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1.
Clinical vaccine immunology ; 17(4): 529-536, apr.2010.
Article in English | Sec. Est. Saúde SP, SESSP-IBPROD, Sec. Est. Saúde SP, SESSP-IBACERVO | ID: biblio-1062098

ABSTRACT

Shiga-like toxin 2 (Stx2)-producing enterohemorrhagic Escherichia coli (referred to as EHEC or STEC) strainsare the primary etiologic agents of hemolytic-uremic syndrome (HUS), which leads to renal failure and highmortality rates. Expression of Stx2 is the most relevant virulence-associated factor of EHEC strains, and toxin neutralization by antigen-specific serum antibodies represents the main target for both preventive and therapeuticanti-HUS approaches. In the present report, we describe two Salmonella enterica serovar Typhimurium aroA vaccine strains expressing a nontoxic plasmid-encoded derivative of Stx2 (Stx2 AB) containing the complete nontoxic A2 subunit and the receptor binding B subunit. The two S. Typhimurium strains differ inthe expression of flagellin, the structural subunit of the flagellar shaft, which exerts strong adjuvant effects. Thevaccine strains expressed Stx2 AB, either cell bound or secreted into the extracellular environment, andshowed enhanced mouse gut colonization and high plasmid stability under both in vitro and in vivo conditions.Oral immunization of mice with three doses of the S. Typhimurium vaccine strains elicited serum anti-Stx2B(IgG) antibodies that neutralized the toxic effects of the native toxin under in vitro conditions (Vero cells) andconferred partial protection under in vivo conditions. No significant differences with respect to gut colonization or the induction of antigen-specific antibody responses were detected in mice vaccinated with flagellated versus nonflagellated bacterial strains. The present results indicate that expression of Stx2 AB by attenuated S. Typhimurium strains is an alternative vaccine approach for HUS control, but additional improvements in the immunogenicity of Stx2 toxoids are still required.


Subject(s)
Humans , Mice , Vero Cells/microbiology , Enteropathogenic Escherichia coli/immunology , Vaccines , Escherichia coli/growth & development , Escherichia coli/isolation & purification , Escherichia coli/metabolism , Salmonella enterica
2.
Clin Vaccine Immunol ; 17(4): 529-36, 2010 Apr.
Article in English | MEDLINE | ID: mdl-20147499

ABSTRACT

Shiga-like toxin 2 (Stx2)-producing enterohemorrhagic Escherichia coli (referred to as EHEC or STEC) strains are the primary etiologic agents of hemolytic-uremic syndrome (HUS), which leads to renal failure and high mortality rates. Expression of Stx2 is the most relevant virulence-associated factor of EHEC strains, and toxin neutralization by antigen-specific serum antibodies represents the main target for both preventive and therapeutic anti-HUS approaches. In the present report, we describe two Salmonella enterica serovar Typhimurium aroA vaccine strains expressing a nontoxic plasmid-encoded derivative of Stx2 (Stx2DeltaAB) containing the complete nontoxic A2 subunit and the receptor binding B subunit. The two S. Typhimurium strains differ in the expression of flagellin, the structural subunit of the flagellar shaft, which exerts strong adjuvant effects. The vaccine strains expressed Stx2DeltaAB, either cell bound or secreted into the extracellular environment, and showed enhanced mouse gut colonization and high plasmid stability under both in vitro and in vivo conditions. Oral immunization of mice with three doses of the S. Typhimurium vaccine strains elicited serum anti-Stx2B (IgG) antibodies that neutralized the toxic effects of the native toxin under in vitro conditions (Vero cells) and conferred partial protection under in vivo conditions. No significant differences with respect to gut colonization or the induction of antigen-specific antibody responses were detected in mice vaccinated with flagellated versus nonflagellated bacterial strains. The present results indicate that expression of Stx2DeltaAB by attenuated S. Typhimurium strains is an alternative vaccine approach for HUS control, but additional improvements in the immunogenicity of Stx2 toxoids are still required.


