The potential role of ABC transporters as factors influencing drug susceptibility in the salmon louse, Lepeophtheirus salmonis (Kroyer, 1837)

Efficient control of sea lice is a major challenge for the sustainable production of farmed Atlantic salmon (Salmo salar (Linnaeus, 1758)). These marine ectoparasites feed on mucus, skin and blood of their hosts, thereby reducing the salmon’s growth rate and overall health. In the northern hemisphere, the most prevalent species is Lepeophtheirus salmonis (Krøyer, 1837). In 2006, global costs of sea lice infections are estimated to have exceeded €300 million, with the majority spent on a limited number of chemical delousing agents. Emamectin benzoate (EMB; SLICE®), an avermectin, has been widely used since its introduction in 2000, due to its convenient administration as an in-feed medication and its high efficacy against all parasitic stages of L. salmonis. However, over-reliance on a single or limited range of medicines favours the emergence of drug resistance and, as a result, the efficacy of this compound in treating L. salmonis has decreased in recent years, as reported from e.g. Chile, Norway, Scotland and Canada. Declining efficacy underlines the need for an improved understanding of the molecular mechanisms underlying EMB drug resistance in L. salmonis. Elucidation of these mechanisms would allow for improved monitoring tools, earlier detection of developing resistance, extended usability of current delousing agents and development of new parasiticides. The work described in this thesis sets out to examine the molecular mechanisms underlying EMB resistance in L. salmonis. In earlier studies, research in nematodes and arthropods has linked drug efflux transporters belonging to the family of ATP-binding cassette (ABC) transporters to ivermectin (IVM) resistance, a parasiticide with high chemical similarity to EMB. ABC transporters such as permeability glycoprotein (P-gp), transport a wide range of substrates, including drugs, and have been suggested to provide a potential molecular mechanism through which EMB resistance might be mediated in sea lice. As an example of such mechanisms, increased expression of P-gp is one of the causative factors for drug resistance in human cancer cells and avermectin resistance in nematode parasites such as Caenorhabditis elegans or Haemonchus contortus. Initial research involved screening for novel salmon lice P-gps that might contribute to EMB resistance. A novel P-gp, SL-PGY1, was discovered using a combined bioinformatic and molecular biological approach. The expression was compared in two well-characterised L. salmonis strains differing in their susceptibility to EMB (S = susceptible, R = resistant). Prior to EMB exposure, mRNA levels did not differ from each other, while, after 24 h exposure, a 2.9-fold increase in SL-PGY1 mRNA expression was observed in the R strain. SL-PGY1 appears not to be a major factor contributing to reduced EMB susceptibility, although it could play a role, as expression levels increased upon exposure to EMB. A further four additional drug transporters (ABC C subfamily) were also discovered showing high homology to multidrug-resistance proteins (MRP). The relative expression levels of each MRP was compared in the strains S and R, before and after exposure to EMB. No significant changes were found in their expression patterns. If ABC drug transporters mediate the efflux of EMB and thereby reduce the intracellular concentrations of the drug in exposed animals, the inhibition of those ABC drug transporters was expected to lead to higher intracellular levels of EMB. This could result in an enhanced toxic effect when EMB is co-administered with an inhibitor. Two known inhibitors of human P-gps and MRPs, cyclosporin A (CSA) and verapamil (VER), were co-administered with EMB. CSA increased the toxic effect of EMB in both tested strains, implying that the targets of CSA are expressed at comparable levels and that they may be part of the mechanism conferring EMB resistance. VER increased the toxic effect of EMB in the R strain, but had no significant effects on the S strain. This implies that the expression of factors inhibited by VER differs between the two L. salmonis strains. It is hypothesised that a number of ABC transporters with distinct, yet overlapping patterns of inhibitor specificity are affected by those inhibitors. The search for drug-resistance conferring genes was complemented with a systematic, genome-wide survey of ABC transporters in L. salmonis to find additional members of this important gene family. Next-generation high-throughput RNA sequencing (RNA-seq) was employed to assemble a reference transcriptome from pooled total RNA of salmon lice at different development stages. The transcriptome was assembled against the L. salmonis genome and annotated. Thirty-nine putative ABC transporters were found. Of further interest were transcripts of the subfamily B, C and G, as they contain drug-transporting ABC proteins. For the ABC B subfamily, one full (SL-PGY1) and three half transporter transcripts were found. Only full transporters are known to transport drugs and SL-PGY1 is apparently not a major factor contributing to EMB resistance. Fourteen ABCC sequences were found – 11 MRPs and 3 homologues to sulfonylurea receptors. Of interest are MRPs, as they contribute to drug detoxification in humans and invertebrates. Four MRPs had been identified previously and their expression ratios did not differ between S and R strain parasites. Seven sequences belonging to ABCG subfamily were found. However, none of the L. salmonis ABCG transcripts identified showed sufficient homology to known drug transporters in other species. With the currently limited understanding of the mechanisms conferring EMB resistance, monitoring the susceptibility of L. salmonis subpopulations is essential. Dose-response bioassays are currently widely used. Tests with pre-adult II or adult parasites requires relatively large numbers of parasites (~150) to conduct this type of bioassay, which may not always be available. Addressing this issue, we tested the feasibility of a single-dose bioassay (requiring fewer test animals than dose-response bioassays) to discriminate between L. salmonis strains with differing EMB susceptibility. This alternative approach uses time-course toxicity analysis, where the toxic effect of EMB is monitored over time. After clearly defining the effect criteria, we found that it is possible to discriminate between those L. salmonis strains. However, while requiring fewer test animals, time course toxicity analysis is more labour-intensive, but the alternative design can be suitable under certain circumstances. The work reported here has provided new knowledge concerning the mechanisms of EMB resistance in sea lice. Several novel putative drug transporters have been identified, an important first step toward unravelling the complex interactions of genes involved in EMB resistance in this commercially important parasite

