scholarly journals Deletion of the Ferric Uptake Regulator Fur Impairs the In Vitro Growth and Virulence of Actinobacillus pleuropneumoniae

2005 ◽  
Vol 73 (6) ◽  
pp. 3740-3744 ◽  
Author(s):  
Ilse Jacobsen ◽  
Jörg Gerstenberger ◽  
Achim D. Gruber ◽  
Janine T. Bossé ◽  
Paul R. Langford ◽  
...  
Keyword(s):  
Deletion Mutant ◽  
Constitutive Expression ◽  
Actinobacillus Pleuropneumoniae ◽  
Infection Model ◽  
Ferric Uptake Regulator ◽  
In Vitro Growth ◽  
Transferrin Binding Proteins ◽  
Aerosol Infection

ABSTRACT In order to investigate the role of the ferric uptake regulator Fur in the porcine lung pathogen Actinobacillus pleuropneumoniae, we constructed an isogenic in-frame deletion mutant, A. pleuropneumoniae Δfur. This mutant showed constitutive expression of transferrin-binding proteins, growth deficiencies in vitro, and reduced virulence in an aerosol infection model.

scholarly journals Activity of the Pore-Forming Virulence Factor Listeriolysin O Is Reversibly Inhibited by Naturally Occurring S-Glutathionylation

2017 ◽  
Vol 85 (4) ◽  
Author(s):  
Jonathan L. Portman ◽  
Qiongying Huang ◽  
Michelle L. Reniere ◽  
Anthony T. Iavarone ◽  
Daniel A. Portnoy
Keyword(s):  
Protein Function ◽  
Inhibitory Effect ◽  
Cysteine Residue ◽  
Infection Model ◽  
Listeriolysin O ◽  
Content Type ◽  
Pore Forming Proteins

ABSTRACT Cholesterol-dependent cytolysins (CDCs) represent a family of homologous pore-forming proteins secreted by many Gram-positive bacterial pathogens. CDCs mediate membrane binding partly through a conserved C-terminal undecapeptide, which contains a single cysteine residue. While mutational changes to other residues in the undecapeptide typically have severe effects, mutation of the cysteine residue to alanine has minor effects on overall protein function. Thus, the role of this highly conserved reactive cysteine residue remains largely unknown. We report here that the CDC listeriolysin O (LLO), secreted by the facultative intracellular pathogen Listeria monocytogenes, was posttranslationally modified by S-glutathionylation at this conserved cysteine residue and that either endogenously synthesized or exogenously added glutathione was sufficient to form this modification. When recapitulated with purified protein in vitro, this modification completely ablated the activity of LLO, and this inhibitory effect was fully reversible by treatment with reducing agents. A cysteine-to-alanine mutation in LLO rendered the protein completely resistant to inactivation by S-glutathionylation, and a mutant expressing this mutation retained full hemolytic activity. A mutant strain of L. monocytogenes expressing the cysteine-to-alanine variant of LLO was able to infect and replicate within bone marrow-derived macrophages indistinguishably from the wild type in vitro, yet it was attenuated 4- to 6-fold in a competitive murine infection model in vivo. This study suggests that S-glutathionylation may represent a mechanism by which CDC-family proteins are posttranslationally modified and regulated and help explain an evolutionary pressure to retain the highly conserved undecapeptide cysteine.

scholarly journals Enzymes Involved in Anaerobic Respiration Appear To Play a Role in Actinobacillus pleuropneumoniae Virulence

2005 ◽  
Vol 73 (1) ◽  
pp. 226-234 ◽  
Author(s):  
Ilse Jacobsen ◽  
Isabel Hennig-Pauka ◽  
Nina Baltes ◽  
Matthias Trost ◽  
Gerald-F. Gerlach
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Anaerobic Respiration ◽  
Clinical Score ◽  
Lavage Fluid ◽  
Actinobacillus Pleuropneumoniae ◽  
Lung Lesion ◽  
Infection Model ◽  
Binding Motif ◽  
Anaerobic Conditions

