Thesis (Ph.D.)--University of Groningen, 2007. / Includes bibliographical references (leaves 106-116). Available also electronically on the Internet.

archives@tulane.edu / During the Deepwater Horizon (DWH) oil spill in 2010, approximately 0.5 billion liters (3.1 million barrels) of oil were released into the northern Gulf of Mexico during the largest marine oil spill in history . A significant portion of the released oil was weathered into residues by physical, photochemical, and biological processes prior to landing on 1773 km of coastline, including 754 km of marsh shoreline in Louisiana. Researchers endeavored to describe effects of oil residues in the soil on salt marsh organisms and communities. Many studies focused on two pillars of salt marsh ecology: the microbial communities through which a large portion of the salt marsh food web is connected and Spartina alterniflora, a foundational species of Gulf Coast salt marshes. In this dissertation I describe how cryptic, or difficult to observe, elements of salt marsh ecology, like microbial communities and plant genetics, respond to oil residues in the environment. Using a suite of field, growth chamber, and greenhouse experiments I show that these microbial communities are difficult to characterize and may respond to other factors more strongly than they do to oil residues. I present evidence that the plant is resilient to oil in the environment, and changes in its microbiome, but exerts a measurable influence on the biodegradation of oil residues and the microbiome in the soil. This dissertation provides a greater understanding of the complexity of the salt marsh response to an oil spill. / 1 / Stephen K. Formel

This research investigated the response of the dominant salt marsh macrophyte Spartina alterniflora var. Loisel to sedimentation from Hurricanes Gustav and Ike. A range of sedimentation was used to track the above and belowground plant response, as well as the volume of mineral and organic matter, in the upper reaches of the marsh soil from September 2008 to April 2009. Where storm sedimentation was greatest (~3.5 cm), there were significant increases in live aboveground biomass. Although live belowground biomass did not change significantly over time, the volumetric contribution of both mineral and soil organic matter in the shallow marsh soil increased. These results indicate that hurricane sedimentation may benefit subsiding coastal marshes by stimulating aboveground vegetation mass as well as soil organic matter volume. However, these results imply that there may be a sediment thickness threshold that must be met before such positive effects are expressed.

Simulated oil spills were created in S. alterniflora sediment-water microcosms to determine the effects of applied crude oil on S.alterniflora during two 90-day studies. In the first experiment, oil dosage was varied at 0-250 mg crude oil/g wet soil to determine the lethal dosage level. In the second experiment, oil type, dosage, and soil type were varied to determine the effects of oil under multiple scales of resolution. A light, medium, and heavy crude oil at dosages ranging from 0-150 mg crude oil/g wet soil were used in addition to an oiled and non-oiled soil. Following the completion of the 90-day experiment, several key findings were observed: (1) The lethal dosage limit was reached at 250 mg crude oil/g wet soil during the first experiment but not the second, by design; (2) At initial dosages of 10 and 50 mg crude oil/g wet soil, the oiled soil (acclimated for 4 months) was more influential in decreasing cumulative biomass growth rates compared to oil applied at the oil-water interface; (3) At the heaviest dosages applied as a simulated oil slick, concentrations of 150 mg crude oil/g wet soil, evapotranspiration rates were negatively affected by the oil (significant at p=0.05 in a one-tailed t-test); (4) Light, heavy, and then medium crude oil showed the lowest biomass growths, in that order, indicating that light crude oil was the most toxic in these microcosm experiments with S. alterniflora; (5) The 10 mg oil/g wet soil out-performed the 0 mg oil/g wet soil in transpiration and biomass growth.

Light, medium and heavy crude oils were studied at three concentrations and with two different sediments in experimental microcosm settings to determine the ability of Spartina alterniflora and associated microbes to breakdown total extractable hydrocarbons (TEH) in the water. It was a baseline experiment designed to quantify the rates of biodegradation under relatively quiescent conditions from different crude oils at moderate doses ranging from 0-150 mg/kg soil. Upon the completion of the experiment there were several key findings: (1) The lethal dosage for Spartina alterniflora was not reached within the 90 day experiment at these dosages, and all plants survived; (2) More than 97% of the total extractable hydrocarbons (TEH) were shown to be degraded by plants and rhizosphere microorganisms within the 90- day experiment; (3) The dose of oil introduced as a slick (simulated spill) on day zero did not significantly affect the results for TEH degradation within the range of dosages from 50-150 mg/g -- these dosages could be degraded by the marsh cord grass system; (4) A sediment type which was acclimated to oil for several months and one which was non-acclimated did not show significantly different results for TEH degradation in the microcosms -- both sediment systems resulted in TEH degradation over the 90-day experiment; and (5) A mathematical model was developed which simulated experiment results quite closely including TEH diffusion from the crude oil slick into the water and subsequent biodegradation.

