Out in the Open: A Geoarchaeological Approach to Open-Air Surface Archaeology in the Semi-Arid Interior of South Africa’s Western Cape

Southern African research into the behavioural evolution of Late Pleistocene human adaptability, flexibility, and innovation is typically pursued through the lens of rock shelter deposits. However, rock shelters only cover a very small, geographically specific area of the subcontinent, distorting our understanding of change in human-environment interaction and demography. While still under-represented and under-explored in regional syntheses, more studies are looking to open-air archaeology to fill this geographic void in Late Pleistocene research. These studies either pursue a landscape approach that prioritises spatial coverage, or site-bound excavation to maximise temporal control. However, few investigate the depositional and erosional phenomena involved in the formation of surface archaeology and its surrounding landscape. To rectify this disparity, this thesis explores the complex spatio-temporal relationship between surface archaeology and the formation history of Uitspankraal (UPK) 7 by combining multiple interdisciplinary methods from the Earth and archaeological sciences: randomised surface survey and sampling, geomorphometry, geophysical survey, granulometry, XRD analysis, OSL dating, artefact mapping, and assemblage composition and artefact condition analysis. UPK7 is located in the semi-arid Doring River valley and yields surface archaeology that implies occupation from the Still Bay to the Historic period. Results show that it is an eroding series of source-bordering dunes draped across a palaeoterrace and a hillslope of bedrock and colluvium. UPK7 formed through rapid but pulsed sediment accumulation over at least the last 80 ka, with periods of surface deflation and exposure that facilitated artefact redistribution. Despite the abundance of Late Pleistocene archaeology at UPK7, erosion currently outpaces deposition and deposit stabilisation. Erosion has accelerated in at least the last 5,000 years and especially within the last 300 years, suggesting feedback between Holocene aridification, an increase in oscillations between wet-dry conditions, and an increase in human-ungulate activity in the study area. Together these conditions have differentially erased younger deposits, exposing the consolidated Late Pleistocene sediment and the more ancient material it preserves. The visibility, spatio-temporal distribution, and preservation of UPK7’s surface artefacts reflect the locality’s topography, the timing of their discard and the duration and process of sediment accumulation and erosion. The spatial patterning and diversity of time-diagnostic and non-diagnostic artefacts is shown to correspond with the depositional age of their underlying substrate in areas where topographic conditions minimize or reduce the impact of surface runoff, but where sediment deflation persists. When artefacts are assessed at the scale of the archaeological epoch the spatial distribution of Middle Stone Age artefacts shows a significant association with the oldest deposit, Lower Red. The spatial distribution of Later Stone Age artefacts is significantly associated with Upper Yellow sediment, as opposed to the older Lower Red substrate and the younger Indurated Sand. The findings presented in this thesis caution against forming behavioural interpretations from spatial patterns in surface material without examining their post-depositional history and without forming an understanding of the coevolution of archaeological and landscape formation. This study underscores the need for incorporating a geoarchaeological approach into Late Pleistocene open-air research to improve southern Africa’s landscape-scale insight into greater Africa’s human behavioural evolution

Systematics of the Relhaniinae (Asteraceae-Gnaphalieae) in southern Africa : geography and evolution in an endemic Cape plant lineage

