Soil microbial community structures are shaped by agricultural systems revealing little temporal variation.

Many studies in soil microbial ecology are undertaken with a single sampling event, with the influence of temporal progression rarely being considered. Under field conditions, soil samples were taken from different agricultural systems; a sown grassland to maize rotation (MC), an intensively managed permanent grassland (INT), as well as extensively managed permanent grasslands with high (EXT_HP), low to sufficient (EXT_LP) and deficient available P (EXT_DP), six times throughout the 2017 growing season. Thus, this study aimed to determine if any differences in soil microbiome structures between both sharply contrasting (MC - INT - EXT), slightly differing (EXT_HP - EXT_DP) and quite similar (EXT_HP - EXT_LP and EXT_LP - EXT_DP) agricultural systems persist through changing growth conditions within the growing season. For both fungal and bacterial community structure, the influence of agricultural system (CV = 0.256, P < 0.001 and CV = 0.145, P < 0.01, respectively) was much greater than that of temporal progression (√CV = 0.065 and 0.042, respectively, both P < 0.001). Importantly, nearly all agricultural systems persistently harbored significantly distinct fungal community structures across each of the six sampling events (all at least P < 0.05). There were not as many pairwise differences in bacterial community structure between the agricultural systems, but some did persist (MC and EXT_HP âˆ¼ EXT_DP, all P < 0.001). Additionally, persistent indicator fungal OTUs (IndVal >0.7, P ≤ 0.05) associated to each agricultural system (except EXT_LP) were found in each of the six sampling events. These results highlight the temporal stability of pairwise differences in soil microbiome structures between established agricultural systems through changing plant growth conditions, even between those with a comparable management regime. This is a highly relevant finding in informing the sampling strategy of studies in soil microbial ecology as well as for designing efficient soil biodiversity monitoring systems.

Small-scale agricultural grassland management can affect soil fungal community structure as much as continental scale geographic patterns.

A European transect was established, ranging from Sweden to the Azores, to determine the relative influence of geographic factors and agricultural small-scale management on the grassland soil microbiome. Within each of five countries (factor 'Country'), which maximized a range of geographic factors, two differing growth condition regions (factor 'GCR') were selected: a favorable region with conditions allowing for high plant biomass production and a contrasting less favorable region with a markedly lower potential. Within each region, grasslands of contrasting management intensities (factor 'MI') were defined: intensive and extensive, from which soil samples were collected. Across the transect, 'MI' was a strong differentiator of fungal community structure, having a comparable effect to continental scale geographic factors ('Country'). 'MI' was also a highly significant driver of bacterial community structure, but 'Country' was clearly the stronger driver. For both, 'GCR' was the weakest driver. Also at the regional level, strong effects of MI occurred on various measures of the soil microbiome (i.e. OTU richness, management-associated indicator OTUs), though the effects were largely regional-specific. Our results illustrate the decisive influence of grassland MI on soil microbial community structure, over both regional and continental scales, and, thus, highlight the importance of preserving rare extensive grasslands.

Potential of legume-based grassland-livestock systems in Europe: a review.

European grassland-based livestock production systems face the challenge of producing more meat and milk to meet increasing world demands and to achieve this using fewer resources. Legumes offer great potential for achieving these objectives. They have numerous features that can act together at different stages in the soil-plant-animal-atmosphere system, and these are most effective in mixed swards with a legume proportion of 30-50%. The resulting benefits include reduced dependence on fossil energy and industrial N-fertilizer, lower quantities of harmful emissions to the environment (greenhouse gases and nitrate), lower production costs, higher productivity and increased protein self-sufficiency. Some legume species offer opportunities for improving animal health with less medication, due to the presence of bioactive secondary metabolites. In addition, legumes may offer an adaptation option to rising atmospheric CO2 concentrations and climate change. Legumes generate these benefits at the level of the managed land-area unit and also at the level of the final product unit. However, legumes suffer from some limitations, and suggestions are made for future research to exploit more fully the opportunities that legumes can offer. In conclusion, the development of legume-based grassland-livestock systems undoubtedly constitutes one of the pillars for more sustainable and competitive ruminant production systems, and it can be expected that forage legumes will become more important in the future.

