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This study investigates the spatial and temporal distributions of 14 key arboreal taxa and their driving forces during the last 22,000 calendar years before ad 1950 (kyr BP) using a taxonomically harmonized and temporally standardized fossil pollen dataset with a 500-year resolution from the eastern part of continental Asia. Logistic regression was used to estimate pollen abundance thresholds for vegetation occurrence (presence or dominance), based on modern pollen data and present ranges of 14 taxa in China. Our investigation reveals marked changes in spatial and temporal distributions of the major arboreal taxa. The thermophilous (Castanea, Castanopsis, Cyclobalanopsis, Fagus, Pterocarya) and eurythermal (Juglans, Quercus, Tilia, Ulmus) broadleaved tree taxa were restricted to the current tropical or subtropical areas of China during the Last Glacial Maximum (LGM) and spread northward since c. 14.5 kyr BP. Betula and conifer taxa (Abies, Picea, Pinus), in contrast, retained a wider distribution during the LGM and showed no distinct expansion direction during the Late Glacial. Since the late mid-Holocene, the abundance but not the spatial extent of most trees decreased. The changes in spatial and temporal distributions for the 14 taxa are a reflection of climate changes, in particular monsoonal moisture, and, in the late Holocene, human impact. The post-LGM expansion patterns in eastern continental China seem to be different from those reported for Europe and North America, for example, the westward spread for eurythermal broadleaved taxa.
We report on the development of an on-chip RPA (recombinase polymerase amplification) with simultaneous multiplex isothermal amplification and detection on a solid surface. The isothermal RPA was applied to amplify specific target sequences from the pathogens Neisseria gonorrhoeae, Salmonella enterica and methicillin-resistant Staphylococcus aureus (MRSA) using genomic DNA. Additionally, a positive plasmid control was established as an internal control. The four targets were amplified simultaneously in a quadruplex reaction. The amplicon is labeled during on-chip RPA by reverse oligonucleotide primers coupled to a fluorophore. Both amplification and spatially resolved signal generation take place on immobilized forward primers bount to expoxy-silanized glass surfaces in a pump-driven hybridization chamber. The combination of microarray technology and sensitive isothermal nucleic acid amplification at 38 °C allows for a multiparameter analysis on a rather small area. The on-chip RPA was characterized in terms of reaction time, sensitivity and inhibitory conditions. A successful enzymatic reaction is completed in <20 min and results in detection limits of 10 colony-forming units for methicillin-resistant Staphylococcus aureus and Salmonella enterica and 100 colony-forming units for Neisseria gonorrhoeae. The results show this method to be useful with respect to point-of-care testing and to enable simplified and miniaturized nucleic acid-based diagnostics.
Background
Nucleic acid amplification is the most sensitive and specific method to detect Plasmodium falciparum. However the polymerase chain reaction remains laboratory-based and has to be conducted by trained personnel. Furthermore, the power dependency for the thermocycling process and the costly equipment necessary for the read-out are difficult to cover in resource-limited settings. This study aims to develop and evaluate a combination of isothermal nucleic acid amplification and simple lateral flow dipstick detection of the malaria parasite for point-of-care testing.
Methods
A specific fragment of the 18S rRNA gene of P. falciparum was amplified in 10 min at a constant 38°C using the isothermal recombinase polymerase amplification (RPA) method. With a unique probe system added to the reaction solution, the amplification product can be visualized on a simple lateral flow strip without further labelling. The combination of these methods was tested for sensitivity and specificity with various Plasmodium and other protozoa/bacterial strains, as well as with human DNA. Additional investigations were conducted to analyse the temperature optimum, reaction speed and robustness of this assay.
Results
The lateral flow RPA (LF-RPA) assay exhibited a high sensitivity and specificity. Experiments confirmed a detection limit as low as 100 fg of genomic P. falciparum DNA, corresponding to a sensitivity of approximately four parasites per reaction. All investigated P. falciparum strains (n = 77) were positively tested while all of the total 11 non-Plasmodium samples, showed a negative test result. The enzymatic reaction can be conducted under a broad range of conditions from 30-45°C with high inhibitory concentration of known PCR inhibitors. A time to result of 15 min from start of the reaction to read-out was determined.
