Dissertations / Theses on the topic 'Microbial Resources Management'

Bois Noir (BN) is a disease of the grapevine yellows (GY) complex associated with ‘Candidatus Phytoplasma solani’ (CaPsol) strains, which causes economic crop losses in viticulture worldwide. The epidemiology of BN is very complex due to the involvement of different herbaceous plants and several insect vectors that transmit CaPsol to grapevine. Therefore, the BN containment is very difficult and require massive efforts for possible spread reduction. The heavy application of chemical insecticides was not successful to control the insect vector presence within the vineyard. The thesis work was framed considering the directives provided by the European council 2009/128/EC regarding the promotion of low use of pesticides in sustainable management approaches. In the present thesis dissertation, CaPsol insect vectors and diseased grapevines were the main targets prioritized for successful containment of BN in organically cultivated vineyards in northern Italy. Since H. obsoletus is the widely distributed insect vector in Europe, the management of the leafhopper population was carefully considered. The use of Vitex agnus-castus as trap plant for H. obsoletus as an indirect control strategy was evaluated. Vitex agnus-castus tended to be a preferred host plant for H. obsoletus, but transmission trials demonstrated its ability to harbor CaPsol and indicated the impossibility of using this plant to avoid BN spread. In addition, the efficacy of different entomopathogenic nematodes and fungi as direct control strategy were evaluated against H. obsoletus nymphs and adults. Their application in a laboratory and semi-field conditions showed a promising killing effect that can be implemented for insect vector control in open field. Due to the very low density of H. obsoletus population in heavily BN infected vineyards questions were raised to figure out the other possible presence of alternative insect vectors. Surveys on Auchenorrhyncha coupled with molecular analyses revealed the presence of numerous putative vectors. Some of them, selected on the basis of their abundance, CaPsol-infection rate and CaPsol strains harbored, went through transmission trials. Eight insects were found able to transmit CaPsol to grapevines. Characterization of the bacterial microbiota associated with H. obsoletus and the alternative insect vectors indicated an interesting perspective regarding the microbial signatures associated with xylem- and phloem-feeding insects, and determinants that could be relevant to establish whether an insect species can be a vector or not, opening up new avenues for developing microbial resource management-based approaches. Moreover, grafting of materials collected from recovered grapevines was conducted in field trials with the aim to evaluate its preventive and curative potentials against BN. Results of symptom observation and CaPsol molecular detection on grafted and non-grafted grapevines showed that grafting of recovered shoots can have a curative effect, increasing the natural recovery. Results obtained in this PhD thesis opened new perspectives to develop integrated sustainable strategies for BN management.

The Water Framework Directive requires reduced environmental impacts from human activities and for the assessment of the non-market benefits of pollution remediation schemes. This policy shift has exacerbated the research problems surrounding the physical, social and economic consequences of the relationship between land use and water quality. This research seeks to quantify the major socio-economic and environmental benefits for people which may arise as riverine pollution is reduced. To achieve these aims this research integrates primary data analyses combining choice experiment techniques with geographical information system based analyses of secondary data concerning the spatial distributions of riverine pollution. Current knowledge on the microbial quality of river water, measured by faecal indicator organism (FIO) concentrations and assessed at catchment scale, is inadequate. This research develops generic regression models to predict base- and high-flow faecal coliform (FC) and enterococci (EN) concentrations, using land cover and population (human and livestock) variables. The resulting models are then used both to predict FIO concentrations in unmonitored watercourses and to evaluate the likely impacts of different land use scenarios, enabling insights into the optimal locations and cost-effective mix of implementation strategies. Valuation experiments frequently conflate respondents’ preferences for different aspects of water quality. This analysis uses stated preference techniques to disaggregate the values of recreation and ecological attributes of water quality, thereby allowing decision makers to better understand the consequences of adopting alternative investment strategies which favour either ecological, recreational or a mix of benefits. The results reveal heterogeneous preferences across society; specifically, latent class analysis identifies three distinct groups, holding significantly different preferences for water quality. From a methodological perspective this research greatly enhances the ongoing synthesis of geographic and economic social sciences and addresses important policy questions which are of interest to a variety of stakeholders, including government departments and the water industry.

