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Concentrate-rich starter feeds are commonly fed to dairy calves to stimulate early solid feed intake and growth performance; yet, starter feeds lacking in forage fiber may jeopardize gut development. This research primarily aimed to test a complete or partial replacement of concentrates with hay of different qualities in the starter feed on nutrient intake, growth performance, apparent total-tract digestibility (ATTD) of nutrients, and blood metabolites in dairy calves. Immediately after birth, 40 Holstein Friesian calves were randomly allocated to 1 of 4 starter diets, which differed in hay quality and concentrate inclusion [MQH = 100% medium-quality hay, 9.4 MJ of metabolizable energy (ME), 149 g of crude protein (CP), 522 g of neutral detergent fiber (NDF)/kg of dry matter (DM); HQH = 100% high-quality hay, 11.2 MJ of ME, 210 g of CP, 455 g of NDF/kg of DM; MQH+C = 30% medium-quality hay + 70% starter concentrate; HQH+C = 30% high-quality hay + 70% starter concentrate]. The concentrate consisted mainly of grains, oilseeds, and mineral supplements (13.5 MJ of ME, 193 g of CP, 204 g of NDF/kg of DM). Calves were used in the experiment from d 1 to 99 of life. During the first 4 wk, all calves were fed acidified whole milk ad libitum, and afterward they were gradually weaned from wk 5 to 12. Calves had ad libitum access to their starter diets and water throughout the experiment. Milk, water, and solid feed intake was recorded daily, live weight was measured once a week, and blood samples were collected on d 1, 3, 7, 21, 49, 77, and 91 and analyzed for selected metabolites. The ATTD was measured in wk 14 of life. Total DM intake and daily weight gain of calves were not affected by the starter feed during the first 8 wk of life. However, from wk 9 to 14, calves fed the MQH diet had lower DM, ME, and CP intake and gained less weight than calves from the other experimental groups. Feeding the HQH diet resulted in similar CP and ME intake and growth performance compared with calves receiving diets containing concentrates. Furthermore, feeding the HQH diet improved the ATTD of NDF, resulting in similar ATTD of organic matter with the HQH+C and MQH+C groups. Interestingly, calves fed the HQH+C diet showed less sorting for concentrate, compared with the MQH+C group. Concentration of blood metabolites, including glucose, lactate, insulin, nonesterified fatty acids, triglycerides, and total protein, did not differ after the first week of life. However, serum β-hydroxybutyrate was higher in calves fed the HQH diet starting from wk 11. Both groups fed the hay-only diets maintained higher cholesterol levels after weaning compared with the groups fed hay-concentrate mixtures. In conclusion, feeding high-quality hay can fully replace starter concentrates in the feeding of dairy calves without adverse effects on performance during the rearing period, while increasing forage fiber intake and utilization, which enhanced ruminal ketogenesis and cholesterogenesis around weaning. Further research is needed to evaluate long-term effects of feeding high-quality hay on health and development of dairy calves, especially in terms of the observed improvements in ruminal ketogenesis and cholesterogenesis around weaning.

International audience; Germplasm from perennial ryegrass (Lolium perenne L.) natural populations is useful for breeding because of its adaptation to a wide range of climates. Climate-adaptive genes can be detected from associations between genotype, phenotype and climate but an integrated framework for the analysis of these three sources of information is lacking. We used two approaches to identify adaptive loci in perennial ryegrass and their effect on phenotypic traits. First, we combined Genome-Environment Association (GEA) and GWAS analyses. Then, we implemented a new test based on a Canonical Correlation Analysis (CANCOR) to detect adaptive loci. Furthermore, we improved the previous perennial ryegrass gene set by de novo gene prediction and functional annotation of 39,967 genes. GEA-GWAS revealed eight outlier loci associated with both environmental variables and phenotypic traits. CANCOR retrieved 633 outlier loci associated with two climatic gradients, characterized by cold-dry winter versus mild-wet winter and long rainy season versus long summer, and pointed out traits putatively conferring adaptation at the extremes of these gradients. Our CANCOR test also revealed the presence of both polygenic and oligogenic climatic adaptations. Our gene annotation revealed that 374 of the CANCOR outlier loci were positioned within or close to a gene. Co-association networks of outlier loci revealed a potential utility of CANCOR for investigating the interaction of genes involved in polygenic adaptations. The CANCOR test provides an integrated framework to analyse adaptive genomic diversity and phenotypic responses to environmental selection pressures that could be used to facilitate the adaptation of plant species to climate change.

This article comments on: Ueno K, Sonoda T, Yoshida M, Shiomi N, Onodera S. 2018. Purification, characterization, and functional analysis of a novel 6G&1-FEH mainly hydrolyzing neokestose from asparagus. Journal of Experimental Botany 69, 4295–4308. ispartof: JOURNAL OF EXPERIMENTAL BOTANY vol:69 issue:18 pages:4227-4231 ispartof: location:England status: published

Perennial iyegrass (Lolium perenne L) is a widely cultivated cool-season grass species because of its high quality for forage and turf. Susceptibility to freezing damage limits its further use in temperate zones. The objective of this study was to identify candidate genes significantly associated with winter survival and spring regrowth in a global collection of 192 perennial ryegrass accessions. Significant differences in winter survival (WS), percentage of canopy green cover (CGC), chlorophyll index (Chl), and normalized difference vegetation index (NDVI) were found among accessions. After controlling population structure, LpLEA3 encoding a late embryogenesis abundant group 3 protein and LpCAT encoding a catalase were associated with CGC and Chl, while LpMnSOD encoding a magnesium superoxide dismutase and LpChl Cu-ZnSOD encoding a chlorophyll copper zinc superoxide dismutase were associated with NDVI or Chl. Significant association was also discovered between C-repeat binding factor LpCBF1b and WS. Three sequence variations identified in LpCAT, LpMnSOD, and LpChl Cu-ZnSOD were synonymous substitutions, whereas one pair of adjacent single nucleotide polymorphisms (SNPs) in LpLEA3 and one SNP in LpCBF1b resulted in amino acid change. The results demonstrated that allelic variation in LpLEA3 and LpCBF1b was closely related to winter survival and spring regrowth in perennial ryegrass. (C) 2015 Elsevier Ireland Ltd. All rights reserved.

SUMMARYSulphur (S) is one of the six main macroelements required to sustain the growth of plants. Sources include soil, fertilizer and atmospheric deposition, which has been reduced by 85% over the last three decades. Risks of S deficiencies are now recognized in high S-demanding species such as Brassica napus L. With the aims of evaluating the risk of excessive or insufficient fertilization and identifying robust relationships that may be used as plant S status indicators, 57 commercial crops of oilseed rape were selected among contrasting soils and along a rainfall gradient that may affect soil S availability. Cultivation practices were investigated and the S and nitrogen (N) concentrations of soil, senescing leaves, stems and seeds were analysed. Despite an excessive organic N supply and large variation in S supply (from 0 to 112 kg S/ha), principal component analysis using 43 parameters indicated that seed yield was poorly related to N and S fertilization rates. While the N and protein-N concentrations in seeds were inversely related to oil and glucosinolate concentrations, they were linked to S and sulphate (SO42−) accumulation in the seeds. Sulphate concentrations in senescing leaves, stems or seeds could be deduced from total S concentrations, as they were positively and highly correlated. Sulphate accounted for on average 0·69 of total S in senescing leaves with minimum and maximum values of 0·007 and 0·94, which revealed conditions of limited and excess supply of S, respectively. This high variation of SO42− concentration in leaves can be interpreted as the result of its mobilization triggered by S deficiency, but cannot be used alone as an indicator of plant S status. A comparison with plants grown in controlled conditions under different S supplies suggests that the intensity of S starvation affects N metabolism, leading to NO3− (nitrate) accumulation. It is further suggested that dual evaluation of SO42− and NO3− concentrations in senescing leaves could be used at the vegetative stage as a field indicator to adjust S fertilization.

