Fire Effects on Soil Properties: Amending Post-Fire Soils with Native Microbial Communities and Biochar to Improve Sagebrush Performance

Within the sagebrush steppe, fire has been shown to affect biogeochemical properties and the microbial community composition in soils. However, there is uncertainty about the magnitude and direction of these effects, since they vary by sites that differ in abiotic and biotic conditions. Moreover, differences in post-fire management strategies are likely to mediate the effect of fire on soil properties, thus further compounding this uncertainty. Any changes in soil biogeochemical properties following fire can prevent successful restoration of Artemisia tridentata sp. wyomingensis (sagebrush), leading to variable outcomes of restoration success in the sagebrush steppe. Previous research has shown that addition of native soil microbes and biochar can improve ecosystem restoration efforts, but the effects of these soil amendments on post-fire soil properties and sagebrush performance across sites are uncertain. With this study, I investigated how fire impacts soil properties (i.e., soil organic matter (SOM), soil structure, carbon (C) and nitrogen (N) concentrations, soil pH, net mineral N, microbial richness and composition) at a variety of sites that differ in time since exposure to fire, post-fire plant communities, and post-fire site management. I then implemented a greenhouse study to evaluate how amending soils with native microbial community inocula and biochar impact soil properties of the post-burn sites and sagebrush germination and growth. Taken together, these findings capture the influences of multiple fires and separate management strategies on soil properties, and how certain soil amendments may redirect soil recovery to aid in sagebrush restoration. In my first chapter, I asked two questions: (1) how does fire affect soil biochemical properties across sites that differ in fire history, post-fire plant communities, and post-fire site management, and (2) how does fire affect soil microbial richness and community composition across sites that differ in post-fire plant communities, and post-fire site management. To assess these questions, soils were collected from three south of Boise, Idaho within the Orchard Combat Training Center (OCTC) that contrasted in fire history, plant community, and post-fire management. The northern part of the Union Fire (180 acres; hereafter: UFN2011) burned in 2011, and was treated with a mix of imazapic and glyphosate during the spring of 2019, after which sagebrush was handplanted 8 months later that yea. The southern part of the Union Fire (160 acres; hereafter: UFS2011) burned in 2011 and seeded with native grass species and planted with sagebrush. The Christmas Fire (hereafter: CF2018) burned in 2018, and was subjected to the same seeding and handplanting treatments as UFS2011. At each site, I selected five locations within the perimeter of the burn, and five locations outside the burn, representing the unburned control plots. In these unburned control plots, the five locations were stratified by sagebrush canopy and interspace microsites separately. At each one of the five locations, I collected four soil cores (10cm depth, 2.5cm diameter). I evaluated differences in soil pH, soil organic matter (SOM), soil carbon (C) and nitrogen (N) contents, soil structure, N cycling, and soil microbial communities between burned and unburned sites. Fire reduced SOM and soil C contents, and these losses were greater in burned areas that received an herbicide treatment. This suggests that suppression of plant growth using herbicides may limit the recovery of soil properties that are foundational to sagebrush steppe ecosystem functioning. Furthermore, I found a loss of arbuscular mycorrhizal fungi (AMF) richness with fire and significant changes in soil microbial community structure when herbicide had been used. Finally, increased soil mineral N concentrations across all burned sites indicate that fire may significantly reduce ecosystem stability and increase the risk of invasion. These changes in soil properties are likely to lead to a persistent ecosystem state-changes in the sagebrush steppe, and future studies should evaluate which management approaches could be used to restore both soils and plant communities. In my second chapter, I investigated two management approaches that may be used to restore the soils and plant communities impacted by fire. I asked (1) How does a live native soil microbial inoculum impact sagebrush performance and soil properties, (2) how do biochar additions impact sagebrush performance and soil properties, and (3) how does prior management (e.g., herbicide) mediate the impact of soil microbial inoculation and biochar amendment on sagebrush performance? A full factorial greenhouse experiment was conducted for three months with soils collected from the three post-burn sites described in Chapter 1. In the greenhouse experiment I incorporated the following treatments: (1) sterile native inoculum [-Inoculum] and no biochar [-Biochar], (2) live native inoculum [+Inoculum] and no biochar [-Biochar], (3) sterile native inoculum [-Inoculum] and biochar [+Biochar], and (4) live native inoculum [+Inoculum] and biochar [+Biochar]. Inocula was derived from sagebrush canopies at unburned sites and either added as live native inocula or autoclaved to sterilize the microbial community. Biochar was crushed into planted, watered daily until cotyledons showed, and continually monitored throughout the growing period. Germination, soil moisture content and pH, above- and below-ground measurements, total mineral N, fungal root colonization proportional abundances, and microbial richness and composition were assessed. I found that inoculations did not significantly benefit sagebrush performance, most likely due to the ratio of inocula administered. In contrast, biochar consistently enhanced soil moisture, pH, sagebrush germination and other performance variables while its effects on total mineral N and fungal root colonization varied by site location. Lastly, presence of herbicide in post-burn soils significantly altered soil bacterial and fungal community composition, and its effects persisted enough to inhibit sagebrush performance. Together, my data show that addition of biochar has a greater positive impact on sagebrush germination and performance than addition of soil microbial inocula, and that herbicide addition has persistent negative impacts on sagebrush performance. My study captured the varying levels at which fire impacts ecosystem structure and function, and how different soil amendments affected sagebrush performance at these post-fire soils. My findings support the notion that soil properties will remain degraded without appropriate management strategies supporting restoration, and herbicide may actually suppress successful restoration, residing longer in the soil than previously documented. When growing sagebrush in post-burn soils within the greenhouse, biochar enabled soil recovery, and this benefited sagebrush performance. However, herbicide impacts persisted and decreased sagebrush biomass even when soil amendments were incorporated. Fire can have profound, yet vastly different, influences on soil properties, and soil amendments may be able to augment soil recovery. Future studies should investigate various soil amendments and their impacts on sagebrush performance in the midst of changing fire regimes, post-fire vegetation shifts, and current post-fire management

