Archaeological otoliths as environmental recorders: high resolution sampling of pacific cod (Gadus macrocephalus) otoliths from Kiska Island, Alaska

Archaeological fish otoliths have the potential to serve as proxies for both season of site occupation and palaeoclimate conditions. By sampling along the distinctive sub-annual seasonal bands of the otolith and completing a stable isotope (δ¹⁸O, δ¹³C) analysis, variations within the fish’s environment can be identified. Through the analysis of cod otoliths from two archaeological sites on Kiska Island, Gertrude Cove (KIS-010) and Witchcraft Point (KIS-005), this research evaluates a micromilling methodological approach to extracting climatic data from archaeological cod otoliths. In addition, δ¹⁸Ootolith data and radiocarbon dates frame a discussion of Pacific cod harvesting, site occupation, and changing climatic conditions on Kiska Island. To aid in the interpretation of the archaeological Pacific cod results, archaeological and modern Atlantic cod otoliths were also analyzed as a component of this study to develop. The Atlantic cod otoliths provided the methodological and interpretative framework for the study, and also served to assess the efficacy of this sampling strategy for archaeological materials and to add time-depth to existing datasets. The δ¹⁸Ootolith values successfully illustrate relative variation in ambient water temperature. The Pacific cod δ¹⁸O values demonstrate a weak seasonal signal identifiable up to year 3, followed by relatively stable values until year 6/7 when values continuously increase. Based on the δ¹⁸O values, the Pacific cod were exposed to the coldest water temperatures immediately prior to capture. The lack of a clear cycle of seasonal variation and the continued increase in values towards the otolith edge obscures the season of capture, and indicates that other behavioural, environmental, or methodological factors influenced the otolith δ¹⁸O values. It is suggested that Pacific cod would have been harvested throughout the year, and the presence of cod remains in Aleutian archaeological sites cannot be used as a reliable indicator of summer occupation. In addition, when the δ¹⁸O otolith values are integrated with radiocarbon dates and known climatic regimes, it is demonstrated that climatic conditions play an integral role in the pattern of occupation at Gertrude Cove. Initial site occupation coincides with the end of a neoglacial cooling period, and the most recent and continuous occupation coincides with the end of a localized warming period and the onset of the Little Ice Age (LIA).

Targeted sequencing for high-resolution evolutionary analyses following genome duplication in salmonid fish : Proof of concept for key components of the insulin-like growth factor axis

Acknowledgements This study was funded by a Natural Environment Research Council grant (NERC, project code: NBAF704). FML is funded by a NERC Doctoral Training Grant (Project Reference: NE/L50175X/1). RLS was an undergraduate student at the University of Aberdeen and benefitted from financial support from the School of Biological Sciences. DJM is indebted to Dr. Steven Weiss (University of Graz, Austria), Dr. Takashi Yada (National Research Institute of Fisheries Science, Japan), Dr. Robert Devlin (Fisheries and Oceans Canada, Canada), Prof. Samuel Martin (University of Aberdeen, UK), Mr. Neil Lincoln (Environment Agency, UK) and Prof. Colin Adams/Mr. Stuart Wilson (University of Glasgow, UK) for providing salmonid material or assisting with its sampling. We are grateful to staff at the Centre for Genomics Research (University of Liverpool, UK) (i.e. NERC Biomolecular Analysis Facility – Liverpool; NBAF-Liverpool) for performing sequence capture/Illumina sequencing and providing us with details on associated methods that were incorporated into the manuscript. Finally, we are grateful to the organizers of the Society of Experimental Biology Satellite meeting 'Genome-powered perspectives in integrative physiology and evolutionary biology' (held in Prague, July 2015) for inviting us to contribute to this special edition of Marine Genomics and hosting a really stimulating meeting.