International audience; Sucrose transport between source and sink tissues is supposed to be a key-step for an efficient regrowth of perennial rye-grass after defoliation and might be altered by light conditions. we assessed the effect of different light regimes (high vs low light applied before or after defoliation) on growth, fructans and sucrose mobilization, as well as on sucrose transporter expression during 14 days of regrowth. Our results reported that defoliation led to a mobilization of C reserves (first sucrose and then fructans), which was parallel to an induction of LpSUT1 sucrose transporter expression in source and sink tissues (i.e. leaf sheaths and elongating leaf bases, respectively) irrespective to light conditions. Light regime (high or low light) had little effects on regrowth and on C reserves mobilization during the first 48 h of regrowth after defoliation. Thereafter, low light conditions, delaying the recovery of photosynthetic capacities, had a negative effect on C reserves re-accumulation (especially sucrose). Surprisingly, high light did not enhance sucrose transporter expression. Indeed, while light conditions had no effect on LpSUT1 expression, LpSUT2 transcripts levels were enhanced for low light grown plants. These results indicate that two sucrose transporter currently identified in Lolium perenne L are differentially regulated by light and sucrose. (C) 2012 Elsevier Masson SAS. All rights reserved.

International audience; The ability of the free ion activity model (FIAM), the terrestrial biotic ligand model (TBLM), the diffusive gradients in thin films (DGT) technique and a plant-based biotest, the RHIZOtest, to predict root copper (Cu) concentration in field-grown durum wheat (Triticum turgidum durum L) was assessed on 44 soils varying in pH (3.9-7.8) and total Cu (32-184 mg kg(-1)). None of the methods adequately predicted root Cu concentration, which was mainly correlated with total soil Cu. Results from DGT measurements and even more so FIAM prediction were negatively correlated with soil pH and over-estimated root Cu concentration in acidic soils. TBLM implementation improved numerically FIAM prediction but still failed to predict adequately root Cu concentration as the TBLM formalism did not considered the rhizosphere alkalisation as observed in situ. In contrast, RHIZOtest measurements accounted for rhizosphere alkalisation and were mainly correlated with total soil Cu.

Fructosyltransferases (FTs) synthesize fructans, fructose polymers accumulating in economically important cool-season grasses and cereals. FTs might be crucial for plant survival under stress conditions in species in which fructans represent the major form of reserve carbohydrate, such as perennial ryegrass (Lolium perenne). Two FT types can be distinguished: those using sucrose (S-type enzymes: sucrose: sucrose 1-fructosyltransferase [1-SST], sucrose: fructan 6-fructosyltransferase) and those using fructans (F-type enzymes: fructan: fructan 1-fructosyltransferase [1-FFT], fructan: fructan 6G-fructosyltransferase [6G-FFT]) as preferential donor substrate. Here, we report, to our knowledge for the first time, the transformation of an F-type enzyme (6G-FFT/1-FFT) into an S-type enzyme (1-SST) using perennial ryegrass 6G-FFT/1-FFT (Lp6G-FFT/1-FFT) and 1-SST (Lp1-SST) as model enzymes. This transformation was accomplished by mutating three amino acids (N340D, W343R, and S415N) in the vicinity of the active site of Lp6G-FFT/1-FFT. In addition, effects of each amino acid mutation alone or in combination have been studied. Our results strongly suggest that the amino acid at position 343 (tryptophan or arginine) can greatly determine the donor substrate characteristics by influencing the position of the amino acid at position 340. Moreover, the presence of arginine-343 negatively affects the formation of neofructan-type linkages. The results are compared with recent findings on donor substrate selectivity within the group of plant cell wall invertases and fructan exohydrolases. Taken together, these insights contribute to our knowledge of structure/function relationships within plant family 32 glycosyl hydrolases and open the way to the production of tailor-made fructans on a larger scale. ispartof: Plant physiology vol:149 issue:1 pages:327-339 ispartof: location:United States status: published

Environmental pollution by heavy metals has become a serious problem in the world. Phytoextraction, which is one of the plant-based technologies, has attracted the most attention for the bioremediation of soils polluted with these contaminants. The aim of this study was to determine whether the multiple-tolerant bacterium. Brevibacterium casei MH8a isolated from the heavy metal-contaminated rhizosphere soil of Sinapis alba L., is able to promote plant growth and enhance Cd, Zn, and Cu uptake by white mustard under laboratory conditions. Additionally, the ability of the rifampicin-resistant spontaneous mutant of MH8a to colonize plant tissues and its mechanisms of plant growth promotion were also examined. In order to assess the ecological consequences of bioaugmentation on autochthonous bacteria, the phospholipid fatty acid (PLFA) analysis was used. The MH8a strain exhibited the ability to produce ammonia, 1-amino-cyclopropane-1-carboxylic acid deaminase, indole 3-acetic acid and HCN but was not able to solubilize inorganic phosphate and produce siderophores. Introduction of MH8a into soil significantly increased S, alba biomass and the accumulation of Cd (208%), Zn (86%), and Cu (39%) in plant shoots in comparison with those grown in non-inoculated soil. Introduced into the soil, MH8a was able to enter the plant and was found in the roots and leaves of inoculated plants thus indicating its endophytic features. PLFA analysis revealed that the MH8a that was introduced into soil had a temporary influence on the structure of the autochthonous bacterial communities. The plant growth-promoting features of the MH8a strain and its ability to enhance the metal uptake by white mustard and its long-term survival in soil as well as its temporary impact on autochthonous microorganisms make the strain a suitable candidate for the promotion of plant growth and the efficiency of phytoextraction.

A cold-regulated gene Lolium perenne LIR1 (LpLIR1) was isolated from perennial ryegrass using a subtractive approach. The gene has strong homology to the Light Induced Rice1 (LIR1) gene and is regulated at the transcriptional level by cold, and by a diurnal rhythm. Expression of LpLIR1 in perennial ryegrass was upregulated by vernalization but did not follow a standard vernalization-responsive expression pattern. LpLIR1 expression was restricted to vegetative tissues and absent in apices during floral induction and in flowers. LpLIR1 mRNA levels displayed diurnal fluctuations, which peaked before dusk and declined during the night. Heterologous expression of LpLIR1 in Arabidopsis led to a significant increase in leaf formation under short days (SD) conditions but only when plants had received a preceding vernalization treatment. Furthermore, dissection of plant development under SD revealed a minor but significant delay of flowering in the transgenic lines compared to wildtype plants.

Timothy is a perennial forage grass grown commonly in Boreal regions. This study explored the effect of vernalization and photoperiod (PP) on flowering and growth characteristics and how this related to changes in expression of three flowering related genes in accessions from different geographic origin. Large variation was found in accessions in their vernalization and PP responses. In southern accessions vernalization response or requirement was not observed, the heading date remained unchanged, and plants flowered without vernalization. On the contrary, northern types had obligatory requirement for vernalization and long PP, but the tiller elongation did not require vernalization at 16-h PP. Longer vernalization or PP treatments reduced the genotypical differences in flowering. Moreover, the vernalization saturation progressed stepwise from main tiller to lateral tillers, and this process was more synchronized in southern accessions. The expression of PpVRN1 was associated with vernalization while PpVRN3 accumulated at long PP. A crucial role for PpVRN3 in the transition to flowering was supported as in southern accession the transcript accumulated in non-vernalized plants after transfer to 16-h PP, and the apices transformed to generative stage. Differences in vernalization requirements were associated with variation in expression levels of PpVRN1 and PpVRN3, with higher expression levels in southern type. Most divergent transcript accumulation of PpMADS10 was found under different vernalization conditions. These differences between accessions can be translated into agronomic traits, such as the tiller composition of canopy, which affects the forage yield. The southern types, with minimal vernalization response, have fast re-growth ability and rapidly decreasing nutritive value, whereas northern types grow slowly and have better quality. This information can be utilized in breeding for new cultivars for longer growing seasons at high latitudes.

