No description available.

<p>Life is cellular. Cells grow and divide to give two new cells; this process is called the cell cycle. The chromosome in a bacterium is replicated into two identical copies before the cell divides. DNA replication is a fundamental process common to all forms of life.</p><p>In my thesis, I have studied control of chromosome and plasmid replication in <i>Escherichia coli</i>, a rod-shaped bacterium. Plasmids are extrachromosomal autonomously replicating DNA molecules. </p><p>I have combined the classical Meselson-Stahl density-shift and DNA hybridisation with theoretical analysis of DNA replication. The minimal time between two successive replications of the same molecule, the eclipse, was determined for both plasmid and chromosome.</p><p>The aim was to investigate the processes ensuring the precise timing of chromosome replication in the cell cycle. In wild-type strains, the chromosomal eclipse was long. Mutations affecting the so-called sequestration process, the superhelicity of the DNA, and the initiation protein, DnaA, reduced the eclipse.</p><p>Fast-growing <i>E. coli</i> has overlapping replicative phases with synchronous initiation from multiple initiation sites, <i>oriC</i>. I have investigated the complex interplay between different control processes by measuring the length of the eclipse and the degree of asynchronous initiation in various mutants.</p><p>I have measured the eclipse period of plasmid R1 during up- and down-shifts in plasmid copy number. The length of the eclipse was found to be determined by structural events as well as by the properties of the copy-number-control system.</p><p>During downshift from very high copy numbers, the rate of plasmid replication started very slowly and gradually increased until the normal copy number was achieved, in accordance with the +<i>n</i> model.</p><p>The CopB system of plasmid R1 was shown to be a rescue system preventing cells with few plasmid copies from losing the plasmid in some of the daughter cells.</p>

Life is cellular. Cells grow and divide to give two new cells; this process is called the cell cycle. The chromosome in a bacterium is replicated into two identical copies before the cell divides. DNA replication is a fundamental process common to all forms of life. In my thesis, I have studied control of chromosome and plasmid replication in Escherichia coli, a rod-shaped bacterium. Plasmids are extrachromosomal autonomously replicating DNA molecules. I have combined the classical Meselson-Stahl density-shift and DNA hybridisation with theoretical analysis of DNA replication. The minimal time between two successive replications of the same molecule, the eclipse, was determined for both plasmid and chromosome. The aim was to investigate the processes ensuring the precise timing of chromosome replication in the cell cycle. In wild-type strains, the chromosomal eclipse was long. Mutations affecting the so-called sequestration process, the superhelicity of the DNA, and the initiation protein, DnaA, reduced the eclipse. Fast-growing E. coli has overlapping replicative phases with synchronous initiation from multiple initiation sites, oriC. I have investigated the complex interplay between different control processes by measuring the length of the eclipse and the degree of asynchronous initiation in various mutants. I have measured the eclipse period of plasmid R1 during up- and down-shifts in plasmid copy number. The length of the eclipse was found to be determined by structural events as well as by the properties of the copy-number-control system. During downshift from very high copy numbers, the rate of plasmid replication started very slowly and gradually increased until the normal copy number was achieved, in accordance with the +n model. The CopB system of plasmid R1 was shown to be a rescue system preventing cells with few plasmid copies from losing the plasmid in some of the daughter cells.