Subject(s)
Enterohemorrhagic Escherichia coli/immunology , Escherichia coli Infections/prevention & control , Escherichia coli Vaccines/immunology , Genetic Vectors , Salmonella typhimurium/genetics , Shiga Toxin 2/immunology , Administration, Oral , Animals , Antibodies, Bacterial/blood , Antibodies, Neutralizing/blood , Antitoxins/blood , Chlorocebus aethiops , Creatinine/blood , Escherichia coli Infections/immunology , Escherichia coli Vaccines/administration & dosage , Escherichia coli Vaccines/genetics , Immunization, Secondary , Mice , Mice, Inbred BALB C , Shiga Toxin 2/biosynthesis , Urea/blood , Vaccines, Attenuated/administration & dosage , Vaccines, Attenuated/genetics , Vaccines, Attenuated/immunology , Vero Cells
3.
FEMS Microbiol Lett ; 269(2): 229-33, 2007 Apr.
Article in English | MEDLINE | ID: mdl-17250759

ABSTRACT

Previous reports have suggested that Escherichia coli K12 mutants defective in the expression of oligogopeptide permease protein A (OppA) exhibit reduced sensitivity to aminoglycosides due to altered permeability of the cell envelope. In this work, the role of the OppA protein, and the oligogopeptide permease (Opp) transport system has been evaluated, in the resistance to aminoglycosides using derivatives of the E. coli K12 SS320 strain selected for triornithine resistance or with a deletion of the complete opp operon. All tested mutants were defective in the uptake of tri- and tetra-peptides but did not expressed resistance to aminoglycosides. Additionally, complementation tests carried out with a plasmid encoding the OppA protein did not affect the sensitivity of the strains to these antibiotics. Taken together, these evidences indicate that the Opp uptake system, as well as the OppA protein, does not play a direct role in the sensitivity to aminoglycosides in E. coli K12.


Subject(s)
Aminoglycosides/pharmacology , Anti-Bacterial Agents/pharmacology , Drug Resistance, Bacterial , Escherichia coli K12/drug effects , Oligopeptides/metabolism , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Carrier Proteins/genetics , Carrier Proteins/metabolism , Escherichia coli K12/enzymology , Escherichia coli K12/genetics , Escherichia coli Proteins/genetics , Escherichia coli Proteins/metabolism , Lipoproteins/genetics , Lipoproteins/metabolism , Membrane Transport Proteins/genetics , Membrane Transport Proteins/metabolism , Microbial Sensitivity Tests , Mutation
4.
Rev. microbiol ; 30(2): 153-6, abr.-jun. 1999. ilus, tab
Article in Portuguese, English | LILACS | ID: lil-257213

ABSTRACT

A spontaneous kanamycin-resistant Escherichia coli mutant, showing cross resitance to five other aminoglycosides and absence of the OppA protein was isolated. [3H]- dihydrostreptomycin uptake is reduced in this mutant, implying that the oligopeptide transport system in involved in accumulation of aminoglucosides, although apparently not related with aminoglycoside permeability alteration due to bacterial adaptation to osmotic changes.


Subject(s)
Oligopeptides/metabolism , Membrane Transport Proteins/metabolism , Periplasm/enzymology , Escherichia coli/isolation & purification , Aminoglycosides/metabolism , Proteins/metabolism , Kanamycin Resistance , Escherichia coli/enzymology , Escherichia coli/genetics , Mutation
5.
Rev. microbiol ; 29(3): 174-8, jul.-set. 1998. ilus, graf
Article in English | LILACS | ID: lil-236203

ABSTRACT

The electrophoretic profiles binding of penicillin binding proteins (PBPs) and outer membrane proteins (OMPs) of Yersinia pestis EV 76 were determined following in vivo growth in diffusion chambers implanted in the peritoneal cavity of mice. In contrast to Y. pestis grown under in vitro conditions which activate the low calcium response (LCR) regulon there was no significant qualitative or quantitative change of the PBP profile of Y. pestis cells during growth in diffusion chambers for up to 72 h following implatation in mice. Three OMPs, with molecular weight of 100, 60 and 58 kDa, were expressed in Y. pestis cells grown for 24 h, but not at 48 h or at 72 h, in diffusion chambers. These results indicate that growth of Y. pestis in intraperitoneal diffusion chambers activates genes which might be relevant to the growth in the mammal host.


Subject(s)
Animals , Mice , Penicillins , Yersinia pestis/cytology , Membrane Proteins , Carrier Proteins , Diffusion Chambers, Culture , Cell Division
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