Thesis (M.A.)--Humboldt State University, 2007. / Includes bibliographical references (leaves 84-101). Also available via Humboldt Digital Scholar.

Thesis (Ph. D.)--Michigan State University. Dept. of Entomology and Program in Ecology, Evolutionary Biology and Behavior, 2008. / Title from PDF t.p. (viewed on July 7, 2009) Includes bibliographical references (p. 185-195). Also issued in print.

The Deilmann Tailings Management Facility (DTMF) at Key Lake in northern Saskatchewan, Canada, is an active deposition site for uranium tailings and it has been in operation since 1996. In terms of geochemical stability of the tailings, a ferrihydrite secondary phase is utilized for the sequestration of contaminants, such as As, Ni, Mo, and Se, under alkaline and highly oxic conditions. Arsenic is highly abundant in the DTMF tailings and the principal environmental concern is the possibility for leaching of ferrihydrite-attached As into the surrounding environment. Microorganisms can proliferate in a broad range of habitats and their activities are key factors in determining fate and transport of contaminants in various environments. This thesis attempts to obtain insights into the biogeochemical processes that may occur during the early phase of the DTMF’s history that could potentially become significant over extended periods of time that run from 100’s to 1000’s of years. Hence, a primary focus was to characterize microbial diversity and extrapolate their potential functional roles as well as their potential to chemically alter the Eh and ferrihydrite, which are the primary controlling conditions within the DTMF tailings and in the mineral secondary phase, respectively. To achieve these goals, two molecular techniques (clone library construction and Ion Torrent sequencing), a range of conventional culture-based techniques, metabolic assays addressing metabolic transformation and resistance to metals/metalloids, microscopic technique (Confocal Laser Scanning Microscope), spectroscopic analyses (Scanning Transmission X-ray Microscope) and bench-scale microcosm assays were carried out. Culture-dependent and -independent methods revealed that the most prevalent microbial groups in the water column, tailings mass and at the tailings-water interface affiliated into phyla (e.g., Proteobacteria, Actinobacteria, Firmicutes and Bacteriodetes) that have previously been detected at uranium-, heavy metal- and complex hydrocarbon-contaminated sites. Phylotypes closely related to well-characterized sulfate-, thiosulfate- and iron-reducing bacteria (e.g., Desulfosporosinus, Dethiobacter, Geoalkalibacter, Ralstonia, Georgfuchsia) were also detected at low frequency, with the exception of the tailings-water interface where sequences closely related to Desulfosporosinus were abundant. The readily culturable heterotrophs (e.g., Pseudomonas, Arthrobacter, Massilia, Hydrogenophaga, Polaromonas, Bacillus) retrieved from the tailings exhibited reducing/oxidizing capabilities as well as high tolerance to metal/metalloids. Bench scale microcosm assays showed that heterotrophs native to the DTMF site could not only reduce ferrihydrite but could also create highly reducing (< -300 mV) conditions within the tailings amenable to strict anaerobic bacteria such as Desulfosporosinus. STXM image analyses confirmed the presence of reduced iron in close proximity to bacterial cells in biofilm grown in situ and in microcosm tailings, strongly suggesting that ferrihydrite served as electron acceptor during microbial processes. Reduced iron detected in situ also indicated that microscale iron reduction could occur even though macroscale DTMF chemistry remained oxidizing. Overall, the nature of microbial community present in the DMTF system strongly indicated that complex hydrocarbons (e.g., kerosene) discharged into the tailings during processing could potentially support microbial processes that involve Fe and S cycling and that this process could become significant over extended period of times, contributing to arsenic escape into the environment.