Diversity-interaction modeling: estimating contributions of species identities and interactions to ecosystem function.

We develop a modeling framework that estimates the effects of species identity and diversity on ecosystem function and permits prediction of the diversity-function relationship across different types of community composition. Rather than just measure an overall effect of diversity, we separately estimate the contributions of different species interactions. This is especially important when both positive and negative interactions occur or where there are patterns in the interactions. Based on different biological assumptions, we can test for different patterns of interaction that correspond to the roles of evenness, functional groups, and functional redundancy. These more parsimonious descriptions can be especially useful in identifying general diversity-function relationships in communities with large numbers of species. We provide an example of the application of the modeling framework. These models describe community-level performance and thus do not require separate measurement of the performance of individual species. This flexible modeling approach can be tailored to test many hypotheses in biodiversity research and can suggest the interaction mechanisms that may be acting.

Plant enemy-derived elicitors increase the foliar tannin concentration of Onobrychis viciifolia without a trade-off to growth.

BACKGROUND AND AIMS: Molecular experiments suggest that the regulation of the biosynthesis of condensed tannin (CT) is sensitive to the presence of plant enemies. The enemy-specific response of CT concentrations to simulated attacks by pathogenic fungi, bacteria or herbivores was studied in Onobrychis viciifolia grown at four levels of nutrient availability. It was hypothesized that CT concentrations increase in response to an attack, and that constitutive and induced levels of CT are higher at low than at high nutrient availability. Investment in CT was also predicted to be negatively related to plant growth. METHODS: Recently discovered substances by which plants recognize their opponents (i.e. elicitors) were used to simulate attacks to Onobrychis viciifolia grown at 0.0027, 0.075, 0.67 or 2 mm phosphorus in the nutrient solution. KEY RESULTS: Relative growth rate and final biomass (P < 0.001) were highest at 0.67 mm of phosphorus. CT concentrations decreased with increasing phosphorus availability, from 94.9 to 69.0 mg g(-1) leaf dry weight (P < 0.001). Compared with unscathed plants, sterile mere mechanical wounding reduced tannin concentrations from 83.8 to 69.3 mg g(-1) leaf dry weight (P < 0.01). Local CT concentrations were higher when wounded leaves were additionally treated with fungal (+15.9 %), bacterial (+19.6 %) or insect (+31.0 %) elicitors (each elicitor; P < 0.05); however, only the insect elicitor (saliva of the lepidopteron Spodoptera littoralis) induced CT concentrations higher than those of unscathed leaves. CONCLUSIONS: CT concentrations were inducible in the vicinity of the wound but the level of induction was unrelated to the nutrient status of the plant. There was no evidence of a growth-defence trade-off. The inverse relationship between CT concentrations and nutrient availability appears to reflect passive growth dilution at high nutrient availability, rather than surplus CT production at low nutrient availability.

Chemotherapeutic strategies against Trypanosoma brucei: drug targets vs. drug targeting.