Genebank accessions stored as seed populations require periodic rejuvenation in order to maintain sufficient numbers of viable seeds. During rejuvenation the genetic composition of accessions may be altered for a variety of reasons, of which variation in pollination rates between plants is the least understood. In the present study, a paternity exclusion analysis was performed on a rejuvenated accession of perennial ryegrass. In addition, flowering data of the 49 parental plants were collected during the flowering season. The aim of the study was to determine how accurate variation in pollination rates between plants can be predicted from data on the spatial and temporal distribution of pollen release. The parental population and a total of 551 offspring from 12 progeny arrays were genotyped by means of molecular analysis. Using 25 microsatellites, paternity was identified for 81.9% of the offspring, while remaining ambiguities were resolved by AFLP analysis, except in four cases. Within the total sample 9 cases of contamination were observed. Mating within the study population was clearly non-random, as 61.9% of the identified pollen donors were located within I m distance from the mother plant. Observed pollination rates were very well described by an inverse quadratic function of inter-plant distance between potential mating pairs. Incorporation of the recorded flowering data in the calculation of expected pollination rates improved the goodness of fit with observed values by only 0.77%. Suggestions to reduce the variance in paternal contributions were presented. However, contamination was considered more threatening to the genetic integrity of perennial ryegrass germplasm than variation in pollination rates between plants, and indicated the need for improved measures to avoid gene flow from other germplasm.

Soil C sequestration may mitigate rising levels of atmospheric CO2. However, it has yet to be determined whether net soil C sequestration occurs in N-rich grasslands exposed to long-term elevated CO2. This study examined whether N-fertilized grasslands exposed to elevated CO2 sequestered additional C. For 10 years, Lolium perenne, Trifolium repens, and the mixture of L. perenne/T. repens grasslands were exposed to ambient and elevated CO2 concentrations (35 and 60 Pa pCO(2)). The applied CO2 was depleted in delta C-13 and the grasslands received low (140 kg ha(-1)) and high (560 kg ha(-1)) rates of N-15-labeled fertilizer. Annually collected soil samples from the top 10 cm of the grassland soils allowed us to follow the sequestration of new C in the surface soil layer. For the first time, we were able to collect dual-labeled soil samples to a depth of 75 cm after 10 years of elevated CO2 and determine the total amount of new soil C and N sequestered in the whole soil profile. Elevated CO2, N-fertilization rate, and species had no significant effect on total soil C. On average 9.4 Mg new C ha(-1) was sequestered, which corresponds to 26.5% of the total C. The mean residence time of the C present in the 0-10 cm soil depth was calculated at 4.6 +/- 1.5 and 3.1 +/- 1.1 years for L. perenne and T. repens soil, respectively. After 10 years, total soil N and C in the 0-75 cm soil depth was unaffected by CO2 concentration, N-fertilization rate and plant species. The total amount of N-15-fertilizer sequestered in the 0-75 cm soil depth was also unaffected by CO2 concentration, but significantly more N-15 was sequestered in the L. perenne compared with the T. repens swards: 620 vs. 452 kg ha(-1) at the high rate and 234 vs. 133 kg ha(-1) at the low rate of N fertilization. Intermediate values of N-15 recovery were found in the mixture. The fertilizer derived N amounted to 2.8% of total N for the low rate and increased to 8.6% for the high rate of N application. On average, 13.9% of the applied N-15-fertilizer was recovered in the 0-75 cm soil depth in soil organic matter in the L. perenne sward, whereas 8.8% was recovered under the T. repens swards, indicating that the N-2-fixing T. repens system was less effective in sequestering applied N than the non-N-2-fixing L. perenne system. Prolonged elevated CO2 did not lead to an increase in whole soil profile C and N in these fertilized pastures. The potential use of fertilized and regular cut pastures as a net soil C sink under long-term elevated CO2 appears to be limited and will likely not significantly contribute to the mitigation of anthropogenic C emissions.

The effects of 8 diploid perennial ryegrass (Lolium perenne L.) cultivars on dry matter (DM) intake, DM digestibility, and milk yield (MY) of dairy cows were evaluated in the summer of 2000 and 2001. Each summer, herbage was harvested daily and stall-fed to 12 dairy cows during six 2-wk periods. Six cultivars were fed in 3 periods (1, 3, and 5) according to a double 3 x 3 Latin square design. In the other periods (2, 4, and 6), 2 cultivars were fed in a repeated measurement design. Herbage mass and leaf blades in the sward canopy varied among cultivars, but differences were not consistent between years. The largest differences in herbage composition were found in water-soluble carbohydrate content, followed by crude protein content. only small differences were found in the neutral detergent fiber (NDF) content. A higher water-soluble carbohydrate content was found in 2 cultivars in both years, whereas ranking of cultivars in crude protein and NDF content was not consistent with years. Dry matter intake and MY were not affected by cultivar. In both years, DM digestiblity was high (>77%), with very small differences among cultivars in 2000 (

This study investigated effects of feeding perennial ryegrass with an elevated concentration of water-soluble carbohydrates (WSC) on dry matter intake (DMI), rumen function, milk production and composition of dairy cows. Twelve Holstein-Friesian dairy cows in mid lactation were stall-fed with fresh herbage of six varieties of perennial ryegrass during three 2-week periods, using a double 3 × 3 Latin square design. In each Latin square, three varieties were fed, one of which was characterised by a higher level of WSC (HSV) of 24 and 31 g/kg DM, respectively, than the lower WSC varieties (LSV). Other chemical components were similar or differed marginally and digestibility was similar. The results prove that it is possible to select for grass varieties with an elevated level of WSC. The DMI was not positively influenced by the increased WSC content, as cows fed HSV had similar DMI to cows fed the LSV (16.2 kg/day versus 16.6 kg/day). Rumen pH, neutral detergent fibre fractional clearance (KclNDF) and fractional degradation (kdNDF) rates were not reduced as a result of feeding HSV. Total VFA concentrations were not changed, nor were the proportions of acetate:propionate:butyrate. Milk urea concentration was reduced (P

To support conservation policies for old Dutch grasslands that are still in agricultural use, morphological variation and AFLP-based (amplified fragment length polymorphism-based) genetic diversity was studied in perennial ryegrass and white clover populations and compared with the diversity in reference varieties. In addition, AFLP variation was also studied in grasslands located in nature reserves. From principal component analysis (PCA), it appeared that date of ear emergence in perennial ryegrass and characters related to plant vigour in white clover were the main morphological characters separating the reference varieties from the old Dutch grassland populations, and some of the grassland populations from each other. In both species, intrapopulation variation was lower for the reference varieties. Lower heterogeneity within the reference varieties was also found in the AFLP analysis. All common AFLP's observed in old Dutch grasslands could also be found in the reference varieties and nature reserves. Only a small number of low-frequency alleles found in old Dutch grasslands were absent from the other two groups. However, band frequencies of markers could vary considerably between populations, which may have been caused by selection. Analysis of the AFLP data by PCA distinguished the majority of reference varieties from the old Dutch grasslands, and showed genetic differentiation only between some grasslands. Comparison of old Dutch grasslands with grasslands in nature reserves indicated that basically the same range of genetic variation is covered by the two groups. Our study indicates that the Netherlands harbour a more or less continuous population for major parts of the diversity of perennial ryegrass and white clover. It was concluded that no specific conservation measures are presently needed to maintain genetic diversity of perennial ryegrass and white clover occurring in old Dutch grasslands

