### Recent Submissions

• Journal Article

#### National Forest Inventories capture the multifunctionality of managed forests in Germany ﻿

Forest Ecosystems. 2021 Jan 27;8(1):5
Background Forests perform various important ecosystem functions that contribute to ecosystem services. In many parts of the world, forest management has shifted from a focus on timber production to multi-purpose forestry, combining timber production with the supply of other forest ecosystem services. However, it is unclear which forest types provide which ecosystem services and to what extent forests primarily managed for timber already supply multiple ecosystem services. Based on a comprehensive dataset collected across 150 forest plots in three regions differing in management intensity and species composition, we develop models to predict the potential supply of 13 ecosystem services. We use those models to assess the level of multifunctionality of managed forests at the national level using national forest inventory data. Results Looking at the potential supply of ecosystem services, we found trade-offs (e.g. between both bark beetle control or dung decomposition and both productivity or soil carbon stocks) as well as synergies (e.g. for temperature regulation, carbon storage and culturally interesting plants) across the 53 most dominant forest types in Germany. No single forest type provided all ecosystem services equally. Some ecosystem services showed comparable levels across forest types (e.g. decomposition or richness of saprotrophs), while others varied strongly, depending on forest structural attributes (e.g. phosphorous availability or cover of edible plants) or tree species composition (e.g. potential nitrification activity). Variability in potential supply of ecosystem services was only to a lesser extent driven by environmental conditions. However, the geographic variation in ecosystem function supply across Germany was closely linked with the distribution of main tree species. Conclusions Our results show that forest multifunctionality is limited to subsets of ecosystem services. The importance of tree species composition highlights that a lack of multifunctionality at the stand level can be compensated by managing forests at the landscape level, when stands of complementary forest types are combined. These results imply that multi-purpose forestry should be based on a variety of forest types requiring coordinated planning across larger spatial scales.
• Journal Article

#### Low Population Differentiation but High Phenotypic Plasticity of European Beech in Germany ﻿

Forests 2020; 11(12) p.1-14: Art. 1354
Drought is increasingly impairing the vitality of European beech (Fagus sylvatica L.) in several regions of its distribution range. In times of climate change, adaptive traits such as plant phenology and frost tolerance are also becoming more important. Adaptive patterns of European beech seem to be complex, as contrasting results regarding the relative effect of phenotypic plasticity and genetic variation in trait variation have been reported. Here, we used a large translocation experiment comprising more than 15,500 seedlings in three regions of Germany to investigate local adaptation and phenotypic plasticity in beech. We found low population differentiation regarding plant survival, and plant height increment, but high phenotypic plasticity for these traits. Survival showed a positive correlation with temperature variables and a less pronounced and negative correlation with precipitation-related variables. This suggests a predominant effect of temperature and growing degree days on the survival of beech seedlings under moderate drought stress. The high phenotypic plasticity may help beech to cope with changing environmental conditions, albeit increasing drought stress may make adaptive changes necessary in the long term.
• Journal Article

#### Establishment of the monomeric yellow-green fluorescent protein mNeonGreen for life cell imaging in mycelial fungi ﻿

AMB Express 2020; 10(1) p.1-10: Art. 222
Abstract The engineered monomeric version of the lancelet Branchiostoma lanceolatum fluorescent protein, mNeonGreen (mNG), has several positive characteristics, such as a very bright fluorescence, high photostability and fast maturation. These features make it a good candidate for the utilization as fluorescent tool for cell biology and biochemical applications in filamentous fungi. We report the generation of plasmids for the expression of the heterologous mNG gene under the control of an inducible and a constitutive promoter in the filamentous ascomycete Sordaria macrospora and display a stable expression of mNG in the cytoplasm. To demonstrate its usefulness for labeling of organelles, the peroxisomal targeting sequence serine-lysine-leucine (SKL) was fused to mNG. Expression of this tagged version led to protein import of mNG into peroxisomes and their bright fluorescence in life cell imaging.
• Journal Article

#### The Role of Socio-Economic Determinants of Horse Farms for Grassland Management, Vegetation Composition and Ecological Value ﻿