During the last decades many studies have been undertaken to investigate life in extreme environments, leading to the discovery of novel organisms and novel habitats previously though to be unapproachable for life. Microbes are key players in a number of ecological processes such as mineral dissolution, soil genesis, plant growth promotion (PGP) and bioremediation of polluted sites and they are the main responsibles for element cycles both in conventional and extreme ecosystems. The biotechnological potential of extremophiles is well recognized, and the aim of this PhD project was to give further insight on the possible exploitation of the microbiome naturally adapted to cope with extreme values of one or more environmental parameters to develop sustainable strategies in agriculture and ecosystem management with a particular focus on arid and saline lands. Mineral-microbe interactions have been studied in detail, particularly regarding the importance of bioweathering bacteria in the ambit of soil fertility promotion in arid lands. Specific sites within the Midtre Lovénbreen glacier moraine (Svalbard, Norway), where pyritic rocks were present, hosted an active acidophilic iron-oxidizing bacterial community involved in the bioweathering of pyrite supplied by the rock disaggregation due to winter freezing. A decreased iron concentration and acidification were observed along the wheathered area departing form the pyrite-rich rock, where the oxidation of ferrous iron led to the accumulation of ferric oxy-hydroxides in the above soil. These ferric compounds were linked to the increase of soil physico-chemical properties that in turn determined a higher water holding capacity (WHC) and nutrient content in the surrounding vegetated area, densely colonized by mosses and small vascular plants. At the outer border of the vegetated area, the rest of the moraine hosted typical first colonizer bacteria, mainly belonging to the class Cyanobacteria, that are capable of nitrogen and carbon fixation. Thus, compared to the rest of the moraine, the enhancement of soil formation processes and plant colonization in the vegetated area was driven by the synergy between acidification and leaching activity of a chemolitotrophic community and the cyanobacteria-mediated primary productivity. A detailed description of the bacterial communities colonizing the weathered area, the vegetated area, and the barren moraine was obtained through the construction of 16S rRNA gene libraries. The statistical ∫-Libshuff analysis indicated these areas as three different ecological niches. The microbiome of the weathered area was dominated by few bacterial taxa due to the low pH value of the biological soil crust (BSC) whereas the vegetated area and the moraine displayed higher biodiversity. The most abundant phylogenetic groups in these BSCs were nevertheless different and in the case of the vegetated area they corresponded to those typical of mature and rhizospheric soils. The ability of microorganisms to interact with minerals is an essential factor that influence plant nutrition by providing nutrients, such as phosphorous, that are generally present in the soils as insoluble forms. The capability to solubilize poorly bioavailable nutrients is one of the PGP activities that have been investigated in the microbiome associated to different plant species living in arid hypersaline soils in Central and South Tunisia (Olea europea and Salicornia spp.) or acid soils located in a volcanic area in Mexico. A large collection of bacterial isolates has been constituted, identified and characterized for the in vitro PGP potential. Halophilic bacteria were isolated from the rhizosphere of Salicornia plants on oligotrophic media enriched with NaCl. The isolates obtained from 15% NaCl enriched media mainly belonged to the Halomonas genus, whereas the bacteria isolated at 10% NaCl showed a higher phylogenetic diversity at the genus level. Most of the bacteria comprised in the halophiles collection exhibited high resistance to drought, temperature and salt stresses. PGP activities were also widespread, especially the ability to produce indol-3-acetic acid (IAA), which promotes lateral roots developement. Furthermore, high percentage of the halophilic bacteria produced ammonium (94%) and were able to solubilize phosphate (64%) while the ability to produce protease, an activity involved in biocontrol processes, was less frequent. The comprehensive study realized on the culturable halophilic fraction of the rhizospheric bacteria associated to Salicornia spp. allowed the identification of 20 isolates as suitable candidate for developing a bacterial inoculum aimed to promote plant growth under saline stress. The diversity of the microbiome inhabiting different fractions of the Olea europea root system was investigated by applying a cultivation-independent method (Denaturing Gradient Gel Electrophoresis). The interior root tissues, the rhizosphere, the root surrounding soil and the bulk soil were colonized by a rich and diverse microbiome, shaped both by interaction with the plant and the environmental parameters of the collection site. Moreover, from these four fractions a bacterial collection was obtained for the screening of ecological and PGP features of culturable bacteria associated to olive tree growing under drought stress. In addition to the abiotic stress resistance, bacterial isolates displayed a variety of PGP activities, such as potential nitrogen fixation, siderophores and exopolysaccharides production and phosphate solubilization. Overall, the obtained dataset highlighted the possibility to use the investigated PGP bacteria associated to olive tree as biofertilizer for supporting olive growth under drought stress. An extremophile plant living at high T (42°C) and low pH (4.1) was collected at El Chichón volcanic system (Mexico). The PGP activities of a collection of rhizobacteria isolated from the plant were explored through in vitro tests. Several strains were able to affect phytohormones balance by the production of indole-3-acetic acid and 1-aminocyclopropane-1-carboxylate (ACC)-deaminase, the latter being involved in the decrease of ethylene level in plants. Remarkable percentage of the isolated bacteria displayed also additional potential PGP activities based on weathering activity. Volcanic