Identifying the Structure and Fate of Wastewater Derived Organic Micropollutants by High-resolution Mass Spectrometry

Human activities represent a significant burden on the global water cycle, with large and increasing demands placed on limited water resources by manufacturing, energy production and domestic water use. In addition to changing the quantity of available water resources, human activities lead to changes in water quality by introducing a large and often poorly-characterized array of chemical pollutants, which may negatively impact biodiversity in aquatic ecosystems, leading to impairment of valuable ecosystem functions and services. Domestic and industrial wastewaters represent a significant source of pollution to the aquatic environment due to inadequate or incomplete removal of chemicals introduced into waters by human activities. Currently, incomplete chemical characterization of treated wastewaters limits comprehensive risk assessment of this ubiquitous impact to water. In particular, a significant fraction of the organic chemical composition of treated industrial and domestic wastewaters remains uncharacterized at the molecular level. Efforts aimed at reducing the impacts of water pollution on aquatic ecosystems critically require knowledge of the composition of wastewaters to develop interventions capable of protecting our precious natural water resources.

The goal of this dissertation was to develop a robust, extensible and high-throughput framework for the comprehensive characterization of organic micropollutants in wastewaters by high-resolution accurate-mass mass spectrometry. High-resolution mass spectrometry provides the most powerful analytical technique available for assessing the occurrence and fate of organic pollutants in the water cycle. However, significant limitations in data processing, analysis and interpretation have limited this technique in achieving comprehensive characterization of organic pollutants occurring in natural and built environments. My work aimed to address these challenges by development of automated workflows for the structural characterization of organic pollutants in wastewater and wastewater impacted environments by high-resolution mass spectrometry, and to apply these methods in combination with novel data handling routines to conduct detailed fate studies of wastewater-derived organic micropollutants in the aquatic environment.

In Chapter 2, chemoinformatic tools were implemented along with novel non-targeted mass spectrometric analytical methods to characterize, map, and explore an environmentally-relevant “chemical space” in municipal wastewater. This was accomplished by characterizing the molecular composition of known wastewater-derived organic pollutants and substances that are prioritized as potential wastewater contaminants, using these databases to evaluate the pollutant-likeness of structures postulated for unknown organic compounds that I detected in wastewater extracts using high-resolution mass spectrometry approaches. Results showed that application of multiple computational mass spectrometric tools to structural elucidation of unknown organic pollutants arising in wastewaters improved the efficiency and veracity of screening approaches based on high-resolution mass spectrometry. Furthermore, structural similarity searching was essential for prioritizing substances sharing structural features with known organic pollutants or industrial and consumer chemicals that could enter the environment through use or disposal.

I then applied this comprehensive methodological and computational non-targeted analysis workflow to micropollutant fate analysis in domestic wastewaters (Chapter 3), surface waters impacted by water reuse activities (Chapter 4) and effluents of wastewater treatment facilities receiving wastewater from oil and gas extraction activities (Chapter 5). In Chapter 3, I showed that application of chemometric tools aided in the prioritization of non-targeted compounds arising at various stages of conventional wastewater treatment by partitioning high dimensional data into rational chemical categories based on knowledge of organic chemical fate processes, resulting in the classification of organic micropollutants based on their occurrence and/or removal during treatment. Similarly, in Chapter 4, high-resolution sampling and broad-spectrum targeted and non-targeted chemical analysis were applied to assess the occurrence and fate of organic micropollutants in a water reuse application, wherein reclaimed wastewater was applied for irrigation of turf grass. Results showed that organic micropollutant composition of surface waters receiving runoff from wastewater irrigated areas appeared to be minimally impacted by wastewater-derived organic micropollutants. Finally, Chapter 5 presents results of the comprehensive organic chemical composition of oil and gas wastewaters treated for surface water discharge. Concurrent analysis of effluent samples by complementary, broad-spectrum analytical techniques, revealed that low-levels of hydrophobic organic contaminants, but elevated concentrations of polymeric surfactants, which may effect the fate and analysis of contaminants of concern in oil and gas wastewaters.

Taken together, my work represents significant progress in the characterization of polar organic chemical pollutants associated with wastewater-impacted environments by high-resolution mass spectrometry. Application of these comprehensive methods to examine micropollutant fate processes in wastewater treatment systems, water reuse environments, and water applications in oil/gas exploration yielded new insights into the factors that influence transport, transformation, and persistence of organic micropollutants in these systems across an unprecedented breadth of chemical space.

