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  • 1
    Electronic Resource
    Electronic Resource
    Survival of a Shiga toxin-encoding bacteriophage in a compost model (2005)
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 245 (2005), S. 0 
    Details
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Bacteriophages that carry the Shiga toxin gene (stx) represent an additional hazard in cattle manure-based fertilizers in that their survival could lead to toxigenic conversion of Escherichia coli and other bacteria post-composting. A Stx-phage in which the Shiga toxin (stx2) gene was inactivated by insertion of a chloramphenicol resistance gene was used in combination with a rifampicin-resistant E. coli host where RecA is constitutively activated so that all infectious phage particles could be enumerated by plaque assay. PCR-based confirmation methods and the additional application of a host enrichment protocol ensured that very low numbers of surviving bacteriophage could be detected and unequivocally identified. Stx-bacteriophage numbers declined rapidly over the first 48 h and none could be detected after 3 days. The host enrichment method was applied after 6 days and no bacteriophages were recovered. While addition of fresh E. coli cells at intervals after the compost temperature had reduced below 40 °C demonstrated that E. coli growth could be supported in the compost, Stx-phages or their lysogens were never detected. Here, we demonstrate that composting animal manure for 40 days during which a temperature of 〉60 °C is maintained for at least 5 days is effective at removing both E. coli and a model infectious Stx-encoding bacteriophage.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1016/j.femsle.2005.03.031
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  • 2
    Electronic Resource
    Electronic Resource
    Meningococcal pilin: a glycoprotein substituted with digalactosyl 2,4-diacetamido-2,4,6-trideoxyhexose (1995)
    Osney Mead, Oxford OX2 0EL, UK : Blackwell Scientific Publications
    Molecular microbiology 17 (1995), S. 0 
    Details
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Neisseria meningitidis pili are filamentous protein structures that are essential adhesins in capsulate bacteria. Pili of adhesion variants of meningococcal strain C311 contain glycosyl residues on pilin (PilE), their major structural subunit. Despite the presence of three potential N-linked glycosylation sites, none appears to be occupied in these pilins. Instead, a novel O-linked trisaccharide substituent, not previously found as a constituent of glycoproteins, is present within a peptide spanning amino acid residues 45 to 73 of the PilE molecule. This structure contains a terminal 1-4-linked digalactose moiety covalently linked to a 2,4-diacetamido-2,4,6-trideoxyhexose sugar which is directly attached to pilin. Pilins derived from galactose epimerase (galE) mutants lack the digalactosyl moiety, but retain the diacetamidotrideoxyhexose substitution. Both parental (#3) pilins and those derived from a hyper-adherent variant (#16) contained identical sugar substitutions in this region of pilin, and galE mutants of #3 were similar to the parental phenotype in their adherence to host cells. These studies have confirmed our previous observations that meningococcal pili are glycosylated and provided the first structural evidence for the presence of covalently linked carbohydrate on pili. In addition, they have revealed a completely novel protein/saccharide linkage.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1365-2958.1995.mmi_17061201.x
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  • 3
    Electronic Resource
    Electronic Resource
    Pilus-facilitated adherence of Neisseria meningitidis to human epithelial and endothelial cells: modulation of adherence phenotype occurs concurrently with changes in primary amino acid sequence and the glycosylation status of pilin (1993)
    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 10 (1993), S. 0 
    Details
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Adherence of capsulate Neisseria meningitidis to endothelial and epithelial cells is facilitated in variants that express pili. Whereas piliated variants of N. meningitidis strain C311 adhered to endothelial cells in large numbers (〈150 bacteria/cell), derivatives containing specific mutations that disrupt pilE encoding the pilin subunit were both non-piliated and failed to adhere to endothelial cells (〈1 bacterium/ cell). In addition, meningococcal pili recognized human endothelial and epithelial cells but not cells originating from other animals. Variants of strain C311 were obtained that expressed pilins of reduced apparent Mr and exhibited a marked increase in adherence to epithelial