habitats can hence be estimated as source of extremophile rhizobacteria potentialy able to help pioneer plants to cope with the severe condition of acidic soils. Microbe-environment interactions have been investigated also in deep hypersaline anoxic basins (DHABs) located in the eastern Mediterranean Sea, model environments to look for bacterial phylotypes specifically adapted to high salinity conditions. Mediterranean DHABs are far below the photic zone and contain brines originated by the dissolution of Messinian evaporites. Compared to other DHABs, Urania has very high concentrations of methane (5.56 mM) and levels of sulfide (up to 16 mM) that make it one of the most sulfidic marine water bodies on Earth. The interface between seawater and the anoxic hypersaline brine of DHABs is an oxic-anoxic interface containing an halocline with layers from seawater to brine typical salinity and is a hot-spot of microbial activity. Methanogenesis activity was detected along the first of the two environmental chemoclines present in the Urania basin and 16S rRNA gene libraries indicated the Euryarchaeota group MSBL1 as the primary candidates for methane production. Cultivation-independent analyses proved that sulfur cycling is a major driver in shaping the microbial communities, though other chemolithoautotrophic processes like manganese oxidation and anaerobic ammonium oxidation (ANAMMOX) are involved. The occurrence of ANAMMOX reaction was verified in other DHABs, namely L’Atalante and Bannock. Labelled dinitrogen gas production in 15N activity test demonstrated that anammox bacteria were active in the chemoclines of both the basins. Fluorescence in situ hybridization and 16S rRNA gene libraries using anammox-specific PCR primers unveiled the presence of the known marine anammox genus ‘Scalindua’, together with putatively novel operational taxonomic units (OTUs) closely affiliated to sequences retrieved in other marine environments where anammox activity were detected. Real Time PCR assay allowed to quantify anammox-related 16S rRNA genes in Bannock basin, which were highly abundant in correspondence of the oxic-anoxic boundary in the salinity range comprised between 6.4 and 12.1%. Cluster analysis of 16S rRNA gene libraries showed that chemoclines of Bannock and L’Atalante basins, having diverse geochemical settings, selected for different anammox phylotypes and that a shift in anammox population could be observed at increasing salinity values. The detection of putative novel phylotypes specifically adapted to peculiar salinity levels represent a key step for designing ad hoc inocula to be used in the remediation of saline wastewaters originated by industrial and agricultural processes. Actually the known freshwater anammox populations could only adapt to salt concentrations up to 3% if salinity is slowly increased, thus the selection of naturally adapted anammox strains would be of primary importance to enhance the exploitation of this process during the removal of nitrogen compounds from wastewater. The occurrence of microbe-plant positive associations was proved in different stressed soils, and their exploitation is likely the most promising approach to avoid or reduce the use of chemical fertilizer and to boost plant growth and crop productivity whitout the use of genetically modified organisms (GMO) in respect of the biodiversity. Similarly, the discovery of novel anammox phylotypes in hypersaline ecosystems shed a new light on the utilization of this functional group of bacteria for the removal of nitrogen from saline wastewaters, a critical step of treatment processes due to the environmental impact of nitrogen compounds and the severe legislation on wastewater discharges.

Waterborne disease is a significant contributor to the global burden of disease, in particular among high-risk populations in developing nations. State-of-the-art methods for the enumeration of microbial pathogens in drinking water sources have important limitations, including high initial cost, 24-48 hour delays in results, high staffing and facility requirements, and training requirements which all become especially problematic in the developing nation context. A number of alternative approaches to microbial water quality testing have been proposed, with the goal of decreasing the required testing time, decreasing overall costs, leveraging appropriate technology approaches, or improving sensitivity or specificity of the water quality testing method. One approach that may offer solutions to some of these limitations involves the deployment of sensor networks using fluorescent spectroscopy to detect intrinsic protein fluorescence in water samples as a proxy for microbial activity. In recent years, a number of researchers have found significant and meaningful correlations between indicator bacteria species and the protein fluorescence of drinking water samples. Additionally, advances in the semiconductor industry could be used to drive down the cost of such sensors. This technology may also be extensible to other water quality parameters, including dissolved organic matter or the presence of fluorescent pollutants. In this thesis, a literature review describes the fundamentals of fluorescence spectroscopy, historical and recent work regarding the fluorescence of the amino acid tryptophan and associated bacterial fluorescence, possible mechanisms for this association, and potential applications of this technology for drinking water quality monitoring and waste water process control. Extensibility of the technology is also discussed. Next, experimental methodology in reproduction of similar results is described. Samples were taken from seven (7) surface water sources and tested using membrane filtration and an off-the-shelf fluorescence spectrometer to help examine the association between the presence of indicator bacteria and the tryptophan fluorescence of the water sample. The results, showing an association of R2 = 0.560, are compared to the results of recent similar experiments. Finally, two prototypes are described, including their design requirements and data from prototype testing. The results of the testing are briefly discussed, and next steps are outlined with the goal of developing a low-cost, in-situ microbial water quality sensor using fluorescence spectroscopy principles.