<p>The primary objective of conventional drinking water treatment and distribution is to deliver to the consumer water that is both aesthetically pleasing and does not constitute a human health risk. To achieve this, water utilities employ a range of physical (i.e. sand and membrane filtration) and chemical (i.e. flocculation and disinfection) barriers in order to reduce the numbers of microorganisms as well as the nutrients that may support their growth within biofilms. In this thesis, biofilms and microbial barriers in water treatment and distribution were therefore examined. The development of biofilms within artificial recharge was investigated in pilot column at Norsborg waterworks in Stockholm. The proportion of active bacteria, measured as numbers of EUB338-positive cells relative to the total number of bacteria enumerated by total direct counts, decreased with time. Through the addition of nutrients however, two to three times more bacteria were able to be active (measured by increase in activity after activation with additional nutrients). By extracting the recalcitrant hydrophilic and hydrophobic fractions of humic substances it was possible to assess the microbiological response to those compounds. It was shown that bacteria more firmly attached to the sand grains preferred the hydrophobic fraction whilst more loosely-associated bacteria preferred the hydrophilic one. The amount of easily degradable matter in raw water (measured as assimilable organic carbon) was generally low. Biofilms were investigated by two different methods for extraction and analysis of microorganisms. Glass slides introduced into the sand material were dominated by α-Proteobacteria, and underestimated loosely-associated bacteria whilst extracts from sand were dominated by γ-Proteobacteria, and also caused variations due to the extraction method employed </p><p>The barrier function of biofilms was investigated in biofilters, also fed with raw water from Gothenburg. The focus here was on particle removal in size-intervals of 1-15 µm (protozoa) and 0.4 - 1 µm (bacteria). In both size fractions, autofluorescent microalgae, which were naturally-occurring in raw water, were also enumerated in parallel. Their removal was 60-90%. In parallel, defined amounts of fluorescent hydrophilic and hydrophobic microspheres (1 µm) were added. They showed a reduction of hydrophobic spheres by 98% and hydrophilic ones by 86%. Removal of viruses was determined by adding a defined dose of bacteriophages and gave lower reduction values of 40 - 61%. Both naturally-occurring particles in defined size intervals and added particles or organisms were shown to provide a clearer picture of barrier function than usually performed measurements of turbidity. </p><p>The efficiency of chemical treatment against viruses was also measured in a pilot-plant in Gothenburg. It was shown that commonly-used MS-2 bacteriophages were much more sensitive than φX174 bacteriophages. Reduction of MS-2 over the entire chemical step (when added after dosing of chemicals) was 5-log10 whilst φX174 was reduced by 1-log10. The latter was shown to be a more conservative model for virus removal. The effects of different steps in the chemical precipitation showed that the primary dosage of chemicals and the development of flocs had great importance for the assessment of removal efficacy. When added before the dosing of chemicals, reduction of φX174 and MS-2 was 3.8-log10 and 6.2-log10, respectively. </p><p>The establishment of biofilm within a distribution system was followed in a 1000 metre long pilot-plant (with parallel lines) at Lovö waterworks as well as in two of Stockholm's main distribution systems (Nockeby and Hässelby). The pilot-plant was shown to satisfactorily represent processes within the distribution systems. The development of biofilms was slow, producing thin biofilms over a one to two month periods. Numbers of bacteria were generally in the range of 104 - 105 per cm2, which is lower than shown in other earlier investigations. The implementation of primary ultra viloet (UV)-treatment in place of chlorination (both being chloraminated prior to distribution) did not considerably change the numbers of bacteria in biofilms. No significant difference could be seen between the system that had UV-treatment as a primary treatment step, and the system that was chlorinated over the whole period. Chlorine residuals were generally low at the distal parts of the distribution systems. Naturally-occurring protozoa were present in distribution systems in numbers ranging from 280 - 3500 protozoa per cm-2. Protozoa may play a significant role as predators of biofilm bacteria, however they can also act as protection for bacteria against external influences i.e. disinfection. Should sudden contamination of a distribution system occur, biofilm can provide protection and act as a site for potential regrowth of introduced microorganisms. Biofilms developed in the pilot-scale that represented water from different distances from waterworks were exposed to fluorescent microspheres, (hydrophobic and hydrophilic, 1 µm) legionellae (as a model for opportunistic bacterial pathogens) and bacteriophages (human enteric virus model) in order to determine their accumulation and persistence within the biofilm, and release to the bulk water. It was shown that introduced model organisms were released continuously, primarily through desorption, and additionally through the influence of disinfection and activity of protozoa. </p><p>Desorption was also assessed in a laboratory experiment under laminar and turbulent flow. Laminar flow conditions that were representative of a distribution system gave a slow and continual release of individual cells, whilst turbulent conditions detached larger aggregates. In conclusion, based on this work an increased understanding was gained both of barrier functions at the different steps of water treatment, their effects on overall biofilm dynamics and structure and the role that biofilm plays within the drinking water system itself.</p>

Thesis (M. Phil.)--University of Queensland, 2002. / Includes bibliographical references.