Sustainability 2020; 12(24) p.1-18: Art. 10641
Socio-economic context is increasingly seen as a decisive factor for sustainable agricultural land use. The high prevalence of part-time farming and frequent lack of formal agricultural education within the equine sector are often seen as reasons why horse-grazed pastures do not fulfill their biodiversity potential. In spite of the substantial variability within horse farming, little is known about the relationship of socio-economic determinants with vegetation characteristics of horse-grazed grasslands. We surveyed 122 horse farms in Germany, classifying them into four socio-economic classes according to farm income type and farm managers’ agricultural education. We recorded farm structure parameters, grassland management practices and vegetation characteristics. Socio-economic class partly explained the great variability in farm structure that we observed. In contrast to our expectation, income type and agricultural education did not distinctly affect grassland management and were neither directly nor indirectly related to vegetation characteristics. Part-time farming and lack of agricultural education thus did not adversely affect the ecological value of horse-grazed grasslands. By contrast, both farm structure and paddock level management affected grassland vegetation and ecological value. Therefore, the socio-economic context of horse farms should be addressed in further research with strategies targeting the development of sustainable grassland management in horse keeping.
• Journal Article

#### Improvement on the genetic engineering of an invasive agricultural pest insect, the cherry vinegar fly, Drosophila suzukii ﻿

BMC Genetics. 2020 Dec 18;21(Suppl 2):139
Background The invasive fly Drosophila suzukii has become an established fruit pest in Europe, the USA, and South America with no effective and safe pest management. Genetic engineering enables the development of transgene-based novel genetic control strategies against insect pests and disease vectors. This, however, requires the establishment of reliable germline transformation techniques. Previous studies have shown that D. suzukii is amenable to transgenesis using the transposon-based vectors piggyBac and Minos, site-specific recombination (lox/Cre), and CRISPR/Cas9 genome editing. Results We experienced differences in the usability of piggyBac-based germline transformation in different strains of D. suzukii: we obtained no transgenic lines in a US strain, a single rare transgenic line in an Italian strain, but observed a reliable transformation rate of 2.5 to 11% in a strain from the French Alps. This difference in efficiency was confirmed by comparative examination of these three strains. In addition, we used an attP landing site line to successfully established φC31-integrase-mediated plasmid integration at a rate of 10% and generated landing site lines with two attP sequences to effectively perform φC31-Recombinase Mediated Cassette Exchange (φC31-RMCE) with 11% efficiency. Moreover, we isolated and used the endogenous regulatory regions of Ds nanos to express φC31 integrase maternally to generate self-docking lines for φC31-RMCE. Besides, we isolated the promoter/enhancer of Ds serendipity α to drive the heterologous tetracycline-controlled transactivator (tTA) during early embryonic development and generated a testes-specific tTA driver line using the endogenous beta-2-tubulin (β2t) promoter/enhancer. Conclusion Our results provide evidence that the D. suzukii strain AM derived from the French Alps is more suitable for piggyBac germline transformation than other strains. We demonstrated the feasibility of using φC31-RMCE in the cherry vinegar fly and generated a set of lines that can be used for highly efficient integration of larger constructs. The φC31-based integration will facilitate modification and stabilization of previously generated transgenic lines that carry at least one attP site in the transgene construction. An early embryo-specific and a spermatogenesis-specific driver line were generated for future use of the binary expression system tet-off to engineer tissue- and stage-specific effector gene expression for genetic pest control strategies.
• Journal Article