Conception et synthèse d'activateurs de la protéine proapoptotique Bax

L'apoptose ou mort cellulaire programmée est indispensable au maintien de l'homéostasie tissulaire. Ses dysfonctionnements, par excès ou par défaut, sont à l'origine de nombreuses pathologies (cancer, maladies neurodégénératives ...). Les protéines de la famille Bcl-2 sont des régulateurs importants du processus apoptotique. Bax est un membre proapoptotique de cette famille : suite à un stimulus, elle subit un changement conformationnel qui permet son insertion dans la membrane mitochondriale puis le déclenchement de l'apoptose. Au cours de cette thèse, nous avons cherché à développer des composés pouvant agir sur le site actif de cette protéine. Ceci est en effet une stratégie de choix pour le développement de nouvelles drogues proapoptotiques. Des études de modélisation moléculaire, basées sur la structure RMN de Bax, ont permis la conception de composés potentiellement inducteurs de l'apoptose de structure tricyclique. Ces nouveaux composés possèdent une structure pyridoquinoléine fonctionnalisée en alpha de la fonction amide par un groupement aromatique et par un bras de type oxypropionique en position 8. Nous avons ensuite développé une voie de synthèse rapide et convergente, nous permettant d'introduire facilement de la diversité structurale lors des dernières étapes de la synthèse dans le but d'établir une étude SAR. Les tests biologiques des premiers composés synthétisés sont actuellement en cours.Apoptosis or programmed cell death plays a vital role in normal development and disruption of the normal apoptotic process is responsible for a variety of human diseases including cancer and autoimmunity. The Bcl-2 family of proteins are important regulators of programmed cell death. Bax is a proapoptotic member of this family : during the apoptotic process it undergoes a conformational change which triggers its insertion from the cytosol into the mitochondrial membranes. During this PhD, we tried to develop some new compounds in order to act on the active site of the protein. This is indeed a promising strategy for the development of new proapoptotic drugs. Some molecular modeling studies based upon the NMR structure of Bax allowed the design of new potential inducers of apoptosis. These molecules have a pyridoquinoline scaffold with an aromatic moiety alpha to the amide bond and an oxypropionic arm linked to the phenol function. We developed a short and convergent synthesis of these molecules, allowing many variations at the last stages in order to establish a SAR study. We have already synthesised a few compounds which have been submitted to biological tests

A Quantitative Description for Optical Mass Measurement of Single Biomolecules

Label-free detection of single biomolecules in solution has been achieved using a variety of experimental approaches over the past decade. Yet, our understanding of the magnitude of the optical contrast and its relationship with the underlying atomic structure as well as the achievable measurement sensitivity and precision remain poorly defined. Here, we use a Fourier optics approach combined with an atomic structure-based molecular polarizability model to simulate mass photometry experiments from first principles. We find excellent agreement between several key experimentally determined parameters such as optical contrast-to-mass conversion, achievable mass accuracy, and molecular shape and orientation dependence. This allows us to determine detection sensitivity and measurement precision mostly independent of the optical detection approach chosen, resulting in a general framework for light-based single-molecule detection and quantification

Preclinical evaluation of NF-kappa B-triggered dendritic cells expressing the viral oncogenic driver of Merkel cell carcinoma for therapeutic vaccination