Trypanosoma brucei rhodesiense and T. b. gambiense are the causative agents of sleeping sickness, a fatal disease that affects 36 countries in sub-Saharan Africa. Nevertheless, only a handful of clinically useful drugs are available. These drugs suffer from severe side-effects. The situation is further aggravated by the alarming incidence of treatment failures in several sleeping sickness foci, apparently indicating the occurrence of drug-resistant trypanosomes. Because of these reasons, and since vaccination does not appear to be feasible due to the trypanosomes' ever changing coat of variable surface glycoproteins (VSGs), new drugs are needed urgently. The entry of Trypanosoma brucei into the post-genomic age raises hopes for the identification of novel kinds of drug targets and in turn new treatments for sleeping sickness. The pragmatic definition of a drug target is, a protein that is essential for the parasite and does not have homologues in the host. Such proteins are identified by comparing the predicted proteomes of T. brucei and Homo sapiens, then validated by large-scale gene disruption or gene silencing experiments in trypanosomes. Once all proteins that are essential and unique to the parasite are identified, inhibitors may be found by high-throughput screening. However powerful, this functional genomics approach is going to miss a number of attractive targets. Several current, successful parasiticides attack proteins that have close homologues in the human proteome. Drugs like DFMO or pyrimethamine inhibit parasite and host enzymes alike--a therapeutic window is opened only by subtle differences in the regulation of the targets, which cannot be recognized in silico. Working against the post-genomic approach is also the fact that essential proteins tend to be more highly conserved between species than non-essential ones. Here we advocate drug targeting, i.e. uptake or activation of a drug via parasite-specific pathways, as a chemotherapeutic strategy to selectively inhibit enzymes that have equally sensitive counterparts in the host. The T. brucei purine salvage machinery offers opportunities for both metabolic and transport-based targeting: unusual nucleoside and nucleobase permeases may be exploited for selective import, salvage enzymes for selective activation of purine antimetabolites.

Biomass allocation is an important determinant of the tannin concentration in growing plants.

BACKGROUND AND AIMS: Condensed tannins (CTs) in the diet affect consumers in a concentration-dependent manner. Because of their importance in plant defence against herbivores and pathogens as well as their potential application against gastrointestinal parasites of ruminants in agronomy, an understanding of the seasonal dynamics of CT concentrations during plant growth is essential. METHODS: Over a vegetation period, CT concentrations in leaves, stems and roots and the biomass proportions between these organs were investigated in Onobrychis viciifolia, Lotus corniculatus and Cichorium intybus. Based on the experimental data, a model has been suggested to predict CT concentrations in harvestable biomass of these species. KEY RESULTS: During the experiment, leaf mass fractions of plants decreased from 85, 64, 85 to 30, 18, 39 % d. wt in Onobrychis, Lotus and Cichorium, respectively, and proportions of stems and roots increased accordingly. While CT concentrations almost doubled in leaves in Onobrychis (from 52 to 86 mg g(-1) d. wt, P<0.001) and Lotus (from 25 to 54 mg g(-1) d. wt, P<0.001), they were stable at low levels in expanding leaves of Cichorium (5 mg g(-1) d. wt) and in stems and roots of all investigated species. Due to an inverse effect of the increasing CT concentrations in leaves and simultaneous dilution from increasing proportions of 'CT-poor' stems, CT concentrations in harvestable biomass were stable over time in all investigated species: 62, 26 and 5 mg g(-1) d. wt for Onobrychis, Lotus and Cichorium, respectively. CONCLUSIONS: As a consequence of the unequal distribution of tannins in different plant parts and due to the changing biomass proportions between them, various herbivores (e.g. a leaf-eating insect and a grazing ruminant) may find not only different concentrations of CT in their diets but also different CT dynamics during the season. For the prediction of seasonal variations of CT concentrations, biomass allocation and accumulation of none-CT plant material are likely to be as important predictors as the knowledge of CT synthesis and its regulation.

Schistocephalus cotti n. sp. (Cestoda: Pseudophyllidea) plerocercoids from bullheads Cottus gobio L. in an Arctic river in Finland, with a key to the plerocercoids of the Palaearctic species of the genus.