Conclusions
Combining the isothermal RPA and the lateral flow detection is an approach to improve molecular diagnostic for P. falciparum in resource-limited settings. The system requires none or only little instrumentation for the nucleic acid amplification reaction and the read-out is possible with the naked eye. Showing the same sensitivity and specificity as comparable diagnostic methods but simultaneously increasing reaction speed and dramatically reducing assay requirements, the method has potential to become a true point-of-care test for the malaria parasite.
The Arabidopsis Kinome
(2014)
Background
Protein kinases constitute a particularly large protein family in Arabidopsis with important functions in cellular signal transduction networks. At the same time Arabidopsis is a model plant with high frequencies of gene duplications. Here, we have conducted a systematic analysis of the Arabidopsis kinase complement, the kinome, with particular focus on gene duplication events. We matched Arabidopsis proteins to a Hidden-Markov Model of eukaryotic kinases and computed a phylogeny of 942 Arabidopsis protein kinase domains and mapped their origin by gene duplication.
Results
The phylogeny showed two major clades of receptor kinases and soluble kinases, each of which was divided into functional subclades. Based on this phylogeny, association of yet uncharacterized kinases to families was possible which extended functional annotation of unknowns. Classification of gene duplications within these protein kinases revealed that representatives of cytosolic subfamilies showed a tendency to maintain segmentally duplicated genes, while some subfamilies of the receptor kinases were enriched for tandem duplicates. Although functional diversification is observed throughout most subfamilies, some instances of functional conservation among genes transposed from the same ancestor were observed. In general, a significant enrichment of essential genes was found among genes encoding for protein kinases.
Conclusions
The inferred phylogeny allowed classification and annotation of yet uncharacterized kinases. The prediction and analysis of syntenic blocks and duplication events within gene families of interest can be used to link functional biology to insights from an evolutionary viewpoint. The approach undertaken here can be applied to any gene family in any organism with an annotated genome.
Pulsar wind nebulae (PWNe) are the most abundant TeV gamma-ray emitters in the Milky Way. The radiative emission of these objects is powered by fast-rotating pulsars, which donate parts of their rotational energy into winds of relativistic particles. This thesis presents an in-depth study of the detected population of PWNe at high energies. To outline general trends regarding their evolutionary behaviour, a time-dependent model is introduced and compared to the available data. In particular, this work presents two exceptional PWNe which protrude from the rest of the population, namely the Crab Nebula and N 157B. Both objects are driven by pulsars with extremely high rotational energy loss rates. Accordingly, they are often referred to as energetic twins. Modelling the non-thermal multi-wavelength emission of N157B gives access to specific properties of this object, like the magnetic field inside the nebula. Comparing the derived parameters to those of the Crab Nebula reveals large intrinsic differences between the two PWNe. Possible origins of these differences are discussed in context of the resembling pulsars.
Compared to the TeV gamma-ray regime, the number of detected PWNe is much smaller in the MeV-GeV gamma-ray range. In the latter range, the Crab Nebula stands out by the recent detection of gamma-ray flares. In general, the measured flux enhancements on short time scales of days to weeks were not expected in the theoretical understanding of PWNe. In this thesis, the variability of the Crab Nebula is analysed using data from the Fermi Large Area Telescope (Fermi-LAT). For the presented analysis, a new gamma-ray reconstruction method is used, providing a higher sensitivity and a lower energy threshold compared to previous analyses. The derived gamma-ray light curve of the Crab Nebula is investigated for flares and periodicity. The detected flares are analysed regarding their energy spectra, and their variety and commonalities are discussed. In addition, a dedicated analysis of the flare which occurred in March 2013 is performed. The derived short-term variability time scale is roughly 6h, implying a small region inside the Crab Nebula to be responsible for the enigmatic flares. The most promising theories explaining the origins of the flux eruptions and gamma-ray variability are discussed in detail.