An object-oriented simulation model was developed to integrate detailed analyses of perennial ryegrass (Lolium perenne) plant part dimensions, and their mass, cell wall content (CWC) and cell wall digestibility (CWD). The model was validated against data from two field experiments, carried out in The Netherlands (52degreesN) and Norway (67degreesN). In the experiments, the effects of temperature x daylength interactions on development, growth and digestibility were determined. Three cultivars differing in heading date were used, and undisturbed growth and regrowth after early (only in The Netherlands) and late cutting were studied. The sensitivity of the model was tested for changes in parameters determining morphological development. A grass plant was modelled as an aggregation of shoot objects. A shoot is an aggregation of phytomers, plus an inflorescence in the reproductive development stage. A phytomer consists of a leaf blade, a leaf sheath, a stem internode and a root. Each phytomer has an axillary bud, which can develop into a new shoot. Morphological development, organic matter (OM) growth and composition were simulated well by the model. Only for regrowing crops in The Netherlands CWC was lower and CWD was higher in the simulations than observed in the field, whereas the differences increased when stem development was more advanced at the moment of cutting. This was attributed to residual stems in the stubble after cutting due to difficulties with machine cutting of lodged crops on the experimental plots. The model can be considered as sufficiently accurate to simulate OM yield and composition of grass cultivars differing in heading date and growing under strongly contrasting temperature and daylength conditions. When the leaf appearance rate, width or length of the leaf blade were increased in the sensitivity analysis, OM yield enhanced, but composition and digestibility of the reproductive grass crops were not systematically affected. (C) 2004 Elsevier B.V. All rights reserved.

Climate change may affect ecosystem functioning through increased temperatures or changes in precipitation patterns. Temperature and water availability are important drivers for ecosystem processes such as photosynthesis, carbon translocation, and organic matter decomposition. These climate changes may affect the supply of carbon and energy to the soil microbial population and subsequently alter decomposition and mineralization, important ecosystem processes in carbon and nutrient cycling. In this study, carried out within the cross-European research project CLIMOOR, the effect of climate change, resulting from imposed manipulations, on carbon dynamics in shrubland ecosystems was examined. We performed a 14C-labeling experiment to probe changes in net carbon uptake and allocation to the roots and soil compartments as affected by a higher temperature during the year and a drought period in the growing season. Differences in climate, soil, and plant characteristics resulted in a gradient in the severity of the drought effects on net carbon uptake by plants with the impact being most severe in Spain, followed by Denmark, with the UK showing few negative effects at significance levels of p ≤ 0.10. Drought clearly reduced carbon flow from the roots to the soil compartments. The fraction of the 14C fixed by the plants and allocated into the soluble carbon fraction in the soil and to soil microbial biomass in Denmark and the UK decreased by more than 60%. The effects of warming were not significant, but, as with the drought treatment, a negative effect on carbon allocation to soil microbial biomass was found. The changes in carbon allocation to soil microbial biomass at the northern sites in this study indicate that soil microbial biomass is a sensitive, early indicator of drought- or temperature-initiated changes in these shrubland ecosystems. The reduced supply of substrate to the soil and the response of the soil microbial biomass may help to explain the observed acclimation of CO2 exchange in other ecosystems.

The effect of salinity on nitrate net uptake rate was studied in the moderately salt tolerant halophyte Festuca rubra L., in relation to changes in relative growth rate, root weight ratio and nitrogen and carbon partitioning. Plants were grown for 21 days on nutrient solution containing 50, 100 and 200 Mol m(-3) sodium chloride. Control plants received no additional sodium chloride. Relative growth rate of Festuca rubra was reduced by exposure to a sodium chloride concentration as low as 50 mol m-3, resulting in a lower nitrogen 'demand' for plant growth and consequently in a lower nitrate net uptake rate. However, the accumulation of reduced nitrogen containing osmotic solutes increased the plant's nitrogen 'demand' slightly The decreased root weight ratio, caused by the stronger inhibition of root growth as compared to shoot growth upon elevated salinity levels complicated the interpretation of the effects of different salinity treatments on nitrate net uptake rates. Utilization of soluble sugars for osmotic adjustment did not divert significant amounts of carbohydrates from the carbon pool. To get insight into the physiological background of the interaction between sodium chloride salinity and nitrate uptake, additional complications in the growth analysis and N nutrition must be considered.

Soils from urban areas often contain enhanced pseudo-total levels of potentially toxic elements (PTEs). Considering the expanding tendency of urban agricultural practices it is necessary to understand if these contaminants are available for plant uptake and if they pose risks to animal and human health. This study showed that estimates of Daily Intakes (DIs) of Cu, Pb and Zn for grazing animals were above animal Acceptable Daily Intakes (ADIs) at specific sites under the influence of an airport, an oil refinery and near highways with high traffic rates in the “Grande Porto” urban area (Portugal). These results suggest that there is a potential for dietary transfer of contaminants associated with the ingestion of both contaminated soil and feed by cows and sheep at unacceptably high concentrations. Furthermore, results showed that 40% of variability of ryegrass shoot contents of Cu, Pb and Ni; 60% for Ba; 70% for Zn; and 80% for Cd can be significantly (p < 0.01) explained by the variability of the corresponding chemical available pools in soils. Since the chemical available pools of PTEs in urban soils were rather low when compared with the corresponding pseudo-total pools (median 0.1–5%) and even when compared with the corresponding reactive pools it is advised to perform further research on the conditions and time span for the limited availability of PTEs in urban soils, and to determine under which conditions PTEs on reactive forms may become available.

International audience; The aim of this study was to derive regression-based soil-plant models to predict and compare metal(loid) (i.e. As, Cd, Cu, Pb and Zn) concentrations in plants (grass Agrostis sp./Poa sp. and nettle Urtica dioica L.) among sites with a wide range of metal pollution and a wide variation in soil properties. Regression models were based on the pseudo total (aqua-regia) and exchangeable (0.01 M CaCl2) soil metal concentrations. Plant metal concentrations were best explained by the pseudo total soil metal concentrations in combination with soil properties. The most important soil property that influenced U. dioica metal concentrations was the clay content, while for grass organic matter (OM) and pH affected the As (OM) and Cu and Zn (pH). In this study multiple linear regression models proved functional in predicting metal accumulation in plants on a regional scale. With the proposed models based on the pseudo total metal concentration, the percentage of variation explained for the metals As, Cd, Cu, Pb and Zn were 0.56%, 0.47%, 0.59%, 0.61%, 0.30% in nettle and 0.46%, 0.38%, 0.27%, 0.50%, 0.28% in grass.

A mechanism of action for the performance of Fe chelates as soil-applied fertilizer has been hypothesized by Lindsay and Schwab (J Plant Nutr 5:821–840, 1982), in which the ligand participates in a cyclic process of delivering Fe at the root surface and mobilizing Fe from the soil. This “shuttle mechanism” seems appealing in view of fertilizer efficiency, but little is known about its performance. The chelate FeEDDHA is a commonly used Fe fertilizer on calcareous soils. In this study, the performance of the shuttle mechanism has been examined for FeEDDHA chelates in soil interaction and pot trial experiments. The specificity of EDDHA ligands for chelating Fe from soils of low Fe availability is limited. Experimental support for a shuttle mechanism in soil-plant systems with FeEDDHA was found: specific metal mobilization only occurred upon FeEDDHA-facilitated Fe uptake. The mobilized metals originated at least in part from the root surface instead of the soil. The results from this study support the existence of a shuttle mechanism with FeEDDHA in soil application. If the efficiency of the shuttle mechanism is however largely controlled by metal availability in the bulk soil, it is heavily compromised by complexation of competing cations: Al, Mn and particularly Cu.

