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  • 1
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Abstract Staphylococcus aureus endo-β-N-acetylglucosaminidase (SaG) has been suggested to function as a virulence determinant which interferes with the host cellular immune response. To further characterize the biological properties of SaG, monoclonal antibodies (mAbs) were raised against purified SaG. Four IgG1 subclass mAbs were obtained, none of which reacted with the reduced, sodium dodecyl sulphate pretreated or boiled enzyme. The ability of the mAbs to react with the enzymes present in supernatants obtained from 197 S. aureus strains indicated that they recognized epitopes which are highly conserved; bacteriolytic enzymes produced by staphylococci other than S. aureus did not show any cross-reactivity. After pretreatment of SaG with mAbs (mAb-SaG molar ratios varying from 1 to 20), it was shown that all selected mAbs caused, at a mAb: SaG molar ratio of 10, a 90% inhibition of SaG bacteriolytic activity and a statistically significant reduction of its ability to interfere with phagocytosis to human polymorphonuclear leukocytes. All selected mAbs reacted with several commercially available exo-β-N-acetylglucosaminidases; mAb C1/10–11 also reacted with chicken and turkey egg muramidases and, at a mAb:SaG molar ratio of 10, inhibited their bacteriolytic activity by 97%. This suggests that one or more epitopes present in the above exo-glucosaminidases and muramidases share some degree of homology with others present in SaG.
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  • 2
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: We have constructed an aromatic amino acid auxotrophic mutant of Bordetella bronchiseptica, harbouring mutations in aroA and trpE to investigate the use of such a strain as a live-attenuated vaccine. B. bronchiseptica aroA trpE was unable to grow in minimal medium without aromatic supplementation. Compared to the parental wild-type strain, the mutant displayed significantly reduced abilities to invade and survive within the mouse macrophage-like cell line J774A.1 in vitro and in the murine respiratory tract following experimental intranasal infection. Mice vaccinated with B. bronchiseptica aroA trpE displayed significant dose-dependent increases in B. bronchiseptica-specific antibody responses, and exhibited increases in the number of B. bronchiseptica-reactive spleen cells in lymphoproliferation assays. Immunised animals were protected against lung colonisation after challenge with the wild-type parental strain. With such a broad host range displayed by B. bronchiseptica, the attenuated strain constructed in this study may not only be used for the prevention of B. bronchiseptica-associated disease, but also for the potential delivery of heterologous antigen.
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  • 3
    Electronic Resource
    Electronic Resource
    s.l. : American Chemical Society
    Journal of agricultural and food chemistry 43 (1995), S. 102-105 
    ISSN: 1520-5118
    Source: ACS Legacy Archives
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition , Process Engineering, Biotechnology, Nutrition Technology
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  • 4
    Electronic Resource
    Electronic Resource
    s.l. : American Chemical Society
    Journal of agricultural and food chemistry 42 (1994), S. 2743-2745 
    ISSN: 1520-5118
    Source: ACS Legacy Archives
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition , Process Engineering, Biotechnology, Nutrition Technology
    Type of Medium: Electronic Resource
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  • 5
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 201 (2001), S. 0 
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Previous studies suggested that the persistence in eukaryotic cells of a Bordetella bronchiseptica mutant carrying an insertion in the locus encoding the response regulator RisAS is impaired. This suggested that ris-dependent products are required for the intracellular survival of bacteria. In this study we demonstrate that ris-regulated products play a role in B. bronchiseptica resistance against both phagosomal acidification and reactive oxygen intermediates.
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  • 6
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Non-motile mutants of Bordetella bronchiseptica were generated after mini-transposon mutagenesis. One non-motile mutant (designated VMM1) was derived from the bvg-positive strain BB7865 and four mutants (designated AMM1–4) were derived from the isogenic bvg-negative strain BB7866. Southern hybridisation analysis indicated that loss of motility was not due to the disruption of the flagellin subunit gene. Western blot and transmission electron microscopic analysis indicated that three of the five mutants expressed neither the flagellin subunit (40 kDa) nor flagella whereas one mutant expressed intact flagella under all conditions tested. One unique bvg-negative mutant, AMM4, exhibited temperature-dependent repression of flagella biosynthesis and motility at 37°C. The ability of AMM4 to invade and survive in HeLa cells was significantly decreased.
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  • 7
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: To investigate the role played by urease during the Bordetella bronchiseptica infection process, the ability to colonise and persist in the mouse respiratory tract of a urease-negative B. bronchiseptica BB7865 and a BB7865 derivative constitutively expressing urease was compared with that of the wild-type strain. The results obtained showed that neither constitutive expression nor abolishment of urease activity had any significant effect on the course of B. bronchiseptica infection. Therefore, under our experimental conditions, urease is not essential for B. bronchiseptica to colonise and persist within the murine host.
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  • 8
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology reviews 25 (2001), S. 0 
    ISSN: 1574-6976
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Bordetella pertussis is the causative agent of whooping cough, a contagious childhood respiratory disease. Increasing public concern over the safety of whole-cell vaccines led to decreased immunisation rates and a subsequent increase in the incidence of the disease. Research into the development of safer, more efficacious, less reactogenic vaccine preparations was concentrated on the production and purification of detoxified B. pertussis virulence factors. These virulence factors include adhesins such as filamentous haemagglutinin, fimbriae and pertactin, which allow B. pertussis to bind to ciliated epithelial cells in the upper respiratory tract. Once attachment is initiated, toxins produced by the bacterium enable colonisation to proceed by interfering with host clearance mechanisms. B. pertussis co-ordinately regulates the expression of virulence factors via the Bordetella virulence gene (bvg) locus, which encodes a response regulator responsible for signal-mediated activation and repression. This strict regulation mechanism allows the bacterium to express different gene subsets in different environmental niches within the host, according to the stage of disease progression.
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  • 9
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 184 (2000), S. 0 
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: The Pas protein plays a key role in the pathogenesis of enterohemorrhagic Escherichia coli (EHEC), being required for the secretion of the Esp proteins. Here, the transcriptional regulation of the pas gene was analyzed through the construction of a pas::lacZ translational fusion. When bacteria were grown in Luria Bertani medium or tissue culture medium supplemented with HEPES, a bimodal activation curve was observed. The early phase of induction was not significantly modified by the incubation temperature (either 25 or 37°C), whereas the second phase, which overlaps with the late exponential growth phase, was enhanced at 37°C. The early phase was also stimulated by growth on tissue culture medium and by the addition of Ca2+, Mn2+or Mg2+ to the M9-glucose minimal medium. Primer extension analysis showed the presence of two major starts of transcription, which were located 58 and 60 bp upstream of the ATG-start codon of the Pas protein, respectively. Although these sites are very close to each other, the transcripts produced during the early induction phase mainly start on the −60 position, whereas the −58 start was activated during the second induction phase.
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  • 10
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 182 (2000), S. 0 
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: A set of expression vectors was constructed which allows the expression of recombinant antigens under the control of Salmonella typhi promoters that are selectively activated after infection of eukaryotic cells. The pUC18Not derivatives contain a promoter downstream of the early transcriptional terminator from phage T7 and followed by a multiple cloning site, whereas the pBluescript II S/K derivatives contain the ribosomal RNA T1 transcriptional terminator and also the strong translation signals of the Escherichia coli atpE gene. The expression cassettes are flanked by NotI or PacI sites to simplify their subcloning where required. The resulting vectors were validated using the S1 subunit of pertussis toxin as a model antigen and Salmonella typhimurium aroA SL7207 as a carrier. The S1 subunit was efficiently expressed by recombinant Salmonella within Henle 407 cells.
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