Eutrophication, particularly due to nitrogen (N) and phosphorus (P) input, has been massively altered by anthropogenic activities. Thus it is important to understand the impact on salt marsh plants; however studies on salt marsh plants within Galveston Bay, Texas are limited. In this study, the effects of repeated nutrient additions in monospecific plots of Spartina alterniflora, Batis maritima¸ and Salicornia virginica as well as mixed plots of B. maritima and S. virginica were studied over 15 months. Results showed that nutrient loading led to an increase in height, biomass, growth rate and percent nitrogen (N) within all three species studied, but were species specific more than dose dependent. Nitrogen content in leaves had a positive correlation with P content in leaves but a negative correlation with carbon (C) content. Nutrient loading lead to a significant increase in total chlorophyll in the fertilized plots of S. alterniflora and S. virginica one month into the study. Nutrient addition to two succulent species, B. maritima and S. virginica in mixed plots did not reveal a distinct superior competitor within the 15 month study in terms of growth and nutrient use efficiencies; however using the maximum growth rates of the monospecific plots, the Monod model was used to determine which species would dominate at high nutrient loads. Based on height data S. alterniflora would dominate, while B. maritima would dominate according to the Monod model based on biomass.

Thesis (Ph. D.)--Florida State University, 2005. / Advisor: Dr. Joel Kostka, Florida State University, College of Arts and Sciences, Dept. of Oceanography. Title and description from dissertation home page (viewed June 22, 2005). Document formatted into pages; contains xi, 108 pages. Includes bibliographical references.

Dissertação (mestrado) - Universidade Federal de Santa Catarina, Centro de Ciências Biológicas. Programa de Pós-Graduação em Biologia Vegetal / Made available in DSpace on 2012-10-20T18:06:06Z (GMT). No. of bitstreams: 1 200047.pdf: 5416566 bytes, checksum: 366692677a5796f5a722a5a189c5b201 (MD5) / Na Ilha de Santa Catarina são encontrados significativos ecossistemas de marismas, em especial na Enseada de Ratones, onde verificam-se densos bancos monotípicos de Spartina alterniflora Loisel. No presente trabalho foram avaliadas a variação da biomassa e da densidade da espécie neste estuário durante a primavera (2000) e outono (2001), bem como a cinética de decomposição da gramínea entre os meses de abril a novembro de 2001. Durante o período avaliado a marisma apresentou 569±128 hastes.m-2. No outono foram verificadas 648±225 hastes.m-2, sendo que destas 32,8% eram senescentes. Na primavera foram registradas 516±80 hastes.m-2, das quais 26,1% estavam em senescência. A média de fitomassa encontrada para a enseada de Ratones foi de 274±46gPS.m-2. Durante a primavera a fitomassa epígea foi de 326±25gPS.m-2 e 197±90gPS.m-2 no outono. Na primavera foi verificada a maior necromassa, tanto em termos absolutos quanto relativos. Durante o outono a fitomassa subterrânea foi muito superior à da fitomassa aérea, resultando em uma baixa relação massa hipógea/epígea. Os sedimentos da marisma de Ratones são orgânicos e de granulometria síltica-argilosa. A cinética de decomposição seguiu um modelo exponencial, e a meia-vida obtida para S. alterniflora foi de 153 dias para o peso seco, com um coeficiente de decomposição de 0,0045 e 52 dias para o peso seco sem cinzas, com um coeficiente de decomposição de 0,0068. O teor de nitrogênio agregado aos detritos, submetidos ao processo de decomposição, indica uma intensa atividade da micro e macrofauna associada a cadeia detritívora. A diversidade da macrofauna encontrada nos detritos de S. alterniflora confirmou sua importância na manutenção da biodiversidade estuarina da enseada de Ratones. Durante a colonização dos detritos de Spartina alterniflora pela macrofauna, houve decréscimo progressivo de crustáceos, paralelo ao acréscimo de outros grupos, principalmente oligoquetas. A variação percentual dos teores de carbono e nitrogênio verificados durante o processo de decomposição de S. alterniflora são indícios da importância da gramínea como fonte de compostos carbonáceos e nitrogenados para o estuário.