Background: Merkel cell carcinoma (MCC) is a rare but very aggressive skin tumor that develops after integration of a truncated form of the large T-antigen (truncLT) of the Merkel cell polyomavirus (MCV) into the host’s genome. Therapeutic vaccination with dendritic cells (DCs) loaded with tumor antigens is an active form of immunotherapy, which intends to direct the immune system towards tumors which express the respective vaccination antigens. Methods: Cytokine-matured monocyte-derived DCs of healthy donors and MCC patients were electroporated with mRNA encoding the truncLT. To permit major histocompatibility complex (MHC) class II next to class I presentation, we used an RNA construct in which the antigen was fused to a DCLamp sequence in addition to the unmodified antigen. To further improve their immunogenicity, the DCs were additionally activated by co-transfection with the constitutively active nuclear factor (NF)-κB activator caIKK. These DCs were used to stimulate autologous CD8 + T-cells or a mixture of CD4 + and CD8 + T-cells. Then the percentage of T-cells, specific for the truncLT, was quantified by interferon (IFN)γ ELISpot assays. Results: Both the truncLT and its DCLamp-fusion were detected within the DCs by flow cytometry, albeit the latter required blocking of the proteasome. The transfection with caIKK upregulated maturation markers and induced cytokine production. After 2–3 rounds of stimulation, the T-cells from 11 out of 13 healthy donors recognized the antigen. DCs without caIKK appeared in comparison less potent in inducing such responses. When using cells derived from MCC patients, we could induce responses for 3 out of 5 patients; however, here the caIKK-transfected DCs did not display their superiority. Conclusion: These results show that optimized DCs are able to induce MCV-antigen-specific T-cell responses. Therapeutic vaccination with such transfected DCs could direct the immune system against MCC

Characterization and differentiation of the tumor microenvironment (TME) of orthotopic and subcutaneously grown head and neck squamous cell carcinoma (HNSCC) in immunocompetent mice

For the development and evaluation of new head and neck squamous cell carcinoma (HNSCC) therapeutics, suitable, well-characterized animal models are needed. Thus, by analyzing orthotopic versus subcutaneous models of HNSCC in immunocompetent mice, we evaluated the existence of adenosine-related immunosuppressive B- and T lymphocyte populations within the tumor microenvironment (TME). Applying the SCC VII model for the induction of HNSCC in immunocompetent C3H/HeN mice, the cellular TME was characterized after tumor initiation over time by flow cytometry. The TME in orthotopic grown tumors revealed a larger population of tumor-infiltrating lymphocytes (TIL) with more B cells and CD4+ T cells than the subcutaneously grown tumors. Immune cell populations in the blood and bone marrow showed a rather distinct reaction toward tumor induction and tumor location compared to the spleen, lymph nodes, or thymus. In addition, large numbers of immunosuppressive B- and T cells were identified within the TME but also in secondary lymphoid organs, independently of the tumor initiation site. The altered immunogenic TME may influence the response to any treatment attempt. Moreover, when analyzing the TME and other lymphoid organs of tumor-bearing mice, we observed conditions reflecting largely those of patients suffering from HNSCC suggesting the C3H/HeN mouse model as a suitable tool for studies aiming to target immunosuppression to improve anti-cancer therapies

NF-κB and its role in checkpoint control

Nuclear factor-κB (NF-κB) has been described as one of the most important molecules linking inflammation to cancer. More recently, it has become clear that NF-κB is also involved in the regulation of immune checkpoint expression. Therapeutic approaches targeting immune checkpoint molecules, enabling the immune system to initiate immune responses against tumor cells, constitute a key breakthrough in cancer treatment. This review discusses recent evidence for an association of NF-κB and immune checkpoint expression and examines the therapeutic potential of inhibitors targeting either NF-κB directly or molecules involved in NF-κB regulation in combination with immune checkpoint blockade

Robustness of the charge-ordered phases in IrTe2{\mathrm{IrTe}}_{2} against photoexcitation

We present a time-resolved angle-resolved photoelectron spectroscopy study of IrTe2, which undergoes two first-order structural and charge-ordered phase transitions on cooling below 270 K and below 180 K. The possibility of inducing a phase transition by photoexcitation with near-infrared femtosecond pulses is investigated in the charge-ordered phases. We observe changes of the spectral function occurring within a few hundreds of femtoseconds and persisting up to several picoseconds, which we interpret as a partial photoinduced phase transition (PIPT). The necessary time for photoinducing these spectral changes increases with increasing photoexcitation density and reaches time scales longer than the rise time of the transient electronic temperature. We conclude that the PIPT is driven by a transient increase of the lattice temperature following the energy transfer from the electrons. However, the photoinduced changes of the spectral function are small, which indicates that the low- temperature phase is particularly robust against photoexcitation. We suggest that the system might be trapped in an out-of-equilibrium state, for which only a partial structural transition is achieved