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Studying the effects of fertilisation on the seed production of grassland species can help understand the vegetation changes and biodiversity losses due to soil eutrophication. The seed production of fifteen grasses and seventeen forbs from a temperate hay meadow was studied under three fertilisation treatments: 0-0-0, 0-54-108 and 192-108-216 kg N, P2O5 and K2O respectively, per year. Fertile shoots collected at the seed maturation stage were analysed for all main traits of the gamic reproduction. On average, forbs produced more ovules and viable seeds per shoot (199 and 65, respectively) than grasses (112 and 35, respectively). Fertilisation increased the number of inflorescences per shoot in both grasses and forbs and had a limited but variable effect on germinability and viability in the two functional groups: viability increased in grasses but often decreased in forbs. This pattern resulted in 55% and 11% increases in viable seed production in grasses and forbs, respectively. At the higher level of fertilisation, shoot density was positively related to the number of viable seeds per shoot in grasses and to the seed size in forbs. These results highlight that the traits of the gamic reproduction can contribute to explain the relationship between soil nutrient richness and grassland species composition and richness.

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In mountainous areas, bare slopes represent often dangerous forms of land degradation that need to be rehabilitated. However, revegetation is usually performed with non-native plants, negatively impacting the natural landscape value. Comprehensive studies, especially on poorly investigated long-term revegetation effects, are therefore essential for the improvement of rehabilitation practices. In this study, four landslides or disused quarries surrounded by well-preserved (semi-) natural vegetation that were revegetated between 1988 and 2002 with non-native only herbaceous or both herbaceous and woody plants were studied 1-9 and 16-31 years after rehabilitation. A total of 111 sampling areas were surveyed for introduced and volunteer species and other important ground cover soil and topography traits. Climatic traits and species' ecological indicators were retrieved from the available databases. The time patterns and ecological spectra of the plant covers were analysed and correlated to the site traits. In the first decade, introduced plants were initially abundant but decreased rapidly. In the long-term, the more stress-tolerant among them still had a considerable cover, especially in fine-textured soils. Native species were established slowly but, due to their higher stress tolerance, dominated in the long-term, especially at more stressful sites. In areas with predominantly herbaceous cover, soil factors, such as gravel content, were more related to the plant cover. In areas with predominantly woody plant cover, the high plant cover was dependent on microclimatic factors, especially aspect. At sites like those studied here, woody species should be always used, but in the form of native plants, as they tend to persist. If herbaceous species are unavailable in native forms, their introduction should be avoided in areas not exposed to erosion, whereas poorly stress-tolerant non-native plants should be used in steep areas as they create a fast but short-lasting cover compensated in the long term by the establishment of native species.

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Livestock farming in large-scale pasture landscapes can fulfil the economic and ecological requirements for sustainable, biodiversity-friendly agriculture of the European Union (EU). However, little is known about the grazing systems that have maintained these pastures for centuries. The traditional grazing methods and their effects on the cattle behaviour were therefore studied at 23 mountain summer farms in the Asiago upland (northeastern Italy). Traditional methods of free-ranging and herding were the only techniques adopted by farmers. Only environmental factors (size and perimeter length of the pastoral unit) were significantly associated with the grazing method used. Free-ranging was more often performed in small and homogeneous farms, while herding was done in large and heterogeneous farms. Herding led to a more homogeneous grazer distribution and better regulated the lengths of the cattle daily routes than free-ranging. The grazing direction and resting sites were almost exclusively affected by environmental factors. Cattle grazed predominantly in the direction from which the wind was blowing, and preferred resting at windy sites in the daytime (escaping from flies) and at open areas close to woods during the night (being protected against cold winds). Other traits shared between free-ranging and herding were: the seasonal organisation of grazing into periods and camps; re-grazing of the same surface within a few days; everyday use of different pasture types; and rhythms in the cattle daily activities. Even if they may have lower productivity compared to rotational stocking, the studied traditional grazing methods, especially herding, have the advantages of respecting spontaneous cattle behaviour, producing more savoury cheeses, and better maintaining heterogeneous and biodiversity-rich landscapes. More attention should be paid to traditional grazing methods within measures aiming to conserve cultural heritage and agricultural biodiversity.

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Semi-natural grasslands are of great importance to European biodiversity. Their restoration in mountain regions often presents specific problems of raw soil stabilisation and difficult climatic conditions. Methods used to tackle these problems include increasing sowing density and fertilisation. However, past studies were performed at only one site or for a short time and do not clearly show how site traits and time affect restoration results. In six field experiments undertaken on raw soils in the Italian Alps, the effects of sowing density, fertilisation, soil, and climate on the development over time of plant density and cover were analysed over three to six years. The seed was collected from nearby grasslands and sown at rates of 335-21,410 seeds m-2. During the first one-two years, the percentage of seeds producing established plants at low seed rates was higher in stonier surfaces; however, at high rates a larger absolute plant number was recorded in finer soils. Two to five years after sowing, climatic conditions became more important. Full vegetation cover was achieved in all experiments. However, at high altitudes, individual plant cover became greater (and plant density decreased), probably because the grasses needed to accumulate more non-structural carbohydrates necessary to survive during the longer winter. Fertilisation accelerated the establishment of full vegetation cover by 1 to 3 years as it increased the mean cover per plant without affecting plant density. Grassland restoration is possible in mountain raw soils by using the seeds collected from nearby donor sites. High sowing densities are not required even in coarse or higher altitude soils because, in the short term, stones on the ground surface increase the percentage of seed producing established plants and, over the long term, plant density is not dependent on the sowing density. In low-fertile soils, fertilisation accelerates the establishment of full vegetation cover.