International audience; In French subalpine grasslands, cessation of mowing promotes dominance of Festuca paniculata, which alters plant diversity and ecosystem functioning. One of the mechanisms underpinning such effects may be linked to simultaneous changes in the abundance of fungal symbionts such as endophytes and arbuscular mycorrhizal fungi. In field conditions, mowing reduced the abundance of the endophyte Neotyphodium sp. in leaves of F. paniculata by a factor of 6, and increased mycorrhizal densities by a factor of 15 in the soil. In greenhouse experiments, the mycorrhizal colonization of Trifolium pratense and Allium porrum increased 3- fold and 3.8- fold respectively in mown vs unmown grassland soil. Significantly reduced growth of the two host plants was also observed on soil from the unmown grassland. Such opposite effects of mowing on the two functional groups of fungal symbionts could suggest interactions between these two groups, which in turn could contribute to structuring plant communities in subalpine grasslands. (c) 2013 Elsevier Ltd and The British Mycological Society. All rights reserved.

International audience; RATIONALE: Plant tissues artificially labeled with C-13 are increasingly used in environmental studies to unravel biogeochemical and ecophysiological processes. However, the variability of C-13-content in labeled tissues has never been carefully investigated. Hence, this study aimed at documenting the variability of C-13-content in artificially labeled leaves. METHODS: European beech and Italian ryegrass were subjected to long-term C-13-labeling in a controlled-environment growth chamber. The C-13-content of the leaves obtained after several months labeling was determined by isotope ratio mass spectrometry. RESULTS: The C-13-content of the labeled leaves exhibited inter- and intra-leaf variability much higher than those naturally occurring in unlabeled plants, which do not exceed a few per mil. This variability was correlated with labeling intensity: the isotope composition of leaves varied in ranges of ca 60 parts per thousand and 90 parts per thousand for experiments that led to average leaf C-13-content of ca +15 parts per thousand and +450 parts per thousand, respectively. CONCLUSIONS: The reported variability of isotope composition in C-13-enriched leaves is critical, and should be taken into account in subsequent experimental investigations of environmental processes using C-13-labeled plant tissues. Copyright (c) 2013 John Wiley & Sons, Ltd.

The importance of aboveground herbivores for modifying belowground ecosystems has prompted numerous studies; however, studies can be biased by context dependent conditions which lead to extremely inconsistent results. So far, the impacts of herbivory intensity by important rice pests on rice paddy soil ecosystems are lacking. The aim of this study was to test the hypothesis that intermediate herbivory intensity of the brown planthopper (Nilaparvata lugens Stal) could promote soil labile resources and microbial biomass, while high intensity would show a reverse pattern, by mediating rice plant growth. This study will also help the development of integrative pest management. Four hopper infestation density treatments (0, 4, 8 and 12 nymphs per rice plant) and two infestation duration treatments (9 and 15 days after N. lugens infestation, DAI 9 and DAI 15) were established in a glasshouse experiment. Soil and plant were sampled destructively from four replicates and analysed for soil labile resources availability, soil microbial biomass and plant performance, respectively. The infestation density significantly affected both shoot and root mass of rice (P

In Ireland, grass is a readily available bioresource. It has previously been established that Green biorefinery (GBR) could become a potential use of Irish grasslands, and a blueprint for a sustainable GBR industry in Ireland has been developed. The objective of this paper is to use scenario analysis to investigate the sensitivity of the profitability of the GBR blueprint to variations in grass quantity and quality as a function of botanical composition, fertiliser application, and biomass availability. As an outcome of these scenario analyses, the price the GBR can offer to farmers above their production costs (€ t −1 dry matter) was calculated. Results of the scenario analyses determined that GBR systems located in a catchment area of permanent pasture ( Lolium perenne > 60%) with annual grass yields in the range of 9–12 t dry matter (DM) ha −1 , and supplied with grass biomass with a fibre content of 500–555 g kg −1 DM and a protein content of 110–130 g kg −1 DM, were viable. The most profitable scenarios were generated when nitrogen fertiliser application was greater than 90 kg ha −1 a −1 . Biomass availability of less than 30% resulted in reduced profitability and for some scenarios resulted in a loss for both the GBR and farmer due to increased transport costs. Within the scenario assumptions of this study, grass feedstock was valued at €4–€56 t −1 dry matter above production costs. However, this value depended on the yields and biomass availability of the GBR catchment area.

New parameterizations for surface–atmosphere exchange of ammonia are presented for application in atmospheric transport models and compared with parameterizations of the literature. The new parameterizations are based on a combination of the results of three years of ammonia flux measurements over a grassland canopy (dominated by Lolium perenne and Poa trivialis) near Wageningen, the Netherlands and existing parameterizations from literature. First, a model for the surface–atmosphere exchange of ammonia that includes the concentration at the external leaf surface is derived and validated. Second, a parameterization for the stomatal compensation point (expressed as Γs, the ratio of [NH4+]/[H+] in the leaf apoplast) that accounts for the observed seasonal variation is derived from the measurements. The new, temperature-dependent Γs describes the observed seasonal behavior very well. It is noted, however, that senescence of plants and field management practices will also influence the seasonal variation of Γs on a shorter timescale. Finally, a relation that links Γs to the atmospheric pollution level of the location through the ‘long-term’ NH3 concentration in the air is proposed.

International audience; Airborne polycyclic aromatic hydrocarbons (PAHs) are deposited on agricultural grasses. In turn, PAHs enter the food chain through animals eating grasses. However, the risk of food contamination, e.g. of milk, is unknown because mechanisms ruling the fate of PAHs during digestion by cows are not understood, especially in the rumen. Here, we studied the disappearance rate of phenanthrene, pyrene and benzo[a]pyrene, n-alkanes, and dry matter from contaminated grass samples in the rumen, the first compartment of the ruminant digestive tract. Three cows fitted with a rumen cannula were used for this study and the grass samples were incubated in sacco for 1, 3, 6, 12, 24 and 48 h. The results show that the disappearance rate of PAHs was fast and higher than that of n-alkanes. On average, 83% of PAHs disappeared after 1 h of incubation. Phenanthrene showed a disappearance rate higher than 90%. The findings suggest a fast desorption of PAHs from the surface of the grass leaves. The digestive hydrolysis of the grass is not required to release PAHs, contrary to other fractions. As a consequence, the bioavailability of PAHs is not limited by the grass.

Quantitative traits, such as size and weight in animals and seed yield in plants, are distributed normally, even within a population of genetically identical individuals. For example, in plants, various factors, such as local soil quality, microclimate, and sowing depth, affect growth differences among individual plants of isogenic populations. Besides these physical factors, also epigenetic components contribute to differences in growth and yield. The network that regulates crop yield is still not well understood. Although this network is expected to have epigenetic elements, it is completely unclear whether it would be possible to shape the epigenome to increase crop yield. Here we show that energy use efficiency is an important factor in determining seed yield in canola ( Brassica napus ) and that it can be selected artificially through an epigenetic feature. From an isogenic canola population of which the individual plants and their self-fertilized progenies were recursively selected for respiration intensity, populations with distinct physiological and agronomical characteristics could be generated. These populations were found to be genetically identical, but epigenetically different. Furthermore, both the DNA methylation patterns as well as the agronomical and physiological characteristics of the selected lines were heritable. Hybrids derived from parent lines selected for high energy use efficiencies had a 5% yield increase on top of heterosis. Our results demonstrate that artificial selection allows the increase of the yield potential by selecting populations with particular epigenomic states.