#### Local Responses and Systemic Induced Resistance Mediated by Ectomycorrhizal Fungi ﻿

Frontiers in Plant Science 2020; 11 p.1-20: Art. 590063
Ectomycorrhizal fungi (EMF) grow as saprotrophs in soil and interact with plants, forming mutualistic associations with roots of many economically and ecologically important forest tree genera. EMF ensheath the root tips and produce an extensive extramatrical mycelium for nutrient uptake from the soil. In contrast to other mycorrhizal fungal symbioses, EMF do not invade plant cells but form an interface for nutrient exchange adjacent to the cortex cells. The interaction of roots and EMF affects host stress resistance but uncovering the underlying molecular mechanisms is an emerging topic. Here, we focused on local and systemic effects of EMF modulating defenses against insects or pathogens in aboveground tissues in comparison with arbuscular mycorrhizal induced systemic resistance. Molecular studies indicate a role of chitin in defense activation by EMF in local tissues and an immune response that is induced by yet unknown signals in aboveground tissues. Volatile organic compounds may be involved in long-distance communication between below- and aboveground tissues, in addition to metabolite signals in the xylem or phloem. In leaves of EMF-colonized plants, jasmonate signaling is involved in transcriptional re-wiring, leading to metabolic shifts in the secondary and nitrogen-based defense metabolism but cross talk with salicylate-related signaling is likely. Ectomycorrhizal-induced plant immunity shares commonalities with systemic acquired resistance and induced systemic resistance. We highlight novel developments and provide a guide to future research directions in EMF-induced resistance.
• Journal Article

#### Predicting Tree Sap Flux and Stomatal Conductance from Drone-Recorded Surface Temperatures in a Mixed Agroforestry System—A Machine Learning Approach ﻿

Remote Sensing 2020; 12(24) p.1-20: Art. 4070
Plant transpiration is a key element in the hydrological cycle. Widely used methods for its assessment comprise sap flux techniques for whole-plant transpiration and porometry for leaf stomatal conductance. Recently emerging approaches based on surface temperatures and a wide range of machine learning techniques offer new possibilities to quantify transpiration. The focus of this study was to predict sap flux and leaf stomatal conductance based on drone-recorded and meteorological data and compare these predictions with in-situ measured transpiration. To build the prediction models, we applied classical statistical approaches and machine learning algorithms. The field work was conducted in an oil palm agroforest in lowland Sumatra. Random forest predictions yielded the highest congruence with measured sap flux (r$^2$ = 0.87 for trees and r$^2$ = 0.58 for palms) and confidence intervals for intercept and slope of a Passing-Bablok regression suggest interchangeability of the methods. Differences in model performance are indicated when predicting different tree species. Predictions for stomatal conductance were less congruent for all prediction methods, likely due to spatial and temporal offsets of the measurements. Overall, the applied drone and modelling scheme predicts whole-plant transpiration with high accuracy. We conclude that there is large potential in machine learning approaches for ecological applications such as predicting transpiration.
• Journal Article

#### Diversity of Expression Types of Ht Genes Conferring Resistance in Maize to Exserohilum turcicum ﻿

Frontiers in Plant Science 2020; 11 p.1-11: Art. 607850
Northern corn leaf blight (NCLB) is an important leaf disease in maize (Zea mays) worldwide and is spreading into new areas with expanding maize cultivation, like Germany. Exserohilum turcicum, causal agent of NCLB, infects and colonizes leaf tissue and induces elongated necrotic lesions. Disease control is based on fungicide application and resistant cultivars displaying monogenic resistance. Symptom expression and resistance mechanisms differ in plants carrying different resistance genes. Therefore, histological studies and DNA quantification were performed to compare the pathogenesis of E. turcicum races in maize lines exhibiting compatible or incompatible interactions. Maize plants from the differential line B37 with and without resistance genes Ht1, Ht2, Ht3, and Htn1 were inoculated with either incompatible or compatible races (race 0, race 1 and race 23N) of E. turcicum. Leaf segments from healthy and inoculated plants were collected at five different stages of infection and disease development from penetration (0–1 days post inoculation - dpi), until full symptom expression (14–18 dpi). Symptoms of resistance responses conveyed by the different Ht genes considerably differed between Ht1 (necrotic lesions with chlorosis), Ht2 (chlorosis and small lesions), Ht3 (chlorotic spots) and Htn1 (no lesions or wilt-type lesions). In incompatible interactions, fungal DNA was only detected in very low amounts. At 10 dpi, DNA content was elevated in all compatible interactions. Histological studies with Chlorazol Black E staining indicated that E. turcicum formed appressoria and penetrated the leaf surface directly in both types of interaction. In contrast to incompatible interactions, however, the pathogen was able to penetrate into xylem vessels at 6 dpi in compatible interactions and strongly colonized the mesophyll at 12 dpi, which is considered the crucial process differentiating susceptible from resistant interactions. Following the distinct symptom expressions, resistance mechanisms conferred by Ht1, Ht2, Ht3, and Htn1 genes apparently are different. Lower disease levels and a delayed progress of infection in compatible interactions with resistant lines imply that maize R genes to E. turcicum are associated with or confer additional quantitative resistance.
• Journal Article