cells. Structural analysis of pilins from two hyper-adherent variants and the parent strain were carried out by DNA sequencing of their pilE genes. Deduced molecular weights of pilins were considerably tower compared with their apparent Mr values on SDS-PAGE. Hyper-adherent pilins shared unique changes in sequence including substitution of Asn-113 for Asp-113 and changes from Asn-Asp-Thr-Asp to Thr-Asp-Ala-Lys at residues 127-130 in mature pilin. Asn residues 113 and 127 of‘parental’pilin both form part of the typical eukaryotic N-glycosylation motif Asn-X-Ser/Thr and could potentially be glycosylated post-translationally. The presence of carbohydrate on pilin was demonstrated and when pilins were deglycosylated, their migration on SDS-PAGE increased, supporting the notion that variable glycosylation accounts for discrepancies in apparent and deduced molecular weights. Functionally distinct pilins produced by two fully piliated variants of a second strain (MC58) differed only in that the putative glycosylation motif Asn-60-Asn-61-Thr-62 in an adherent variant was replaced with Asp-60-Asn-61-Ser-62 in a non-adherent variant. Fully adherent backswitchers obtained from the non-adherent variant always regained Asn-60 but retained Ser-62. We propose, therefore, that functional variations in N. meningitidis pili may be modulated in large part by primary amino acid sequence changes that ablate or create N-linked glycosylation sites on the pilin subunit.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1365-2958.1993.tb00972.x
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  • 4
    Electronic Resource
    Electronic Resource
    Development and application of molecular tools in the study of IncN-related plasmids from lakewater sediments (2000)
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 186 (2000), S. 0 
    Details
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Homology to IncN, P, Q and W inc regions was investigated amongst 114 Hg2+-resistant or antibiotic-resistant bacteria isolated from lakewater sediments. No hybridisation signals were found with Inc P, Q and W probes, and only one plasmid, pLV1402, hybridised to the IncN probe. PCR primers designed to conserved regions in the replicon of the IncN plasmid pCU1 and the related beta replicon from pGSH500 were used to amplify a 978-bp fragment from pLV1402, with sequence analysis showing a close relationship (99.2% identity) between their replication genes. A 387-bp region from the pLV1402 rep gene was used to re-screen the isolates and identified another related plasmid, pLV1403. A 3.7-kb probe containing the alpha replicon from pGSH500 hybridised to both pLV1402 and pLV1403, suggesting that both are multi-replicon plasmids. The PCR primers and probes described will be useful in future studies of plasmid diversity.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1574-6968.2000.tb09105.x
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  • 5
    Electronic Resource
    Electronic Resource
    Posttranslational Modifications of Meningococcal Pili: Identification of a Common Trisaccharide Substitution on Variant Pilins of Strain C311 (1996)
    Oxford, UK : Blackwell Publishing Ltd
    Annals of the New York Academy of Sciences 797 (1996), S. 0 
    Details
    ISSN: 1749-6632
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Natural Sciences in General
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1749-6632.1996.tb52949.x
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  • 6
    Electronic Resource
    Electronic Resource
    The role of bacterial motility in the survival and spread of Pseudomonas fluorescens in soil and in the attachment and colonisation of wheat roots (2001)
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology ecology 36 (2001), S. 0 
    Details
    ISSN: 1574-6941
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Motile and non-motile strains of Pseudomonas fluorescens SBW25 were constructed using different combinations of the lacZY, xylE and aph marker genes which allowed their detection and differentiation in soil, root and seed samples. The survival of motile and non-motile strains was investigated in both non-competitive and competitive assays in water and non-sterile soil. Although there was no difference between strains in water, the motile strain survived in significantly greater numbers than the non-motile strain after 21 days in soil. There was no significant difference between competitive assays, where motile and non-motile cells were co-inoculated into soil, and non-competitive assays where strains were inoculated separately. Bacterial survival decreased as matric potential increased from −224 to −17 kPa but matric potential had no significant effect on motile compared to non-motile strains. Vertical spread of both