The interactions between parasites, hosts, and the ecosystems they live in are complex and diverse. This can make predicting how they will respond to pressures like nutrient enrichment difficult. However, this endeavor is important, due to the far-reaching effects it has on ecosystems functioning, industries like aquaculture, and even human health. Even with the significance of this topic, there are key parts of the picture that have received little research attention. For this reason, I chose to study the effects of nutrient enrichment on parasite-host interactions at two different scales, first investigating the interaction directly between parasites and their host, and then manipulating experimental ponds to look at trophic effects. Using the theory of ecological stoichiometry as a conceptual framework allowed for comparison across the two investigations. In both studies, I used largemouth bass and the macro parasites of their digestive tract as my study system, due to the importance of largemouth bass as sport fish, their high trophic status, and diverse parasite assemblages. My first study examined the effects of eutrophication on parasite-host interactions occurring within the host. It centered on four hypotheses: 1) the nutrient content of different host tissues (infection sites) varies within and across hosts, 2) the nutrient contents of parasite genera differ from that of their host tissue(s), 3) the nutrient contents of parasite genera differ from one another, and 4) nutrient availability within specific host tissues is related to the nutrient content and abundance of parasite genera within tissues. I quantified the nutrient content (carbon (C), nitrogen (N), and phosphorous (P) and their ratios) of parasitic infracommunities in the tissues of the liver, stomach, pyloric caeca, intestines, and visceral mesentery of 30 largemouth bass (Micropterus salmoides). I found stoichiometric variation across host tissues and that fish collection site explained variation in pyloric caeca N:P ratios. Parasitic genera differed in their nutrient content with actively feeding parasite forms having higher %N and lower C:N ratios than encysted/non-reproductive forms. In addition, the %C of both actively feeding parasites varied across organs, and, for one genus, this variation reflected differences in host tissue %C. Finally, I found that the total number of actively feeding parasites in the pyloric caeca increased with that tissue’s N:P ratio. My results suggest that parasites encounter significant variation in nutrient availability within and across hosts, and that this variation can influence the nutrient content and abundance of some actively feeding parasites within specific tissues. To determine if the effects from my first study were a result of nutrient changes to host tissues or if they were an artifact of changes occurring at other trophic levels caused by nutrient addition, I conducted an experimental manipulation of a short trophic system. I applied two treatments, a fertilized treatment and a control, to 14 ponds stocked with largemouth bass. I hypothesized that adding a low N:P fertilizer to the ponds would result in fewer parasites in the pyloric caeca of the bass in accordance with findings from my previous work. Alternatively, I hypothesized that indirect changes to intermediate hosts would be responsible for changes in parasite abundance. To measure the effects of nutrient additions on multiple trophic levels, samples of fine particulate organic matter (FPOM), benthic invertebrates, and vertebrate prey were taken from each pond. Stocked bass were retrieved and dissected to obtain samples for nutrient analysis from their pyloric caeca, and to quantify parasite abundances. My treatment was able to affect the %P, C:N, C:P, and N:P of the FPOM in the ponds. Benthic invertebrate biomass was negatively correlated with FPOM %P. I did not find a direct effect of benthic invertebrates on vertebrate prey species, but fertilized ponds had a smaller decrease in the number of vertebrate prey over the course of the experiment. Counterintuitively, the %N of the pyloric caeca of fish in the fertilized ponds decreased, and the %C and C:N increased in relation to the number of vertebrate prey, but %P did not change. The treatment status of the pond and the nutrients of the pyloric caeca did not affect the number of parasites that were found, but parasite numbers were positively associated with prey fish. This result would suggest the number of intermediate hosts, in this case, prey fish, is the determining factor for parasitic loads in the bass I collected. The indirect effects of trophic interaction and the direct effects of changes to host and parasite dynamics both play important roles in predicting the outcome of eutrophication on disease. The conflicting results of these two studies highlight the need for in-depth knowledge of the systems being affected by nutrient additions.