#### Environmental Concern and Urbanization in India: Towards Psychological Complexity ﻿

Sustainability 2020; 12(24) p.1-25: Art. 10402
Urbanizing social-ecological systems often experience environmental degradation, especially in the Global South. Traditional urban psychology has attributed this to decreasing environmental concern due to weakening connections to nature. However, urban psychological research has barely considered how predictions may improve when including psychological complexity, exemplified by context, in the urbanization-concern link. In this work, we test for sensitivity of a loss of nature connection to cultural context, for substitution by additional southern urban features, and for the emergence of aggregate preferences based on the feedback between these mediators in regard to the overall relationship. Our structural equations model is calibrated using original survey data from the globalized southern megacity Bangalore, India. The spatial explicitness of our data allows for representative sampling from its rich urban variation. Spatial lags of exogenous variables provide instrumental variables to control for endogeneity arising from feedback. The results suggest that modernization-induced value change is the main policy leverage that facilitates pro-environmental preferences within a uniquely Indian interplay of various urban psychological effects.
• Journal Article

#### Current State and Drivers of Arable Plant Diversity in Conventionally Managed Farmland in Northwest Germany ﻿

Diversity 2020; 12(12) p.1-15: Art. 469
Agricultural intensification has led to dramatic diversity losses and impoverishment of the arable vegetation in much of Europe. We analyzed the status of farmland phytodiversity and its determinants in 2016 in northwest Germany by surveying 200 conventionally managed fields cultivated with seven crops. The study was combined with an analysis of edaphic (soil yield potential), agronomic (crop cover, fertilizer and herbicide use) and landscape factors (adjacent habitats). In total, we recorded 150 non-crop plant species, many of them nitrophilous generalist species, while species of conservation value were almost completely absent. According to a post-hoc pairwise comparison of the mixed model results, the cultivation of rapeseed positively influenced non-crop plant species richness as compared to winter cereals (wheat, barley, rye and triticale; data pooled), maize or potato. The presence of grassy strips and ditch margins adjacent to fields increased plant richness at field edges presumably through spillover effects. In the field interiors, median values of non-crop plant richness and cover were only 2 species and 0.5% cover across all crops, and at the field edges 11 species and 4% cover. Agricultural intensification has wiped out non-crop plant life nearly completely from conventionally managed farmland, except for a narrow, floristically impoverished field edge strip.
• Journal Article

#### Mucilage Polysaccharide Composition and Exudation in Maize From Contrasting Climatic Regions ﻿

Frontiers in Plant Science 2020; 11 p.1-13: Art. 587610
Mucilage, a gelatinous substance comprising mostly polysaccharides, is exuded by maize nodal and underground root tips. Although mucilage provides several benefits for rhizosphere functions, studies on the variation in mucilage amounts and its polysaccharide composition between genotypes are still lacking. In this study, eight maize (Zea mays L.) genotypes from different globally distributed agroecological zones were grown under identical abiotic conditions in a randomized field experiment. Mucilage exudation amount, neutral sugars and uronic acids were quantified. Galactose (∼39–42%), fucose (∼22–30%), mannose (∼11–14%), and arabinose (∼8–11%) were the major neutral sugars in nodal root mucilage. Xylose (∼1–4%), and glucose (∼1–4%) occurred only in minor proportions. Glucuronic acid (∼3–5%) was the only uronic acid detected. The polysaccharide composition differed significantly between maize genotypes. Mucilage exudation was 135 and 125% higher in the Indian (900 M Gold) and Kenyan (DH 02) genotypes than in the central European genotypes, respectively. Mucilage exudation was positively associated with the vapor pressure deficit of the genotypes’ agroecological zone. The results indicate that selection for environments with high vapor pressure deficit may favor higher mucilage exudation, possibly because mucilage can delay the onset of hydraulic failure during periods of high vapor pressure deficit. Genotypes from semi-arid climates might offer sources of genetic material for beneficial mucilage traits.
• Journal Article

