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Doctor of Philosophy / Dental caries continues to be a significant public health problem affecting mankind in many parts of the world. Microbial activities include the progressive localised destruction of teeth that without treatment, would eventually result in infection of the dental pulp and surrounding periapical tissues. Although the bacteria responsible for caries initiation and early caries progression have been extensively studied, the microbiology of dentine caries is reported to show considerable diversity and has not yet been fully identified. Few studies have analysed the microbiology of deep caries or examined the relationship between the microflora and the histopathy of chronic pulpits in symptomatic teeth. Matched carious dentine samples and dental pulps were obtained from teeth without evidence of periodontal disease but with coronal caries and symptoms of pulpits. Bacteria were cultured from the carious dentine samples under both anaerobic and microaerophilic conditions. Real-time polymerase chain reaction (PCR) technology was also used to identify and enumerate the bacteria. Development of the techniques for the efficient extraction of bacterial DNA from both Gram-negative and Gram-positive bacteria found in carious dentine was an essential prerequisite for molecular analysis. In addition, the dental pulps were processed and categorised into one of four groups on the basis of dominant pathology of the tissue (minimal inflammation, soft tissue degeneration, hard tissue degeneration, inflammatory degeneration). Analysis of the culture data indicated a predominance of Gram-positive bacteria, particularly lactobacilli, while Gram-negative bacteria were also present in significant numbers with Prevotella species the most numerous anaerobic group cultured. Real-time PCR indicated a greater anaerobic microbial load than that determined by colony counting. The total number of anaerobes detected by PCR was 41-fold greater, while Prevotella spp. and Fusobacterium ssp. were 82-fold and 2.4-fold greater respectively. PCR also identified the presence of Micromonas micros, Porphyromonas endodontalis and Porphyromonas gingivalis in 71%, 60% and 52% of carious dentine samples, respectively. Correlation matrices from the real-time PCR data revealed significant multiple associations involving Fusobacterium spp. in combination with P. endodontalis, M. micros and/or Prevotella in the tissue response categories of minimal inflammation, soft and hard disuse degeneration. A positive correlation was also observed between M. micros and P. endodontalis for the category of inflammatory degeneration of the dental pulp. These anaerobes have been strongly implicated in the endodontic infections that occur as sequelae to carious pulpitis. Accordingly, the data suggest that the presence of threshold levels of these bacteria in carious dentine may be indicative of irreversible pulpitis. Knowledge of the microbial predictors associated with irreversible pulpitis creates potential for the development of a diagnostic tool, and for restorative materials with antimicrobial properties.

Thesis (Ph.D. Biological Sciences)--University of Waikato, 2007. / Title from PDF cover (viewed March 6, 2008) Includes bibliographical references (p. 111-134)

Thesis (Ph. D.)--Institute of Dental Research, Faculty of Dentistry, University of Sydney, 2002. / Title from title screen (viewed Apr. 23, 2009) Submitted in fulfilment of the requirements for the degree of Doctor of Philosophy to the Faculty of Dentistry. Includes bibliography. Also available in print form.

Three different methods were developed to evaluate nutritional status: stable isotopes, near infra-red reflectance spectroscopy (NIRS), and microbial populations. In the first method, hair samples were collected, analyzed, stable isotope content greatly differed (P < 0.01) between pre- and post-weaning samples. The second method used NIRS spectral patterns and analyzed fecal samples, and a difference was found around 2100 to 2250 nm where diets containing milk showed a peak, and non-milk diets showed a plateau. Finally, the third method involved profiling microbial populations from fecal samples as calves experienced dietary shifts. Total anaerobes, enterics, and clostridium were different (P<0.05) from pre to post-weaning states. Total anaerobe samples differed (P < 0.005) from milk, milk-grain diets to grain and grain-hay diets. Total aerobes, streptococci, and clostridium samples differed (P <0.001) from the milk diet to the other diets. In summary, all three methods showed differences when comparing pre- to post-weaning states.

Arsenic (As) is a problem with rice production systems throughout the world as high As concentrations are reported in rice grains originating from several parts of the world. This characteristic is mainly due to the flooded conditions utilized in rice culture. We hypothesized that the soluble As concentrations in the rice rhizosphere can be decreased by growing rice more aerobically through intermittent flooding. Intermittent water management practices might also change microbial populations in the rice rhizosphere that might potentially impact As chemistry and bioavailability. Two field-scale experiments were conducted over two years to study the impact of intermittent and continuous flooding on As speciation and microbial populations in the rice rhizosphere. As levels and speciation in the rhizosphere soil, root-plaque and pore-water were determined using a high performance liquid chromatography-inductively coupled plasmamass spectroscopy (HPLC-ICP-MS). The microbial populations were assessed from the rhizosphere soil and root-plaque samples using quantitative polymerase chain reaction (qPCR) and 16S rRNA sequencing. Pore-water and root-plaque total-As concentrations significantly decreased in the intermittent compared to the continuous flood plots. Inorganic arsenite (iAsIII) was predominant in pore-water and inorganic arsenate (iAsV) in root-plaque and soil. Rootplaque sequestered significantly higher levels of As (almost tenfold higher) than the adjacent rhizosphere soil. Grain As concentrations also decreased by 35 to 45 percent in the intermittent compared to the continuously flooded plots. Organic As species, monomethyl and dimethyl arsenate were detected in the rhizosphere with relative increases and decreases among the treatments. Bacteria were the predominant group (91 to 94 percent and 48 to 78 percent of total community in root-plaque and rhizosphere soils, respectively). Archaea were also a major component of rhizosphere soil with their populations being higher under continuous flooding. The relative abundance of iron-reducing bacteria was around 3 to 6 percent of the total community in root-plaque and around 6 to 6 percent in soil, with significantly lower abundance in the intermittent compared to the continuously flooded plots. Results of these studies demonstrated that intermittent flooding could be a potential management option to reduce grain As in rice cultivated on fields with moderate to high As concentrations.

