Aigarchaeota, a deeply branching archaeal lineage with no cultivated representatives, is found in geothermal and hydrothermal systems worldwide and consists of at least 9 genus-level groups, each predicted to have diverse metabolic capabilities. This candidate archaeal phylum is part of the TACK superphylum, members of which possess eukaryotic-signature proteins, thus suggesting that they may represent evolutionary steps along the way to the genesis of the first eukaryotic cells. Cultivating members of Aigarchaeota would elucidate how eukaryotes arose in evolutionary history and provide biotechnological applications. Aigarchaeota Group 4 (AigG4), one genus in Aigarchaeota, was previously found to be abundant in corn stover in situ enrichments in Great Boiling Spring (GBS). AigG4 has been maintained in mixed laboratory cultures, where it composes ~ 0.5-1% of the community. However, these cultures could only be maintained when GBS water, which contains ~300 nM tungsten, was included in the medium. In addition, AigG4 metagenome bins from the in situ enrichments and laboratory cultures contained multiple genes encoding putative tungsten-containing aldehyde:ferredoxin oxidoreductases (TAORs). These observations led to the hypothesis that tungsten was the key component in GBS water that allowed for growth of AigG4. The requirement of tungsten for AigG4 long-term maintenance in mixed culture was tested using three different approaches: (1) Assessing the phylogeny of tungsten transporters and TAORs across the Aigarchaeota lineage, followed by confirmation of transcription of hypothesized AigG4 tungsten-associated genes in lab cultures, (2) Measuring tungsten levels in Great Boiling Spring (GBS) using ICP-OES (Inductively coupled plasma-optical emission spectroscopy), and (3) determining minimum amount of tungsten for long-term AigG4 maintenance in corn stover and in a defined mix of sugars (1% glucose, 1% xylose, 1% D-arabinose, 1% L-arabinose, 1% mannose). In addition, FISH (Fluorescent in situ hybridization) probes were designed to target the AigG4 lineage in the hopes that in conjunction with nanometer-scale secondary ion mass spectroscopy (Nano-SIMS), this would test whether tungsten has indirect or direct effects on AigG4 to track carbon substrate intake. Most Aigarchaeota lineages possess a tungsten transporter complex (TTC). AigG4 TTC groups most closely with the Tup family while other Aigarchaeota group with the Wtp family. Group 4 and Group 5 Aigarchaeota contained TAORs that grouped to other hypothesized TAORs but not to characterized counterparts suggesting diverse functional capabilities. Group 4 and Group 5 TAORs clustered together suggesting that these are conserved within these lineages. Gene expression of predicted AigG4 tungsten-associated enzymes was detected in culture. Tungsten was detected in GBS water as previously observed in 2005. In both corn stover and sugar mix, 1 nM tungsten was sufficient for long term AigG4 maintenance. In corn stover, AigG4 decreased to levels below detection after three 3-week transfer periods in 0 nM and 20 nM tungsten. In sugar mix, AigG4 abundance levels varied wildly in 0 nM tungsten after the fifth transfer period suggesting tungsten contamination. Two newly designed FISH probes exhibited lower fluorescence signal intensity than the previously designed FISH probe suggesting issues with either target site accessibility or conjugation of the fluorescent moiety to oligonucleotide probes.

The specific identification of a megalurous cercaria found in the snail Oxytrema plicifera was accomplished by completing the life cycle in the laboratory. This species is compared with the eastern Philophthalmus megalurus and P. gralli with which it was once confused. The eggs, miracidia, and rediae of the Oregon species were found to be similar to those of the above species. The body and organ sizes and sucker ratios for the cercariae and adult stages obviously indicate that the Oregon species is Philophthalmus megalurus.

The field of viral ecology is still relatively new and many processes by which viruses influence hosts are still widely unknown. One problem is that there are few standardized techniques in virus ecology, making comparisons of data very difficult. To better understand the methodology, we first set out to make a video showing the process for the viral production assay using the “dilution and reoccurrence” method, which has become the standard to analyze production rates in aquatic ecosystems. Using this method, we also determined the production rates of viruses during a seasonal pelagic phytoplankton bloom during a cruise off the coast of the north island of New Zealand in the subtropical Pacific Ocean. Other biotic and abiotic parameters were also compared throughout the bloom. Production rates were within normal ranges, but showed that viruses were very important for the remobilization of nutrients in the nitrogen-limited system. It is well known that the cyanobacterial genera Synechococcus and Prochlorococcus thrive in the world’s oceans with Synechococcus and other cyanobacterial species also succeeding in freshwater ecosystems. Cyanophages are viruses which infect cyanobacteria and many studies have investigated their diversity using the portal vertex g20 gene in the Cyanomyoviridae family. Although we know that there is significant genetic richness in these phage in marine and freshwater environments, information on their numerical distributions is rare. Using quantitative PCR with the g20 gene, we determined that cyanomyoviruses are ubiquitous and abundant in the Atlantic and Pacific Oceans as well as within Lake Erie. Using statistical analyses we were able to find correlations between cyanomyoviruses and other biotic and abiotic parameters: in the Sargasso Sea, cyanomyovirus abundance correlated well to biology, but in the other systems there was no significant correlation to biological abundances. This suggests that the constraints of this group of viruses may be different in different aquatic realms.