#### Infection Patterns and Fitness Effects of Rickettsia and Sodalis Symbionts in the Green Lacewing Chrysoperla carnea ﻿

Insects 2020; 11(12) p.1-17: Art. 867
Endosymbionts are widely distributed in insects and can strongly affect their host ecology. The common green lacewing (Chrysoperla carnea) is a neuropteran insect which is widely used in biological pest control. However, their endosymbionts and their interactions with their hosts have not been very well studied. Therefore, we screened for endosymbionts in natural and laboratory populations of Ch. carnea using diagnostic PCR amplicons. We found the endosymbiont Rickettsia to be very common in all screened natural and laboratory populations, while a hitherto uncharacterized Sodalis strain was found only in laboratory populations. By establishing lacewing lines with no, single or co-infections of Sodalis and Rickettsia, we found a high vertical transmission rate for both endosymbionts (>89%). However, we were only able to estimate these numbers for co-infected lacewings. Sodalis negatively affected the reproductive success in single and co-infected Ch. carnea, while Rickettsia showed no effect. We hypothesize that the fitness costs accrued by Sodalis infections might be more tolerable in the laboratory than in natural populations, as the latter are also prone to fluctuating environmental conditions and natural enemies. The economic and ecological importance of lacewings in biological pest control warrants a more profound understanding of its biology, which might be influenced by symbionts.
• Journal Article

#### Animal Welfare Programs in Germany—An Empirical Study on the Attitudes of Pig Farmers ﻿

Agriculture 2020; 10(12) p.1-17: Art. 609
In Europe, there is ongoing social criticism of conventional pig farming and demands for higher farm animal welfare standards. This applies primarily to products from pig production, as consumers criticize, among other things, the animals’ housing conditions, tail docking, neutering, or keeping them on slatted floors. Various animal welfare programs have tried to meet the consumers’ demands. Pig farmers are directly involved in the production process and are therefore key stakeholders for the successful implementation of animal welfare programs such as the German Initiative Animal Welfare. The Initiative Animal Welfare was founded in 2015 and serves as an example in this study, as it has been established for two rounds and involves high numbers of participants. However, little is known about the attitudes of pig farmers towards this specific animal welfare program. Thus, the aim of this study is to investigate these attitudes towards animal welfare programs using the example of German pig producers and identify group differences. Based on an online survey of German conventional pig farmers, four clusters were formed which differ in their attitude to the Initiative Animal Welfare. Overall, all farmers, regardless of the cluster, feel publicly pressured by politics and the media. In addition, all farmers are skeptical about the effort involved in participating in the Initiative Animal Welfare (IAW), especially with regard to the additional documentation requirements and unannounced controls. The findings can provide guidance for the design of animal welfare programs taking into account the needs of farmers.
• Journal Article

#### Stress-related changes in leukocyte profiles and telomere shortening in the shortest-lived tetrapod, Furcifer labordi ﻿

BMC Evolutionary Biology 2020; 20(1) p.1-11: Art. 160
Abstract Background Life history theory predicts that during the lifespan of an organism, resources are allocated to either growth, somatic maintenance or reproduction. Resource allocation trade-offs determine the evolution and ecology of different life history strategies and define an organisms’ position along a fast–slow continuum in interspecific comparisons. Labord’s chameleon (Furcifer labordi) from the seasonal dry forests of Madagascar is the tetrapod species with the shortest reported lifespan (4–9 months). Previous investigations revealed that their lifespan is to some degree dependent on environmental factors, such as the amount of rainfall and the length of the vegetation period. However, the intrinsic mechanisms shaping such a fast life history remain unknown. Environmental stressors are known to increase the secretion of glucocorticoids in other vertebrates, which, in turn, can shorten telomeres via oxidative stress. To investigate to what extent age-related changes in these molecular and cellular mechanisms contribute to the relatively short lifetime of F. labordi, we assessed the effects of stressors indirectly via leukocyte profiles (H/L ratio) and quantified relative telomere length from blood samples in a wild population in Kirindy Forest. We compared our findings with the sympatric, but longer-lived sister species F. cf. nicosiai, which exhibit the same annual timing of reproductive events, and with wild-caught F. labordi that were singly housed under ambient conditions. Results We found that H/L ratios were consistently higher in wild F. labordi compared to F. cf. nicosiai. Moreover, F. labordi already exhibited relatively short telomeres during the mating season when they were 3–4 months old, and telomeres further shortened during their post-reproductive lives. At the beginning of their active season, telomere length was relatively longer in F. cf. nicosiai, but undergoing rapid shortening towards the southern winter, when both species gradually die off. Captive F. labordi showed comparatively longer lifespans and lower H/L ratios than their wild counterparts. Conclusion We suggest that environmental stress and the corresponding accelerated telomere attrition have profound effects on the lifespan of F. labordi in the wild, and identify physiological mechanisms potentially driving their relatively early senescence and mortality.
• Journal Article

