An investigation of Clostridium difficile has shown : 1. that the organism is resistant to cresols and phenolq and may be grown from material grossly contaminated with other organisms by culture in media containing these substances; strains isolated before this method was evised are also resistant to cresols and phenol; 2. that the organism divides into four daughter cells; 3. that each strain examined has a different strain specific agglutinogen, and that the strains are otherwise antigenically complex; 4. that the sugar reactions are fairly consistent 5. that gelatin is liquefied by all available strains in three weeks; and that apart from this there is no proteolytic activity 6. that the hyaluronidase produced by all strains is antigenically uniform? and so is the lethal toxin; 7. and that the 'haemolysin' is almost certainly complex.

Propionibacterium acnes, a resident micro-organism of human skin, is thought to be involved in the development of inflammatory acne, which is an exclusive human disease affecting more than 80% of the whole population. Quorum sensing is the regulation of gene expression in response to cell density. As it is often involved in pathogenecity of bacteria, its signal transduction pathway has been suggested as potential target for new drug development. This project identified a putative unique quorum sensing system of P. acnes, consisting of a putative signalling peptide, and divergently transcribed histidine kinase and response regulator. The aim of this project is to investigate the relationship among these three components as being elements of a putative quorum sensing system. Using purified proteins and in vitro phosphorylation assays, the histidine kinase was demonstrated to phosphorylate the response regulator indicating they may constitute a legitimate pair of a two-component system, despite being encoded by divergently transcribed genes. By mapping transcriptional start site, it was found that the signalling peptide and histidine kinase were co-transcribed from the same start site, suggesting that the signalling peptide was associated with the two-component system. Gene expression analysis also revealed these three genes were co-regulated during the growth of P.acnes, which is consistent with these three genes functioning together as a part of quorum sensing system. The results of this project suggested that the signalling peptide, histidine kinase and response regulator are associated with each other and may constitute a quorum sensing system.

In nature, bacteria usually exist as aggregates, in order to withstand changes in environmental conditions. Bacterial aggregation is of great significance in the field of biotechnology, environmental studies and medicine. Bacterial aggregation is thought to be governed by physical forces such Van der Waals and electrostatic interaction. However, extracellular polymeric substances (EPS), and the ability. of bacteria ·to participate in cell-to-cell communication via quorum sensing molecules have also been implicated in the bacterial aggregation. process. Despite the wealth of knowledge available, a detailed understanding of bacterial aggregation still remains unclear. The overall aim of this work therefore, is to understand bacterial aggregation at the cellular and sub-cellular level using existing colloidal characterisation techniques and post genomic methods. This will enable both the biological and the physical aspects of aggregation to be studied together. E.coli strains (AB1157, MG1655 and MG1655 luxS) were cultivated in Luria-Bertani (LB) medium at 30°C supplemented with or without 0.5 w/v (%) glucose at the beginning of growth phase. Depletion aggregation studies were carried out using E.coli AB1157 and E.coli MG1655 harvested at different growth phases using a nonadsorbing polymer, sodium polystyrene sulphonate (SPS) and biological produced extracellular polymeric substances (EPS) respectively. The content of EPS produced by E.coli MG1655 during their growth in different media w~s quantified and characterized using Fourier transformation infrared spectroscopy (FTIR), SDS-PAGE and an electrospray ionization-tandem mass spectrometry. The changes in cell surface properties of E.coli strains during growth, changes in media composition and quorum sensing were elucidated using potentiometric titration, FTIR and electrophoretic mobility. Neither quorum sensing, nor the addition of 0.5 w/v (%) glucose affected the growth pattern for the strains. However, the addition of 0.5 w/v (%) glucose to the medium affected the measurable amount of quorum sensing molecule present in the supernatant. Aggregation of E. coli was found to be dependent on the concentration and type of polymer used, as well as the surface chemistry of the cell. The cell surface functional groups, such as such as, hydroxyl, phosphoryl, amines and carboxylate groups varied with respect to different growth phase and changes in media. The protein content of free-EPS was found to significantly increase due to changes in growth phase and media composition. The growth phase, changes in media and quorum sensing all affected the cell surface properties and hence played a role in the aggregation capability of E.coli.