Includes bibliographical references (p. 161-194).The Greater Cape Floristic Region (GCFR) houses a flora unique for its diversity and high endemicity. A large amount of the diversity is housed in just a few lineages, presumed to have radiated in the region. For many of these lineages there is no robust phylogenetic hypothesis of relationships, and few Cape plants have been examined for the spatial distribution of their population genetic variation. Such studies are especially relevant for the Cape where high rates of species diversification and the ongoing maintenance of species proliferation is hypothesised. Subtribe Relhaniinae of the daisy tribe Gnaphalieae is one such little-studied lineage. The taxonomic circumscription of this subtribe, the biogeography of its early diversification and its relationships to other members of the Gnaphalieae are elucidated by means of a dated phylogenetic hypothesis. Molecular DNA sequence data from both chloroplast and nuclear genomes are used to reconstruct evolutionary history using parsimony and Bayesian tools for phylogeny estimation. The subtribe is found to be nonmonophyletic, and three independent Cape-centred clades are identified. These constitute the earliest-diverging lineages in the Gnaphalieae. Ancestral areas are reconstructed onto the tree using maximum likelihood and indicate a Cape origin and diversification for the Gnaphalieae, with subsequent, multiple dispersals out of southern Africa. A relaxed Bayesian clock is used to estimate dates for important events, calibrated using a combination of secondary age estimates, fossil pollen and geological events. The single inferred dispersal to Australasia, which may be responsible for the large diversity of the tribe there, is compared with other dated trans-Indian Ocean disjunctions, and results in a hypothesis of direct Miocene longdistance trans-oceanic dispersal facilitated by the West Wind Drift. Important directions for future biogeographic and systematic studies on the Gnaphalieae are identified. One of the Cape clades identified in the above study, the Stoebe clade, is investigated using the same molecular markers and an additional chloroplast DNA region, in order to reconstruct relationships amongst the species and test generic circumscription. One genus, Amphiglossa, is found to be non-monophyletic and a recommendation is made to resurrect the genus Pterothrix in order to maintain monophyly. Relationships amongst subclades in the Stoebe clade can not be resolved, possibly due to a paucity of chloroplast DNA polymorphisms. Ancestral-state reconstruction using maximum likelihood indicates a high degree of homoplasy in all the macro-morphological characters used by previous workers to delimit the genera that fall within this polytomy (Disparago, Elytropappus and Stoebe). The evolution of these leaf, synflorescence and floral morphological characters is examined in order to shed light on the degree of homoplasy exhibited by each, and the patterns of character-state change across the tree. The analysis provides insight into patterns of morphological evolution, laying the groundwork for re-evaluation of characters previously thought to be homologous. The need for a rigorous, systematic anatomical study including all members of the clade is highlighted and some previously little-explored characters are identified that might provide greater future taxonomic insight. Due to a poor palaeoclimatic record, the impact of Pleistocene climatic fluctuations on the palaeodistributions of plant species in the Greater Cape Floristic Region is largely unknown. One of the members of the Stoebe clade, the common and widespread shrub Elytropappus rhinocerotis, is examined using population genetic tools in order to explore the spatial distribution of genetic variation and to infer the degree and pattern of responses (range shifts, population extinction) to recent (Quaternary) climatic changes. A molecular fingerprinting tool, inter-simple sequence repeat (ISSR) PCR is used to examine populations from across the range of E. rhinocerotis in order to compare patterns of population genetic diversity with the long-noted and strong spatial patterns of plant species diversity in the CFR. A large amount of genetic variation is detected, which is apportioned largely amongst individuals within populations rather than amongst populations or regions. This is to be expected for an outcrossing and well-dispersed plant species. However, there is significant spatial structure and a very uneven distribution of diversity across the geographic range of E. rhinocerotis. Although areas that have high species-diversity also generally exhibited high genetic diversity, the eastern margin of the range and the Kamiesberg highlands both have a far higher relative genetic diversity in E. rhinocerotis than would be predicted by their relative species diversity. This indicates that the processes producing greater leves of species diversity in some parts of the CFR are different than the processes responsible for high genetic variation within E. rhinocerotis populations. Geographic distance is a poor predictor of genetic distance between localities, especially towards the east of the range. This may be due to range alteration over the time-scale reflected by ISSR polymorphism. Inter-SSR variation declined from south to north in the western arm of the range, consistent with the prediction of Holocene aridification starting first and being most extreme in the north. Areas shown by the marker to harbour populations with high levels of variability include most parts of the eastern arm of the range, and the Kamiesberg highlands. The present study constitutes a demonstration of the range of evolutionary questions that can be addressed using a range of molecular phylogenetic and population genetic techniques, and the elucidation of both deep and shallow evolutionary history of a single Cape floral lineage