#### Studies into Fungal Decay of Wood In Ground Contact—Part 1: The Influence of Water-Holding Capacity, Moisture Content, and Temperature of Soil Substrates on Fungal Decay of Selected Timbers ﻿

Forests 2020; 11(12) p.1-19: Art. 1284
This article presents the results from two separate studies investigating the decay of wood in ground contact using adapted versions of laboratory-based terrestrial microcosm (TMC) tests according to CEN/TS 15083-2:2005. The first study (A) sought to isolate the effect of soil water-holding capacity (WHC$_{\text{soil}}$ [%]) and soil moisture content (MC$_{\text{soil}}$ [%WHC$_{\text{soil}}$]) on the decay of five commercially important wood species; European beech (Fagus sylvatica), English oak heartwood (Quercus robur), Norway spruce (Picea abies), Douglas-fir heartwood (Pseudotsuga menziesii), and Scots pine sapwood (Pinus sylvestris), while keeping soil temperature (Tsoil) constant. Combinations of soil mixtures with WHC$_{\text{soil}}$ of 30%, 60%, and 90%, and MC$_{\text{soil}}$ of 30%, 70%, and 95%WHC$_{\text{soil}}$ were utilized. A general trend showed higher wood decay, measured in oven-dry mass loss (ML$_{\text{wood}}$ [%]), for specimens of all species incubated in soils with WHC$_{\text{soil}}$ of 60% and 90% compared to 30%. Furthermore, drier soils (MC$_{\text{soil}}$ of 30 and 70%WHC$_{\text{soil}}$) showed higher MLwood compared to wetter soils (95%WHC$_{\text{soil}}$). The second study (B) built on the first’s findings, and sought to isolate the effect of Tsoil and MC$_{\text{soil}}$ on the decay of European beech wood, while keeping WHC$_{\text{soil}}$ constant. The study used constant incubation temperature intervals (T$_{\text{soil}}$), 5–40 °C, and alternating intervals of 10/20, 10/30, and 20/30 °C. A general trend showed drier MCsoil (60%WHC$_{\text{soil}}$), and T$_{\text{soil}}$ of 20–40 °C, delivered high wood decay (ML$_{\text{wood}}$ > 20%). Higher MC$_{\text{soil}}$ (90%WHC$_{\text{soil}}$) and T$_{\text{soil}}$ of 5–10 °C, delivered low wood decay (ML$_{\text{wood}}$ < 5%). Alternating T$_{\text{soil}}$ generally delivered less ML$_{\text{wood}}$ compared to their mean constant T$_{\text{soil}}$ counterparts (15, 20, 25 °C). The results suggest that differences in wood species and inoculum potential (WHC$_{\text{soil}}$) between sites, as well as changes in MC$_{\text{soil}}$ and T$_{\text{soil}}$ attributed to daily and seasonal weather patterns can influence in-ground wood decay rate.
• Journal Article

#### N$_2$O isotope approaches for source partitioning of N$_2$O production and estimation of N$_2$O reduction – validation with the $^{15}$N gas-flux method in laboratory and field studies ﻿