We compared plerocercoids of Schistocephalus Creplin, 1829 from Cottus gobio (n = 57) and Gasterosteus aculeatus f. semiarmatus (n = 45) from the River Utsjoki, Finland, taken only from single worm infections. Segment numbers in the two populations were distinct (G. aculeatus range 55-107, average 74 (SE 1.66), median 73; C. gobio range 122-189, average 146 (SE 1.78); median 144). The mean difference between populations, 71.47, t = 28.76 with 100 degrees of freedom, two-tailed p value <0.001, was considered extremely significant. Amplification of microsatellite loci that were originally designed for Schistocephalus from G. aculeatus was positive for all larvae from G. aculeatus (n = 20), whereas in no plerocercoids from C. gobio (n = 20) were any of the six microsatellites amplified, indicating that plerocercoids from G. aculeatus and C. gobio were two distinct genetic populations of Schistocephalus. The material from C. gobio is described as S. cotti n. sp. Plerocercoids of the Palaearctic species of Schistocephalus are identified as follows: S. nemachili Dubinina, 1959 with 228-235 or more segments, specific to Barbatula spp. (Balitoridae); S. pungitii Dubinina, 1959 with 62-92 (usually 70-80) segments, specific to Pungitius pungitius; S. solidus (Müller, 1776) in two forms, one in G. aculeatus f. leiurus and f. semiarmatus, with 48-100 (usually 65-75) segments, and the other in G. aculeatus f. trachurus, with 99-138 (usually 112-122) segments; and S. cotti n. sp. with 103-189 (usually 130-159) segments, probably specific to cottids. Nearctic Schistocephalus were not considered owing to the uncertain status of some North American records. Some other species of Schistocephalus of highly doubtful status were briefly noted. Cross-infection experiments and molecular studies are recommended to further elucidate the interrelationships between the various species of Schistocephalus.

Simultaneous hermaphrodites reproducing in pairs self-fertilize some of their eggs: an experimental test of predictions of mixed-mating and Hermaphrodite's Dilemma theory.

Theory predicts (1) that mixed-mating systems (i.e. reproduction through both selfing and outcrossing) should usually not evolve and (2) that reproducing simultaneous hermaphrodites should be in a conflict over the preferred sexual role (The Hermaphrodite's Dilemma). In an in vitro system with the endoparasitic cestode Schistocephalus solidus, a simultaneous hermaphrodite, we tested predictions of both the mixed-mating and the Hermaphrodite's Dilemma theory. Using microsatellite markers, we measured the proportion of selfed offspring and the total reproductive output of each worm within pairs varying in mean weight and weight difference. Worms produced more outbred offspring not only with increasing total weight of the pair, but also with decreasing weight difference between the two paired worms. These results suggest: (1) that this parasite species reproduces by mixed-mating, which may be maintained by stochastic density fluctuations in the definitive host and hence unpredictability of self reproduction and (2) reproductive conflict may prevent worm pairs from achieving an optimal intermediate selfing rate.

Arbuscular mycorrhiza infection enhances the growth response of Lolium perenne to elevated atmospheric pCO(2).

Elevated atmospheric pCO(2) increases the C-availability for plants and thus leads to a comparable increase in plant biomass production and nutrient demand. Arbuscular mycorrhizal fungi (AMF) are considered to play an important role in the nutrient uptake of plants as well as to be a significant C-sink. Therefore, an increased colonization of plant roots by AMF is expected under elevated atmospheric pCO(2). To test these hypotheses, Lolium perenne L. plants were grown from seeds in a growth chamber in pots containing a silica sand/soil mixture for 9 weeks with and without inoculation with Glomus intraradices (Schenck and Smith). The growth response of plants at two different levels of N fertilization (1.5 or 4.5 mM) combined with ambient (35 Pa) and elevated atmospheric pCO(2) (60 Pa) was compared. The inoculation with G. intraradices, the elevated atmospheric pCO(2) and the high N fertilization treatment all led to an increased plant biomass production of 16%, 20% and 49%, respectively. AMF colonization and high N fertilization increased the plant growth response to elevated atmospheric pCO(2); the plant growth response to high N fertilization was also increased by AMF colonization. The root/shoot ratio was reduced by high N fertilization or elevated atmospheric pCO(2), but was not affected by AMF colonization. The unchanged specific leaf area indicated that if AMF colonization represented an increased C-sink, this was fully covered by the plant. Elevated atmospheric pCO(2) strongly increased AMF colonization (60%) while the high N fertilization had a slightly negative effect. AMF colonization neither improved the N nor P nutrition status, but led to an improved total P uptake. The results underline the importance of AMF for the response of grassland ecosystems to elevated atmospheric pCO(2).

Force transducer design: a new approach combining nonlinear finite element analysis and robust design.