It has been reported that globally we have achieved Millennium Development Goal (MDG) Target 7C, to halve the proportion of the population without access to safe drinking water; however, there is a major flaw with this statement. While Target 7C calls for access to `safe' drinking water, what is actually being measured and reported is access to an `improved' water source. The World Health Organization (WHO) maintains that they must use this proxy measure because the methods for water quality testing are too expensive and logistically complicated, but by doing so, they may be over reporting safe water coverage. This was shown to be true in Tamatave, Madagascar, where thermotolerant coliforms were detected in water from a type of `improved' source, the Pitcher Pump system. This research looked at several parameters - Pitcher Pump system depth, sampling neighborhood, requirement of pump priming, frequency that the system was repaired, distance from on-site sanitation, and number of users - to see if they were influencing water quality. Of all the parameters tested, only priming was found to be significantly associated with the levels of thermotolerant coliforms detected (Fisher exact test p = 0.03). Using a Mann-Whitney U test, it was shown that the median thermotolerant coliform concentration was significantly higher in primed wells (41.3 cfu/100 ml) than unprimed wells (3.5) (p = 0.01 cfu/100 ml). A pilot study was conducted to look at only the effect of depth and to determine if a depth could be identified that could provide safe drinking water. The result of the pilot study showed that, while thermotolerant coliform concentration did decrease with increasing depth, even at the deepest well of 9.4 m, levels were still above 100 cfu/100 ml. Additional research was conducted to investigate the performance and cost of three test kits for both total coliform and Escherichia coli quantification for water quality analysis in developing countries. IDEXX Colilert Quanti-trays[reg] (Colilert), Micrology Laboratories Coliscan[reg] Membrane Filtration tests (Coliscan MF) and a modified method for 3-M PetrifilmTM Coliform/E. coli plates (modified 3-M) were compared with standard membrane filtration (standard MF) methods under a range of incubation temperature conditions (22.0, 35.0 and 44.5[deg]C). Each test method was also performed by inexperienced volunteers, with the results compared to those of an experienced technician. At non-standard temperatures, Coliscan MF proved to be the most accurate when compared to standard methods, with a significant difference with only total coliforms at 44.5[deg]C. Modified 3-M had the poorest correlation with standard MF over the range of temperatures tested, with significant differences noted for all the temperatures except for E. coli at 44.5[deg]C. Inexperienced university volunteers found Colilert easiest to use, but Coliscan MF produced E. coli results that were most similar to the experts. Coliscan MF was found to have the overall best performance and lowest cost in this study; however, it did produce high numbers of false positive results.

Reliable access to safe drinking water is one necessity for humans to live without concern for major health risks. The overall goal of this research is to improve the public health, through improved drinking water, for communities in the Rakai District in Uganda, directly, and other communities in the world, indirectly, via dissemination of knowledge. This study specifically assessed the knowledge of drinking water quality in regards to public health, their sanitation measures, and water treatment methods for users of Brick by Brick rainwater harvesting tanks in the Rakai District (N = 28) by using a knowledge, attitudes, and practice survey and a sanitary inspection; tested the water quality of the Brick by Brick rainwater harvesting tanks (N = 33) in the Rakai District for physical, chemical, and microbial parameters; and piloted a sustainable treatment technology called the chulli system that uses excess heat from a cookstove to treat water. Twenty of the participants identified contaminated water as a cause of diarrheal disease (N = 28). Participants perceived boiling (1), chlorine (2), and filtering (3) as the best three methods of treating water. The average score for the sanitary inspection was 2.27±2.31, which falls between the low and medium expected risk score categories. Fourteen of the thirty-three samples showed detectable levels of colony forming units for coliforms, and two of the thirty-three samples showed detectable levels of colony forming units for E. coli. A demonstration chulli system was constructed for St. Andrew’s Primary School in Rakai District and operated successfully. The research supports that the chulli system along with proper sanitation measures identified in the sanitary inspections can be a sustainable option for users of Brick by Brick rainwater harvesting tanks in the Rakai District.

Microalgae are considered one of the most promising feedstocks for biofuel production for the future. The most efficient way to produce vast amounts of algal biomass is the use of closed tubular photobioreactors (PBR). The heat requirement for a given system is a major concern since the best algae growth rates are obtained between 25-30 °C, depending on the specific strain. A procedure to determine temperature influence on algal growth rates was developed for a lab-scale PBR system using the species Chlorella. A maximum growth rate of 1.44 doublings per day at 29 °C (optimal temperature) was determined. In addition, a dynamic mathematical model was developed to simulate heating and cooling energy requirements of tubular PBRs for any desired location. Operating the model with hourly weather data as input, heating and cooling loads can be calculated early in the planning stage of a project. Furthermore, the model makes it possible to compare the operation inside a greenhouse to the outdoor operations, and consequently provides fundamental information for an economic feasibility study. The best configuration for a specific location can be evaluated easily. The model was exemplary tested for a hypothetical 100,000 l photobioreactor located in San Luis Obispo, California, U.S.A. Average algae productivity rates of 23% and 67% for outdoor and indoor PBR operations, respectively, were obtained. Actual energy loads (heating and cooling) needed to maintain the PBR at optimal temperature were determined and compared. Sensitivity analyses had been performed for abrupt temperature and solar radiation steps, PBR row distances, ground reflectivities, and ventilation rates of the greenhouse. An optimal row distance of 0.75 m was determined for the specific PBR. The least amount of energy was needed for a ground reflectivity of 20%. The ventilation rate had no major influence on the productivity rate of the system. Results demonstrated the importance of a simulation model as well as the economic impact of a sophisticated heat management system. Energy savings due to an optimized heat management system will eventually increase proficiency of the systems, which will support a new sustainable industry and future developmental potential.