High-moisture ear corn (HMEC) was untreated, treated with propionic acid (PA) or inoculated with a mixture of Lactobacillus plantarum and Streptococcus faecium, and then ensiled in both tower and laboratory silos. Ensiled HMEC was evaluated for fermentation quality, silage microbiology, and aerobic stability. In control and inoculated ensiled HMEC, maximum pH reduction was observed within 7 d; such pH reduction was observed only after 21 d of ensiling with PA-treated HMEC. Irrespective of treatment, ammonia concentration increased with storage time. The lactic acid content increased up to 42 d of ensilage; between 138 d and 202 d of ensilage, there was a substantial reduction in lactic acid with all treatments. Throughout ensilage, there was no difference in water soluble carbohydrate (WSC) content between control and inoculated HMEC. Secondary fermentation was minimized by both inoculation and PA treatment. Regardless of treatment, the population of lactic acid bacteria increased within 7 d of ensilage and reached a maximum at 21 d of ensilage. Populations of yeasts and moulds decreased with fermentation time up to 42 d, then increased as the ensilage progressed. Bacterial inoculation increased aerobic stability of ensiled HMEC. PA was more effective than inoculation in reducing the disappearance of lactic acid and the rise in pH. Irrespective of treatment, the population of yeasts and moulds and other microorganisms increased in aerated HMEC. Both PA treatment and bacterial inoculation reduced aerobic spoilage of ensiled HMEC. (Abstract shortened by UMI.)

Determining the factors responsible for the origin and maintenance of diversity remains a difficult problem in evolutionary biology. There is extensive theoretical work which suggests that environmental heterogeneity plays a major role. This theory argues that diversification is ultimately due to divergent natural selection for alternative resources. In this thesis I investigate adaptation and the evolution of diversity in experimental populations of the asexual bacterium Pseudomonas fluorescens. In all experiments I introduce clonal isolates of Pseudomonas to a novel environment and allow evolution to occur through the substitution of random mutations. Adaptation can then be quantified by comparing evolved genotypes to the ancestor. These experiments show that when Pseudomonas is selected in a complex environment containing several resources, sympatric genotypes adapt to use different resources, leading to the evolution of genetically diverse populations. In environments containing just a single resource, most genotypes adapt to use the same resource and no such diversity is observed. Adaptation in the experimental populations is caused by the fixation of beneficial mutations of intermediate fitness effect. My results highlight the value of microbial model systems for answering evolutionary questions and provide strong evidence for the role of ecological factors in the origin of diversity.

Understanding the origins of biological diversity is a fundamental goal of evolutionary biology. A large body of theory attributes ecological and genetic diversification to divergent natural selection for resource specialization. This thesis examines adaptive radiation in response to selection for resource specialization in microcosm populations of the asexual bacterium Pseudomonas fluorescens. The general protocol for these experiments is to introduce a clonal population of Pseudomonas into a novel environment and to allow evolution to occur through the spontaneous appearance of novel genotypes carrying beneficial mutations. Adaptation can then be quantified through direct comparisons between evolved populations and their clonal ancestors. These experiments show that resource heterogeneity generates divergent natural selection for specialization on alternative resources, irrespective of the spatial structure of the environment. Adaptive radiation is possible in sympatry because of genetic trade-offs in the ability to exploit different resources, but these trade-offs are often not the result of antagonistic pleiotropy among loci that determine fitness on alternative resources. The rate of phenotypic diversification declines during adaptive radiation, apparently because the ecological opportunities required to support specialist lineages disappear as a consequence of initial diversification. The ultimate outcome of repeated instances of adaptive radiation is the evolution of a community of ecologically equivalent specialists that share similar adaptive traits, despite differences in the underlying genetic basis of specialization in replicate radiations. Comparisons with the literature on experimental evolution in microbial populations illustrate the results of this thesis are well-supported by experiments in a wide range of microbial microcosms.

Thesis (Ph. D. in Mathematics)--Vanderbilt University, Dec. 2005. / Title from title screen. Includes bibliographical references.