Designing a strain gage based force transducer is considered more of an art than an engineering process. Only general guidelines are available, and many trial and error iterations are needed to optimize the geometry and minimize the errors caused by nonlinear behavior of the structure. A new method, based on nonlinear finite element analysis and robust design principles, is proposed. A matrix of experiments considers relevant geometric and loading parameters. The behavior of the structure under different combinations of these parameters is determined by calculating the strain at different locations (suitable for strain gage installation) using finite element models. The nonlinear behavior of the transducer is identified by comparing the results of the nonlinear finite element analysis with those obtained using a linear finite element analysis. A signal-to-noise ratio is defined to quantify the nonlinearities and how the considered parameters affect them. An analysis of variance is employed to determine their relative influence. Based on the results of the statistical analysis, it is possible to identify the best value for each geometric parameter that would reduce, if not eliminate the nonlinearities. Once these optimal geometric parameters are chosen, a prototype can be built, instrumented with strain gages, and tested, to validate the obtained design. To illustrate this new proposed methodology, and appreciate its advantages over current practice of designing a force transducer, an example of the step-by-step procedure is illustrated considering a thin-wall cylindrical transducer.

Evidence that P deficiency induces N feedback regulation of symbiotic N2 fixation in white clover (Trifolium repens L.).

Trifolium repens L. was grown to test the following hypotheses: when P is deficient (i) N2 fixation decreases as a result of the plant's adaptation to the low N demand, regulated by an N feedback mechanism, and (ii) the decrease in the photosynthetic capacity of the leaves does not limit N2 fixation. Severe P deficiency prevented nodulation or stopped nodule growth when the P deficiency occurred after the plants had formed nodules. At low P, the proportion of whole-plant-N derived from symbiotic N2 fixation decreased, whereas specific N2 fixation increased and compensated partially for poor nodulation. Leaf photosynthesis was reduced under P deficiency due to low Vc,max and Jmax. Poor growth or poor performance of the nodules was not due to C limitation, because (i) the improved photosynthetic performance at elevated pCO2 had no effect on the growth and functioning of the nodules, (ii) starch accumulated in the leaves, particularly under elevated pCO2, and (iii) the concentration of WSC in the nodules was highest under P deficiency. Under severe P deficiency, the concentrations of whole-plant-N and leaf-N were the highest, indicating that the assimilation of N exceeded the amount of N required by the plant for growth. This was clearly demonstrated by a strong increase in asparagine concentrations in the roots and nodules under low P supply. This indicates that nodulation and the proportion of N derived from symbiotic N2 fixation are down-regulated by an N feedback mechanism.

Long-term responsiveness to free air CO2 enrichment of functional types, species and genotypes of plants from fertile permanent grassland.

To test inter- and intraspecific variability in the responsiveness to elevated CO2, 9-14 different genotypes of each of 12 perennial species from fertile permanent grassland were grown in Lolium perenne swards under ambient (35 Pa) and elevated (60 Pa) atmospheric partial pressure of CO2 (pCO2) for 3 years in a free air carbon dioxide enrichment (FACE) experiment. The plant species were grouped according to their functional types: grasses (L. perenne, L. multiflorum, Arrhenatherum elatius, Dactylis glomerata, Festuca pratensis, Holcus lanatus, Trisetum flavescens), non-legume dicots (Rumex obtusifolius, R. acetosa, Ranunculus friesianus), and legumes (Trifolium repens, T. pratense). Yield (above a cutting height of 4.5 cm) was measured three times per year. The results were as follow. (1) There were highly significant differences in the responsiveness to elevated pCO2 between the three functional types; legumes showed the strongest and grasses the weakest yield increase at elevated pCO2. (2) There were differences in the temporal development of responsiveness to elevated pCO2 among the functional types. The responsiveness of the legumes declined from the first to the second year, while the responsiveness of the non-legume dicots increased over the 3 years. During the growing season, the grasses and the non-legume dicots showed the strongest response to elevated pCO2 during reproductive growth in the spring. (3) There were no significant genotypic differences in responsiveness to elevated pCO2. Our results suggest that, due to interspecific differences in the responsiveness to elevated pCO2, the species proportion within fertile temperate grassland may change if the increase in pCO2 continues. Due to the temporal differences in the responsiveness to elevated pCO2 among species, complex effects of elevated pCO2 on competitive interactions in mixed swards must be expected. The existence of genotypic variability in the responsiveness to elevated pCO2, on which selection could act, was not found under our experimental conditions.