Biogeosciences 2020; 17(22) p.5513-5537
The approaches based on natural abundance N$_2$O stable isotopes are often applied for the estimation of mixing proportions between various N$_2$O-producing pathways as well as for estimation of the extent of N$_2$O reduction to N$_2$. But such applications are associated with numerous uncertainties; hence, their limited accuracy needs to be considered. Here we present the first systematic validation of these methods for laboratory and field studies by applying the $^{15}$N gas-flux method as the reference approach. Besides applying dual-isotope plots for interpretation of N$_2$O isotopic data, for the first time we propose a three dimensional N2O isotopocule model based on Bayesian statistics to estimate the N$_2$O mixing proportions and reduction extent based simultaneously on three N$_2$O isotopic signatures (δ$^{15}$N, δ$^{15}$N$^{\text{SP}}$, and δ$^{18}$O). Determination of the mixing proportions of individual pathways with N2O isotopic approaches often appears imprecise, mainly due to imperfect isotopic separation of the particular pathways. Nevertheless, the estimation of N$_2$O reduction is much more robust, when applying an optimal calculation strategy, typically reaching an accuracy of N$_2$O residual fraction determination of about 0.15.
• Journal Article

#### Insulin signaling represents a gating mechanism between different memory phases in Drosophila larvae ﻿

PLOS Genetics 2020; 16(10) p.1-25: Art. e1009064
The ability to learn new skills and to store them as memory entities is one of the most impressive features of higher evolved organisms. However, not all memories are created equal; some are short-lived forms, and some are longer lasting. Formation of the latter is energetically costly and by the reason of restricted availability of food or fluctuations in energy expanses, efficient metabolic homeostasis modulating different needs like survival, growth, reproduction, or investment in longer lasting memories is crucial. Whilst equipped with cellular and molecular pre-requisites for formation of a protein synthesis dependent long-term memory (LTM), its existence in the larval stage of Drosophila remains elusive. Considering it from the viewpoint that larval brain structures are completely rebuilt during metamorphosis, and that this process depends completely on accumulated energy stores formed during the larval stage, investing in LTM represents an unnecessary expenditure. However, as an alternative, Drosophila larvae are equipped with the capacity to form a protein synthesis independent so-called larval anaesthesia resistant memory (lARM), which is consolidated in terms of being insensitive to cold-shock treatments. Motivated by the fact that LTM formation causes an increase in energy uptake in Drosophila adults, we tested the idea of whether an energy surplus can induce the formation of LTM in the larval stage. Suprisingly, increasing the metabolic state by feeding Drosophila larvae the disaccharide sucrose directly before aversive olfactory conditioning led to the formation of a protein synthesis dependent longer lasting memory. Moreover, formation of this memory component is accompanied by the suppression of lARM. We ascertained that insulin receptors (InRs) expressed in the mushroom body Kenyon cells suppresses the formation of lARM and induces the formation of a protein synthesis dependent longer lasting memory in Drosophila larvae. Given the numerical simplicity of the larval nervous system this work offers a unique prospect to study the impact of insulin signaling on the formation of protein synthesis dependent memories on a molecular level.
• Journal Article

#### Frameshift Variant in Novel Adenosine-A1-Receptor Homolog Associated With Bovine Spastic Syndrome/Late-Onset Bovine Spastic Paresis in Holstein Sires ﻿

