Nutrient concentrations and fibre contents of plant community biomass reflect species richness patterns along a broad range of land-use intensities among agricultural grasslands

Understanding changes in biodiversity in agricultural landscapes in relation to land-use type and intensity is a major issue in current ecological research. In this context nutrient enrichment has been identified as a key mechanism inducing species loss in Central European grassland ecosystems. At the same time, insights into the linkage between agricultural land use and plant nutrient status are largely missing. So far, studies on the relationship between chemical composition of plant community biomass and biodiversity have mainly been restricted to wetlands and all these studies neglected the effects of land use. Therefore, we analyzed aboveground biomass of 145 grassland plots covering a gradient of land-use intensities in three regions across Germany. In particular, we explored relationships between vascular plant species richness and nutrient concentrations as well as fibre contents (neutral and acid detergent fibre and lignin) in the aboveground community biomass.We found the concentrations of several nutrients in the biomass to be closely linked to plant species richness and land use. Whereas phosphorus concentrations increased with land-use intensity and decreased with plant species richness, nitrogen and potassium concentrations showed less clear patterns. Fibre fractions were negatively related to nutrient concentrations in biomass, but hardly to land-use measures and species richness. Only high lignin contents were positively associated with species richness of grasslands. The N:P ratio was strongly positively related to species richness and even more so to the number of endangered plant species, indicating a higher persistence of endangered species under P (co-)limited conditions. Therefore, we stress the importance of low P supply for species-rich grasslands and suggest the N:P ratio in community biomass to be a useful proxy of the conservation value of agriculturally used grasslands.     

Organic vs. Conventional Grassland Management: Do 15N and 13C Isotopic Signatures of Hay and Soil Samples Differ?

Distinguishing organic and conventional products is a major issue of food security and authenticity. Previous studies successfully used stable isotopes to separate organic and conventional products, but up to now, this approach was not tested for organic grassland hay and soil. Moreover, isotopic abundances could be a powerful tool to elucidate differences in ecosystem functioning and driving mechanisms of element cycling in organic and conventional management systems. Here, we studied the δ¹⁵N and δ¹³C isotopic composition of soil and hay samples of 21 organic and 34 conventional grasslands in two German regions. We also used Δδ¹⁵N (δ¹⁵N plant - δ¹⁵N soil) to characterize nitrogen dynamics. In order to detect temporal trends, isotopic abundances in organic grasslands were related to the time since certification. Furthermore, discriminant analysis was used to test whether the respective management type can be deduced from observed isotopic abundances.Isotopic analyses revealed no significant differences in δ¹³C in hay and δ¹⁵N in both soil and hay between management types, but showed that δ¹³C abundances were significantly lower in soil of organic compared to conventional grasslands. Δδ¹⁵N values implied that management types did not substantially differ in nitrogen cycling. Only δ¹³C in soil and hay showed significant negative relationships with the time since certification. Thus, our result suggest that organic grasslands suffered less from drought stress compared to conventional grasslands most likely due to a benefit of higher plant species richness, as previously shown by manipulative biodiversity experiments. Finally, it was possible to correctly classify about two third of the samples according to their management using isotopic abundances in soil and hay. However, as more than half of the organic samples were incorrectly classified, we infer that more research is needed to improve this approach before it can be efficiently used in practice.     

Naphthoquinone Derivatives Exert Their Antitrypanosomal Activity via a Multi-Target Mechanism

Background and Methodology

Recently, we reported on a new class of naphthoquinone derivatives showing a promising anti-trypanosomatid profile in cell-based experiments. The lead of this series (B6, 2-phenoxy-1,4-naphthoquinone) showed an ED₅₀ of 80 nM against Trypanosoma brucei rhodesiense, and a selectivity index of 74 with respect to mammalian cells. A multitarget profile for this compound is easily conceivable, because quinones, as natural products, serve plants as potent defense chemicals with an intrinsic multifunctional mechanism of action. To disclose such a multitarget profile of B6, we exploited a chemical proteomics approach.

Principal Findings

A functionalized congener of B6 was immobilized on a solid matrix and used to isolate target proteins from Trypanosoma brucei lysates. Mass analysis delivered two enzymes, i.e. glycosomal glycerol kinase and glycosomal glyceraldehyde-3-phosphate dehydrogenase, as potential molecular targets for B6. Both enzymes were recombinantly expressed and purified, and used for chemical validation. Indeed, B6 was able to inhibit both enzymes with IC₅₀ values in the micromolar range. The multifunctional profile was further characterized in experiments using permeabilized Trypanosoma brucei cells and mitochondrial cell fractions. It turned out that B6 was also able to generate oxygen radicals, a mechanism that may additionally contribute to its observed potent trypanocidal activity.