ABSTRACT Actinobacillus pleuropneumoniae, the etiological agent of porcine pleuropneumonia, is able to survive on respiratory epithelia, in tonsils, and in the anaerobic environment of encapsulated sequesters. It was previously demonstrated that a deletion of the anaerobic dimethyl sulfoxide reductase gene (dmsA) results in attenuation in acute disease (N. Baltes, S. Kyaw, I. Hennig-Pauka, and G. F. Gerlach, Infect. Immun. 71:6784-6792, 2003). In the present study, using two-dimensional polyacrylamide gel electrophoresis and quadrupole time-of-flight mass spectrometry, we identified an aspartate ammonia-lyase (AspA) which is upregulated upon induction with bronchoalveolar lavage fluid (BALF). This enzyme is involved in the production of fumarate, an alternative electron acceptor under anaerobic conditions. The coding gene (aspA) was cloned and shown to be present in all A. pleuropneumoniae serotype reference strains. The transcriptional start point was identified downstream of a putative FNR binding motif, and BALF-dependent activation of aspA was confirmed by construction of an isogenic A. pleuropneumoniae mutant carrying a chromosomal aspA::luxAB transcriptional fusion. Two aspA deletion mutants, A. pleuropneumoniae ΔaspA and A. pleuropneumoniae ΔaspAΔdmsA, were constructed, both showing reduced growth under anaerobic conditions in vitro. Pigs challenged with either of the two mutants in an aerosol infection model showed a lower lung lesion score than that of the A. pleuropneumoniae wild-type (wt) controls. Pigs challenged with A. pleuropneumoniae ΔaspAΔdmsA had a significantly lower clinical score, and this mutant was rarely reisolated from unaltered lung tissue; in contrast, A. pleuropneumoniae ΔaspA and the A. pleuropneumoniae wt were consistently reisolated in high numbers. These results suggest that enzymes involved in anaerobic respiration are necessary for the pathogen's ability to persist on respiratory tract epithelium and play an important role in A. pleuropneumoniae pathogenesis.

scholarly journals Enhancement of In Vitro Growth of Pathogenic Bacteria by Norepinephrine: Importance of Inoculum Density and Role of Transferrin

2006 ◽  
Vol 72 (7) ◽  
pp. 5097-5099 ◽  
Author(s):  
Phyllis M. O'Donnell ◽  
Hernan Aviles ◽  
Mark Lyte ◽  
Gerald Sonnenfeld
Keyword(s):  
Bacterial Growth ◽  
Pathogenic Bacteria ◽  
Inoculum Density ◽  
In Vitro Growth ◽  
Stress Hormone ◽  
Serum Free ◽  
Free Media

ABSTRACT Norepinephrine is a stress hormone that enhances bacterial growth. We examined the effects of a small inoculum on the norepinephrine-induced growth of species previously reported to be unaffected by norepinephrine. The results indicated that a reduced inoculum density is essential for observing norepinephrine-induced effects. Additional studies using serum-free media suggested that transferrin plays a role in norepinephrine-induced growth.

scholarly journals Assessing the role of deoD gene in Mycobacterium tuberculosis in vitro growth and macrophage infection

2018 ◽  
Vol 119 ◽  
pp. 60-64 ◽  
Author(s):  
Pedro Ferrari Dalberto ◽  
Valnês Rodrigues-Junior ◽  
Virginia Carla Almeida Falcão ◽  
Antônio Frederico Michel Pinto ◽  
Bruno Lopes Abbadi ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
In Vitro Growth ◽  
Macrophage Infection

Challenges with Wound Infection Models in Drug Development

2020 ◽  
Vol 21 (13) ◽  
pp. 1301-1312 ◽  
Author(s):  
Sandeep K. Shukla ◽  
Ajay K. Sharma ◽  
Vanya Gupta ◽  
Aman Kalonia ◽  
Priyanka Shaw
Keyword(s):  
Wound Healing ◽  
Wound Infection ◽  
Chronic Wound ◽  
Wound Care ◽  
Infection Model ◽  
In Vivo Models ◽  
Infection Models

