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Leibniz Institute for Science and Mathematics Education, Kiel

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  • 1
    ISSN: 1365-2761
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Significant numbers of perch, Perca fluviatilis, raised on a pilot fish farm in Switzerland presented focal skin lesions on the lateral sides and fin rot. Mortality rates reached levels of up to 1% of the total fish on the farm per day. Virtually pure cultures of Aeromonas sobria were isolated from the liver, kidney, spleen and skin lesions of affected fish. Aeromonas sobria isolated from the farmed perch had a haemolytic effect on sheep and trout erythrocytes, autoaggregated, was cytotoxic for cultured fish cells and possessed genes involved in type III protein secretion. Experimental infection of naïve perch with a single colony isolate of A. sobria from an affected farm fish resulted in the development of clinical signs identical to those seen on the farm. The results indicate that A. sobria can act as a primary pathogen of perch.
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  • 2
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The cpb2 gene of β2-toxigenic Clostridium perfringens isolated from horses, cattle, sheep, human and pigs was sequenced. The cpb2 gene of equine and other non-porcine isolates differed from porcine isolates by the absence of an adenine in a poly A tract immediately downstream of the start codon in all non-porcine C. perfringens strains. This deletion involved formation of a cryptic gene harbouring a premature stop codon after only nine amino acid codons, while the full β2-toxin protein consists of 265 amino acids. Immunoblots carried out with antibodies directed against a recombinant β2-toxin showed the absence of expression of the β2-toxin in equine and the other non-porcine strains under standard culture conditions. However, treatment of C. perfringens with the aminoglycosides gentamicin or streptomycin was able to induce expression of the cpb2 gene in a representative equine strain of this group, presumably by frameshifting. The presence of the β2-toxin was revealed by immunohistology in tissue samples of small and large intestine from horses with severe typhlocolitis that had been treated before with gentamicin. This result may explain the finding that antibiotic treatment of horses affected by β2-toxigenic C. perfringens leads to a more accentuated and fatal progression of equine typhlocolitis. Clinical observations show a reduced appearance of strong typhlocolitis in horses with intestinal complications admitted to hospital care since the standard use of gentamicin has been abandoned. This is the first report on expression of a bacterial toxin gene by antibiotic-induced ribosomal frameshifting.
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  • 3
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Enteropathogenic Escherichia coli (EPEC) is an intestinal attaching and effacing pathogen that utilizes a type III secretion system (T3SS) for the delivery of effectors into host cells. The chaperone CesT has been shown to bind and stabilize the type III translocated effectors Tir and Map in the bacterial cytoplasm prior to their delivery into host cells. In this study we demonstrate  a  role  for  CesT  in  effector  recruitment  to the membrane embedded T3SS. CesT-mediated effector recruitment was dependent on the presence of the T3SS membrane-associated ATPase EscN. EPEC ΔcesT carrying a C-terminal CesT variant, CesT(E142G), exhibited normal cytoplasmic Tir stability function, but was less efficient in secreting Tir, further implicating CesT in type III secretion. In vivo co-immunoprecipitation studies using CesT-FLAG containing EPEC lysates demonstrated that CesT interacts with Tir and EscN, consistent with the notion of CesT recruiting Tir to the T3SS. CesT was also shown to be required for the efficient secretion of several type III effectors encoded within and outside the locus of enterocyte effacement (LEE) in addition to Tir and Map. Furthermore, a CesT affinity column was shown to specifically retain multiple effector proteins from EPEC culture supernatants. These findings indicate that CesT is centrally involved in recruiting multiple type III effectors to the T3SS via EscN for efficient secretion, and functionally redefine the role of CesT in multiple type III effector interactions.
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  • 4
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Enterohaemorrhagic Escherichia coli (EHEC) O157:H7 uses a specialized protein translocation apparatus, the type III secretion system (TTSS), to deliver bacterial effector proteins into host cells. These effectors interfere with host cytoskeletal pathways and signalling cascades to facilitate bacterial survival and replication and promote disease. The genes encoding the TTSS and all known type III secreted effectors in EHEC are localized in a single pathogenicity island on the bacterial chromosome known as the locus for enterocyte effacement (LEE). In this study, we performed a proteomic analysis of proteins secreted by the LEE-encoded TTSS of EHEC. In addition to known LEE-encoded type III secreted proteins, such as EspA, EspB and Tir, a novel protein, NleA (non-LEE-encoded effector A), was identified. NleA is encoded in a prophage-associated pathogenicity island within the EHEC genome, distinct from the LEE. The LEE-encoded TTSS directs translocation of NleA into host cells, where it localizes to the Golgi apparatus. In a panel of strains examined by Southern blot and database analyses, nleA was found to be present in all other LEE-containing pathogens examined, including enteropathogenic E. coli and Citrobacter rodentium, and was absent from non-pathogenic strains of E. coli and non-LEE-containing pathogens. NleA was determined to play a key role in virulence of C. rodentium in a mouse infection model.