In the technical part of this work, a new analysis framework is presented. The introduced software, called gammalib/ctools, is currently being developed for the future CTA observa- tory. The analysis framework is extensively tested using data from the H. E. S. S. experiment. To conduct proper data analysis in the likelihood framework of gammalib/ctools, a model describing the distribution of background events in H.E.S.S. data is presented. The software provides the infrastructure to combine data from several instruments in one analysis. To study the gamma-ray emitting PWN population, data from Fermi-LAT and H. E. S. S. are combined in the likelihood framework of gammalib/ctools. In particular, the spectral peak, which usually lies in the overlap energy regime between these two instruments, is determined with the presented analysis framework. The derived measurements are compared to the predictions from the time-dependent model. The combined analysis supports the conclusion of a diverse population of gamma-ray emitting PWNe.
Magnetite is an iron oxide, which is ubiquitous in rocks and is usually deposited as small nanoparticulate matter among other rock material. It differs from most other iron oxides because it contains divalent and trivalent iron. Consequently, it has a special crystal structure and unique magnetic properties. These properties are used for paleoclimatic reconstructions where naturally occurring magnetite helps understanding former geological ages. Further on, magnetic properties are used in bio- and nanotechnological applications –synthetic magnetite serves as a contrast agent in MRI, is exploited in biosensing, hyperthermia or is used in storage media.
Magnetic properties are strongly size-dependent and achieving size control under preferably mild synthesis conditions is of interest in order to obtain particles with required properties. By using a custom-made setup, it was possible to synthesize stable single domain magnetite nanoparticles with the co-precipitation method. Furthermore, it was shown that magnetite formation is temperature-dependent, resulting in larger particles at higher temperatures. However, mechanistic approaches about the details are incomplete.
Formation of magnetite from solution was shown to occur from nanoparticulate matter rather than solvated ions. The theoretical framework of such processes has only started to be described, partly due to the lack of kinetic or thermodynamic data. Synthesis of magnetite nanoparticles at different temperatures was performed and the Arrhenius plot was used determine an activation energy for crystal growth of 28.4 kJ mol-1, which led to the conclusion that nanoparticle diffusion is the rate-determining step.
Furthermore, a study of the alteration of magnetite particles of different sizes as a function of their storage conditions is presented. The magnetic properties depend not only on particle size but also depend on the structure of the oxide, because magnetite oxidizes to maghemite under environmental conditions. The dynamics of this process have not been well described. Smaller nanoparticles are shown to oxidize more rapidly than larger ones and the lower the storage temperature, the lower the measured oxidation. In addition, the magnetic properties of the altered particles are not decreased dramatically, thus suggesting that this alteration will not impact the use of such nanoparticles as medical carriers.
Finally, the effect of biological additives on magnetite formation was investigated. Magnetotactic bacteria¬¬ are able to synthesize and align magnetite nanoparticles of well-defined size and morphology due to the involvement of special proteins with specific binding properties. Based on this model of morphology control, phage display experiments were performed to determine peptide sequences that preferably bind to (111)-magnetite faces. The aim was to control the shape of magnetite nanoparticles during the formation. Magnetotactic bacteria are also able to control the intracellular redox potential with proteins called magnetochromes. MamP is such a protein and its oxidizing nature was studied in vitro via biomimetic magnetite formation experiments based on ferrous ions. Magnetite and further trivalent oxides were found.
This work helps understanding basic mechanisms of magnetite formation and gives insight into non-classical crystal growth. In addition, it is shown that alteration of magnetite nanoparticles is mainly based on oxidation to maghemite and does not significantly influence the magnetic properties. Finally, biomimetic experiments help understanding the role of MamP within the bacteria and furthermore, a first step was performed to achieve morphology control in magnetite formation via co-precipitation.