The environmental processes and conditions controlling productivity and organic matter (OM) accumulation/preservation as well as bottom–water redox conditions in the lower black shale (LBS) and upper black shale (UBS) members of the Devonian-Mississippian (D–M) Bakken Formation were evaluated utilizing trace metal (TM) concentrations, degree of pyritization (DOPT), enrichment factors (EF) of TMs, bi–metal ratios (V/Cr, V/(V+Ni), Ni/Co, U/Th), total sulfur (ST) vs. iron (Fe), total organic carbon (TOC), carbon–sulfur–iron relationships (C–S–Fe), as well as Mo–TOC and Mo EF–U EF relationships. High-resolution (1- to 3-cm scale) chemostratigraphic records were generated for twelve drill cores, four of which closely flank the N–S-trending axis of the Nesson Anticline, proximal to the center of the Williston Basin in northwest North Dakota, USA. Furthermore, five of the twelve drill cores were selected (sample selection was based on down–core spacing and TM concentrations) for petrographic and Rock-Eval analysis to assess variations in kerogen type, quantity, quality, and thermal maturity (based on solid bitumen reflectance (%SBRo), vitrinite reflectance equivalence (%VRE), Rock–Eval Tmax–derived vitrinite reflectance (%Ro)) from immature to condensate, wet gas hydrocarbon generation windows. Degree of pyritization (DOPT) values (0.25 to 1.0) indicate that bottom waters were frequently dysoxic (> 60%) with intermittent aerobic and anoxic/euxinic conditions which is in agreement with C–S–Fe and total ST vs. Fe assessments of paleoredox conditions and sedimentological evidence. Furthermore, using published Mo–TOC relationships from modern anoxic-euxinic basins, it is estimated that renewal time of the sub-chemoclinal water mass during accumulation of the LBS and UBS approximated 10 and 30 yrs., respectively. Agreement is also seen between Mo/TOC and Mo EF/U EF where both suggest the Bakken shales were deposited under relatively unrestricted water mass conditions resulting in consistent renewal of TMs into the basin. However, bi–metal ratios suggest > 80% of samples were deposited under suboxic to anoxic/euxinic conditions. Trace metal concentrations for the Bakken Fm. show considerable range for Co (0–10324 ppm), Mo (0–2018 ppm), Ni (0–1574 ppm), U (0–1604 ppm), and V (0–3194 ppm), and bi–metal ratios for the Bakken Fm. are up to 5x greater than those reported for other D–M black shale formations. The Bakken black shales represent a unique sedimentary system where the EF of various TMs such as Cu (6.2–7.7), Mo (219.7–237.8), Ni (9.4–10.2), U (20.6–29.3), V (9.9–14.2), and Zn (10.4–12.2) as well as total organic carbon contents (LBS = 10.80 and UBS = 11.80 avg. wt.%) are considerably higher than other Devonian–Mississippian black shales. In this study, raw distributions of elemental concentrations combined with bivariate and principal component analysis (PCA) were used to elucidate the processes that could have contributed to the high EF of TMs in the Bakken shales. Total organic carbon shares heavier PCA component loadings (>0.445) and stronger correlation coefficients (r) with Cu, Mo, Ni, U, V, and Zn rather than with pyrite-associated (As, Co, Fe, and S) elements, suggesting that TOC played a primary role in the scavenging and accumulation of TMs in the sediments. Reducing conditions within bottom waters or sediment pore waters may have accelerated the accumulation of redox-sensitive Cu, Mo, Ni, V, and Zn introduced into the sediments via primarily an organic matter (OM) detritus host and most likely played a secondary role in the enrichment of TMs. The high EF of TMs observed in the Bakken shales may be the result of the frequent resupply of TMs into basin waters, enhanced primary productivity that is necessary in scavenging TMs from the water column, the presence of H2S within sediment pore or bottom waters, or possibly secondary processes associated with basin-wide fluid and hydrocarbon migration. Factors controlling TM accumulation during time of deposition (e.g., TM availability, bottom-water redox conditions, adsorption onto organic matter) and during diagenesis and catagenesis (e.g., alteration and break down of organic matter, movement of fluid hydrocarbons or other basinal fluids) likely contribute to the lack of agreement between redox proxies, and subsequently, the lack of applicability of bi–metal ratios (i.e., V/Cr, V/(V+Ni), Ni/Co, U/Th) in assessing bottom–water conditions for the Bakken shales. Solid bitumen (SB), a secondary organic matter formed as a residue after hydrocarbon generation (through either sufficient thermal maturation or microbial degradation) and expulsion, is primarily dispersed within the mineral matrix and increases in quantity with increasing thermal maturity. Rock-Eval II and HAWK analyzers were used to measure and estimate the hydrogen index (HI; avg. 201 mg HC/g TOC), oxygen index (OI; avg. 7mg CO2/g TOC), S1 (free hydrocarbons; avg. 8.0 mg HC/g rock), S2 (hydrocarbons generated after cracking kerogen; avg. 24.3 mg HC/g rock), and %Ro (0.60–1.03%; estimated from Tmax). The HI and OI values are calculated from TOC as well as S2 and S3 (oxygen bonded to hydrocarbons). Plots of HI vs. Tmax (ºC) and HI vs. OI as well as S2 vs. S3 ratio were utilized to determine the type of kerogen, primary OM that is insoluble in organic solvents. However, these relationships are not in agreement with kerogen typing based on petrographic observations, where samples from more thermally mature cores plot as Type III (vitrinite) kerogen instead of observed Type I/II (marine algae) kerogen. This