Sanitation, renewable energy, and food security are among the most pressing global development needs of the century, especially for small cities with rapid population growth. Currently, 53% of the world’s population either lacks access to improved sanitation or discharges fecal waste to the environment without treatment. Furthermore, 80% of food consumed in developing regions is produced by 500 million small farms, and while many of them are still rain-fed, irrigated agriculture is increasing. The post-2015 Sustainable Development Goals, recently adopted by the United Nations, include targets to address the water-energy-food nexus. Wastewater reuse in agriculture can be an important solution for these goals, if it is done safely. Globally, 18 – 20 million hectares of agricultural land are irrigated with wastewater, but much is untreated, unregulated, or unsanctioned, causing concerns and uncertainty about health risks. There is a need to better understand pathogen removal in natural and non-mechanized wastewater treatment systems, such as waste stabilization ponds (WSPs) and upflow anaerobic sludge blanket (UASB) reactors, which are commonly used in small cities and towns. Riverbank filtration (RBF) is also a natural technique used by farmers in developing countries to treat surface water polluted with untreated sewage, but pathogen removal in these systems has seldom been assessed in developing countries. The focus of this dissertation is on pathogen removal in natural and non-mechanized wastewater treatment and reuse systems, to evaluate the health implications of water reuse for irrigation, with the following three objectives: 1) assess the current understanding of virus removal in WSP systems through a systematic review of the literature; 2) measure the removal of viruses and their association with particles in systems with WSPs, UASB reactors, or both; and 3) assess the fate and transport of pathogens and fecal indicators in wastewater treatment systems with direct and indirect reuse for irrigation to estimate microbial risks. To advance the understanding of virus removal in WSP systems, a comprehensive analysis of virus removal reported in the literature from 71 different WSP systems revealed only a weak to moderate correlation of virus removal with theoretical hydraulic retention time (HRT). For each log10 reduction of viruses a geometric mean of 14.5 days of retention was required, but the 95th percentile of the data analyzed was 54 days. Also, whereas virus-particle association and subsequent sedimentation has been assumed to be an important removal mechanism for viruses in WSPs, the literature review revealed a lack of evidence to confirm the validity of this assumption. The association of human adenovirus (AdV) with wastewater particles was assessed in five full-scale wastewater treatment systems in Bolivia, Brazil, and the United States (two with only WSPs, two with a UASB reactor and WSPs, and one with only UASB reactors). A mesocosm study was also conducted with WSP water from one of the full-scale systems, and some samples were also analyzed for pepper mild mottle virus (PMMoV), F+ coliphage, culturable enterovirus (EV), norovirus (NoV), and rotavirus (RV). Results indicate that WSPs and UASB reactors affect virus-particle associations in different ways, which may differ for different viruses. In UASB reactor effluent, PMMoV was more associated with particles <180 >µm, showed no indication of settling in subsequent ponds, and appeared to degrade in pond sediments after 5 days. In contrast, AdV in UASB reactor effluent was associated with small and large particles, and in subsequent ponds, particle-associated AdV showed evidence of possible settling or more rapid decay at the water surface. AdV and culturable EV were also more volumetrically-concentrated in UASB reactor sludge than they were in untreated sewage, WSP water, UASB effluent, and WSP sediments, indicating that the reactors may cause these viruses to become entrapped and concentrated in granular sludge. Some viruses may be removed in the sludge, but others exit the reactors in solution and attached to particles. The resuspension of pellets from centrifuged UASB reactor sludge samples in an eluant buffer indicated reversible AdV association with granular sludge, but some associations with particles in solution may not be reversible. The fate and transport of pathogens and fecal indicators was assessed in Bolivia for two WSP systems with direct reuse for irrigation, and one on-farm RBF system used to treat surface water polluted by untreated sewage. In the WSP systems, despite HRTs of 10 days, pathogen and fecal indicator removal was generally ≤1-log10, possibly due to overloading and short-circuiting from sludge accumulation. The RBF system provided removals on the order of 2-log10 for protozoan parasites, 3-log10 or more for viruses, and 4-log10 or more for bacteria. The use of RBF also reduced cumulative estimated health burdens associated with irrigated lettuce. Irrigation of lettuce with untreated river water caused an estimated disease burden that represents 37% of the existing burden from acute diarrhea in Bolivia; when RBF was used, this decreased to only 1.1%, which is not epidemiologically-significant, and complies with the World Health Organization guidelines. Ratios of concentrations of microorganisms in irrigation water to their respective concentrations in soil or crops were calculated, to assess transfer from irrigation water to soil or crops. These ratios (with units mL g-1) were generally < 0.1 mL g-1 for coliphage, between 1 and 100 mL g-1 for Giardia and Cryptosporidium, and generally between 100 and 1,000 mL g-1 for helminth eggs. Higher ratios could indicate more efficient transfer from water to soil or crops, longer persistence in soil or on crops, or slower leaching away from soil or crops. The results from this research demonstrate that pathogen removal in full-scale natural wastewater treatment systems happens via complex mechanisms that vary with respect to pathogen type, treatment systems configuration, and other environmental and operational parameters. Future research and innovation efforts should focus on the use of a combination of natural and non-mechanized technologies, surface-flow systems (e.g., WSPs) and subsurface systems (e.g., RBF), applied at both semi-centralized (e.g., wastewater treatment plant) and decentralized levels (e.g., on farms), to evaluate how this affects the efficiency and resiliency of pathogen removal. Also, future research is needed to further elucidate reasons for the observed differences in virus-particle associations in natural wastewater treatment systems.