A dramatic efficiency improvement of bulk heterojunction solar cells based on electron-donating conjugated polymers in combination with soluble fullerene derivatives has been achieved over the past years. Certified and reported power conversion efficiencies now reach over 9% for single junctions and exceed the 10% benchmark for tandem solar cells. This trend brightens the vision of organic photovoltaics becoming competitive with inorganic solar cells including the realization of low-cost and large-area organic photovoltaics. For the best performing organic materials systems, the yield of charge generation can be very efficient. However, a detailed understanding of the free charge carrier generation mechanisms at the donor acceptor interface and the energy loss associated with it needs to be established. Moreover, organic solar cells are limited by the competition between charge extraction and free charge recombination, accounting for further efficiency losses. A conclusive picture and the development of precise methodologies for investigating the fundamental processes in organic solar cells are crucial for future material design, efficiency optimization, and the implementation of organic solar cells into commercial products.
In order to advance the development of organic photovoltaics, my thesis focuses on the comprehensive understanding of charge generation, recombination and extraction in organic bulk heterojunction solar cells summarized in 6 chapters on the cumulative basis of 7 individual publications.
The general motivation guiding this work was the realization of an efficient hybrid inorganic/organic tandem solar cell with sub-cells made from amorphous hydrogenated silicon and organic bulk heterojunctions. To realize this project aim, the focus was directed to the low band-gap copolymer PCPDTBT and its derivatives, resulting in the examination of the charge carrier dynamics in PCPDTBT:PC70BM blends in relation to by the blend morphology. The phase separation in this blend can be controlled by the processing additive diiodooctane, enhancing domain purity and size. The quantitative investigation of the free charge formation was realized by utilizing and improving the time delayed collection field technique. Interestingly, a pronounced field dependence of the free carrier generation for all blends is found, with the field dependence being stronger without the additive. Also, the bimolecular recombination coefficient for both blends is rather high and increases with decreasing internal field which we suggest to be caused by a negative field dependence of mobility. The additive speeds up charge extraction which is rationalized by the threefold increase in mobility.
By fluorine attachment within the electron deficient subunit of PCPDTBT, a new polymer F-PCPDTBT is designed. This new material is characterized by a stronger tendency to aggregate as compared to non-fluorinated PCPDTBT. Our measurements show that for F-PCPDTBT:PCBM blends the charge carrier generation becomes more efficient and the field-dependence of free charge carrier generation is weakened. The stronger tendency to aggregate induced by the fluorination also leads to increased polymer rich domains, accompanied in a threefold reduction in the non-geminate recombination coefficient at conditions of open circuit. The size of the polymer domains is nicely correlated to the field-dependence of charge generation and the Langevin reduction factor, which highlights the importance of the domain size and domain purity for efficient charge carrier generation. In total, fluorination of PCPDTBT causes the PCE to increase from 3.6 to 6.1% due to enhanced fill factor, short circuit current and open circuit voltage. Further optimization of the blend ratio, active layer thickness, and polymer molecular weight resulted in 6.6% efficiency for F-PCPDTBT:PC70BM solar cells.
Interestingly, the double fluorinated version 2F-PCPDTBT exhibited poorer FF despite a further reduction of geminate and non-geminate recombination losses. To further analyze this finding, a new technique is developed that measures the effective extraction mobility under charge carrier densities and electrical fields comparable to solar cell operation conditions. This method involves the bias enhanced charge extraction technique. With the knowledge of the carrier density under different electrical field and illumination conditions, a conclusive picture of the changes in charge carrier dynamics leading to differences in the fill factor upon fluorination of PCPDTBT is attained. The more efficient charge generation and reduced recombination with fluorination is counterbalanced by a decreased extraction mobility. Thus, the highest fill factor of 60% and efficiency of 6.6% is reached for F-PCPDTBT blends, while 2F-PCPDTBT blends have only moderate fill factors of 54% caused by the lower effective extraction mobility, limiting the efficiency to 6.5%.