Phenotypic plasticity is the ability of a given genotype to exhibit different phenotypes in response to environmental variables, which can impact population level processes. Plasticity of ecologically-relevant traits is important to an organism’s environmental response; however, the underlying mechanisms of plasticity are largely unknown. Ecological epigenetics may offer mechanisms (e.g. DNA methylation) underlying phenotypic plasticity. Epigenetics can be defined as the underlying molecular mechanisms that allow one genotype to exhibit different phenotypes. Differential DNA methylation is one epigenetic mechanism that has been correlated with a number of ecologically-relevant traits; including, differential herbivory in Viola cazorlensis, spinescence in Ilex aquifolium, flower morphology in Linaria vulgaris, and fitness in Arabidopsis thaliana. The epigenetic correlations with traits found in these studies are interesting, but they are also partially confounded by a potential correlation between genetic and epigenetic variation. Teasing apart the correlation between genetic and epigenetic variation is one of the challenges within ecological epigenetics. This correlation has resulted in epigenetic variation being partitioned into three types by researchers: obligate, facilitated, and pure. Changes in obligate epigenetic variation are directly correlated with genetic variation. Changes in pure epigenetic variation are completely independent from genetic variation. Changes in facilitated epigenetic variation are partially dependent on genetic variation, but the outcome of the phenotype is context-dependent based on environmental conditions. Since our predictions about the outcome of phenotypic variation are driven largely by population genetics theories, which make no room for variation that operates in non-Mendelian ways, epigenetics research needs to utilize unique ways to tease apart the interaction between genetic and epigenetic variation where facilitated or pure epigenetic variation exists outside of the realm of population genetics theory. To address these issues, I performed a literature review and two research-based studies. In Chapter 1 I performed a literature review on the topic of population epigenetics addressing the correlation with genetic variation and recommending an extension to the Modern Synthesis to accommodate the non-Mendelian nature of DNA methylation. While population genetics has approximately 85-years of data to support it, epigenetics is beginning to show some of the limitations associated with predictions made using populations genetics models. One of these limitations is that population genetics as defined by the Modern Synthesis does not allow for violations of Mendelian genetics (i.e. random assortment and segregation of alleles). This limitation does not allow for phenotypic variation that is directly due to environmental conditions; however, recent ecological epigenetics data shows that this can, indeed, occur. Within this review I propose epigenetic questions that we should focus on at the population level, and I make recommendations for how to approach these questions in future studies. In the second and third research-based chapters, I investigated whether an independent component of epigenetic variation was correlated with habitat, while controlling for a correlation with genetic variation, for Spartina alterniflora and Borrichia frutescens, respectively. Previous work has shown that there is no consistent genetic response to environment in these species. I, therefore, hypothesized that there would be a significant epigenetic correlation with habitat instead. To test this hypothesis, I collected leaf samples from five different sites for each species on Sapelo Island, GA. Within each site I established three 10m transects (n=20 for each microhabitat) in low, middle, and high marsh microhabitats, respectively. Plants of both species exhibit different phenotypes for height (tall, intermediate, short, respectively) based on their location within the marsh. I screened AFLP and methylation-sensitive AFLP (MS-AFLP) markers for genetic and epigenetic variation, respectively. I used a variety of statistical tests to attempt to tease apart a potential correlation between genetic and epigenetic variation and found that when genetic population structure is controlled for, significant epigenetic population structure persists across all populations for S. alterniflora and within 3 of 5 populations for B. frutescens. These results suggest that regulation of certain genomic elements via DNA methylation may play an important role in dealing with environmental variables. To fully determine the significance of these findings, future studies should examine the interaction between environmentally-mediated epigenetic variation and gene expression to determine its importance to phenotypic plasticity and habitat differentiation. The body of work I produced supports that epigenetics may play a role in environmental response in populations within relatively small spatial scales. I used a combination of statistical tests to control for potential correlations with genetic variation which allowed me to see patterns that may normally be hidden. These findings expand upon traditional views of evolution by suggesting that environment can play a role in phenotypic variation, and other research supports that the variation due to epigenetic mechanisms can be inherited in future generations. Much of the current epigenetic research is based upon studies involving model species in highly controlled studies. While this research is been incredibly informative about some of the mechanisms underlying epigenetics, to fully understand the role of epigenetics to environmental response and evolution we must pair these data with field studies of non-model organisms. Only then will we begin to see the full role of epigenetics in organisms.