Pushing the limits of two-dimensional mass spectrometry

Two-dimensional Fourier transform ion cyclotron resonance mass spectrometry (2D FT-ICR MS) is a data independent tandem mass spectrometry technique that allows direct correlation between precursor and fragment ions without the necessity of any sort of isolation prior to fragmentation. Two-dimensional mass spectrometry (2D MS) experiments were conceived in the 1980s from inspiration of two-dimensional nuclear magnetic resonance (2D-NMR), but the development of the technique was stopped because of the insufficient computational capabilities of that time. In 2010, with the progress in computer technology and the improvements in FT-ICR MS instrumentation and methods, the interest in developing 2D MS was renewed. Since then, 2D FT-ICR MS showed to be a valuable platform for the analysis of small molecules, macromolecules such as peptides and glycopeptides, and complex mixtures deriving from tryptic digestion of proteins of increasing sizes. All the proteomics studies performed with 2D MS used the bottom-up (BUP) approach. The main goal of this Ph.D. research work was to develop two-dimensional mass spectrometry for the challenges of contemporary mass spectrometry, therefore pushing the limits of the technique towards the most interesting, cutting-edge research grounds, by optimising the technique, developing 2D MS analysis methods, and finally constructing improved instrumentation act to better perform 2D MS experiments. On this purpose, the rising importance of top-down proteomics (TDP) led to the development of 2D FT-ICR MS for the analysis of intact proteins. To realise this first project, 2D MS was optimised for TDP analysis using calmodulin (CaM) as a model, and infrared multiphotondissociation (IRMPD) as fragmentation technique. The study compared the two-dimensional BUP and TDP analysis of CaM with standard one-dimensional tandem mass spectrometry, and showed that the use of 2D MS allowed to obtain comparable cleavage coverages, with a consistent saving in sample and time. The study affirmed the suitability of 2D MS for top-down proteomics, still leaving space for more developments allowing its optimisation. The total cleavage coverage of CaM was later implemented with the use of BUP 2D electron-capture dissociation (ECD) MS. In order to improve the cleavage coverages obtained in top-down two-dimensional mass spectrometry, another technique was developed, called MS/2D MS. The technique makes use of an extra fragmentation step before two-dimensional mass spectrometry, achieved through external isolation of a charge state of interest, and collisional fragmentation in a hexapole-based collision cell before the ions are sent to the ICR-cell. The output of MS/2D MS is a single 2D mass spectrum with information equivalent to MS3 about the entire mixture. The technique has been successfully developed using ubiquitin (Ubi) and calmodulin as models, and showed new uses of the features exclusive of 2D MS, such as the use of precursor ion scans to assign the protein terminus of many precursors ions at once, and the analysis of internal fragments formed by the first dissociation. Furthermore, as long as extra fragmentation steps can be performed, a deeper investigation of the analytes can be achieved, in an experiment collectively called MSn /2D MS. After the development of 2D MS for TDP, another method was developed, implementing the use of two-dimensional mass spectrometry for another type of complex mixtures: polymers. An extensive 2D MS study was performed on a simple mixture of homopolymers such as PMMA, and on two complex mixtures commonly used as drug excipients, polysorbate 80 and TPGS. The study demonstrated again that single 2D MS experiments give results equivalent to multiple MS/MS experiments targeted at the different precursors. Furthermore, it allowed the high resolution tandem mass spectrometry of polysorbate 80, representing the first kind of study of that type because of the challenges of isolating single analytes from the polymer mixture without the use of a previous separation, such as chromatography. The use of neutral-loss lines was particularly useful in this study, allowing to discriminate between oleated, linoleated, and non-esterified species. Finally, 2D mass spectra of polymers showed characteristic diagonal lines, specific for each polymeric mixture, representing some sort of fingerprint of the mixture. The last part of the thesis is dedicated to the optimisation of 2D FT-ICR MS for ECD MS, the application of the technique to another class of compounds, glycans and glycan-derivatives, and finally to the development of a more capacious ICR-cell. The research performed in this thesis showed that 2D FT-ICR MS is a viable data independent mass spectrometry technique for top-down proteomics and polymer analysis; that the technique is fast, and it possesses unique features that improve ion assignments, such as precursor ion scans and neutral-loss lines. Finally, it opens for two-dimensional mass spectrometry the path to glycomics and to an improved 2D MS analysis of even more complex mixtures, thanks to the development of a novel ICR-cell

Patterns of population structure and adaptive genetic variation in alpine populations of Picea abies (L.) Karst.