Variations in the inulin contents have been detected in rhizophores of Vernonia herbacea during the phenological cycle. These variations indicate the occurrence of active inulin synthesis and depolymerization throughout the cycle and a role for this carbohydrate as a reserve compound. 1-Fructan exohydrolase (1-FEH) is the enzyme responsible for inulin depolymerization, and its activity has been detected in rhizophores of sprouting plants. Defoliation and low temperature are enhancer conditions of this 1-FEH activity. The aim of the present work was the cloning of this enzyme. Rhizophores were collected from plants induced to sprout, followed by storage at 5C. A full length 1-FEH cDNA sequence was obtained by PCR and inverse PCR techniques, and expressed in Pichia pastoris. Cold storage enhances FEH gene expression. Vh1-FEH was shown to be a functional 1-FEH, hydrolyzing predominantly -2,1 linkages, sharing high identity with chicory FEH sequences, and its activity was inhibited by 81 in the presence of 10 mM sucrose. In V. herbacea, low temperature and sucrose play a role in the control of fructan degradation. This is the first study concerning the cloning and functional analysis of a 1-FEH cDNA of a native species from the Brazilian Cerrado. Results will contribute to understanding the role of fructans in the establishment of a very successful fructan flora of the Brazilian Cerrado, subjected to water limitation and low temperature during winter. ispartof: Plant & Cell Physiology vol:49 issue:8 pages:1185-1195 ispartof: location:Japan status: published

Received for publication October 12, 2007. The total metal content of the soil or total metal concentration in the soil solution is not always a good indicator for metal availability to plants. Therefore, several speciation techniques have been developed that measure a defined fraction of the total metal concentration in the soil solution. In this study the Donnan Membrane Technique (DMT) was used to measure free metal ion concentrations in CaCl2 extractions (to mimic the soil solution, and to work under standardized conditions) of 10 different soils, whereas diffusive gradients in thin-films (DGT) and scanning chronopotentiometry (SCP) were used to measure the sum of free and labile metal concentrations in the CaCl2 extracts. The DGT device was also exposed directly to the (wetted) soil (soil-DGT). The metal concentrations measured with the speciation techniques are related to the metal adsorption at the root surface of ryegrass (Lolium perenne L.), to be able to subsequently predict metal uptake. In most cases the metal adsorption related pH-dependently to the metal concentrations measured by DMT, SCP, and DGT in the CaCl2 extract. However, the relationship between metal adsorption at the root surface and the metal concentrations measured by the soil-DGT was not¿or only slightly¿pH dependent. The correlations between metal adsorption at the root surface and metal speciation detected by different speciation techniques allow discussion about rate limiting steps in biouptake and the contribution of metal complexes to metal bioavailability.

17 páginas, 3 figuras, 3 tablas., Grass-based diets are of increasing social-economic importance in dairy cattle farming, but their low supply of glucogenic nutrients may limit the production of milk. Current evaluation systems that assess the energy supply and requirements are based on metabolisable energy (ME) or net energy (NE). These systems do not consider the characteristics of the energy delivering nutrients. In contrast, mechanistic models take into account the site of digestion, the type of nutrient absorbed and the type of nutrient required for production of milk constituents, and may therefore give a better prediction of supply and requirement of nutrients. The objective of the present study is to compare the ability of three energy evaluation systems, viz. the Dutch NE system, the agricultural and food research council (AFRC) ME system, and the feed into milk (FIM) ME system, and of a mechanistic model based on Dijkstra et al. [Simulation of digestion in cattle fed sugar cane: prediction of nutrient supply for milk production with locally available supplements. J. Agric. Sci., Cambridge 127, 247-60] and Mills et al. [A mechanistic model of whole-tract digestion and methanogenesis in the lactating dairy cow: model development, evaluation and application. J. Anim. Sci. 79, 1584-97] to predict the feed value of grass-based diets for milk production. The dataset for evaluation consists of 41 treatments of grass-based diets (at least 0.75 g ryegrass/g diet on DM basis). For each model, the predicted energy or nutrient supply, based on observed intake, was compared with predicted requirement based on observed performance. Assessment of the error of energy or nutrient supply relative to requirement is made by calculation of mean square prediction error (MSPE) and by concordance correlation coefficient (CCC). All energy evaluation systems predicted energy requirement to be lower (6-11%) than energy supply. The root MSPE (expressed as a proportion of the supply) was lowest for the mechanistic model (0.061), followed by the Dutch NE system (0.082), FIM ME system (0.097) and AFRCME system(0.118). For the energy evaluation systems, the error due to overall bias of prediction dominated the MSPE, whereas for the mechanistic model, proportionally 0.76 of MSPE was due to random variation. CCC analysis confirmed the higher accuracy and precision of the mechanistic model compared with energy evaluation systems. The error of prediction was positively related to grass protein content for the Dutch NE system, and was also positively related to grass DMI level for all models. In conclusion, current energy evaluation systems overestimate energy supply relative to energy requirement on grass-based diets for dairy cattle. The mechanistic model predicted glucogenic nutrients to limit performance of dairy cattle on grass-based diets, and proved to be more accurate and precise than the energy systems. The mechanistic model could be improved by allowing glucose maintenance and utilization requirements parameters to be variable. (C) 2007 Elsevier B.V. All rights reserved.

Soil C sequestration may mitigate rising levels of atmospheric CO2. However, it has yet to be determined whether net soil C sequestration occurs in N-rich grasslands exposed to long-term elevated CO2. This study examined whether N-fertilized grasslands exposed to elevated CO2 sequestered additional C. For 10 years, Lolium perenne, Trifolium repens, and the mixture of L. perenne/T. repens grasslands were exposed to ambient and elevated CO2 concentrations (35 and 60 Pa pCO(2)). The applied CO2 was depleted in delta C-13 and the grasslands received low (140 kg ha(-1)) and high (560 kg ha(-1)) rates of N-15-labeled fertilizer. Annually collected soil samples from the top 10 cm of the grassland soils allowed us to follow the sequestration of new C in the surface soil layer. For the first time, we were able to collect dual-labeled soil samples to a depth of 75 cm after 10 years of elevated CO2 and determine the total amount of new soil C and N sequestered in the whole soil profile. Elevated CO2, N-fertilization rate, and species had no significant effect on total soil C. On average 9.4 Mg new C ha(-1) was sequestered, which corresponds to 26.5% of the total C. The mean residence time of the C present in the 0-10 cm soil depth was calculated at 4.6 +/- 1.5 and 3.1 +/- 1.1 years for L. perenne and T. repens soil, respectively. After 10 years, total soil N and C in the 0-75 cm soil depth was unaffected by CO2 concentration, N-fertilization rate and plant species. The total amount of N-15-fertilizer sequestered in the 0-75 cm soil depth was also unaffected by CO2 concentration, but significantly more N-15 was sequestered in the L. perenne compared with the T. repens swards: 620 vs. 452 kg ha(-1) at the high rate and 234 vs. 133 kg ha(-1) at the low rate of N fertilization. Intermediate values of N-15 recovery were found in the mixture. The fertilizer derived N amounted to 2.8% of total N for the low rate and increased to 8.6% for the high rate of N application. On average, 13.9% of the applied N-15-fertilizer was recovered in the 0-75 cm soil depth in soil organic matter in the L. perenne sward, whereas 8.8% was recovered under the T. repens swards, indicating that the N-2-fixing T. repens system was less effective in sequestering applied N than the non-N-2-fixing L. perenne system. Prolonged elevated CO2 did not lead to an increase in whole soil profile C and N in these fertilized pastures. The potential use of fertilized and regular cut pastures as a net soil C sink under long-term elevated CO2 appears to be limited and will likely not significantly contribute to the mitigation of anthropogenic C emissions.