Frontiers in Genetics 2020; 11 p.1-13: Art. 591794
Since their first description almost 100 years ago, bovine spastic paresis (BSP) and bovine spastic syndrome (BSS) are assumed to be inherited neuronal-progressive diseases in cattle. Affected animals are characterized by (frequent) spasms primarily located in the hind limbs, accompanied by severe pain symptoms and reduced vigor, thus initiating premature slaughter or euthanasia. Due to the late onset of BSP and BSS and the massively decreased lifespan of modern cattle, the importance of these diseases is underestimated. In the present study, BSP/BSS-affected German Holstein breeding sires from artificial insemination centers were collected and pedigree analysis, genome-wide association studies, whole genome resequencing, protein–protein interaction network analysis, and protein-homology modeling were performed to elucidate the genetic background. The analysis of 46 affected and 213 control cattle revealed four significantly associated positions on chromosome 15 (BTA15), i.e., AC_000172.1:g.83465449A>G (–log$_{10}$P = 19.17), AC_000172.1:g.81871849C>T (–log$_{10}$P = 8.31), AC_000172.1:g.81872621A>T (–log$_{10}$P = 6.81), and AC_000172.1:g.81872661G>C (–log$_{10}$P = 6.42). Two additional loci were significantly associated located on BTA8 and BTA19, i.e., AC_000165.1:g.71177788T>C and AC_000176.1:g.30140977T>G, respectively. Whole genome resequencing of five affected individuals and six unaffected relatives (two fathers, two mothers, a half sibling, and a full sibling) belonging to three different not directly related families was performed. After filtering, a homozygous loss of function variant was identified in the affected cattle, causing a frameshift in the so far unknown gene locus LOC100848076 encoding an adenosine-A1-receptor homolog. An allele frequency of the variant of 0.74 was determined in 3,093 samples of the 1000 Bull Genomes Project.
• Journal Article

#### Polymorphism of bis(1,3-benzothiazol-2-yl) trithiocarbonate ﻿

Acta Crystallographica Section E Crystallographic Communications 2020; 76(7) p.1126-1130
The polymorphism of the title compound, C15H8N2S5, is reported, in which the (syn,syn) and (syn,anti) conformers simultaneously crystallized from a chloroform solution. The complete molecule of the (syn,syn) form is generated by a crystallographic twofold axis. The geometries of both conformers are compared in detail, revealing no significant differences in bond lengths, despite different bond angles because of the conformational changes. Analysis of the intermolecular interactions, aided by Hirshfeld surfaces, shows distinctive SS and SN contacts only for the (syn,anti) conformer. Aromatic –-stacking interactions are found for both conformers, which occur for the (syn,anti) conformer between pairs of molecules, but are continuous stacks in the (syn,syn) conformer. Non merohedral twinning was found for the crystal of the (syn,anti) conformer used for data collection.
• Journal Article

#### Transgenerational effect of drug-mediated inhibition of LSD1 on eye pigment expression in Drosophila ﻿

BMC Ecology 2020; 20(1) p.1-12: Art. 62
Abstract Background The Drosophila melanogaster mutant white-mottled is a well-established model for position-effect variegation (PEV). Transposition of the euchromatic white gene into the vicinity of the pericentric heterochromatin caused variegated expression of white due to heterochromatin spreading. The establishment of the euchromatin-heterochromatin boundary and spreading of silencing is regulated by mutually exclusive histone modifications, i.e. the methylations of histone H3 at lysine 9 and lysine 4. Demethylation of H3K4, catalysed by lysine-specific demethylase LSD1, is required for subsequent methylation of H3K9 to establish heterochromatin. LSD1 is therefore essential for heterochromatin formation and spreading. We asked whether drug-mediated inhibition of LSD affects the expression of white and if this induced change can be transmitted to those generations that have never been exposed to the triggering signal, i.e. transgenerational epigenetic inheritance. Results We used the lysine-specific demethylase 1 (LSD1)-inhibitor Tranylcypromine to investigate its effect on eye colour expression in consecutive generations by feeding the parental and F1 generations of the Drosophila melanogaster mutant white-mottled. Quantitative Western blotting revealed that Tranylcypromine inhibits H3K4-demethylation both in vitro in S2 cells as well as in embryos when used as feeding additive. Eye colour expression in male flies was determined by optical measurement of pigment extracts and qRT-PCR of white gene expression. Flies raised in the presence of Tranylcypromine and its solvent DMSO showed increased eye pigment expression. Beyond that, eye pigment expression was also affected in consecutive generations including F3, which is the first generation without contact with the inhibitor. Conclusions Our results show that feeding of Tranylcypromine and DMSO caused desilencing of white in treated flies of generation F1. Consecutive generations, raised on standard food without further supplements, are also affected by the drug-induced alteration of histone modifications. Although eye pigment expression eventually returned to the basal state, the observed long-lasting effect points to a memory capacity of previous epigenomes. Furthermore, our results indicate that food compounds potentially affect chromatin modification and hence gene expression and that the alteration is putatively inherited not only parentally but transgenerationally.