Animal and human wastes introduce pathogens into rivers and streams, creating human health and economic burdens. While direct monitoring for pathogens is possible, it is impractical due to the sporadic distribution of pathogens, cost to identify, and health risks to laboratory workers. To overcome these issues, fecal indicator organisms are used to estimate the presence of pathogens. Although fecal indicators generally protect public health, they fall short in their utility because of difficulties in public health risk characterization, inconsistent correlations with pathogens, weak source identification, and their potential to persist in environments with no point sources of fecal pollution. This research focuses on characterizing the ecology of fecal indicators using both modeling and metabolic indicators to better understand the processes that drive fecal pollution. Fecal indicator impairment was modeled in Sinking Creek, a 303 (d) listed stream in Northeast Tennessee, using the ecological niche model, Maxent, for two different fecal indicators. While the use of Maxent has been well demonstrated at the macroscale, this study introduces its application to ecological niches at the microscale. Stream impairment seasonality was exhibited in two different indicators over multiple years and different resolutions (quarterly versus monthly sampling programs). This stresses the need for multiple year and month sampling to capture heterogeneity in fecal indicator concentrations. Although discharge is strongly associated with dissolved solutes, fecal indicator impairment was governed by other ecological factors such as populations of heterotrophic bacteria, enzyme activity, nutrient conditions, and other metabolic indicators. This research also incorporated metabolic indicators to characterize spatiotemporal variability in microbial community function, making connections to fecal and other pollution gradients. Communities differed in their ability to use a wide variety of substrates, and metabolic inhibition in sediments captured most of the interaction of aquatic and benthic communities. Sediment substrate activity was also indicative of degrees of pollution, suggesting that sediment is a potential reservoir for Escherichia coli in this stream, and there is possibility for resuspension, extended residence times, and increased duration for exposure. This research highlights the benefit of using models and other microbial indicators to better understand how environment shapes the niche of fecal indicators.

Packed-bed digesters are an alternative to covered lagoon digesters for methane production and anaerobic treatment of dilute wastewaters such as dairy barn flush water. The physical media of packed-beds retain biofilms, often allowing increased treatment rates. Previous studies have evaluated several types of media for digestion of dilute wastewaters, but cost and media fouling have setback commercial development. A major operational cost has been effluent recirculation pumping. In the present effort, a novel approach to anaerobic digestion of flush dairy water was developed at pilot-scale: broken walnut shells were used as a low-cost packed-bed medium and effluent recirculation was replaced by reciprocation mixing to decrease pumping costs and the risk of media clogging. Three packed-bed digesters containing walnut shells as media were constructed at the on-campus dairy and studied for about six months. Over that time, several organic loading rates (OLRs), measured as both chemical oxygen demand (COD) and volatile solids (VS) were applied to the new packed-bed digesters to allow modeling of methane production. The influence of temperature on methane production was also investigated. Additionally, the study measured solids accumulation in the walnut shell packed-bed as well as the effectiveness and durability of walnut shells as packing media. Finally, a simple economic analysis was developed from the methane model to predict the financial feasibility of packed-bed digesters at flush water dairies under similar OLR conditions. Three methane production models were developed from organic loading: saturation-type (following the form of the Monod equation), power and linear. The models were evaluated in terms of regression analysis and the linearity of experimental to predicted methane production. The best model was then chosen to develop the economic predictions. Economic predictions for packed-bed digesters were calculated as internal rate of return (IRR) using the methane models along with additional input variables. Comparisons of IRRs were made using electric retail rates of $0.10 to $0.20 per kilowatt-hour and capital cost subsidies from zero to 50%. Sludge accumulation in the packed-bed was measured via change in porosity, and walnut shell durability was measured as the change in mass of representative walnut shells over the course of the study. The linear-type model of methane production from volatile solids OLR best represented this data set. Digester temperature was not found to influence methane production in this study, likely due to the small daily average ambient temperature range experienced (14°C to 24°C) and the greater influence of organic loading. Porosity of the walnut shell packed-bed decreased from 0.70 at startup to 0.34±0.06 at the end of the six-month study, indicating considerable media fouling. Sludge accumulated in each digester from zero at startup to 281±46 liters at termination. Walnut shells in the packed-bed lost on average 31.4±6.3% mass during the study period which may be attributed to degradation of more readily bio-degradable cellulose and hemi-cellulose within the walnut shells. Given the predicted methane production and media life, at present, the economic outlook for packed-bed digesters at commercial dairies is quite dependent on utility electrical rates, available subsidies and future improvements to packed-bed digester technology. The predicted IRRs ranged from below 0% (at 0% capital subsidy and $0.10/kWh) up to 25% (at 50% capital subsidy and $0.20/kWh) at large dairies (3000 milking cows). Increases in organic loading were not shown to necessarily increase IRR, particularly at OLRs above 10 g/Lliquid-d (as COD or VS). Ultimately, to better assess the value of packed-bed digesters for flush dairies, additional study is needed on topics such as sludge accumulation prevention, long-term walnut shell degradation, dairy barn flush water mixing, and more detailed economic analysis.

