No description available.

No description available.

European legislation has increased pressure on the use of land to represent the major disposal option for sewage sludge. Owing to their importance in regulating soil fertility, much research has been conducted into the effects of sewage sludge application on soil microorganisms. However, little knowledge is known about its effects on community dynamics of the beta-subgroup ammonia oxidising bacteria. This is despite the fact that nitrification activity of these bacteria plays an important role in soil nitrogen cycling. This investigation aimed to examine community dynamics of the beta-subgroup ammonia oxidising bacteria in sewage sludge amended soil by employing recently developed molecular based techniques. Firstly, a soil DNA extraction protocol was identified that allowed routine nested PCR amplification of 16S rDNA using beta-subgroup ammonia oxidiser directed primers. Reproducibility observed in denaturant gradient gel electrophoresis (DGGE) profiles suggested that 0.5 g samples used for DNA extraction allowed consistent detection of dominant beta-subgroup ammonia oxidiser community members. The effects of applying primary treatment sewage sludge to a variety of contrasting soils on (3-subgroup ammonia oxidiser community structure was unclear. This was partially hampered by lack of specificity of PCR primers for non ammonia oxidiser 16S rDNA sequences. Further, through measurement of net nitrification, there was no indication that sewage sludge addition stimulated the activity of nitrifier populations. Nevertheless, this study indicated the usefulness of DGGE for screening multiple environmental samples. A set of hierarchical oligonucleotide probes exhibiting specificity at the group, genus and cluster level were optimised using a non-radioactive system. These probes were used to assess the effects of application of aerobically digested sewage sludge to soil for 4 y on beta-subgroup ammonia oxidiser community structure. Despite significant changes in measurable soil parameters including net nitrification activity no changes were observed in beta-subgroup ammonia oxidiser community structure. This indicated resilience of these communities to change. A final study was conducted examining the effects of application of anaerobically digested sewage sludge to soil on beta-subgroup ammonia oxidiser community structure. Inhibition of nitrification by acetylene indicated the presence of a viable population of ammonia oxidising bacteria in continuously shaken samples of sludge. DGGE and oligonucleotide probing analysis provided evidence that sewage populations had the potential to outcompete indigenous soil populations of beta-subgroup ammonia oxidising bacteria. This was despite the fact that MPN enumeration suggested that soil populations of these bacteria were in some cases 10-fold greater than sludge populations. Evidence was also provided that suggested net nitrification to be an unreliable indicator of ammonia oxidiser activity in soil. It is possible to conclude that community structure of the beta-subgroup ammonia oxidising bacteria may be altered by application of sewage sludge to soil. The effects on community structure are likely to be influenced by both the type and level of sludge applied to soil.

Streptococcus pneumoniae is a highly diverse and adaptable opportunistic pathogen that can infect and colonise different niches within the human host to cause a wide range of invasive disease (sepsis and meningitis) and noninvasive disease (pneumonia, otitis media and sinusitis). The molecular mechanisms that contribute to the different patterns of pneumococcal infection remain largely unknown. This thesis aims to determine the physiological and proteomic responses that allow the pneumococcus to survive and adapt to invasive and non-invasive sites. The comparative proteomic analyses of clinical S. pneumoniae isolates recovered from blood cultures (classified as invasive site isolates) and mucosal surfaces such as sputum, skin and ear swabs (classified as non-invasive site isolates) was initiated. The pneumococci were grown in vitro under standard conditions and the total cellular bacterial proteins extracted and analysed using both gel based and non-gel based proteomic approaches. Analysis of the pneumococcal isolates by two-dimensional polyacrylamide gel electrophoresis (2DGE) revealed that a high degree of heterogeneity existed between the pneumococcal isolates particularly among isolates in the invasive site isolates. Differential patterns of protein synthesis were observed that discriminated the pneumococcal isolates according to their sites of isolation. These were proposed to be associated with the bacterial adaptation to invasive and non-invasive sites of infection. Mass spectrometry was used to identify selected significant (ANOVA, p < 0.05) protein spots, which were further categorised into functional groups by Gene Ontology analysis. An extension of the 2DGE data using an integrated approach comprising bioinformatics, surfome analysis and a shotgun proteomic workflow provided a comprehensive qualitative and quantitative analyses of the pneumococcal intracellular and cell-surface proteomes. Proteins potentially involved in pneumococcal niche-specific adaptation and surface proteins with potential for further investigation and inclusion in the pipeline of vaccine candidates were identified. Quantitative regulation of proteins involved in energy metabolism, genetic competence, stress response, surface adhesion and virulence were considered important for pneumococcal adaptation to invasive and non-invasive sites. The anatomical sites colonised by the pneumococcus vary in their V availability for iron. The 2DGE method was also used on selected pneumococcal isolates from the two sites of infection to define the proteome variability linked to the effect of iron starvation that may contribute to the different disease outcomes associated with pneumococcal infections. The iron restricted condition was generated by cation depletion of the growth medium using Chelex-100. Quantitative differences in protein abundance were demonstrated that correlated with pneumococcal adaptation to iron restriction. The identification of selected significant spots by liquid chromatography-mass spectrometry and systems biology analysis of the identified proteins contributed to the elucidation of the molecular mechanisms underlying pneumococcal survival under iron limitation. The expression/repression of proteins functionally associated with metal ion binding, oxidative stress response, translation and virulence mainly constituted the pneumococcal adaptive responses to growth under conditions of limited iron availability. The data presented in this thesis extended our understanding of the molecular events underlying pneumococcal physiological adaptation and provide the basis of future work in this area.