Clostridium perfringens may be the most widely occurring bacterial pathogen and is responsible for a variety of diseases in both humans and animals. The virulence of this organism is associated with the ability to produce an estimated 17 potential exotoxins. The production of one or more of the five major toxins (α,β,ε, and ι) is the basis for placing isolates into five toxigenic types, A through E. Enterotoxin (CPE), is not used in typing but is considered a major virulence factor. A multiplex PCR genotyping assay was developed, utilizing primers derived from sequences of cpa, cpb, etx, iA, and cpe, yielding products of 324, 196, 655, 446, and 233 bp, respectively. Template for this assay was derived from individual colonies suspended in 200 μl of HPLC-grade water, boiled for 20 min or heated in a microwave oven for 10 min at 700 W. Included in the 50 μl reaction volume was 10 μl of template, 0.15 to 0.7 μM of each primer, 0.1 mM dNTPs, 2 mM MgCl₂, and 2 units of Taq DNA polymerase. The PCR products were examined by electrophoresis in a 1.5% agarose gel stained with EtBr. Correlation of genotype with toxin phenotype in strains examined by mouse inoculation was excellent, and it was possible to provide results rapidly, usually in < 4 h. An ELISA procedure was established for detection of β toxin produced by C. perfringens types B and C. The ELISA was used to differentiate Cpb⁺ from Cpb⁻ isolates grown in overnight broth cultures and to measure β toxin in commercial fermentations of type C organisms. In addition to the above assays, preliminary work was initiated on the development of a PCR procedure for quantitation of C. perfringens in clinical or environmental samples, and involved the construction of a 233 bp homologous, competitive mimic from a restriction digest of a 323 bp PCR product generated from cpa.

No description available.

Single-strain cultures of Streptococcus cremoris, Streptococcus lactis, Streptococcus diacetilactis, and Leuconostoc citrovorum produced little or no acetone and no dimethyl sulfide when grown in milk culture. These organisms had little or no ability to decarboxylate antexogenous source of acetoacetic acid nor were they capable of producing dimethyl sulfide from methyl methionine sulfonium chloride. The dimethyl sulfide content of milk was increased by heating which indicated that a heat labile dimethyl sulfide precursor was present in milk. The precursor remained in the skimmilk fraction and was dialyzable. The precursor was identified as a methyl methionine sulfonium salt on the basis of its thin-layer chromatographic mobility and the heat instability of the compound. Heating of samples caused the disappearance of the precursor compound with a subsequent increase in the content of homoserine and dimethyl sulfide. Single strain cultures of Pseudomonas fragi, Pseudomonas fluorescens, Pseudomonas putrefaciens, and two marine Pseudomonas species reduced acetaldehyde, propionaldehyde, and butyraldehyde to the corresponding alcohols at 21°C. All species studied reduced propionaldehyde at 6°C. P. fragi and the marine species reducted butanone and/or acetone at both 6 and 21°C. Under aerobic conditions a strain of P. fragi quantitatively reduced added propionaldehyde to n-propanol. The quantities of acetaldehyde and ethanol produced by single-strain cultures of Lactobacillus brevis, Lactobacillus casei, Lactolactis, and Lactobacillus plantarum differed significantly both between species and between strains of a species on incubation at both their optimum growth temperature and 8°C. Growth and production of these Compounds were very slow at 8°C. All organisms studied were capable of reducing acetaldehyde and propionaldehyde to the corresponding alcohol. L. brevis strains alone reduced added butanone to 2-butanol. A strain of L. brevis produced n-propanol as a normal metabolite when grown in milk culture. Single-strain cultures of L. casei and L. plantarum accumulated diacetyl when grown in milk culture at both 8 and 30°C, but strains of L. lactis and L. brevis did not. Diacetyl reductase activity was demonstrated in single-strain cultures of L. casei, L. brevis, and L. lactis. Diacetyl reductase could be induced in L. plantarum by growth in the presence of citrate. Growth in milk medium supplemented with citrate resulted in a stimulation of diacetyl reductase activity with L. casei. / Graduation date: 1968

No description available.

No description available.