The Department of Water Affairs and Forestry (DW AF) is the custodian of South Africa's water resources and its primary role is to maintain the fitness for use of water on a sustained basis. DW AF recognized that management and assessment of fitness for use can only be based on reliable monitoring data. For this purpose DWAF has already for a number of years operated a national programme which collects data on the chemical and physical quality of South Africa's water resources. The microbial quality of surface water is of growing concern in a number of areas in South Africa. Water of poor microbial quality has serious implications for domestic, recreational and agricultural use due to the risk of water-borne diseases. DW AF acknowledged the need for information on the microbial quality of South Africa's water resources to assess and manage the potential health risk to water users. As an initial step the development of a national microbial monitoring programme to assess the faecal pollution of surface waters was initiated. This study describes the development of the conceptual design of such a programme and demonstrates how a novel area prioritization procedure enhanced the design. The focus of the programme was to be areas where human health might be severely impacted by the microbial quality of surface water. To identify such areas, a procedure for the identification and prioritization of specific areas of concern was developed and used as part of the design approach. Two factors were identified for the quantification of the potential health risk. They were the threat of microbial pollution of water (the result ofland use) and the exposure of consumers to the water (sensitivity of water uses). A number of land and water uses information sources therefore served as the basis for determining priority among the different areas. The described approach to identify and prioritize specific areas of concern has a number of benefits. Primarily, the approach assists in focusing the monitoring efforts on problem areas without a need for extensive historical microbial water quality data. The approach could be used to optimize the spatial distribution of sampling stations and assist in determining their national distribution. The approach also allows for phased implementation of the programme which facilitates the development of skills and capacity, as well as required infrastructure needed for the large scale operation of the programme. The approach to focus on impacted areas is generic enough not to be restricted to the design of microbial water quality monitoring systems. Other monitoring objectives could also be dealt with in the same manner. During evaluation of the design on a pilot scale the conceptual design was found to meet the set information objectives. The conceptual design for the programme also deals effectively with constraints and changes in the external environment in which it has to operate. Implementation of the national programme has started and plans to expand the programme are progressing well. The concept of high risk areas and the procedure to identify and prioritize such areas as developed during this study is a critical component of the overall design. The programme appears to address a significant information need on an important aspect of water resources management and to do so in an efficient and effective manner.
Thesis (PhD (Water Resource Management))--University of Pretoria, 2005.
Microbiology and Plant Pathology
unrestricted

abstract: This study was designed to provide insight into microbial transport kinetics which might be applied to bioremediation technology development and prevention of groundwater susceptibility to pathogen contamination. Several pilot-scale experiments were conducted in a saturated, 2 dimensional, packed porous media tank to investigate the transport of Escherichia coli bacteria, P22 bacteriophage, and a visual tracer and draw comparisons and/or conclusions. A constructed tank was packed with an approximate 3,700 cubic inches (in3) of a fine grained, homogeneous, chemically inert sand which allowed for a controlled system. Sampling ports were located at 5, 15, 25, and 25 vertical inches from the base of the 39 inch saturated zone and were used to assess the transport of the selected microorganisms. Approximately 105 cells of E. coli or P22 were injected into the tank and allowed to move through the media at approximately 10.02 inches per day. Samples were collected intermittently after injection based off of an estimated sampling schedule established from the visual tracer. The results suggest that bacteriophages pass through soil faster and with greater recovery than bacteria. P22 in the tank reservoir experienced approximately 1 log reduction after 36 hours. After 85 hours, P22 was still detected in the reservoir after experiencing a 2 log reduction from the start of the experiment. E. coli either did not reach the outlet or died before sampling, while P22 was able to be recovered. Bacterial breakthrough curves were produced for the microbial indicators and illustrate the peak concentrations found for each sampling port. For E. coli, concentrations at the 5 inch port peaked at a maximum of 5170 CFU/mL, and eventually at the 25 inch port at a maximum of 90 CFU/mL. It is presumed that E. coli might have experienced significant filtration, straining and attachment, while P22 might have experienced little adsorption and instead was transported rapidly in long distances and was able to survive for the duration of the experiment.