Conclusions and Significance

Overall, B6 showed a multitarget mechanism of action, which provides a molecular explanation of its promising anti-trypanosomatid activity. Furthermore, the forward chemical genetics approach here applied may be viable in the molecular characterization of novel multitarget ligands.     

Francisella tularensis: No Evidence for Transovarial Transmission in the Tularemia Tick Vectors Dermacentor reticulatus and Ixodes ricinus

Background

Tularemia is a zoonosis caused by the Francisella tularensis, a highly infectious Gram-negative coccobacillus. Due to easy dissemination, multiple routes of infection, high environmental contamination and morbidity and mortality rates, Francisella is considered a potential bioterrorism threat and classified as a category A select agent by the CDC. Tick bites are among the most prevalent modes of transmission, and ticks have been indicated as a possible reservoir, although their reservoir competence has yet to be defined. Tick-borne transmission of F. tularensis was recognized in 1923, and transstadial transmission has been demonstrated in several tick species. Studies on transovarial transmission, however, have reported conflicting results.

Objective

The aim of this study was to evaluate the role of ticks as reservoirs for Francisella, assessing the transovarial transmission of F. tularensis subsp. holarctica in ticks, using experimentally-infected females of Dermacentor reticulatus and Ixodes ricinus.

Results

Transmission electron microscopy and fluorescence in situ hybridization showed F. tularensis within oocytes. However, cultures and bioassays of eggs and larvae were negative; in addition, microscopy techniques revealed bacterial degeneration/death in the oocytes.

Conclusions

These results suggest that bacterial death might occur in oocytes, preventing the transovarial transmission of Francisella. We can speculate that Francisella does not have a defined reservoir, but that rather various biological niches (e.g. ticks, rodents), that allow the bacterium to persist in the environment. Our results, suggesting that ticks are not competent for the bacterium vertical transmission, are congruent with this view.     

Fern and bryophyte endozoochory by slugs

Endozoochory plays a prominent role for the dispersal of seed plants, and dispersal vectors are well known. However, for taxa such as ferns and bryophytes, endozoochory has only been suggested anecdotally but never tested in controlled experiments. We fed fertile leaflets of three ferns and capsules of four bryophyte species to three slug species. We found that, overall, spores germinated from slug feces in 57.3 % of all 89 fern and in 51.3 % of all 117 bryophyte samples, showing that the spores survived gut passage of slugs. Moreover, the number of samples within which spores successfully germinated did not differ among plant species but varied strongly among slug species. This opens new ecological perspectives suggesting that fern and bryophyte endozoochory by gastropods is a so-far-overlooked mode of dispersal, which might increase local population sizes of these taxa by spore deposition on suitable substrates.     

Interactive effects of mycorrhizae and a root hemiparasite on plant community productivity and diversity

Plant communities can be affected both by arbuscular mycorrhizal fungi (AMF) and hemiparasitic plants. However, little is known about the interactive effects of these two biotic factors on the productivity and diversity of plant communities. To address this question, we set up a greenhouse study in which different AMF inocula and a hemiparasitic plant (Rhinanthus minor) were added to experimental grassland communities in a fully factorial design. In addition, single plants of each species in the grassland community were grown with the same treatments to distinguish direct AMF effects from indirect effects via plant competition. We found that AMF changed plant community structure by influencing the plant species differently. At the community level, AMF decreased the productivity by 15–24%, depending on the particular AMF treatment, mainly because two dominant species, Holcus lanatus and Plantago lanceolata, showed a negative mycorrhizal dependency. Concomitantly, plant diversity increased due to AMF inoculation and was highest in the treatment with a combination of two commercial AM strains. AMF had a positive effect on growth of the hemiparasite, and thereby induced a negative impact of the hemiparasite on host plant biomass which was not found in non-inoculated communities. However, the hemiparasite did not increase plant diversity. Our results highlight the importance of interactions with soil microbes for plant community structure and that these indirect effects can vary among AMF treatments. We conclude that mutualistic interactions with AMF, but not antagonistic interactions with a root hemiparasite, promote plant diversity in this grassland community. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Implementing large-scale and long-term functional biodiversity research: The Biodiversity Exploratories