Research on natural environments, over the last decade, is replete with microbial diversity studies that used culture-independent approaches. The cloning and sequencing of the 16S rRNA genes has been the driving force in the expansion of awareness about the great diversity of previously undiscovered microorganisms. Well-known uncultured groups of microorganisms are numerous, and half of the known phylogenetic divisions of bacteria are not represented in any culture collection. It is no longer assumed that cultures acquired from an environment represent the dominant or physiologically important organisms from that environment. A high throughput culturing (HTC) technique was developed in an attempt to bring into culture some of these widespread and uncultured microorganisms. Over the course of 3 years, 2,484 culturing attempts were screened for microbial growth from sample collections off the coast of Oregon and 576 attempts from groundwater at McClellan Air Force Base (MAFB). However, using the HTC approach up to 14% of the microorganisms counted by direct microscopy were cultured. In contrast, less than 1% of the microorganisms from natural environments that are observed under a microscope can be grown using standard agar plating techniques. This newly developed technique was successful at bringing into culture 11 previously uncultured or undescribed Proteobacteria. Four were isolated from the marine environment including, members of the SAR11 clade (alpha subclass), OM43 (beta-subclass), SAR92 (gamma subclass), and OM60/OM241 (gamma subclass). SAR11 was transiently cultured in this study but was later successfully brought into culture using these HTC techniques by Mike Rapp��. Eight were isolated from a trichioroethene (TCE) and cis-dichloroethene (cis-DCE) contaminated aquifer, including members of the MHP14 clade (alpha subclass), 4-Org1-14 dade (alpha subclass), Herbaspirillum/Oxalobacter clade (beta subclass), HTCC333 (beta subclass), HTCC410 (beta subclass), PM1 clade (beta subclass), Boom-7m-04 clade (beta subclass) and OM43 clade (beta subclass). Culturing microorganisms is an important step towards understanding their physiology and ecology, and in most cases is necessary for the formal systematic description of a new species. For microorganisms of global significance, such as the major uncultured bacterioplankton and soil microbiota, obtaining cultures is a prerequisite for obtaining complete genome sequences and understanding the relevance of these microorganisms to biogeochemical cycles. / Graduation date: 2003

It is recognized that microbial transformations are the primary mechanism of organic contaminant removal in natural and constructed wetland systems. However, not much is known about urban wetland microbial communities or their functional capacity to process contaminants. The objective of this research was to first characterize the physiological and phylogenetic diversity of microbial communities of different urban wetland types using the BIOLOG™ method and through DGGE of 16S rRNA sequences. The capacity of urban wetlands to attenuate model chlorinated aromatic compounds (2,4-D and 3-CBA) was assessed by UPLC biodegradation and 14C mineralization experiments. Toxicity tests were conducted to assess microbial tolerance to pollutant addition. In general, results indicate that urbanization has a homogenizing effect on microbial community structure and distribution within urban wetland systems, regardless of type. Urban wetlands also appear to have a limited capacity to remove chlorinated organic pollutants. Microbial community tolerance to chlorinated organic pollutants is relatively high, whereas heavy metal tolerance was found to coincide with history of contaminant exposure. / UOIT

The microbial diversity of branches of Kaoping River at Lao-Nong Basin with altitudes from 100-2204m and Nantzuhien Basin with altitudes from 100-654m were studied. We used combining methods of bacterial, chemical and biotechnological to explore microbial diversity at the two basins to provide a reference for the environmental ecology of Kapoing River. It is the foundation for practicable environmental care. The results show the following: (1) The temperature in Tianchin at Lao-Nong Basin was lower(16.3¢J) than the rest of sampling sites(22-28¢J). (2) The temperature in Minsheng at Nantzuhsien Basin is lower(21.5¢J)than the of sampling sites(24-33¢J); other factors such as pH value, total organic carbon, and total organic nitrogen, varied according to different locations. In the microbacterial composition quantitatively, among all microorganisms, the bacteria content was the highest. Each gram of soil contain about 106~109 CFU at Lao-Nong Basin and 105~109 CFU at Nantzuhsien Basin, where as others such as the contents of actinomycetes and fungi were lower. Using 16s rDNA DGGE (denaturing gradient gel electrophoresis)analysis, the soil bacterial community composition at the two basin districts had discoverd that the microbial diversites at Nantzuhsine Basin were more abundant than those at Lao-Nong Basin. In regard to seasonal changes, our data agreed with traditional data. In the warm season, the microbial content is higher, and the community composition is also more abundant. The weather change in a short time period, such as a rainstorm and a quick shift of temperature seriously affected the number of bacteria. When the rain pours steadily down, it caused some degree of drop in bacteria number. However, when the rain stop, the microbacterial content and community composition gradually retured to original forms.

Thesis (Ph.D.)--Montana State University--Bozeman, 2007. / Typescript. Chairperson, Graduate Committee: Gill G. Geesey. Includes bibliographical references.