Forest trees dominate many alpine landscapes and are currently exposed to changing climate. Picea abies (L.) Karst (Norway spruce) is one of the most important conifer species of the Italian Alps due to its ecological and economical relevance. Natural populations of this species are found across steep environmental gradients with large differences in temperature and moisture availability. Steep environmental gradients represent interesting models to study the interaction between natural selection and gene flow, especially when aiming to understand adaptation processes under global change. The present work aims to understand adaptive responses to changing climate by determining and quantifying patterns of genetic diversity in natural population of P.abies. a wide array of potential candidate genes was tested, by means of Single Nucleotide Polymorphisms (SNPs), for correlation with climatic and environmental parameters at different spatial scales: i) a geographical scale corresponding to the natural distribution of P.abies across the Italian Alps and ii) at a regional scale on the Eastern Italian Alps. Weak population structure was revealed at the geographical scale with only one population clearly divergent from the unique major genetic cluster identified. At the regional scale, hierarchical analyses of molecular variance revealed that most of the genetic variability was found within populations (ca. 99%), and small but significant variation was also found due landscape features (ca. 0.38%). In order to detect potentially adaptive markers, classical FST outlier approaches were first applied and five outlier loci were revealed at broad scale, while contrasting results were obtained at the regional scale according to the model used. Subsequently, environmental association analysis were performed: at the geographical scale temperature and precipitation were found to influence allelic variation at seven polymorphic loci, while at the regional scale, the Alpine topography resulted a potential adaptive determinants at 19 polymorphic loci, thus considered of ecological relevance. The results obtained in this study may provide relevant information for forestry management and genetic conservation, to understand and quantify the effect of climate change on conifer species as well as their adaptive potential

Bayesian networks for spatio-temporal integrated catchment assessment

Includes abstract.Includes bibliographical references (leaves 181-203).In this thesis, a methodology for integrated catchment water resources assessment using Bayesian Networks was developed. A custom made software application that combines Bayesian Networks with GIS was used to facilitate data pre-processing and spatial modelling. Dynamic Bayesian Networks were implemented in the software for time-series modelling

Patterns of population structure and adaptive genetic variation in alpine populations of Picea abies (L.) Karst.

Forest trees dominate many alpine landscapes and are currently exposed to changing climate. Picea abies (L.) Karst (Norway spruce) is one of the most important conifer species of the Italian Alps due to its ecological and economical relevance. Natural populations of this species are found across steep environmental gradients with large differences in temperature and moisture availability. Steep environmental gradients represent interesting models to study the interaction between natural selection and gene flow, especially when aiming to understand adaptation processes under global change. The present work aims to understand adaptive responses to changing climate by determining and quantifying patterns of genetic diversity in natural population of P.abies. a wide array of potential candidate genes was tested, by means of Single Nucleotide Polymorphisms (SNPs), for correlation with climatic and environmental parameters at different spatial scales: i) a geographical scale corresponding to the natural distribution of P.abies across the Italian Alps and ii) at a regional scale on the Eastern Italian Alps. Weak population structure was revealed at the geographical scale with only one population clearly divergent from the unique major genetic cluster identified. At the regional scale, hierarchical analyses of molecular variance revealed that most of the genetic variability was found within populations (ca. 99%), and small but significant variation was also found due landscape features (ca. 0.38%). In order to detect potentially adaptive markers, classical FST outlier approaches were first applied and five outlier loci were revealed at broad scale, while contrasting results were obtained at the regional scale according to the model used. Subsequently, environmental association analysis were performed: at the geographical scale temperature and precipitation were found to influence allelic variation at seven polymorphic loci, while at the regional scale, the Alpine topography resulted a potential adaptive determinants at 19 polymorphic loci, thus considered of ecological relevance. The results obtained in this study may provide relevant information for forestry management and genetic conservation, to understand and quantify the effect of climate change on conifer species as well as their adaptive potential