Molecular composition of organic aerosols in central Amazonia: an ultra-high-resolution mass spectrometry study

The Amazon Basin plays key role in atmospheric chemistry, biodiversity and climate change. In this study we applied nanoelectrospray (nanoESI) ultra-high-resolution mass spectrometry (UHRMS) for the analysis of the organic fraction of PM2.5 aerosol samples collected during dry and wet seasons at a site in central Amazonia receiving background air masses, biomass burning and urban pollution. Comprehensive mass spectral data evaluation methods (e.g. Kendrick mass defect, Van Krevelen diagrams, carbon oxidation state and aromaticity equivalent) were used to identify compound classes and mass distributions of the detected species. Nitrogen- and/or sulfur-containing organic species contributed up to 60 % of the total identified number of formulae. A large number of molecular formulae in organic aerosol (OA) were attributed to later-generation nitrogen- and sulfur-containing oxidation products, suggesting that OA composition is affected by biomass burning and other, potentially anthropogenic, sources. Isoprene-derived organosulfate (IEPOX-OS) was found to be the most dominant ion in most of the analysed samples and strongly followed the concentration trends of the gas-phase anthropogenic tracers confirming its mixed anthropogenic–biogenic origin. The presence of oxidised aromatic and nitro-aromatic compounds in the samples suggested a strong influence from biomass burning especially during the dry period. Aerosol samples from the dry period and under enhanced biomass burning conditions contained a large number of molecules with high carbon oxidation state and an increased number of aromatic compounds compared to that from the wet period. The results of this work demonstrate that the studied site is influenced not only by biogenic emissions from the forest but also by biomass burning and potentially other anthropogenic emissions from the neighbouring urban environments.

Time-resolved dosimetric verification of respiratory-gated radiotherapy exposures using a high-resolution 2D ionisation chamber array

The aim of this work was to track and verify the delivery of respiratory-gated irradiations, performed with three versions of TrueBeam linac, using a novel phantom arrangement that combined the OCTAVIUS® SRS 1000 array with a moving platform. The platform was programmed to generate sinusoidal motion of the array. This motion was tracked using the real-time position management (RPM) system and four amplitude gating options were employed to interrupt MV beam delivery when the platform was not located within set limits. Time-resolved spatial information extracted from analysis of x-ray fluences measured by the array was compared to the programmed motion of the platform and to the trace recorded by the RPM system during the delivery of the x-ray field. Temporal data recorded by the phantom and the RPM system were validated against trajectory log files, recorded by the linac during the irradiation, as well as oscilloscope waveforms recorded from the linac target signal. Gamma analysis was employed to compare time-integrated 2D x-ray dose fluences with theoretical fluences derived from the probability density function for each of the gating settings applied, where gamma criteria of 2%/2 mm, 1%/1 mm and 0.5%/0.5 mm were used to evaluate the limitations of the RPM system. Excellent agreement was observed in the analysis of spatial information extracted from the SRS 1000 array measurements. Comparisons of the average platform position with the expected position indicated absolute deviations of  <0.5 mm for all four gating settings. Differences were observed when comparing time-resolved beam-on data stored in the RPM files and trajectory logs to the true target signal waveforms. Trajectory log files underestimated the cycle time between consecutive beam-on windows by 10.0  ±  0.8 ms. All measured fluences achieved 100% pass-rates using gamma criteria of 2%/2 mm and 50% of the fluences achieved pass-rates  >90% when criteria of 0.5%/0.5 mm were used. Results using this novel phantom arrangement indicate that the RPM system is capable of accurately gating x-ray exposure during the delivery of a fixed-field treatment beam.