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Brown apical necrosis (BAN) is a most recently described disease affecting English (Persian) walnut fruit. BAN was only recorded in intensively managed walnut orchards and was found to be a disease complex mainly caused by Fusarium species. All fungi associated with this disease are polyphagous and ubiquitous, not specific to walnut. Consequently, BAN occurrence is more strictly dependent, than generally, on the interaction between pathological features and environmental conditions. Environmental variables identified with regression analysis showed that maximum temperature, angle of main wind direction versus tree row orientation, and orchard distance to the closest river/canal, all representative of climatic conditions occurring in the orchard, were related to fruit drop. The factor displaying the highest influence on severity of BAN fruit drop was maximum temperature and only subordinately factors are associated with relative humidity. BAN symptoms were reproduced with in planta artificial inoculation, and fruit drop of symptomatic fruit was significantly higher than that of the noninoculated trees for each type of inoculum (Fusarium semitectum, F. graminearum, and Alternaria spp.). F. semitectum and F. graminearum were more aggressive than Alternaria species, and the earliest artificial inoculations in mid-May resulted in the highest fruit drop. The extension of walnut fruit susceptibility and the conducive environmental factors to BAN are discussed.

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A reliable and species-specific real-time quantitative polymerase chain reaction (qPCR) assay was developed for detection of the complex soilborne anamorphic fungus Fusarium oxysporum. The new primer pair, designed on the translation elongation factor 1-α gene with an amplicon of 142 bp, was highly specific to F. oxysporum without cross reactions with other Fusarium spp. The protocol was applied to grafted melon plants for the detection and quantification of F. oxysporum f. sp. melonis, a devastating pathogen of this cucurbit. Grafting technologies are widely used in melon to confer resistance against new virulent races of F. oxysporum f. sp. melonis, while maintaining the properties of valuable commercial varieties. However, the effects on the vascular pathogen colonization have not been fully investigated. Analyses were performed on 'Charentais-T' (susceptible) and 'Nad-1' (resistant) melon cultivars, both used either as rootstock and scion, and inoculated with F. oxysporum f. sp. melonis race 1 and race 1,2. Pathogen development was compared using qPCR and isolations from stem tissues. Early asymptomatic melon infections were detected with a quantification limit of 1 pg of fungal DNA. The qPCR protocol clearly showed that fungal development was highly affected by host-pathogen interaction (compatible or incompatible) and time (days postinoculation). The principal significant effect (P ≤ 0.01) on fungal development was due to the melon genotype used as rootstock, and this effect had a significant interaction with time and F. oxysporum f. sp. melonis race. In particular, the amount of race 1,2 DNA was significantly higher compared with that estimated for race 1 in the incompatible interaction at 18 days postinoculation. The two fungal races were always present in both the rootstock and scion of grafted plants in either the compatible or incompatible interaction.

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BACKGROUND: Fusarium oxysporum f. sp. melonis Snyd. & Hans. (FOM) causes Fusarium wilt, the most important infectious disease of melon (Cucumis melo L.). The four known races of this pathogen can be distinguished only by infection on appropriate cultivars. No molecular tools are available that can discriminate among the races, and the molecular basis of compatibility and disease progression are poorly understood. Resistance to races 1 and 2 is controlled by a single dominant gene, whereas only partial polygenic resistance to race 1,2 has been described. We carried out a large-scale cDNA-AFLP analysis to identify host genes potentially related to resistance and susceptibility as well as fungal genes associated with the infection process. At the same time, a systematic reisolation procedure on infected stems allowed us to monitor fungal colonization in compatible and incompatible host-pathogen combinations. RESULTS: Melon plants (cv. Charentais Fom-2), which are susceptible to race 1,2 and resistant to race 1, were artificially infected with a race 1 strain of FOM or one of two race 1,2 w strains. Host colonization of stems was assessed at 1, 2, 4, 8, 14, 16, 18 and 21 days post inoculation (dpi), and the fungus was reisolated from infected plants. Markedly different colonization patterns were observed in compatible and incompatible host-pathogen combinations. Five time points from the symptomless early stage (2 dpi) to obvious wilting symptoms (21 dpi) were considered for cDNA-AFLP analysis. After successful sequencing of 627 transcript-derived fragments (TDFs) differentially expressed in infected plants, homology searching retrieved 305 melon transcripts, 195 FOM transcripts expressed in planta and 127 orphan TDFs. RNA samples from FOM colonies of the three strains grown in vitro were also included in the analysis to facilitate the detection of in planta-specific transcripts and to identify TDFs differentially expressed among races/strains. CONCLUSION: Our data suggest that resistance against FOM in melon involves only limited transcriptional changes, and that wilting symptoms could derive, at least partially, from an active plant response.We discuss the pathogen-derived transcripts expressed in planta during the infection process and potentially related to virulence functions, as well as transcripts that are differentially expressed between the two FOM races grown in vitro. These transcripts provide candidate sequences that can be further tested for their ability to distinguish between races.Sequence data from this article have been deposited in GenBank, Accession Numbers: HO867279-HO867981.

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