In the summers of 2000 and 2001, the effect of six diploid perennial ryegrass (Lolium perenne L.) cultivars on the N utilisation by 12 high productive dairy cows was determined. Experiments were conducted according to a double 33 Latin square design; within each Latin square, three cultivars were fed to six cows during three periods of 2 weeks each. Two cultivars had a higher water-soluble carbohydrate (WSC) content than the other cultivars in both years. A higher WSC content was more compensated by a lower neutral detergent fibre (NDF) content than by a lower crude protein content. In both years, dry matter intake (DMI) and milk production (MP) varied only slightly among cultivars and no differences among cultivars were found in the excretion of N in milk (in g/day and as % of N intake). Furthermore, cows fed the two cultivars with elevated WSC content had a lower N intake and this was associated with a lower milk urea N content and urinary N excretion. The small differences in chemical composition among cultivars did not result in significant differences in DMI, MP and N utilisation in dairy cows fed indoors. The results suggest that there is little scope to improve N utilisation of dairy cows at high N fertilization levels by breeding new cultivars. D 2005 Elsevier B.V. All rights reserved.

The effects of 4 diploid perennial ryegrass cultivars that differed in water-soluble carbohydrate (WSC) concentrations on milk yield and nitrogen (N) utilization in dairy cows were evaluated in a 2-yr grazing experiment. Twelve lactating dairy cows were assigned to 1 cultivar for a 2-wk period in a 4 x 4 Latin square design with 3 replicates. Each year, the experiment lasted 8 wk. Swards were in a vegetative stage throughout the experiment. Herbage constituents were determined, and DM intake was estimated with the n-alkane technique. Nitrogen utilization was calculated as N excreted in milk divided by N intake, assuming a zero N retention. Two cultivars had consistently higher WSC concentrations and slightly lower neutral detergent fiber concentrations than the other 2 cultivars. The ranking of the cultivars in chemical composition traits in both years was rather consistent. Cows grazing the cultivar with the lowest concentration of WSC had the lowest herbage DM intake, N intake, milk yield, and milk N yield in 2002, but with a similar difference in WSC concentration, no differences among cultivars were found in 2003. In both years, milk urea N concentration was slightly higher for cows grazing the cultivar with the lowest WSC concentration, although it was significant only in 2003. Nitrogen utilization (N milk:N intake, g/g) varied between 0.241 and 0.246 in 2002 and between 0.190 and 0.209 in 2003, and in both years there was no effect of cultivar. At relatively high N concentrations in grass and only small differences among cultivars in neutral detergent fiber concentrations, cultivars with an elevated WSC concentration did not increase N utilization in grazing dairy cows.

The net flux of soil C is determined by the balance between soil C input and microbial decomposition, both of which might be altered under prolonged elevated atmospheric CO 2 . In this study, we determined the effect of elevated CO 2 on decomposition of grass root material ( Lolium perenne L.). 14 C-labeled root material, produced under ambient (35 Pa pCO 2 ) or elevated CO 2 (70 Pa pCO 2 ) was incubated in soil for 64 days. The soils were taken from a pasture ecosystem which had been exposed to ambient (35 Pa pCO 2 ) or elevated CO 2 (60 Pa pCO 2 ) under FACE-conditions for 10 years and two fertilizer N rates: 140 and 560 kg N ha −1 year −1 . In soil exposed to elevated CO 2 , decomposition rates of root material grown at either ambient or elevated CO 2 were always lower than in the control soil exposed to ambient CO 2 , demonstrating a change in microbial activity. In the soil that received the high rate of N fertilizer, decomposition of root material grown at elevated CO 2 decreased by approximately 17% after incubation for 64 days compared to root material grown at ambient CO 2 . The amount of 14 CO 2 respired per amount of 14 C incorporated in the microbial biomass ( q 14 CO 2 ) was significantly lower when roots were grown under high CO 2 compared to roots grown under low CO 2 . We hypothesize that this decrease is the result of a shift in the microbial community, causing an increase in metabolic efficiency. Soils exposed to elevated CO 2 tended to respire more native SOC, both with and without the addition of the root material, probably resulting from a higher C supply to the soil during the 10 years of treatment with elevated CO 2 . The results show the importance of using soils adapted to elevated CO 2 in studies of decomposition of roots grown under elevated CO 2 . Our results further suggest that negative priming effects may obscure CO 2 data in incubation experiments with unlabeled substrates. From the results obtained, we conclude that a slower turnover of root material grown in an ‘elevated-CO 2 world’ may result in a limited net increase in C storage in ryegrass swards.

To support conservation policies for old Dutch grasslands that are still in agricultural use, morphological variation and AFLP-based (amplified fragment length polymorphism-based) genetic diversity was studied in perennial ryegrass and white clover populations and compared with the diversity in reference varieties. In addition, AFLP variation was also studied in grasslands located in nature reserves. From principal component analysis (PCA), it appeared that date of ear emergence in perennial ryegrass and characters related to plant vigour in white clover were the main morphological characters separating the reference varieties from the old Dutch grassland populations, and some of the grassland populations from each other. In both species, intrapopulation variation was lower for the reference varieties. Lower heterogeneity within the reference varieties was also found in the AFLP analysis. All common AFLP's observed in old Dutch grasslands could also be found in the reference varieties and nature reserves. Only a small number of low-frequency alleles found in old Dutch grasslands were absent from the other two groups. However, band frequencies of markers could vary considerably between populations, which may have been caused by selection. Analysis of the AFLP data by PCA distinguished the majority of reference varieties from the old Dutch grasslands, and showed genetic differentiation only between some grasslands. Comparison of old Dutch grasslands with grasslands in nature reserves indicated that basically the same range of genetic variation is covered by the two groups. Our study indicates that the Netherlands harbour a more or less continuous population for major parts of the diversity of perennial ryegrass and white clover. It was concluded that no specific conservation measures are presently needed to maintain genetic diversity of perennial ryegrass and white clover occurring in old Dutch grasslands

In laboratory experiments, bacterioplankton were incubated under different nutrient conditions, and the percentage of bacteria exhibiting a polysaccharidic capsule (capsulated bacteria) and that of CTC (cyanotetrazolium chloride)-positive and therefore metabolically highly active bacteria were determined. In these seawater cultures amended with nutrients more than 95% of the CTC-positive cells exhibited a capsule. During two cruises, one to the North Atlantic and one to the North Sea, we investigated the distribution of capsulated bacteria throughout the water column. Capsulated bacteria were generally more abundant in eutrophic surface waters than in deeper layers or more oligotrophic regions. In the upper 100 m of the North Atlantic, about 6–14% of the total bacterioplankton community was capsulated, while in the layers below 100 m depth, 97% of the bacteria lacked a visible capsule. The percentage of capsulated bacteria correlated with bacterial abundance and production, and chlorophyll a concentration. Also, the bioavailability of DOC (dissolved organic carbon), estimated by the ratio between bacterial production and DOC concentration, significantly correlated with the percentage of capsulated bacteria. In the North Sea, the contribution of capsulated bacteria to the total number of bacteria decreased from the surface (3 m depth) to the near-bottom (25–35 m) layers from 20% to 14% capsulated bacteria. In the nearshore area of the North Sea, about 27% of the bacteria exhibited a capsule. Overall, a pronounced decrease in the contribution of capsulated bacteria to the total bacterial abundance was detectable from the eutrophic coastal environment to the open North Atlantic. Using this epifluorescence-based technique to enumerate capsulated bacterioplankton thus allowed us to routinely assess the number of capsulated bacteria even in the oceanic water column. Based on the data obtained in this study we conclude that almost all metabolically highly active bacteria exhibit a capsule, but also some of the metabolically less active cells express a polysaccharide capsule detectable with this method.