To understand the details of the charge generation mechanism and the related losses, we evaluated the yield and field-dependence of free charge generation using time delayed collection field in combination with sensitive measurements of the external quantum efficiency and absorption coefficients for a variety of blends. Importantly, both the yield and field-dependence of free charge generation is found to be unaffected by excitation energy, including direct charge transfer excitation below the optical band gap. To access the non-detectable absorption at energies of the relaxed charge transfer emission, the absorption was reconstructed from the CT emission, induced via the recombination of thermalized charges in electroluminescence. For a variety of blends, the quantum yield at energies of charge transfer emission was identical to excitations with energies well above the optical band-gap. Thus, the generation proceeds via the split-up of the thermalized charge transfer states in working solar cells. Further measurements were conducted on blends with fine-tuned energy levels and similar blend morphologies by using different fullerene derivatives. A direct correlation between the efficiency of free carrier generation and the energy difference of the relaxed charge transfer state relative to the energy of the charge separated state is found. These findings open up new guidelines for future material design as new high efficiency materials require a minimum energetic offset between charge transfer and the charge separated state while keeping the HOMO level (and LUMO level) difference between donor and acceptor as small as possible.
Research in rodents has shown that dietary vitamin A reduces body fat by enhancing fat mobilisation and energy utilisation; however, their effects in growing dogs remain unclear. In the present study, we evaluated the development of body weight and body composition and compared observed energy intake with predicted energy intake in forty-nine puppies from two breeds (twenty-four Labrador Retriever (LAB) and twenty-five Miniature Schnauzer (MS)). A total of four different diets with increasing vitamin A content between 5.24 and 104.80 mu mol retinol (5000-100 000 IU vitamin A)/4184 kJ (1000 kcal) metabolisable energy were fed from the age of 8 weeks up to 52 (MS) and 78 weeks (LAB). The daily energy intake was recorded throughout the experimental period. The body condition score was evaluated weekly using a seven-category system, and food allowances were adjusted to maintain optimal body condition. Body composition was assessed at the age of 26 and 52 weeks for both breeds and at the age of 78 weeks for the LAB breed only using dual-energy X-ray absorptiometry. The growth curves of the dogs followed a breed-specific pattern. However, data on energy intake showed considerable variability between the two breeds as well as when compared with predicted energy intake. In conclusion, the data show that energy intakes of puppies particularly during early growth are highly variable; however, the growth pattern and body composition of the LAB and MS breeds are not affected by the intake of vitamin A at levels up to 104.80 mu mol retinol (100 000 IU vitamin A)/4184 kJ (1000 kcal).
The complementary advantages of high-rate Global Positioning System (GPS) and accelerometer observations for measuring seismic ground motion have been recognised in previous research. Here we propose an approach of tight integration of GPS and accelerometer measurements. The baseline shifts of the accelerometer are introduced as unknown parameters and estimated by a random walk process in the Precise Point Positioning (PPP) solution. To demonstrate the performance of the new strategy, we carried out several experiments using collocated GPS and accelerometer. The experimental results show that the baseline shifts of the accelerometer are automatically corrected, and high precision coseismic information of strong ground motion can be obtained in real-time. Additionally, the convergence and precision of the PPP is improved by the combined solution.
Bats are important components in tropical mammal assemblages. Unravelling the mechanisms allowing multiple syntopic bat species to coexist can provide insights into community ecology. However, dietary information on component species of these assemblages is often difficult to obtain. Here we measuredstable carbon and nitrogen isotopes in hair samples clipped from the backs of 94 specimens to indirectly examine whether trophic niche differentiation and microhabitat segregation explain the coexistence of 16 bat species at Ankarana, northern Madagascar. The assemblage ranged over 4.4% in delta N-15 and was structured into two trophic levels with phytophagous Pteropodidae as primary consumers (c. 3% enriched over plants) and different insectivorous bats as secondary consumers (c. 4% enriched over primary consumers). Bat species utilizing different microhabitats formed distinct isotopic clusters (metric analyses of delta C-13-delta N-15 bi-plots), but taxa foraging in the same microhabitat did not show more pronounced trophic differentiation than those occupying different microhabitats. As revealed by multivariate analyses, no discernible feeding competition was found in the local assemblage amongst congeneric species as compared with non-congeners. In contrast to ecological niche theory, but in accordance with studies on New and Old World bat assemblages, competitive interactions appear to be relaxed at Ankarana and not a prevailing structuring force.