Herbivores and pathogens pose a consistent threat to plant productivity. In response, plants invest in structural and/or chemical defenses that minimize damage caused by these biotic stressors. In salt marshes along the Atlantic coast of the United States, a facultative mutualism between snails (Littoraria irrorata) and multiple species of fungi exert intense top-down control of the foundation grass species Spartina alterniflora. Since exposure to herbivores and pathogens are tightly coupled in this system, I investigated whether S. alterniflora utilizes chemical and/or structural defenses to deter both snails and fungi, and examined how plant defenses varied among S. alterniflora individuals and populations. I also assessed how other marsh plants prevent snails from establishing farms, and considered whether interspecific variation in plant chemical defenses influences marsh community structure. Initial experiments revealed that S. alterniflora chemical defenses inhibited L. irrorata and two fungi that snails commonly farm. A caging experiment determined that production of chemical defenses could not be induced in the presence of snails and fungi, nor relaxed in their absence. Through separations chemistry guided by ecological assays, I isolated two distinct classes of chemical defenses from short form S. alterniflora, one of which inhibited fungal growth and the other decreased plant palatability. In a community context, the chemical defenses produced by S. alterniflora were relatively weak compared to those of four other salt marsh plant species, which produced compounds that completely inhibited L. irrorata grazing and strongly hindered fungal growth in lab assays. Nutritional and structural differences among marsh plants did not influence feeding preferences, suggesting that plant secondary chemistry was the primary driver for food selection by snails. It appears that S. alterniflora produces weak chemical defenses that slow down or limit fungal growth and snail herbivory, and may compensate for tissue losses by producing new growth. In contrast, less abundant marsh plants express chemical defenses that completely inhibit fungal farming and deter snail grazing, but doing so may come at a cost to growth or competitive ability. As marsh dieback continues with rising herbivore densities and compounding abiotic stressors, the ecosystem services that salt marshes provide may be lost. Therefore, understanding how and under what conditions salt marsh plants resist losses to herbivores and pathogens will help predict which marsh communities are most likely to be threatened in the future. Initial experiments revealed that S. alterniflora chemical defenses inhibited L. irrorata and two fungi that snails commonly farm. A caging experiment determined that production of chemical defenses could not be induced in the presence of snails and fungi, nor relaxed in their absence. Through separations chemistry guided by ecological assays, I isolated two distinct classes of chemical defenses from short form S. alterniflora, one of which inhibited fungal growth and the other decreased plant palatability. In a community context, the chemical defenses produced by S. alterniflora were relatively weak compared to those of four other salt marsh plant species, which produced compounds that completely inhibited L. irrorata grazing and strongly hindered fungal growth in lab assays. Nutritional and structural differences among marsh plants did not influence feeding preferences, suggesting that differences in plant chemistry were the primary driver for food selection by snails. It appears that S. alterniflora produces weak chemical defenses that slow down or limit fungal growth and snail herbivory, and may compensate for tissue losses by producing new growth. In contrast, less abundant marsh plants express chemical defenses that completely inhibit fungal farming and deter snail grazing, but doing so may come at a cost to growth or competitive ability against S. alterniflora. As marsh dieback continues with rising herbivore densities and compounding abiotic stressors, the ecosystem services that salt marshes provide may be lost. Therefore, understanding how and under what conditions salt marsh plants resist losses to herbivores and pathogens will help predict which marsh communities are most likely to be threatened in the future.