Dissertation/Thesis
Masters Thesis Civil, Environmental and Sustainable Engineering 2017

abstract: Understanding how microorganisms adapt and respond to the microgravity environment of spaceflight is important for the function and integrity of onboard life support systems, astronaut health and mission success. Microbial contamination of spacecraft Environmental Life Support Systems (ECLSS), including the potable water system, are well documented and have caused major disruption to spaceflight missions. The potable water system on the International Space Station (ISS) uses recycled wastewater purified by multiple processes so it is safe for astronaut consumption and personal hygiene. However, despite stringent antimicrobial treatments, multiple bacterial species and biofilms have been recovered from this potable water system. This finding raises concern for crew health risks, vehicle operations and ECLSS system integrity during exploration missions. These concerns are further heightened given that 1) potential pathogens have been isolated from the ISS potable water system, 2) the immune response of astronauts is blunted during spaceflight, 3) spaceflight induces unexpected alterations in microbial responses, including growth and biofilm formation, antimicrobial resistance, stress responses, and virulence, and 4) different microbial phenotypes are often observed between reductionistic pure cultures as compared to more complex multispecies co-cultures, the latter of which are more representative of natural environmental conditions. To advance the understanding of the impact of microgravity on microbial responses that could negatively impact spacecraft ECLSS systems and crew health, this study characterized a range of phenotypic profiles in both pure and co-cultures of bacterial isolates collected from the ISS potable water system between 2009 and 2014. Microbial responses profiled included population dynamics, resistance to silver, biofilm formation, and in vitro colonization of intestinal epithelial cells. Growth characteristics and antibiotic sensitivities for bacterial strains were evaluated to develop selective and/or differential media that allow for isolation of a pure culture from co-cultures, which was critical for the success of this study. Bacterial co-culture experiments were performed using dynamic Rotating Wall Vessel (RWV) bioreactors under spaceflight analogue (Low Shear Modeled Microgravity/LSMMG) and control conditions. These experiments indicated changes in fluid shear have minimal impact on strain recovery. The antimicrobial efficacy of silver on both sessile co-cultures, grown on 316L stainless steel coupons, and planktonic co-cultures showed that silver did not uniformly reduce the recovery of all strains; however, it had a stronger antimicrobial effect on biofilm cultures than planktonic cultures. The impact of silver on the ability of RWV cultured planktonic and biofilm bacterial co-cultures to colonize human intestinal epithelial cells showed that, those strains which were impacted by silver treatment, often increased adherence to the monolayer. Results from these studies provide insight into the dynamics of polymicrobial community interactions, biofilm formation and survival mechanisms of ISS potable water isolates, with potential application for future design of ECLSS systems for sustainable human space exploration.
Dissertation/Thesis
Masters Thesis Molecular and Cellular Biology 2019

abstract: Biological soil crusts (biocrust) are photosynthetic communities of organisms forming in the top millimeters of unvegetated soil. Because soil crusts contribute several ecosystem services to the areas they inhabit, their loss under anthropogenic pressure has negative ecological consequences. There is a considerable interest in developing technologies for biocrust restoration such as biocrust nurseries to grow viable inoculum and the optimization of techniques for field deployment of this inoculum. For the latter, knowledge of the natural rates of biocrust dispersal is needed. Lateral dispersal can be based on self-propelled motility by component microbes, or on passive transport through propagule entrainment in runoff water or wind currents, all of which remain to be assessed. I focused my research on determining the capacity of biocrust for lateral self-propelled dispersal. Over the course of one year, I set up two greenhouse experiments where sterile soil substrates were inoculated with biocrusts and where the lateral advancement of biocrust and their cyanobacteria was monitored using time-course photography, discrete determination of soil chlorophyll a concentration, and microscopic observations. Appropriate uninoculated controls were also set up and monitored. These experiments confirm that cyanobacterial biological soil crusts are capable of laterally expanding when provided with presumably optimal watering regime similar to field conditions and moderate temperatures. The maximum temperatures of Sonoran Desert summer (up to 42 °C), exacerbated in the greenhouse setting (48 °C), caused a loss of biomass and the cessation of lateral dispersal, which resumed as temperature decreased. In 8 independent experiments, biocrust communities advanced laterally at an average rate of 2 cm per month, which is half the maximal rate possible based on the instantaneous speed of gliding motility of the cyanobacterium Microcoleus vaginatus. In a span of three months, populations of M. vaginatus, M. steenstrupii, and Scytonema spp. advanced 1 cm/month on average. The advancing crust front was found to be preferentially composed of hormogonia (differentiated, fast-gliding propagules of cyanobacteria). Having established the potential for laterally self-propelled community dispersal (without wind or runoff contributions) will help inform restoration efforts by proposing minimal inoculum size and optimal distance between inoculum patches.
Dissertation/Thesis
Masters Thesis Biology 2017