·0. This mutant strain also showed undetectable synthesis of an invasion gene product, SipC, at pH 6·0 and reduced cell invasion capacity – as low as 20 % of that of the parent. In this mutant, the reduction in hilA expression was much less marked at pH 8·0 than at pH 6·0 – no less than 50 % of that in the parent, and no significant reduction was observed in either SipC synthesis or cell invasion rate, compared to the parent. Unexpectedly, a Salmonella cpxR mutant strain and the parent showed no apparent difference in all three characteristics described above at either pH. These results indicate that in Salmonella, the sensor kinase CpxA activates hilA, and consequently, invasion genes and cell invasion capacity at pH 6·0. At pH 8·0, however, CpxA does not seem to have a large role in activation of these factors. Further, the results show that this CpxA-mediated activation does not require its putative cognate response regulator, CpxR. This suggests that CpxA may interact with regulator(s) other than CpxR to achieve activation at low pH.

Two new variants of the Rhodococcus equi virulence plasmid, 90 kb type III and type IV, recovered from a foal in Japan

2001 ◽  
Vol 82 (4) ◽  
pp. 373-381 ◽  
Author(s):  
Shinji Takai ◽  
Norihisa Murata ◽  
Ryudai Kudo ◽  
Nobuhiro Narematsu ◽  
Tsutomu Kakuda ◽  
...  
Keyword(s):  
Rhodococcus Equi ◽  
Virulence Plasmid ◽  
Type Iii ◽  
Type Iv

scholarly journals Molecular Epidemiology of Rhodococcus equi Based on traA, vapA, and vapB Virulence Plasmid Markers

2007 ◽  
Vol 196 (5) ◽  
pp. 763-769 ◽  
Author(s):  
Alain A Ocampo-Sosa ◽  
Deborah A Lewis ◽  
Jesús Navas ◽  
Frances Quigley ◽  
Raquel Callejo ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Molecular Typing ◽  
Rhodococcus Equi ◽  
Virulence Plasmid ◽  
Chain Reaction ◽  
Gene Markers ◽  
Typing System ◽  
New Type ◽  
Polymerase Chain ◽  
Related Plasmid

AbstractMolecular typing of the actinomycete Rhodococcus equi is insufficiently developed, and little is known about the epidemiology and transmission of this multihost pathogen. We report a simple, reliable polymerase chain reaction typing system for R. equi based on 3 plasmid gene markers: traA from the conserved conjugal transfer machinery and vapA and vapB, found in 2 different plasmid subpopulations. This “TRAVAP” typing scheme classifies R. equi into 4 categories: traA+/vapA+B−, traA+/vapA−B+, traA+/vapAB−, and traA−/vapAB− (plasmidless). A TRAVAP survey of 215 R. equi strains confirmed the strong link between vapA (traA+/vapA+B− plasmids) and horse isolates and revealed other host-related plasmid associations: between traA+/vapA−B+ and pigs and between traA+/vapAB−—a new type of R. equi plasmid—and cattle. Plasmidless strains were more frequent among isolates from nonpathological specimens. All plasmid categories were common in human isolates, which possibly reflects the predominantly opportunistic nature of R. equi infection in this host and a zoonotic origin.

scholarly journals Clearance of Virulent but Not Avirulent Rhodococcus equi from the Lungs of Adult Horses Is Associated with Intracytoplasmic Gamma Interferon Production by CD4+ and CD8+ T Lymphocytes

2003 ◽  
Vol 10 (2) ◽  
pp. 208-215 ◽  
Author(s):  
Stephen A. Hines ◽  
Diana M. Stone ◽  
Melissa T. Hines ◽  
Debby C. Alperin ◽  
Donald P. Knowles ◽  
...  
Keyword(s):  
T Lymphocytes ◽  
Virulent Strain ◽  
Lavage Fluid ◽  
Rhodococcus Equi ◽  
Gamma Interferon ◽  
Effective Vaccine ◽  
Virulence Plasmid ◽  
Flow Cytometric Method ◽  
Ifn Γ ◽  
Rhodococcal Pneumonia

ABSTRACT Rhodococcus equi is a gram-positive bacterium that infects alveolar macrophages and causes rhodococcal pneumonia in horses and humans. The virulence plasmid of R. equi appears to be required for both pathogenicity in the horse and the induction of protective immunity. An understanding of the mechanisms by which virulent R. equi circumvents protective host responses and by which bacteria are ultimately cleared is important for development of an effective vaccine. Six adult horses were challenged with either virulent R. equi or an avirulent, plasmid-cured derivative. By using a flow cytometric method for intracytoplasmic detection of gamma interferon (IFN-γ) in equine bronchoalveolar lavage fluid (BALF) cells, clearance of the virulent strain was shown to be associated with increased numbers of pulmonary CD4+ and CD8+ T lymphocytes producing IFN-γ. There was no change in IFN-γ-positive cells in peripheral blood, suggesting that a type 1 recall response at the site of challenge was protective. The plasmid-cured strain of R. equi was cleared in horses without a significant increase in IFN-γ-producing T lymphocytes in BALF. In contrast to these data, a previous report in foals suggested an immunomodulating role for R. equi virulence plasmid-encoded products in downregulating IFN-γ expression by equine CD4+ T lymphocytes. Intracytoplasmic detection of IFN-γ provides a method to better determine whether modulation of macrophage-activating cytokines by virulent strains occurs uniquely in neonates and contributes to their susceptibility to rhodococcal pneumonia.