No description available.

A unique opportunity to study both benthic microbial mats and modern stromatolites from a common niche is presented by the hypersaline environment of Hamelin Pool, Shark Bay, Australia. However, prior to this study, the microbial mat communities have not been well characterised. To investigate the taxonomic and functional diversity of Hamelin Pool pustular and smooth mats, and their similarity to Hamelin Pool stromatolites, culturing, culture-independent, and lipid analysis methods were employed. The cultured isolates obtained included heterotrophic bacteria similar to those obtained from other hypersaline environments, and 19 strains of cyanobacteria including potentially novel species. For the first time archaeal isolates were obtained from the pustular and smooth mats, and further characterisation of two strains indicated they might represent novel species of the genus Haloferax. Total DNA was extracted from the mats and 16S rRNA gene clone libraries were generated targeting the bacteria, cyanobacteria, archaea and eukarya. Both the pustular and smooth mat bacterial clone libraries were highly diverse, with 11 bacterial divisions represented, and Chao1 estimates of total species richness indicating ~3000 ??? 6000 species. Cyanobacterial and archaeal clone libraries revealed unique phylotypes associated with sediments of differing morphology. Statistically significant differences between the mat populations and Hamelin Pool stromatolite communities were identified using !-LIBSHUFF, a program designed to compare two 16S rRNA gene libraries. Signature lipid biomarkers were assessed for the pustular and smooth mats and an intertidal stromatolite from Hamelin Pool. Fatty acids (as methyl esters), wax esters, hydrocarbons, ether-bound lipids, hopanoids and sterols indicated the presence of oxygenic phototrophs, anoxygenic phototrophs, sulfate-reducing bacteria, sulfur-oxidising bacteria, heterotrophic bacteria and archaea in each of the sediment types. Limited contributions from diatoms, bivalves and their dinoflagellate symbionts, and from higher plant aerosols were also detected. Significantly, 2-methyl hopanoids and eight pseudohomologous series of branched alkanes with quaternary carbon centers were identified in the mats and stromatolites. Although differing in lithification status and precise microbial composition, the pustular and smooth mats are excellent analogues for the extant stromatolites of Hamelin Pool, and by corollary, provide a fascinating link to Precambrian microbial communities.

The objectives in this research were to assess whether Klebsiella planticola SDF20 could survive in soil and result in ecological effects to soil foodweb organisms and plant growth. Four experiments were conducted using soil microcosms. Klebsiella planticola SDF20 has been genetically engineered to produce ethanol from agricultural waste for use in alternative fuels. Theoretically, after ethanol is removed from fermentors, the remaining residue that includes SDF20 would be spread onto crop fields as organic amendments. The parent strain SDF15 and genetically engineered strain SDF20 were added to sandy and clay soils with varying organic matter content. Alterations to soil foodweb organisms and plant growth were assessed using direct methods. These alterations were considered to be ecological effects if changes in nutrient cycling processes and plant growth would result. Ethanol produced by SDF20 was detected in the headspace of microcosms that demonstrated that SDF20 can survive and express its novel function in high organic matter clay soil. Soil containing higher organic matter and higher clay content may have increased the survival of SDF20 due to less competition with indigenous microbiota for substrates and protection from bacterial predators in clay soil with smaller pore sizes, thereby allowing SDF20 to produce a detectable concentration of ethanol. Significant changes to soil foodweb organisms were not detected using this soil type. However, significant increases in soil nematodes and significant decreases in vescular-arbuscular mycorrhizal colonization of plant roots were detected after the addition of SDF20 to low organic matter clay, low organic matter sandy and high organic matter sandy soils. Significant changes in soil foodweb organisms associated with SDF20 occurred only when living plant roots were present. This indicated the importance of having biotic interactions in test systems to elucidate ecological effects. The effects associated with SDF20 varied with the chemical, physical and biological properties of soils and indicated the importance of assessing the release of genetically engineered microorganisms on a case by case basis. / Graduation date: 1996

Thesis (Ph. D.)--University of Missouri-Columbia, 2000. / Typescript. Vita. Includes bibliographical references (leaves 67-78). Also available on the Internet.

Assignment (MCur)--University of Stellenbosch, 2008. / Bibliography. Also available via the Internet.

Thesis (Ph. D.)--University of Missouri-Columbia, 2000. / Typescript. Vita. Includes bibliographical references (leaves 67-78). Also available on the Internet.