High-resolution electrophoretic procedures for the identification of five Eimeria species from chickens, and detection of population variation

To overcome limitations of conventional approaches for the identification of Eimeria species of chickens, we have established high resolution electrophoretic procedures using genetic markers in ribosomal DNA. The first and second internal transcribed spacer (ITS-1 and ITS-2) regions of ribosomal DNA were amplified by polymerase chain reaction (PCR) from genomic DNA samples representing five species of Eimeria (E. acervulina, E. brunetti, E. maxima, E. necatrix and E. tenella), denatured and then subjected to denaturing polyacrylamide gel electrophoresis (D-PAGE) or single-strand conformation polymorphism (SSCP) analysis. Differences in D-PAGE profiles for both the ITS-1 and ITS-2 fragments (combined with an apparent lack of variation within individual species) enabled the unequivocal identification of the five species, and SSCP allowed the detection of population variation between some isolates representing E. acervulina, which remained undetected by D-PAGE. The establishment of these approaches has important implications for controlling the purity of laboratory lines of Eimeria, for diagnosis and for studying the epidemiology of coccidiosis.

Climate variability of the last 1000 years in the NW Pacific: high resolution, multi-biomarker records from Lake Toyoni

The East Asian Monsoon (EAM) is an active component of the global climate system and has a profound social and economic impact in East Asia and its surrounding countries. Its impact on regional hydrological processes may influence society through industrial water supplies, food productivity and energy use. In order to predict future rates of climate change, reliable and accurate reconstructions of regional temperature and rainfall are required from all over the world to test climate models and better predict future climate variability. Hokkaido is a region which has limited palaeo-climate data and is sensitive to climate change. Instrumental data show that the climate in Hokkaido is influenced by the East Asian Monsoon (EAM), however, instrumental data is limited to the past ~150 years. Therefore down-core climate reconstructions, prior to instrumental records, are required to provide a better understanding of the long-term behaviour of the climate drivers (e.g. the EAM, Westerlies, and teleconnections) in this region. The present study develops multi-proxy reconstructions to determine past climatic and hydrologic variability in Japan over the past 1000 years and aid in understanding the effects of the EAM and the Westerlies independently and interactively. A 250-cm long sediment core from Lake Toyoni, Hokkaido was retrieved to investigate terrestrial and aquatic input, lake temperature and hydrological changes over the past 1000-years within Lake Toyoni and its catchment using X-Ray Fluorescence (XRF) data, alkenone palaeothermometry, the molecular and hydrogen isotopic composition of higher plant waxes (δD(HPW)). Here, we conducted the first survey for alkenone biomarkers in eight lakes in the Hokkaido, Japan. We detected the occurrence of alkenones within the sediments of Lake Toyoni. We present the first lacustrine alkenone record from Japan, including genetic analysis of the alkenone producer. C37 alkenone concentrations in surface sediments are 18µg C37 g−1 of dry sediment and the dominant alkenone is C37:4. 18S rDNA analysis revealed the presence of a single alkenone producer in Lake Toyoni and thus a single calibration is used for reconstructing lake temperature based on alkenone unsaturation patterns. Temperature reconstructions over the past 1000 years suggest that lake water temperatures varies between 8 and 19°C which is in line with water temperature changes observed in the modern Lake Toyoni. The alkenone-based temperature reconstruction provides evidence for the variability of the EAM over the past 1000 years. The δD(HPW) suggest that the large fluctuations (∼40‰) represent changes in temperature and source precipitation in this region, which is ultimately controlled by the EAM system and therefore a proxy for the EAM system. In order to complement the biomarker reconstructions, the XRF data strengthen the lake temperature and hydrological reconstructions by providing information on past productivity, which is controlled by the East Asian Summer monsoon (EASM) and wind input into Lake Toyoni, which is controlled by the East Asian Winter Monsoon (EAWM) and the Westerlies. By combining the data generated from XRF, alkenone palaeothermometry and the δD(HPW) reconstructions, we provide valuable information on the EAM and the Westerlies, including; the timing of intensification and weakening, the teleconnections influencing them and the relationship between them. During the Medieval Warm Period (MWP), we find that the EASM dominated and the EAWM was suppressed, whereas, during the Little Ice Age (LIA), the influence of the EAWM dominated with time periods of increased EASM and Westerlies intensification. The El Niño Southern Oscillation (ENSO) significantly influenced the EAM; a strong EASM occurred during El Niño conditions and a strong EAWM occurred during La Niña. The North Atlantic Oscillation, on the other hand, was a key driver of the Westerlies intensification; strengthening of the Westerlies during a positive NAO phase and weakening of the Westerlies during a negative NAO phase. A key finding from this study is that our data support an anti-phase relationship between the EASM and the EAWM (e.g. the intensification of the EASM and weakening of the EAWM and vice versa) and that the EAWM and the Westerlies vary independently from each other, rather than coincide as previously suggested in other studies.