Disentangle the price dispersion of residential solar photovoltaic systems: Evidence from Germany

Although Germany has the largest capacity of installed residential photovoltaic (PV) systems in Europe, comprehensive evidence on transparent pricing information remains missing. This study disentangles why PV quote prices are subject to significant dispersion and analyzes which factors influence particularly low- and high-priced systems in Germany. We create a comprehensive cross-sectional dataset of 19 561 PV system quotes from 2011 to 2022 and use regression analyses to investigate the effects of system characteristics, installation scope, and location-related parameters on quoted prices. Our results reveal highly volatile annual price dispersion consistent over 11 years and large price differences despite similar system characteristics. Applying hedonic regression techniques, we reveal spatially fine-resolved price heterogeneity with up to 20 % difference in the German PV market. System characteristics such as battery usage, installation scope, and system capacity have the most significant effect sizes and are instead control variables. More insightful, the installer density shows price-lowering effects, whereas more PV installations per region, higher solar radiation, and higher labor wages cause price-increasing effects. Quantile regression results reveal that installer density promotes the price reduction of high-priced systems more. Scaffolding, AC installation, and elevation are significant price-increasing factors but with small effect sizes. Finally, DC optimizers affect the levels of high-priced systems more than low-priced ones

A Spatiotemporal Study and Location-Specific Trip Pattern Categorization of Shared E-Scooter Usage

This study analyzes the temporally resolved location and trip data of shared e-scooters over nine months in Berlin from one of Europe’s most widespread operators. We apply time, distance, and energy consumption filters on approximately 1.25 million trips for outlier detection and trip categorization. Using temporally and spatially resolved trip pattern analyses, we investigate how the built environment and land use affect e-scooter trips. Further, we apply a density-based clustering algorithm to examine point of interest-specific patterns in trip generation. Our results suggest that e-scooter usage has point of interest related characteristics. Temporal peaks in e-scooter usage differ by point of interest category and indicate work-related trips at public transport stations. We prove these characteristic patterns with the statistical metric of cosine similarity. Considering average cluster velocities, we observe limited time-saving potential of e-scooter trips in congested areas near the city center