Studies were made in relation to the theory of panspermia, in particular, the version known as cometary panspermia, which suggests that life on Earth a) originated from space (from comets) and b)continues to arrive from this source (Le. neopanspermia). The following conclusions were arrived at; I) Evidence is provided to support/recent findings showing that viable bacteria are present ill the stratosphere at a height of 41 km (circa 25 miles). These bacteria appear in clumps of a size in excess of 10-20 microns. It is argued that it is unlikely that bacterial clumps of this size could be elevated from Earth to 41 km and that as result these clumps must be incoming from space. It is suggested that these may be unculturable and may make up the majority of unculturable bacteria that are found in the environments of Earth (this suggestion is however, remains conjectural). 2) Studies were made on the Red Rain that recently fell on Kerala in India. It was confirmed that Red Rain is made up of microbial cells of morphology similar to that of algae and fungal spores. Another type of Red Rain, sampled in England, is algal in appearance, although morphologically it is completely different from the Red Rain of Kerala. 3) The chemical composition of a comet was simulated. The simulated comct was shown to be capa~le of supporting the growth or bacteria directly sampled from the atmosphere on Earth. Bacteria grew in the presence and absence of the PAH, naphthalene. Thc results suggest that Earth bacteria could grow in the nutrient conditions provided by a comet. 4) Sulphuric acid at a concentration in excess of those likely to be found in the stratosphere did not completely sterilise a culture of Bacillus subtilis, showing that this acid, although present ill the stratosphere, is unlikely to completely inhibit bacterial survival in this environment. 5) A study was made of the microbiology of rocks (i.e. amber, coals, limestone and clay). A large numbcr of bacteria were isolated from clay (mainly species of Bacillus) While two bacteria were isolated from amber and one from a coal sample, no organisms were isolated from limestonc. Obviously, the theory of geopanspermia, the view that bacteria arc transmitted through space in rocks, requires such a demonstration that rocks on Earth do in fc1ct harbour bacteria.

No description available.

No description available.

In their natural habitat bacteria predominantly exist as biofilms in which bacteria differ physiologically and metabolically from free-living cells. The aim of the study was to identify and characterize genes that are involved in biofilm maturation. The crystal violet (CV) assay is widely used to identify early stage biofilm mutants. However, it is not sufficiently sensitive to isolate late stage mutants that fail to develop mature biofilms. Using tetrazolium salt XTT (sodium 3'-{ l-[(phenylamino)-carbonyl]-3,4tetrazolium}- bis (4- methoxy-6-nitro) benzenesulfonic acid hydrate) this study therefore developed a more sensitive assay, named the XTT assay, to identify smaller changes in the quantity of biofilm. Using a combination of the CV and the XTT assays, a pool of transposon mediated mutant cells was screened and four genes (yhjN, adiA, bglX and glpX) were identified as potentially being involved in biofilm maturation. glpX was selected for further study. GlpX is a largely uncharacterised protein that had been shown to have FBPase activity in vitro. FBPase activity is required for the synthesis of colanic acid, a polysaccharide previously shown to playa key role in the development of the three-dimensional structure of the fully formed biofilm. The findings of this study 4 indicate that glpX-encoded FBPase activity is involved in colanic acid biosy~thesis in E. coli. A preliminary molecular analysis was made of the expression of glpX.

Of >500 bacterial isolates from seaweed, namely Fucus serratus, algae (Halimeda sp, Sargassum sp), sponges (Siphonochalina sp, Leucetta 'chgosensis), sea cucumber, (Holothuria atra), star fish (Acanthaster planci), mangrove (Avecenia marina) roots, sea anemone, (Heteractis magnifica), ,jellyfish (Disambiguation), sediment and seawater from the Scottish and Saudi Arabian coasts, a culture of Bacillus licheniformis produced a novel protein with antibacterial activity against methicillin-resistant Staphylococcus aureus, vancomycin-resistant enterococci and Listeria monocytogenes. The antibacterial activity was maximal in cultures prepared in Columbia broth -containing pieces of synthetic polyurethane sponge and shaken at 210-230 rpm. Antibacterial activity was not found in cultures grown statically or with different speeds of rotary shaking. Reduced activity was apparent in supernatants prepared from marine . 2216E broth and tryptone soya broth with or without 1% (w/v) sodium chloride. The antibacterial compound was sensitive to proteinase K, pronase and trypsin, but was not affected by Tween 20, 40, 60 or 80, or a- or ~-amylase. Activity was not adversely affected by heating to 40°C or treatment at pH 5-14. The bioactive compound was purified by gel filtration and ion exchange, and classified by using MALDI-TOF determined as a protein of 30.7 kDa, which had homology to the YbdN protein of B. licheniformis ATCC 14580. The gene encoding YbdN was isolated and transferred to Escherichia coli XL BLUE, and the product was purified by SDS-PAGE giving a single band with a molecular size 30.7 kDa.

No description available.

No description available.