Turbulent Pipe Flow - High Resolution Measurements in CICLoPE

The thesis deals with the experimental investigation of turbulent pipe flow at high Reynolds number. Wall-bounded turbulence is an extremely relevant topic for engineering and natural science applications and yet many aspects of the physics are not clear due to the difficulty in performing high Re experiments. To overcome these difficulties the CICLoPE Laboratory was developed, the main element of which is the Long Pipe wind tunnel. The facility is unique in its kind, as thanks to its large scale it delivers a flow quality and resolution that can not be achieved elsewhere at these Reynolds number. Reported here are the results from the first experimental campaign performed in the facility. A first part of the results presented concerns the characterization of this new facility. Flow quality and stability are assessed, particular attention is given to the characterization of pressure drop. The scaling of velocity fluctuations is analysed. The magnitude of the inner peak of the streamwise normal stress shows an increasing trend up to the highest Reynolds number examined, while no outer peak was clearly distinguishable from present measurements. Scaling of coherent motions is investigated via spectral analysis. An inner and outer spectral peaks are identified, with the former scaling in inner units while the latter neither following inner nor outer scaling, and increasing in magnitude with Re. Analysis of the spectra at y+ ≈ 15 shows how the increase of Reynolds normal stress is related to the influence of large scales in the inner wall region. Quadrant analysis was carried out on streamwise and wall-normal velocity fluctuations. The results show the important role in contribution to Reynolds shear stress of highly intermittent and strong events like ejections, that assume an even more intermittent and dominant role with the increase of Reynolds number.

Unveiling the unknown of cool stars with high-resolution spectroscopy

Cool giant and supergiant stars are among the brightest populations in any stellar system and they are easily observable out to large distances, especially at infrared wavelengths. These stars also dominate the integrated light of star clusters in a wide range of ages, making them powerful tracers of stellar populations in more distant galaxies. High-resolution near-IR spectroscopy is a key tool for quantitatively investigating their kinematic, evolutionary and chemical properties. However, the systematic exploration and calibration of the NIR spectral diagnostics to study these cool stellar populations based on high-resolution spectroscopy is still in its pioneering stage. Any effort to make progress in the field is innovative and of impact on stellar archaeology and stellar evolution. This PhD project takes the challenge of exploring that new parameter space and characterizing the physical properties, the chemical content and the kinematics of cool giants and supergiants in selected disc fields and clusters of our Galaxy, with the ultimate goal of tracing their past and recent star formation and chemical enrichment history. By using optical HARPS-N and near-infrared GIANO-B high-resolution stellar spectra in the context of the large program SPA-Stellar Population Astrophysics: the detailed, age-resolved chemistry of the Milky Way disk” (PI L. Origlia), an extensive study of Arcturus, a standard calibrator for red giant stars, has been performed. New diagnostics of stellar parameters as well as optimal linelists for chemical analysis have been provided. Then, such diagnostics have been used to determine evolutionary properties, detailed chemical abundances of almost 30 different elements and mixing processes of a homogeneous sample of red supergiant stars in the Perseus complex.