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  • 5
    ISSN: 1365-3040
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: The aim of this study was to show the potential of Thlaspi caerulescens in the cleaning-up of a moderately Zn -contaminated soil and to elucidate tolerance mechanisms at the cellular and subcellular level for the detoxification of the accumulated metal within the leaf. Measured Zn concentrations in shoots were high and reached a maximum value of 83 mmol kg−1 dry mass, whereas total concentrations of Zn in the roots were lower (up to 13 mmol kg−1). In order to visualize and quantify Zn at the subcellular level in roots and leaves, ultrathin cryosections were analysed using energy-dispersive X-ray micro-analysis. Elemental maps of ultrathin cryosections showed that T. caerulescens mainly accumulated Zn in the vacuoles of epidermal leaf cells and Zn was almost absent from the vacuoles of the cells from the stomatal complex, thereby protecting the guard and subsidiary cells from high Zn concentrations. Observed patterns of Zn distribution between the functionally different epidermal cells were the same in both the upper and lower epidermis, and were independent of the total Zn content of the plant. Zinc stored in vacuoles was evenly distributed and no Zn-containing crystals or deposits were observed. From the elemental maps there was no indication that P, S or Cl was associated with the high Zn concentrations in the vacuoles. In addition, Zn also accumulated in high concentrations in both the cell walls of epidermal cells and in the mesophyll cells, indicating that apoplastic compartmentation is another important mechanism involved in zinc tolerance in the leaves of T. caerulescens.
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  • 6
    ISSN: 1365-3040
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Compartmentation of heavy metals on or within mycorrhizal fungi may serve as a protective function for the roots of forest trees growing in soils containing elevated concentrations of metals such as Cd and Zn. In this paper we present the first quantitative measurements by X-ray microanalysis of heavy metals in high-pressure frozen and cryosectioned ectomycorrhizal fungal hyphae. We used this technique to analyse the main sites of Cd and Zn in fungal cells of mantle and Hartig net hyphae and in cortical root cells of symbiotic Picea abies – Hebeloma crustuliniforme associations to gain new insights into the mechanisms of detoxification of these two metals in Norway spruce seedlings. The mycorrhizal seedlings were exposed in growth pouches to either 1 mM Cd or 2 mM Zn for 5 weeks. The microanalytical data revealed that two distinct Cd- and Zn-binding mechanisms are involved in cellular compartmentation of Cd and Zn in the mycobiont. Whereas extracellular complexation of Cd occurred predominantly in the Hartig net hyphae, both extracellular complexation and cytosolic sequestration of Zn occurred in the fungal tissue. The vacuoles were presumed not to be a significant pool for Cd and Zn storage. Cadmium was almost exclusively localized in the cell walls of the Hartig net (up to 161 mmol kg−1 DW) compared with significantly lower concentrations in the cell walls of mantle hyphae (22 mmol kg−1 DW) and in the cell walls of cortical cells (15 mmol kg−1 DW). This suggests that the apoplast of the Hartig net is a primary accumulation site for Cd. Zinc accumulated mainly in the cell walls of the mantle hyphae (111 mmol kg−1 DW), the Hartig net hyphae (130 mmol kg−1 DW) and the cortical cells (152 mmol kg−1 DW). In addition, Zn occurred in high concentrations in the cytoplasm of the fungal mantle hyphae (up to 164 mmol kg−1 DW) suggesting that both the cell walls and the cytoplasm of fungal tissue are the main accumulation sites for Zn in P. abies resulting in decreased Zn transfer from the fungus to the root.
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  • 7
    ISSN: 1365-3040
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: The consequences of manipulating abscisic acid (ABA) biosynthesis rates on stomatal response to drought were analysed in wild-type, a full-deficient mutant and four under-producing transgenic lines of N. plumbaginifolia. The roles of ABA, xylem sap pH and leaf water potential were investigated under four experimental conditions: feeding detached leaves with varying ABA concentration; injecting exogenous ABA into well-watered plants; and withholding irrigation on pot-grown plants, either intact or grafted onto tobacco. Changes in ABA synthesis abilities among lines did not affect stomatal sensitivity to ABA concentration in the leaf xylem sap ([ABA]xyl), as evidenced with exogenous ABA supplies and natural increases of [ABA]xyl in grafted plants subjected to drought. The ABA-deficient mutant, which is uncultivable under normal evaporative demand, was grafted onto tobacco stock and then presented the same stomatal response to [ABA]xyl as wild-type and other lines. This reinforces the dominant role of ABA in controlling stomatal response to drought in N. plumbaginifolia whereas roles of leaf water potential and xylem sap pH were excluded under all studied conditions. However, when plants were submitted to soil drying onto their own roots, stomatal response to [ABA]xyl slightly differed among lines. It is suggested, consistently with all the results, that an additional root signal of soil drying modulates stomatal response to [ABA]xyl.
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