scholarly journals Involvement of the HP0165-HP0166 Two-Component System in Expression of Some Acidic-pH-Upregulated Genes of Helicobacter pylori

2006 ◽  
Vol 188 (5) ◽  
pp. 1750-1761 ◽  
Author(s):  
Yi Wen ◽  
Jing Feng ◽  
David R. Scott ◽  
Elizabeth A. Marcus ◽  
George Sachs
Keyword(s):  
Helicobacter Pylori ◽  
Sequence Similarity ◽  
Low Ph ◽  
Ph Sensitive ◽  
Cytoplasmic Ph ◽  
Two Component System ◽  
Mobility Shift ◽  
Component System ◽  
Two Component

ABSTRACT About 200 genes of the gastric pathogen Helicobacter pylori increase expression at medium pHs of 6.2, 5.5, and 4.5, an increase that is abolished or much reduced by the buffering action of urease. Genes up-regulated by a low pH include the two-component system HP0165-HP0166, suggesting a role in the regulation of some of the pH-sensitive genes. To identify targets of HP0165-HP0166, the promoter regions of genes up-regulated by a low pH were grouped based on sequence similarity. Probes for promoter sequences representing each group were subjected to electrophoretic mobility shift assays (EMSA) with recombinant HP0166-His6 or a mutated response regulator, HP0166-D52N-His6, that can specifically determine the role of phosphorylation of HP0166 in binding (including a control EMSA with in-vitro-phosphorylated HP0166-His6). Nineteen of 45 promoter-regulatory regions were found to interact with HP0166-His6. Seven promoters for genes encoding α-carbonic anhydrase, omp11, fecD, lpp20, hypA, and two with unknown function (pHP1397-1396 and pHP0654-0675) were clustered in gene group A, which may respond to changes in the periplasmic pH at a constant cytoplasmic pH and showed phosphorylation-dependent binding in EMSA with HP0166-D52N-His6. Twelve promoters were clustered in groups B and C whose up-regulation likely also depends on a reduction of the cytoplasmic pH at a medium pH of 5.5 or 4.5. Most of the target promoters in groups B and C showed phosphorylation-dependent binding with HP0166-D52N-His6, but promoters for ompR (pHP0166-0162), pHP0682-0681, and pHP1288-1289 showed phosphorylation-independent binding. These findings, combined with DNase I footprinting, suggest that HP0165-0166 is an acid-responsive signaling system affecting the expression of pH-sensitive genes. Regulation of these genes responds either to a decrease in the periplasmic pH alone (HP0165 dependent) or also to a decrease in the cytoplasmic pH (HP0165 independent).

scholarly journals A novel method to generate unmarked gene deletions in the intracellular pathogen Rhodococcus equi using 5-fluorocytosine conditional lethality

2008 ◽  
Vol 36 (22) ◽  
pp. e151-e151 ◽  
Author(s):  
R. van der Geize ◽  
W. de Jong ◽  
G. I. Hessels ◽  
A. W. F. Grommen ◽  
A. A. C. Jacobs ◽  
...  
Keyword(s):  
Rhodococcus Equi ◽  
Intracellular Pathogen ◽  
Gene Deletions ◽  
Novel Method

Safety and immunogenicity of a live-attenuated auxotrophic candidate vaccine against the intracellular pathogen Rhodococcus equi

Vaccine ◽  
2008 ◽  
Vol 26 (7) ◽  
pp. 998-1009 ◽  
Author(s):  
A.M. Lopez ◽  
H.G.G. Townsend ◽  
A.L. Allen ◽  
M.K. Hondalus
Keyword(s):  
Rhodococcus Equi ◽  
Candidate Vaccine ◽  
Intracellular Pathogen

scholarly journals Rhodococcus equi Infections in Dogs

2016 ◽  
Vol 54 (1) ◽  
pp. 159-163 ◽  
Author(s):  
L. K. Bryan ◽  
S. D. Clark ◽  
J. Díaz-Delgado ◽  
S. D. Lawhon ◽  
J. F. Edwards
Keyword(s):  
At Risk ◽  
Polymerase Chain Reaction ◽  
Companion Animals ◽  
Immunosuppressive Drugs ◽  
Rhodococcus Equi ◽  
Virulence Plasmid ◽  
The Novel ◽  
Chain Reaction ◽  
Polymerase Chain

Five cases of Rhodococcus equi infection in dogs were identified from 2003 to 2014. Three of the dogs had severe, internal lesions attributable to R. equi that have not been previously described: endophthalmitis, endocarditis, and suppurative pleuropneumonia. Isolates from 4 of the dogs were analyzed by polymerase chain reaction for Rhodococcus virulence-associated plasmid (vap) genes. One isolate was vapA-positive, 2 lacked a virulence plasmid, and 1 carried the novel vapN-associated plasmid (pVAPN) recently characterized in bovine isolates. The pVAPN plasmid has not been described in isolates cultured from companion animals. Four of the dogs either were receiving immunosuppressive drugs or had endocrinopathies. R. equi has the potential to cause significant infections in dogs, and immunocompromised animals should be considered at risk for infection.

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