Antibodies to Core Lipopolysaccharide Determinants: Absence of Cross-reactivity with Heterologous Lipopolysaccharides

Using monoclonal antibodies directed against defined epitopes of endotoxin core, this study demonstrated that the presentation of lipopolysaccharide (LPS) to antibodies is critical for measuring the specific binding of antibodies to LPS structures. False cross-reactive reactions apparently were observed when free core LPS or lipid A were used as antigens in ELISA, whereas coating with complexes of high-density lipoproteins with core LPS increased both the sensitivity and the specificity of the test compared with coating with free core LPS, so that nonspecific binding of antibodies was largely avoided. Using this technique, it was not possible to find broadly cross-reactive core LPS antibodies after immunization of rabbits and humans with rough mutants of gram-negative bacteria. These observations underscore the need for careful evaluation of the potential for cross-reactivity of antisera and of monoclonal antibodies directed against endotoxin cor

Individually optimized commercial road transport: A decision support system for customizable routing problems

The Vehicle Routing Problem (VRP) in its manifold variants is widely discussed in scientific literature. We investigate related optimization models and solution methods to determine the state of research for vehicle routing attributes and their combinations. Most of these approaches are idealized and focus on single problem-tailored routing applications. Addressing this research gap, we present a customizable VRP for optimized road transportation embedded into a Decision Support System (DSS). It integrates various model attributes and handles a multitude of real-world routing problems. In the context of urban logistics, practitioners of different industries and researchers are assisted in efficient route planning that allows for minimizing driving distances and reducing vehicle emissions. Based on the design science research methodology, we evaluate the DSS with computational benchmarks and real-world simulations. Results indicate that our developed DSS can compete with problem-tailored algorithms. With our solution-oriented DSS as final artifact, we contribute to an enhanced economic and environmental sustainability in urban logistic applications

Irreversibilitaet der aeroben Spaltkraft bei optimaler Sauerstoffversorgung der Zelle

Mit der Entdeckung der Krebsglykolyse durch Warburg sind zwei differente Arten von Spaltungsstoffwechsel bekannt geworden. Die aerob irreversible Spaltkraft ist nach Warburg fuer alle malignen Zellen symptomatisch. Aber auch die Hefen und Milchsaeurebakterien in der Natur verfuegen, wie im folgenden nachgewiesen wird, ueber die Faehigkeit der aeroben Zuckerspaltung. Die gaerfaehige Torulopsis, von Meyerhof als Prototyp der aerob nicht-gaerenden Hefen herausgestellt, bildet oxy- wie anoxybiotisch annaehernd die gleichen Mengen Alkohol. Bei der zymagenen Umzuechtung von urspruenglich nur atmenden Torulazeen ist die Spaltkraft bereits in statu nascendi aerob irreversibel konstituiert. Die Meyerhofsche Korrelationstheorie von der aeroben Dominanz der Atmung ueber die Spaltung ist demnach in ihrem unitarischen Anspruch unsubstantiiert und zu reduzieren. Die Hefezelle, die im ausgereiften, ruhenden Zustand keiner O(2)-Beeinflussung unterliegt, ist vegetativ O(2)-abhaengig, sowohl in bezug auf die Proliferation selbst, als auch hinsichtlich der potentiellen Ausbildung der Stoffwechselkraefte. Im vegetativen Stadium tritt die Pasteursche "consequence de la vie sans air" voll in Erscheinung. Fuer die fertig ausgebildete, nicht in Vermehrung befindliche Hefezelle gibt es indessen weder eine Pasteursche Konsequenz der Anaerobiose, noch eine Meyerhofsche Korrelation zwischen Oxy- und Anoxybiose