Functional biodiversity research explores drivers and functional consequences of biodiversity changes. Land use change is a major driver of changes of biodiversity and of biogeochemical and biological ecosystem processes and services. However, land use effects on genetic and species diversity are well documented only for a few taxa and trophic networks. We hardly know how different components of biodiversity and their responses to land use change are interrelated and very little about the simultaneous, and interacting, effects of land use on multiple ecosystem processes and services. Moreover, we do not know to what extent land use effects on ecosystem processes and services are mediated by biodiversity change. Thus, overall goals are on the one hand to understand the effects of land use on biodiversity, and on the other to understand the modifying role of biodiversity change for land-use effects on ecosystem processes, including biogeochemical cycles. To comprehensively address these important questions, we recently established a new large-scale and long-term project for functional biodiversity, the Biodiversity Exploratories (www.biodiversity-exploratories.de). They comprise a hierarchical set of standardized field plots in three different regions of Germany covering manifold management types and intensities in grasslands and forests. They serve as a joint research platform for currently 40 projects involving over 300 people studying various aspects of the relationships between land use, biodiversity and ecosystem processes through monitoring, comparative observation and experiments. We introduce guiding questions, concept and design of the Biodiversity Exploratories – including main aspects of selection and implementation of field plots and project structure – and we discuss the significance of this approach for further functional biodiversity research. This includes the crucial relevance of a common study design encompassing variation in both drivers and outcomes of biodiversity change and ecosystem processes, the interdisciplinary integration of biodiversity and ecosystem researchers, the training of a new generation of integrative biodiversity researchers, and the stimulation of functional biodiversity research in real landscape contexts, in Germany and elsewhere.     

Structure-based optimization of cephalothin-analogue boronic acids as beta-lactamase inhibitors

Boronic acids have proved to be promising selective inhibitors of beta-lactamases, acting as transition state analogues. Starting from a previously described nanomolar inhibitor of AmpC beta-lactamase, three new inhibitors were designed to gain interactions with highly conserved residues, such as Asn343, and to bind more tightly to the enzyme. Among these, one was obtained by stereoselective synthesis and succeeded in placing its anionic group into the carboxylate binding site of the enzyme, as revealed by X-ray crystallography of the complex inhibitor/AmpC. Nevertheless, it failed at improving affinity, when compared to the lead from which it was derived. The origins of this structural and energetic discrepancy are discussed.     

Structure and dynamics of CTX-M enzymes reveal insights into substrate accommodation by extended-spectrum beta-lactamases

Oxyimino-cephalosporin antibiotics, such as ceftazidime, escape the hydrolytic activity of most bacterial β-lactamases. Their widespread use prompted the emergence of the extended-spectrum β-lactamases CTX-Ms, which have become highly prevalent. The C7 β-amino thiazol-oxyimino-amide side chain of ceftazidime has a protective effect against most CTX-M β-lactamases. However, Asp240Gly CTX-M derivatives demonstrate enhanced hydrolytic activity against this compound. In this work, we present the crystallographic structures of Asp240Gly-harboring enzyme CTX-M-16 in complex with ceftazidime-like glycylboronic acid (resolution 1.80 Å) and molecular dynamics simulations of the corresponding acyl-enzyme complex. These experiments revealed breathing motions of CTX-M enzymes and the role of the substitution Asp240Gly in the accommodation of ceftazidime. The substitution Asp240Gly resulted in insertion of the C7β side chain of ceftazidime deep in the catalytic pocket and orchestrated motions of the active serine Ser70, the β3 strand and the omega loop, which favored the key interactions of the residues 237 and 235 with ceftazidime.     

Structures of ceftazidime and its transition-state analogue in complex with AmpC beta-lactamase: implications for resistance mutations and inhibitor design

Third-generation cephalosporins are widely used beta-lactam antibiotics that resist hydrolysis by beta-lactamases. Recently, mutant beta-lactamases that rapidly inactivate these drugs have emerged. To investigate why third-generation cephalosporins are relatively stable to wild-type class C beta-lactamases and how mutant enzymes might overcome this, the structures of the class C beta-lactamase AmpC in complex with the third-generation cephalosporin ceftazidime and with a transition-state analogue of ceftazidime were determined by X-ray crystallography to 2.0 and 2.3 A resolution, respectively. Comparison of the acyl-enzyme structures of ceftazidime and loracarbef, a beta-lactam substrate, reveals that the conformation of ceftazidime in the active site differs from that of substrates. Comparison of the structures of the acyl-enzyme intermediate and the transition-state analogue suggests that ceftazidime blocks formation of the tetrahedral transition state, explaining why it is an inhibitor of AmpC. Ceftazidime cannot adopt a conformation competent for catalysis due to steric clashes that would occur with conserved residues Val211 and Tyr221. The X-ray crystal structure of the mutant beta-lactamase GC1, which has improved activity against third-generation cephalosporins, suggests that a tandem tripeptide insertion in the Omega loop, which contains Val211, has caused a shift of this residue and also of Tyr221 that would allow ceftazidime and other third-generation cephalosporins to adopt a more catalytically competent conformation. These structural differences may explain the extended spectrum activity of GC1 against this class of cephalosporins. In addition, the complexed structure of the transition-state analogue inhibitor (K(i) 20 nM) with AmpC reveals potential opportunities for further inhibitor design.