motile and non-motile strains was detected 6.4 mm from the inoculum zone after 14 days in the absence of percolating water. There was no significant difference, for either strain, in distance moved from the inoculum zone after 14, 26 or 40 days. The motile strain had a significant advantage in attachment to sterile wheat roots in both non-competitive and competitive studies. When the spatial colonisation of wheat root systems was assessed in non-sterile soil, there was no significant difference between the motile and non-motile strain from either seed or soil inoculum. However, when the whole root system was assessed as one sample unit, differences could be detected. Bacterial motility could contribute to survival in soil and the initial phase of colonisation, where attachment and movement onto the root surface are important.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1574-6941.2001.tb00822.x
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  • 7
    Electronic Resource
    Electronic Resource
    Detection and distribution of insertion sequence 1 (IS1)-containing bacteria in the freshwater environment (2000)
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology ecology 34 (2000), S. 0 
    Details
    ISSN: 1574-6941
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: The distribution of insertion sequence 1 (IS1)-containing bacteria was investigated in Windermere (Cumbria, UK), a freshwater body impacted by treated sewage discharge and run-off from the surrounding catchment. Culturable IS1-containing bacteria were recovered from the water column at three depths in Windermere North Basin (WNB) and South Basin (WSB), and from sediment at both sites (at the sediment surface in WSB and to a depth of 12–13 cm in WNB). Polymerase chain reaction amplification of IS1 and the Escherichia coli/Shigella sp. specific gene uidA, from community DNA from shallow sediments, extended the detection limit beyond that of culture at both sites. This detection was extended further into deep sediment extracted from WNB as IS1 and uidA were detected in sub-samples to a depth of 4.7 and 2.3 m, respectively. Analysis of a representative subset of 90 IS1-carrying isolates recovered from water and sediment at both sites demonstrated 21 heterogeneous IS1 profiles with estimated copy numbers ranging from 1 to 16. Identification of the host bacteria showed that the element was confined mainly to Enterobacter spp. However, this study showed IS1 to be present in Citrobacter freundii for the first time. Plasmids were carried by 75.3% of enterobacterial isolates and four plasmids (2.6%) carried IS1. DNA sequence analysis of five IS1 clones demonstrated that IS1 isoforms from this study were similar (〉89% nucleotide identity) to known IS1 isoforms. Two isoforms of IS1 from a single Enterobacter cloacae isolate differed by 6.7% at the nucleotide level suggesting that they had been acquired independently.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1574-6941.2000.tb00757.x
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  • 8
    Electronic Resource
    Electronic Resource
    Direct molecular biological analysis of ammonia oxidising bacteria populations in cultivated soil plots treated with swine manure (1997)
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology ecology 23 (1997), S. 0 
    Details
    ISSN: 1574-6941
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: The application of pig slurry, high in ammonia-nitrogen concentration, to agricultural land is a practice whose effect on soil microbial communities is poorly understood. The autotrophic ammonia-oxidising bacteria are an integral component of the nitrogen cycle in soil, and their activity will be affected by addition of nitrogenous fertilizer. Molecular biological techniques were applied to the direct detection and identification of ammonia-oxidiser populations in cultivated soil plots treated with different amounts of pig slurry. Members of the genus Nitrosospira were shown by 16S rDNA-directed PCR to be present in both unamended and amended soils, regardless of the quantity of pig slurry applied. In contrast, members of the genus Nitrosomonas were detected by the same approach only in those soil plots that had received high loadings of slurry. The fidelity of amplification products was always confirmed by oligonucleotide probing. In addition, we used high stringency PCR and confirmatory gene probing to detect the presence of the ammonia monooxygenase gene (amoA) of Nitrosomonas europaea directly in all amended soil samples, with hybridization signal intensities that increased with the amount of pig slurry applied to plots. Nitrosomonas europaea amoA DNA could not be detected in soil from the untreated plot. These data support the view that nitrosospiras are ubiquitous as important members of nitrifying populations in the environment. The direct detection of nitrosomonad DNA only in amended soils supports the hypothesis that these nitrosomonads become highly competitive under conditions analogous to laboratory enrichment cultures.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1574-6941.1997.tb00390.x