: Wound research is an evolving science trying to unfold the complex untold mechanisms behind the wound healing cascade. In particular, interest is growing regarding the role of microorganisms in both acute and chronic wound healing. Microbial burden plays an important role in the persistence of chronic wounds, ultimately resulting in delayed wound healing. It is therefore important for clinicians to understand the evolution of infection science and its various etiologies. Therefore, to understand the role of bacterial biofilm in chronic wound pathogenesis, various in vitro and in vivo models are required to investigate biofilms in wound-like settings. Infection models should be refined comprising an important signet of biofilms. These models are eminent for translational research to obtain data for designing an improved wound care formulation. However, all the existing models possess limitations and do not fit properly in the model frame for developing wound care agents. Among various impediments, one of the major drawbacks of such models is that the wound they possess does not mimic the wound a human develops. Therefore, a novel wound infection model is required which can imitate the human wounds. : This review article mainly discusses various in vitro and in vivo models showing microbial colonization, their advantages and challenges. Apart from these models, there are also present ex vivo wound infection models, but this review mainly focused on various in vitro and in vivo models available for studying wound infection in controlled conditions. This information might be useful in designing an ideal wound infection model for developing an effective wound healing formulation.

scholarly journals Role of Protein Kinase G in Growth and Glutamine Metabolism of Mycobacterium bovis BCG

2005 ◽  
Vol 187 (16) ◽  
pp. 5852-5856 ◽  
Author(s):  
Liem Nguyen ◽  
Anne Walburger ◽  
Edith Houben ◽  
Anil Koul ◽  
Stefan Muller ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Mycobacterium Bovis ◽  
Glutamine Metabolism ◽  
Protein Kinase G ◽  
In Vitro Growth ◽  
Wild Type ◽  
Putative Operon ◽  
Glutamine Uptake

ABSTRACT The survival of pathogenic mycobacteria in macrophages requires the eukaryotic enzyme-like serine/threonine protein kinase G. This kinase with unknown specificity is secreted into the cytosol of infected macrophages and inhibits phagosome-lysosome fusion. The pknG gene is the terminal gene in a putative operon containing glnH, encoding a protein potentially involved in glutamine uptake. Here, we report that the deletion of pknG did not affect either glutamine uptake or intracellular glutamine concentrations. In vitro growth of Mycobacterium bovis BCG lacking pknG was identical to that of the wild type. We conclude that in M. bovis BCG, glutamine metabolism is not regulated by protein kinase G.

scholarly journals The galU Gene of Pseudomonas aeruginosa Is Required for Corneal Infection and Efficient Systemic Spread following Pneumonia but Not for Infection Confined to the Lung

2004 ◽  
Vol 72 (7) ◽  
pp. 4224-4232 ◽  
Author(s):  
Gregory P. Priebe ◽  
Charles R. Dean ◽  
Tanweer Zaidi ◽  
Gloria J. Meluleni ◽  
Fadie T. Coleman ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Severe Pneumonia ◽  
Infection Model ◽  
Eye Infections ◽  
Corneal Infection ◽  
Systemic Spread ◽  
Type Strains ◽  
Lethal Doses

ABSTRACT Acute pneumonias and corneal infections due to Pseudomonas aeruginosa are typically caused by lipopolysaccharide (LPS)-smooth strains. In cystic fibrosis patients, however, LPS-rough strains of P. aeruginosa, which lack O antigen, can survive in the lung and cause chronic infection. It is not clear whether an LPS-rough phenotype affects cytotoxicity related to the type III secretion system (TTSS). We previously reported that interruption of the galU gene in P. aeruginosa results in production of a rough LPS and truncated LPS core. Here we evaluated the role of the galU gene in the pathogenesis of murine lung and eye infections and in cytotoxicity due to the TTSS effector ExoU. We studied galU mutants of strain PAO1, of its cytotoxic variant expressing ExoU from a plasmid, and of the inherently cytotoxic strain PA103. The galU mutants were more serum sensitive than the parental strains but remained cytotoxic in vitro. In a corneal infection model, the galU mutants were significantly attenuated. In an acute pneumonia model, the 50% lethal doses of the galU mutants were higher than those of the corresponding wild-type strains, yet these mutants could cause mortality and severe pneumonia, as judged by histology, even with minimal systemic spread. These findings suggest that the galU gene is required for corneal infection and for efficient systemic spread following lung infection but is not required for infection confined to the lung. Host defenses in the lung appear to be insufficient to control infection with LPS-rough P. aeruginosa when local bacterial levels are high.

Sign in / Sign up

Export Citation Format

Share Document