Submitted in complete fulfilment for the Degree of Master of Technology: Biotechnology, Durban University of Technology, 2013. / The continual growth of the global biodiesel industry has resulted in a proportional increase in crude glycerol production. The by-production of glycerol waste during the manufacture of biodiesel has, with recent research, proven to hold use as a feedstock for the production of several commodity chemicals. The conversion of glycerol may be carried out by both chemical as well as biological means. The biological conversion of glycerol surpasses chemical conversion with respect to higher yield and selectivity, normal reaction conditions and the use of cheaper biological catalysts. Many microorganisms are known to convert glycerol to different value added products. This study involved the isolation of bacteria from soil and crude glycerol from a local biodiesel plant. Isolates were then used to convert crude glycerol supplemented with salts and a nitrogen source into commercially viable products. Isolates which successfully degraded glycerol were then identified via 16S PCR. A strain of Klebsiella pneumoniae, which is a known producer of 1,3-propanediol (1,3-PDO), was isolated from soil and two strains of Sphingomonas sp., which is a known gellan producer, was isolated from biodiesel waste. Gellan is an exopolysaccharide used in the food, cosmetic and pharmaceutical industries sold commercially as a product known as Gelrite or Gelzan while 1,3-PDO is an important component of fuels and polyesters (used widely in the petroleum industry) and is currently chemically produced. Using crude glycerol for producing 1,3-PDO is a good solution from an economic as well as ecological point of view. K. pneumoniae, Sphingomonas psueudosanguinis and Sphingomonas yabuuchiae were subjected to a series of shake flask fermentations in order to determine optimal growth conditions. This microoganism was able to successfully produce significant amounts of 1,3-PDO and lactic acid using crude glycerol (80 g/l), without pre-treatment (37 and 6.8 g/l respectively). S psueudosanguinis and S. yabuuchiae were both able to produce two of the highest amounts of gellan gum than that reported by other studies using crude glycerol (80 g/l) as a sole carbon source in a minimal medium (50.9 and 52.6 g/l respectively). / National Research Foundation

Leafy greens like lettuce and spinach are a common vehicle for foodborne illness in United States. It is unknown if native plant epiphytic bacteria may play a role in the establishment of enteric pathogens on leaf surfaces. The objective of this study was to characterize the bacterial communities of fresh and packaged spinach leaves and to explore interactions with E. coli O157:H7. We assessed the bacterial diversity present on the spinach leaf surfaces and how parameters such as spinach cultivar, field conditions, post-harvest operations and the presence of E. coli O157:H7 affected its diversity. Differences in bacterial population size and species richness were associated with differences in plant topography; flat leaves had smaller bacterial populations than savoy leaves, which correlated with larger number of stomata and trichomes in savoy leaves. During spinach growing season shifts in environmental conditions affected richness and population size of the spinach bacterial community. Decreases in the overall soil and ambient temperature and increased rainfall decreased richness and bacterial population size. Fresh spinach richness and composition assessed by parallel pyrosequencing of 16S rRNA elucidated 600 operational taxonomic units, with 11 different bacterial phyla. During postharvest operations diversity indexes and evenness tended to decrease, likely attributed to storage at low temperature and time of storage (4°C and 10°C), that promoted the dominance of g-Proteobacteria. Bacteria isolated from fresh spinach elicited growth inhibition of E. coli O157:H7 in vitro, which was associated with nutrient competition. In contrast growth enhancement produced by epiphytes was associated to low correlations in carbon source utilization and the ability of E. coli O157:H7 to rapidly utilize carbon resources. In packaged spinach, E. coli O157:H7 altered the composition of the bacterial community and its growth was promoted on packaged spinach when a disinfection and temperature abuse occurred, removal of the epiphytic bacteria resulted in significant increases in numbers of E. coli O157:H7 at 10°C and was associated with increased expression of E. coli O157:H7 virulence and stress response genes. The large diversity present on the surface of spinach leaves significantly impacted the ecology of enteric pathogens like E. coli O157:H7 on the phyllosphere. / Ph. D.

Leafy greens such as spinach have been the object of several recent food-borne pathogen outbreaks. The purpose of this study was to isolate bacteria spinach epiphytic bacteria that inhibit growth of E. coli O157:H7, which we describe as antagonism. The mechanism of antagonism was investigated and we attempted to improve the antagonistic potential in vitro and on spinach leaves when cellobiose, a carbon source utilized by the antagonists but not E. coli O157:H7, was added. There were larger culturable populations of bacteria on the leaves of savoyed cultivars compared to flat. From the isolated colonies, 47 displayed antagonism towards E.coli O157:H7, and were identified as members of 11 different genera and sixteen species. A representative isolate from each species was evaluated for three possible mechanisms of antagonism: acid production, secretion of an inhibitory compounds or secreted protease. The majority (14/16) produced at least a moderate level of acid. Two of these strains, Paenibacillus polymyxa and Pseudomonas espejiana, were found to secrete a non- protease antagonistic compound. These antagonists varied in their reduction of E.coli O157:H7 numbers in vitro, but all significantly reduced numbers in 48 hours of co-culturing in nutrient rich media. Five antagonists resulted in a significant reduction in E.coli O157:H7 populations when co-cultured on spinach leaves. Application of cellobiose did not improve the amount of antagonism in vitro or on the leaf surface after 24 hours. / Master of Science