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  • 9
    Electronic Resource
    Electronic Resource
    Micro-ecology of peat: minimally invasive analysis using confocal laser scanning microscopy, membrane inlet mass spectrometry and PCR amplification of methanogen-specific gene sequences (1998)
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology ecology 25 (1998), S. 0 
    Details
    ISSN: 1574-6941
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: A peat monolith (15 cm diameter, 35 cm length) from Ellergower Moss (New Galloway, Scotland), kept outdoors and maintained water-saturated, was investigated for the distributions of gases (O2, CO2, CH4), micro-organisms, total archaeal DNA and methanogen DNA. From the water table (at the surface of the Sphagnum), a steep oxycline gave 〈0.25 μM O2 at 2 cm depth (as shown by membrane inlet mass spectrometry and oxygen electrode methods) and 〈10 nM O2 at 6 cm depth (photobacterium gas diffusion probe). Redox potential measurements indicated a steep decline between 6 cm and 13 cm to a value of −90 mV. At the oxic surface of the peat, CO2 measured 0.5 mM and CH4 〈1 μM. Below 7 cm both gases increased to plateaux at 2 mM and 550 μM, respectively; CH4 concentrations also indicated two distinct zones (7 μM to 2.5 cm depth, then to 28 μM at between 3 and 6 cm). Confocal laser scanning microscopy using the fluorophores 5-cyano-2,3-ditolyl tetrazolium chloride or 3,3-dihexyloxacarbocyanine iodide was used to image micro-organisms with redox active electron transport activities or transmembrane electrochemical potentials, respectively. Samples from 1–5 cm depth showed the presence of active aerobic organisms, whereas those from 10 and 20 cm depth were more active anaerobically, and especially so under H2. Archaeal DNA was present throughout the core; strongest hybridisation was below 9 cm. Two methanogen-specific primers, ME1 and ME2 (which amplify a region of the α-subunit of methyl coenzyme M reductase), hybridised with DNA extracted from below 9 cm depth. Here we describe the concerted application of a number of techniques providing direct information on the precise location and activities of microbes involved in the flux of gases from peatlands.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1574-6941.1998.tb00471.x
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  • 10
    Electronic Resource
    Electronic Resource
    Transfer of plasmid pBC16 between Bacillus thuringiensis strains in non-susceptible larvae (2002)
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology ecology 40 (2002), S. 0 
    Details
    ISSN: 1574-6941
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Plasmid transfer was investigated in larvae of insects of the orders Coleoptera, Diptera, and Lepidoptera. The effects of introducing Bacillus thuringiensis strains in live non-susceptible larvae, and in the presence of added insecticidal toxins to kill the larvae, were examined. Plasmid transfer was not detected as the strains passed through non-susceptible live larvae, but was detected when the larvae were toxin-killed. The results indicate that growth of B. thuringiensis and plasmid transfer between strains while simply passing through an insect gut system is an infrequent event. In toxin-killed larvae, a more complex picture was recorded. B. thuringiensis subsp. kurstaki transferred pBC16 at a lower rate in killed Phaedon cochleriae larvae compared to previous work studying transfer with this strain in susceptible Lacanobia oleracea larvae. Similarly, B. thuringiensis subsp. tenebrionis transferred pBC16 in killed L. oleracea larvae, while no transfer in susceptible P. cochleriae larvae was detected. The results indicate that gene transfer was more frequent in killed L. oleracea larvae. When both B. thuringiensis strains were studied in Aedes aegypti, transfer of pBC16 was detected in toxin-killed larvae. This was surprising since in similar studies with strain B. thuringiensis subsp. israelensis that kills mosquitoes, transfer of pBC16 was not detected in mosquito cadavers. The improved transfer frequency of B. thuringiensis subsp. kurstaki and subsp. tenebrionis compared to B. thuringiensis subsp. israelensis in laboratory broth culture could account for this difference in detection of transfer within killed insects.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1574-6941.2002.tb00951.x
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