is largely due to the abundant presence of SB in the more thermally mature section of the Bakken (Rock-Eval Ro = 0.83–1.03%) as SB is known to have a lower HI content than Type II kerogen. Petrographic evidence shows greater abundance of alginite and amorphous organic matter (AOM or bituminite) in the thermally less mature (Rock-Eval Ro = 0.60–0.83%) section of the Bakken compared to the greater abundance of dispersed SB in the more thermally mature section where AOM is absent. Early research on the Bakken Fm. reported lower than expected vitrinite reflectance values attributed to vitrinite “suppression". The overall lack of vitrinite and abundance of solid bitumen in these shales suggests that these early attempts likely reported solid bitumen reflectance rather than vitrinite reflectance. More recent attempts to assess the thermal maturity of the Bakken Fm. black shales have measured and converted SBRo to vitrinite reflectance equivalent (VRE). However, samples selected for SBRo by some previous workers have included heterogenous, granular as well as high reflecting SB samples, which introduce error in the measurements. As such, reported reflectance values are most likely lower than they would be if smooth, homogenous solid bitumen with no inclusions were measured. For this project, smooth and homogenous SB was measured to produce consistent and reliable VRE values to assess the thermal maturity gradient from the Bakken Fm. basin margins to the depocenter. Blue-light fluorescence petrography was done to support thermal maturity assessments. Results from SBRo, Rock-Eval Ro, VRE, and blue-light fluorescence observations suggest that cores from the current study range from early oil window into condensate, wet gas.

Cyanobacteria, also known as blue-green algae for a long time, are the most ancient and problematic bloom-forming phylum on earth. An alert levels framework has been established by World Health Organization(WHO) to prevent the potential harmful cyanobacterial blooms. Normally, low cyanobacteria levels are found in surface water. 2000 cyanobacterial cells/mL and 100,000 cyanobacterial cells/mL are established for WHO Alert Level 1 and 2, respectively. However, eutrophication, climate change and other factors may promote the spread of cyanobacteria and increase the occurrence of harmful cyanobacterial blooms in water on a global scale. Hence, a rapid real time cyanobacteiral monitoring system is required to protect public health from the cyanotoxins produced by toxic cyanobacterial species. Current methods to control or prevent the development of harmful cyanobacterial blooms are either expensive, time consuming or not effective in the long term. The best method to control the blooms is to prevent the formation of the blooms at the very beginning. Although emerging advanced autofluorescence-based sensors, imaging flow cytometry applications, and remote sensing have been utilized for rapid real-time enumeration and classification of cyanobacteria, the need to accurately monitor low-level cyanobacterial species in water remains unsolved. Microflow cytometry has been employed as a functional cell analysis technique in past decades, and it can provide real-time, accurate results. The autofluorescence of cyanobacterial pigments can be used for determination and counting of cyanobacterial density in water. A pre-concentration system of an automated cyanobacterial concentration and recovery system (ACCRS) based on tangential flow filtration and back-flushing technique was applied to reduce the sample assay volume and increase the concentration of target cells for further cell capture and detection. In this project, a microflow cytometry platform with a microfluidic device and an automated pre-concentration system was established to monitor cyanobacteria and provide early warning alerts for potential harmful blooms. In this work, quantification of low-level cyanobacterial samples (∼ 5 cyanobacterial cells/mL) in water has been achieved by using a microflow cytometer together with a pre-concentration system (ACCRS). Meanwhile, this platform can also provide early warning alerts for potential harmful cyanobacterial blooms at least 15 days earlier before reaching WHO Alert Level 1. Results have shown that this platform can be applied for rapid determination of cyanobacteria and early warning alerts can be triggered for authorities to protect the public and the environment. / Thesis / Doctor of Engineering (DEng) / Harmful cyanobacterial blooms have been a rising risk to the public heath across the world in recent decades. Alert levels of cyanobacteria in water has also been established. In this case, a rapid on-side monitoring system for cyanobacteria is required. In this thesis, a microflow cytometer platform combined with a bacterial concentration and recovery system was built to quickly monitor the relatively low level of cyanobacteria for early warning alerts. A pre-enrichment system based on tangential flow filtration and back-flushing technique was applied to increase the concentration levels of microbial samples and a microfluidic device capable of collecting phycocyanin fluorescence was designed to count cyanobacterial cells. The limit of quantification for cyanobacterial concentration based on the microflow cytometry platform was as low as ∼ 5 cells/mL. We can claim that the microflow cytometry platform can provide useful early warning alerts for the decision-makers to control the potential harmful cyanobacterial blooms at the very early stage and protect the aquatic animals and public health.