The effect of salinity on nitrate net uptake rate was studied in the moderately salt tolerant halophyte Festuca rubra L., in relation to changes in relative growth rate, root weight ratio and nitrogen and carbon partitioning. Plants were grown for 21 days on nutrient solution containing 50, 100 and 200 Mol m(-3) sodium chloride. Control plants received no additional sodium chloride. Relative growth rate of Festuca rubra was reduced by exposure to a sodium chloride concentration as low as 50 mol m-3, resulting in a lower nitrogen 'demand' for plant growth and consequently in a lower nitrate net uptake rate. However, the accumulation of reduced nitrogen containing osmotic solutes increased the plant's nitrogen 'demand' slightly The decreased root weight ratio, caused by the stronger inhibition of root growth as compared to shoot growth upon elevated salinity levels complicated the interpretation of the effects of different salinity treatments on nitrate net uptake rates. Utilization of soluble sugars for osmotic adjustment did not divert significant amounts of carbohydrates from the carbon pool. To get insight into the physiological background of the interaction between sodium chloride salinity and nitrate uptake, additional complications in the growth analysis and N nutrition must be considered.

The effects of grazing regime (cattle, sheep and mixed cattle + sheep) on white clover content (clover dry matter as a percentage of total dry matter) were measured in a permanent pasture over a 7 year period in the west of Ireland (53°17′N 8°47′E). Rotational grazing was simulated by grazing for short intense periods of 3–4 d at 3 to 5 week intervals. In general, cattle grazing resulted in higher clover content (13.5%) compared with mixed (9.5%) or sheep (4.9%) grazing. The ranking of clover contents (cattle > mixed > sheep) which developed rapidly in 1990 persisted until 1996. Clover contents under mixed grazing tracked those under cattle grazing during the first 4 years, and sheep grazing during the final 3 years. Within-year relativities in clover content among grazing regimes that existed at the start of the grazing season persisted throughout the year. There was an indication that differences in clover content between grazing regimes at the end of grazing persisted until the following spring. A regression analysis of clover content in each grazing period showed strong effects of grazing regime, generally positive relationships with mean air temperature in the period and clover content in the preceding period, and an interaction between air temperature and clover content in the previous period. Implications for pasture management and experimentation are discussed.

Fungal endophytes infect several grass species. The fungus can alter the growth, physiological and morphological characteristics of the infected plant. A greenhouse experiment was designed to determine the effect of the fungal endophyte Neotyphodium coenophialum on the nutrient content of a selected ecotype of tall fescue (Festuca arundinacea), when growing at two nutrient supply levels. Stem dry matter, averaged over all harvests and nutrient supply treatments, was higher in non-infected (E-) than in endophyte infected (E+) plants. We found a significant interaction between endophyte infection and nutrient supply level and/or harvest date on the N, P, Ca and Mg plant tissue concentrations. Only for Ca leaf concentrations the effect of infection status was not influenced by harvest date or nutrient supply treatment and Ca concentration was higher in E- than in E+ plants. Differences between E+ and E- plants in P concentration were significant at the beginning of the experiment, and for N and Mg were observed at the end of the experiment (16-18 weeks). Endophyte infected plants had higher stem N and leaf Mg concentrations in the high nutrient supply but significantly lower concentrations at the low nutrient supply treatment. Phosphorus 2 concentration in leaf and aboveground plant tissue was significantly higher in E- than in E+ plants at the high nutrient fertility treatment at week 12, but in the low fertility treatment differences were not significant. Differences in K, Fe, Mn, Zn and Cu concentration between infected and non-infected plants were not statistically significant. Our results suggest that endophyte infection of tall fescue appears to increase the content of nutrients related to protein synthesis processes (N and Mg). This work was supported by the Spanish Interministerial Commission of Science and Technology (CICYT) AGF96-0423. B.R. Vázquez de Aldana was supported by a research contract from the Spanish Ministry of Education and Culture, and I. Zabalgogeazcoa by a postdoctoral fellowship from the INIA.

An allotriploid (ALA, 2n=3 x=36) BC(1) plant was obtained by backcrossing a diploid F(1) interspecific hybrid (LA, 2n=2 x=24), derived from a Lilium longiflorum (L genome) and an Asiatic hybrid (A genome), to the latter parent. This allotriploid was backcrossed to a diploid Asiatic hybrid (2n=2 x=24) and to an allotetraploid (LLAA, 2n=4 x=48) LA hybrid. A total of 25 plants of these crosses were examined for ploidy level, and 12 individuals were analyzed for their genome constitution through genomic in situ hybridization (GISH). In most cases the progenies from the triploid-diploid (3 x-2 x) crosses consisted of aneuploids. Further more, there was evidence for the formation of near-haploid (x=12+2) to triploid (3 x=36) gametes in the allotriploid BC(1) plant. The progenies of triploid-tetraploid (3 x-4 x) cross also consisted of mostly aneuploids but in this case the triploid female parent had contributed predominantly near-triploid (2n) gametes for the origin of BC(2) progenies. The different ploidy levels observed between 3 x-2 x and 3 x-4 x crosses are possibly caused by preferential fertilization or survival resulting in a different ratio of chromosome numbers between the embryo and endosperm. Though Lilium has a tetrasporic, eight-nucleate type of embryo sac formation (Fritillaria type), the observed difference between the progeny types in 3 x-2 x and 3 x-4 x crosses is comparable to that of observed in monosporic eight nucleate types (Polygonum type) that predominate in most genera of Angiosperms. An important feature of the genome constitution of the progenies was that the homoeologous recombinant chromosomes were transmitted intact from BC(1) to BC(2) progenies in variable numbers. In addition, there was evidence for the occurrence of new homoeologous recombinations in the triploid BC(1). Of the two euploid BC(2) plants one had originated through the parthenogenetic development of a 2n egg and the other had originated through indeterminate meiotic restitution (IMR).

Drought events can strongly affect ecosystem functioning by modifying relationship between plants, microbes and soil chemistry, with consequent impacts on nutrient cycling. However, the potential impacts of a soil moisture reduction on the nitrogen (N) and phosphorus (P) cycling in grasslands remain poorly understood, especially in regard to. forage production., To fill this knowledge gap, a drought experiment was carried out using rainout shelters in two permanent grasslands, characterized by similar vegetation communities but contrasted soil nutrient limitations. Drought treatments were applied during two months, either when plant growth was highest (Early-season drought) or after the peak of biomass production (Late-season drought). Dry matter production, forage N status (NNI) and P content as well as N and P contents in microbial biomass and soil were determined., Both early and late-season drought significantly reduced soil moisture during the vegetation growth period. Forage yield was also reduced by drought, but only when it occurred late in the season. Using a structural equation model, we showed that soil moisture reduction had a direct effect on forage N status, suggesting that water shortage induced lower transpiration and water fluxes. Soil moisture reduction also affected forage P by reducing the availability of soil P. However, other mechanisms played a larger role and were site-specific. At the more fertile site, reduction in soil moisture directly impaired forage P, suggesting that water stress mainly resulted in lower diffusion rates to roots, while at the less fertile site, an indirect reduction of forage P through a pathway implying microbes (decrease in microbial P) was detected., Our results suggest that the two grasslands suffered mainly from water shortage per se, but also from drought induced nutrient deficiency (mainly P), which amplified yield losses and further decreased forage quality. Overall, our findings emphasize the need for further research on the plant-soil-microbe system functioning, in order to secure a sustainable and resilient forage production in the context of climate change.