High-resolution stratigraphy and seismic site characterization of late Quaternary paleovalley systems (Adriatic coastal plain, Italy)

In highly urbanized coastal lowlands, effective site characterization is crucial for assessing seismic risk. It requires a comprehensive stratigraphic analysis of the shallow subsurface, coupled with the precise assessment of the geophysical properties of buried deposits. In this context, late Quaternary paleovalley systems, shallowly buried fluvial incisions formed during the Late Pleistocene sea-level fall and filled during the Holocene sea-level rise, are crucial for understanding seismic amplification due to their soft sediment infill and sharp lithologic contrasts. In this research, we conducted high-resolution stratigraphic analyses of two regions, the Pescara and Manfredonia areas along the Adriatic coastline of Italy, to delineate the geometries and facies architecture of two paleovalley systems. Furthermore, we carried out geophysical investigations to characterize the study areas and perform seismic response analyses. We tested the microtremor-based horizontal-to-vertical spectral ratio as a mapping tool to reconstruct the buried paleovalley geometries. We evaluated the relationship between geological and geophysical data and identified the stratigraphic surfaces responsible for the observed resonances. To perform seismic response analysis of the Pescara paleovalley system, we integrated the stratigraphic framework with microtremor and shear wave velocity measurements. The seismic response analysis highlights strong seismic amplifications in frequency ranges that can interact with a wide variety of building types. Additionally, we explored the applicability of artificial intelligence in performing facies analysis from borehole images. We used a robust dataset of high-resolution digital images from continuous sediment cores of Holocene age to outline a novel, deep-learning-based approach for performing automatic semantic segmentation directly on core images, leveraging the power of convolutional neural networks. We propose an automated model to rapidly characterize sediment cores, reproducing the sedimentologist's interpretation, and providing guidance for stratigraphic correlation and subsurface reconstructions.

High-resolution Transcript Profiling Of The Atypical Biotrophic Interaction Between Theobroma Cacao And The Fungal Pathogen Moniliophthora Perniciosa.

Witches' broom disease (WBD), caused by the hemibiotrophic fungus Moniliophthora perniciosa, is one of the most devastating diseases of Theobroma cacao, the chocolate tree. In contrast to other hemibiotrophic interactions, the WBD biotrophic stage lasts for months and is responsible for the most distinctive symptoms of the disease, which comprise drastic morphological changes in the infected shoots. Here, we used the dual RNA-seq approach to simultaneously assess the transcriptomes of cacao and M. perniciosa during their peculiar biotrophic interaction. Infection with M. perniciosa triggers massive metabolic reprogramming in the diseased tissues. Although apparently vigorous, the infected shoots are energetically expensive structures characterized by the induction of ineffective defense responses and by a clear carbon deprivation signature. Remarkably, the infection culminates in the establishment of a senescence process in the host, which signals the end of the WBD biotrophic stage. We analyzed the pathogen's transcriptome in unprecedented detail and thereby characterized the fungal nutritional and infection strategies during WBD and identified putative virulence effectors. Interestingly, M. perniciosa biotrophic mycelia develop as long-term parasites that orchestrate changes in plant metabolism to increase the availability of soluble nutrients before plant death. Collectively, our results provide unique insight into an intriguing tropical disease and advance our understanding of the development of (hemi)biotrophic plant-pathogen interactions.

Application Of Laser Microdissection Icp-ms For High Resolution Elemental Mapping In Mouse Brain Tissue: A Comparative Study With Laser Ablation Icp-ms.

Mapping of elements in biological tissue by laser induced mass spectrometry is a fast growing analytical methodology in life sciences. This method provides a multitude of useful information of metal, nonmetal, metalloid and isotopic distribution at major, minor and trace concentration ranges, usually with a lateral resolution of 12-160 µm. Selected applications in medical research require an improved lateral resolution of laser induced mass spectrometric technique at the low micrometre scale and below. The present work demonstrates the applicability of a recently developed analytical methodology - laser microdissection associated to inductively coupled plasma mass spectrometry (LMD ICP-MS) - to obtain elemental images of different solid biological samples at high lateral resolution. LMD ICP-MS images of mouse brain tissue samples stained with uranium and native are shown, and a direct comparison of LMD and laser ablation (LA) ICP-MS imaging methodologies, in terms of elemental quantification, is performed.