Lead contamination threatens human life and the environment. The biological reduction of Pb(II) to metallic Pb is an attractive solution for Pb(II) pollution. Delftia acidovorans, Azonexus caeni, and Comamonas testosteroni were isolated and studied for their capabilities to utilize Pb(II) as a terminal electron acceptor. D. acidovorans strain Pb11 and A. caeni strain Pb2 cultures showed a 5.2- and 8.1-fold growth at 10.0 mg-Pb(II)/L in 3 d, respectively. Petroleum-based plastics are another emergent environmental concern. Polyhydroxyalkanoates (PHAs), a sustainable alternative to conventional plastics, are biodegradable polymers produced by PHA accumulators under autotrophic or heterotrophic conditions. In this thesis, the growth and enrichment of PHA accumulators, such as Plasticicumulans acidivorans and Cupriavidus necator, were investigated as they can accumulate 90% of their cell weight as PHA. An energetic model was developed to calculate theoretical PHA yields. The true autotrophic and heterotrophic PHB yields were estimated as 2.97 (g_PHB/ 〖mol〗_(H_2 )) and 0.66 (g_PHB/ g_acetate), respectively. Moreover, the growth of C. necator was investigated in lab-scale experiments under various autotrophic, heterotrophic, and mixotrophic conditions. When C. necator was cultivated in two-stage systems, high optical densities were attained in less than 24 h. In addition, a mathematical model for the competition between PHA and non-PHA accumulators in the feast-famine enrichments was developed. The calibrated and validated model for P. acidivorans suggested that microbial diversity in mixed cultures impacted the enrichment process. Another aspect of this thesis was to propose an innovative method for enriching PHA accumulators in mixed cultures. By applying autotrophic and autotrophic-heterotrophic enrichment strategies, C. necator dominated the mixed cultures (> 90%) in less than five days. Based on this thesis findings, it can be concluded that biotechnology applications in Pb(II) remediation and PHA production could reduce the severe impacts of Pb contamination, petrochemical plastics, and climate change due to elevated CO2 levels. / Thesis / Doctor of Philosophy (PhD) / This thesis aimed to provide sustainable biotechnological solutions to three environmental challenges: lead contamination, petroleum-based plastics, and elevated CO2 levels in the atmosphere. Certain metal-reducing bacteria can grow by consuming toxic Pb(II) ions from aqueous environments and thus reduce their toxicity. Furthermore, various microorganisms can store biodegradable polymers, known as polyhydroxyalkanoates (PHAs), in their cells. The stored PHA polymers can be extracted and processed to produce biodegradable plastics. PHA accumulators can produce significant amounts of PHA by utilizing organic substrates or CO2. Therefore, PHA-based plastics can reduce environmental deterioration due to non-degradable plastics and elevated CO2 levels. Lab-scale experiments and mathematical modeling can provide a better understanding of the growth and enrichment of PHA accumulators in engineered PHA-production systems. Research findings in this thesis will allow cost-effective and sustainable production of biodegradable plastics from organic wastes and flue gas.

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