Bioremediation of chlorinated contaminants in groundwater can be achieved by reductive dechlorination by anaerobic dechlorinating cultures. However, at sites impacted with multiple different chlorinated contaminants, reductive dechlorination is often inhibited by co-contaminants. The inhibitory effects of trichloroethene, cis-dichloroethene and vinyl chloride on chloroform dechlorination were studied using a Dehalobacter-containing chloroform dechlorinating mixed culture (ACT-3). The inhibitory effect of chloroform on chlorinated ethene dechlorination was studied in a Dehalococcoides-containing trichloroethene dechlorinating mixed culture (KB-1). Vinyl chloride was found to be the strongest inhibitor of chloroform in ACT-3. Chloroform exerted a complex and strong inhibitory effect on chlorinated ethene dechlorination in KB-1. The potential for microbial reductive defluorination was also examined in the enrichment cultures. Quantifying the substrates and substrate interactions in dechlorinating cultures is necessary to most efficiently use these cultures to remediate contaminated sites.

Bioremediation of chlorinated contaminants in groundwater can be achieved by reductive dechlorination by anaerobic dechlorinating cultures. However, at sites impacted with multiple different chlorinated contaminants, reductive dechlorination is often inhibited by co-contaminants. The inhibitory effects of trichloroethene, cis-dichloroethene and vinyl chloride on chloroform dechlorination were studied using a Dehalobacter-containing chloroform dechlorinating mixed culture (ACT-3). The inhibitory effect of chloroform on chlorinated ethene dechlorination was studied in a Dehalococcoides-containing trichloroethene dechlorinating mixed culture (KB-1). Vinyl chloride was found to be the strongest inhibitor of chloroform in ACT-3. Chloroform exerted a complex and strong inhibitory effect on chlorinated ethene dechlorination in KB-1. The potential for microbial reductive defluorination was also examined in the enrichment cultures. Quantifying the substrates and substrate interactions in dechlorinating cultures is necessary to most efficiently use these cultures to remediate contaminated sites.

Enhanced biological phosphorus removal (EBPR) is a biological wastewater treatment process facilitated by polyphosphate-accumulating organisms (PAO). The absence of isolates that have the PAO phenotype has limited the scope of studies into the physiology of these industrially significant and metabolically unique organisms. This thesis outlines findings into the physiology and ecology of EBPR in mixed microbial cultures, which contribute to the fundamental understanding of the process. The first experimental approach used in these studies was to investigate the microbial abundance of identified PAOs and GAOs in full-scale and lab-scale EBPR processes, and correlate these data with chemical monitoring methods both at a “macroscale” and “microscale”. The “macroscale” studies consisted of process optimisation experiments that found propionate to be a more effective and stable carbon source than acetate. The “microscale” study investigated the activity of Competibacter, growing in dense aggregates. This study discovered that the structure of the granules affected the distribution of activity by limiting the supply of oxygen and that the activity of the Competibacter in turn affected the structure of the aggregate. The second experimental approach was to target key facets of the microbial physiology of PAOs and GAOs at a molecular level. Environmental gene expression studies were used to investigate the stimulus for the expression of a putative Accumulibacter polyphosphate kinase gene (ppk). This study found that the expression of this gene was repressed by high external phosphate concentrations, which suggests that the pho regulon is functioning in Accumulibacter. In another study, previously published models were integrated and elaborated to develop a model for the membrane transport processes in PAOs and GAOs, which give them the unique ability to sequester VFA without an electron acceptor. These studies confirmed that the proton-motive force (PMF) drives the uptake of VFA by both PAOs and GAOs and postulated fundamental differences in the molecular mechanisms that PAOs and GAOs use to create a PMF in the absence of respiratory electron transport. The studies also explain the molecular basis for findings in other studies that PAOs have a competitive advantage over GAOs at increased pH. The third experimental approach was to attempt to isolate organisms significant to EBPR. Some measure of success was achieved: colonies of Competibacter were obtained in pure culture but the growth could not be sustained further than the growth of micro-colonies just visible to the eye. EBPR microbiology, like many other subjects of inquiry in environmental microbiology, has benefited greatly from developments in molecular methods to identify and describe microbial communities. However, the investigation of microbial physiology in the environment remains a challenge; this thesis has taken up that challenge. Discoveries regarding the benefits of propionate as a carbon source and the basis for the competitive advantage that PAOs derive from an increased pH have potential application for practitioners of EBPR plants. Furthermore the findings make a contribution to the fundamental understanding of the physiology of EBPR organisms that may in the future lead to entirely novel approaches to EBPR optimisation.

The goal of this study was to assess the epiphytic diatom community structure of two host species along a karst gradient in the upper Green River, Kentucky to a gain a better understanding of the role of diatoms in the food web. The host species studied were Podostemum ceratophyllum and Cladophora. Percent cover of P. ceratophyllum and Cladophora were quantified in the four study reaches. The host species were sampled near-shore and mid-channel in each reach in September and October of 2013. After diatoms were extracted from the host and enumerated the density and diversity were quantified. Twelve genera were identified with > 91% of the community in each reach being Cocconeis. The second most abundant genus was Achnanthes or Navicula depending on the reach. The density and diversity of diatoms increased longitudinally going downstream. Exceptions to this trend occurred when high flow events disturbed the community. Within reaches there were no differences in diatom diversity in near-shore and mid-channel habitats. Diatom density in near-shore and mid-channel habitats was only different in the most downstream reach. Cladophora had a community twice as dense as P. ceratophyllum, but less diverse. The results of this study indicate that there are longitudinal differences in diatom communities in the upper Green River and host species are an important factor in determining the community composition. The importance of epiphytic diatoms in the food web, however, remains unclear.

No description available.

No description available.

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