Does nitrogen nutrition restrict the CO2 response of fertile grassland lacking legumes?

The extent of the response of plant growth to atmospheric CO2 enrichment depends on the availability of resources other than CO2. An important growth-limiting resource under field conditions is nitrogen (N). N may, therefore, influence the CO2 response of plants. The effect of elevated CO2 (60 Pa) partial pressure (pCO2) on the N nutrition of field-grown Lolium perenne swards, cultivated alone or in association with Trifolium repens, was investigated using free air carbon dioxide enrichment (FACE) technology over 3 years. The established grassland ecosystems were treated with two N fertilization levels and were defoliated at two frequencies. Under elevated pCO2, the above-ground plant material of the L. perenne monoculture showed a consistent and significant decline in N concentration which, in general, led to a lower total annual N yield. Despite the decline in the critical N concentration (minimum N concentration required for non-N-limited biomass production) under elevated pCO2, the index of N nutrition (ratio of actual N concentration and critical N concentration) was lower under elevated pCO2 than under ambient pCO2 in frequently defoliated L. perenne monocultures. Thus, we suggest that reduced N yield under elevated pCO2 was evoked indirectly by a reduction of plant-available N. For L. perenne grown in association with T. repens and exposed to elevated pCO2, there was an increase in the contribution of symbiotically fixed N to the total N yield of the grass. This can be explained by an increased apparent transfer of N from the associated N2-fixing legume species to the non-fixing grass. The total annual N yield of the mixed grass/legume swards increased under elevated pCO2. All the additional N yielded was due to symbiotically fixed N. Through the presence of an N2-fixing plant species more symbiotically fixed N was introduced into the system and consequently helped to overcome N limitation under elevated pCO2.

Stimulation of Symbiotic N2 Fixation in Trifolium repens L. under Elevated Atmospheric pCO2 in a Grassland Ecosystem.

Symbiotic N2 fixation is one of the main processes that introduces N into terrestrial ecosystems. As such, it may be crucial for the sequestration of the extra C available in a world of continuously increasing atmospheric CO2 partial pressure (pCO2). The effect of elevated pCO2 (60 Pa) on symbiotic N2 fixation (15N-isotope dilution method) was investigated using Free-Air-CO2-Enrichment technology over a period of 3 years. Trifolium repens was cultivated either alone or together with Lolium perenne (a nonfixing reference crop) in mixed swards. Two different N fertilization levels and defoliation frequencies were applied. The total N yield increased consistently and the percentage of plant N derived from symbiotic N2 fixation increased significantly in T. repens under elevated pCO2. All additionally assimilated N was derived from symbiotic N2 fixation, not from the soil. In the mixtures exposed to elevated pCO2, an increased amount of symbiotically fixed N (+7.8, 8.2, and 6.2 g m-2 a-1 in 1993, 1994, and 1995, respectively) was introduced into the system. Increased N2 fixation is a competitive advantage for T. repens in mixed swards with pasture grasses and may be a crucial factor in maintaining the C:N ratio in the ecosystem as a whole.

Distribution of acid ribonuclease and other enzymes in stratified Acetabularia.

A method for producing stratified and viable subcells, consisting of segments of the stalk, has been worked out in Acetabularia mediterranea (LmX). Centrifugation at low speed results in the formation of a cytoplasmic moiety at the centrifugal pole which contains ca. 90% of the aldolase activity that is localized in the chloroplasts and in the cytosol. The vacuolar moiety, which occupies 80-92% of the cell volume, stains with neutral red, i.e. is viable. It contains ca. 76% of the total RNase and 72% of the total phosphotase activity. It is concluded that a large proportion of these hydrolase activities is present in the vacuole.