Faecal nutrient deposition of domestic and wild herbivores in an alpine grassland.

The contribution of herbivores to ecosystem nutrient fluxes through dung deposition has the potential to, directly and indirectly, influence ecosystem functioning. This process can be particularly important in nutrient-limited ecosystems such as alpine systems. However, herbivore dung content (carbon, C; nitrogen, N; phosphorus, P; potassium, K) and stoichiometry (C/N) may differ among species due to differences in diet, seasonality, body type, feeding strategy, and/or digestive system with consequences for soil biogeochemistry. Here we explore how species, body size, and seasonality may result in differences in dung stoichiometry for four alpine herbivores (chamois, sheep, horse, and cattle). We found that herbivore dung nutrient content often varies among species as well as with body size, with the dung of small herbivores having larger C, N, and P faecal content. Seasonality also showed marked effects on faecal nutrient content, with a general pattern of decreasing levels of faecal P, N and an increase of C/N as the summer progresses following the loss of nutrient value of the vegetation. Moreover, we showed how herbivores play an important role as natural fertilizers of C, N, and P in our study area, especially cattle. Our study highlights the importance of considering the relative contribution of different herbivores to ecosystem nutrient fluxes in management practices, especially with ongoing changes in wild and domestic herbivore populations in alpine ecosystems.

Feeding Ecology of the Cuvier's Gazelle (Gazella cuvieri, Ogilby, 1841) in the Sahara Desert.

Knowledge of the feeding ecology of ungulates in arid biomes offers an interesting model for understanding the drought resistance of large desert-adapted herbivores, a crucial issue in the face of increasing desertification due to climate change. To assess the feeding ecology of the endangered Cuvier's gazelle (Gazella cuvieri) in the Sahara desert, we used a multi-method approach combining faecal samples, direct observations, and the recording of indirect signs of feeding. We hypothesised that browser behaviour is the best foraging strategy for species living in hyper-arid environments, mainly due to long periods without grazing opportunities. Complementarily, we explored the effects of the main environmental descriptors (rainfalls and NDVI) on feeding patterns and diet quality. We found that Cuvier's diets are based mainly on acacias (Vachellia tortilis, V. flava) and occasionally on the annual forb Anastatica hierochuntica. In total, eighteen species (five trees, nine shrubs, three herbs, and one grass) belonging to fifteen families were recorded. Our result confirmed the browsers' characteristic of this species, reaffirming its ability to settle in a hostile environment. Acacias stand out as key species consumed at the southernmost limit of their range; hence, future conservation plans and strategies should take this into account for the survival of Cuvier's gazelle in desert environments.

Positive effect of spring advance on the diet quality of an alpine herbivore.

Changes in vegetation phenology related to global warming are having alarming effects on the life history traits of many herbivore species. Such changes are particularly critical in alpine ecosystems, where strong climate limitations on plant growth make seasonal synchronization imperative for the growth, reproduction and survival of herbivores. However, despite the pivotal role of resource-use strategies on the performances of such species, few studies have explicitly assessed the mechanistic impact of climate change on their diets. We aimed to fill this gap by studying the effect of spring onset on the dietary composition and quality of a medium-size alpine herbivore while considering density-dependent processes and age- and sex-specific differences in foraging behavior. Using an exceptional, long-term (24 years) direct individual-based dietary monitoring of a Pyrenean chamois population (Rupicapra pyrenaica pyrenaica), we showed that ongoing earlier onsets of spring are leading to an earlier access to high-quality forage and therefore a higher diet quality at a fixed date, without apparent changes in diet composition. We also showed that at high densities, intraspecific competition reduced diet quality by driving animals to feed more on woody plants and less on nutritious forbs and graminoids. By assessing the mechanistic effects of global warming on the dietary patterns of species at the center of trophic networks, this study is an essential step for predictive models aiming at understanding the ongoing ecosystem consequences of the global climatic crisis.

Grazing influences biomass production and protein content of alpine meadows.

Alpine grasslands are essential for carbon sequestration and food supply for domestic and wild herbivores inhabiting mountainous areas worldwide. These biomes, however, are alternatively threatened by the abandonment of agricultural and livestock practices leading to a fast-growing shrubification process while other mountain grasslands are suffering from the impacts of overgrazing. The functioning of alpine meadow ecosystems is primarily driven by climatic conditions, land-use legacies and grazing. However, although it is critically important, the role of large herbivores on the aboveground biomass and protein content of palatable plants is poorly understood for most alpine meadows. In this work, we explore the effects of grazing on grassland vegetation at two different spatial and temporal scales in the Eastern Pyrenees, Spain. Remote sensing was used to assess the effect of high and moderate grazing (HG and MG respectively) on grass biomass using the leaf area index (LAI) at the meso-scale (patches between 2.3 and 38.7 ha). We also explored the impact of null (NG), overgrazing (MO, mimicked overgrazing) and high (HG) grazing intensities at local scale setting eighteen 1 m2 exclusion boxes in six meadows (three boxes each) commonly used by domestic and wild ungulates. Historical satellite data showed that LAI values are greater in high than in low grazed areas (HG, mean = 0.66, LG, mean = 0.55). Along the same lines, high and moderate grazing pressures improved biomass production at the local-scale (HG, mean = 590.3 g/m2, MO, mean = 389.3 g/m2 and NG, mean = 110.8 g/m2). Crude protein content reached higher values under MO pressure than under HG pressure. Our results confirm that grazing intensity exerts significant changes on the above-ground biomass production and the protein content of plants consumed by domestic (cattle and horses) and wild ungulates (Southern Chamois, Rupicapra pyrenaica). We can conclude that ungulates sustain biomass and nutritive values of grass exerting a negligible effect on biomass and protein content of woody vegetation. Our results will inform management guidelines to support profitable grazing activities and promote conservation of the open landscapes in the alpine ecosystems under the current global change scenario.

Near Infrared Reflectance Spectroscopy Analysis to Predict Diet Composition of a Mountain Ungulate Species.

The diet composition of ungulates is important to understand not only their impact on vegetation, but also to understand the consequences of natural and human-driven environmental changes on the foraging behavior of these mammals. In this work, we evaluated the use of near infrared reflectance spectroscopy analysis (NIRS), a quick, economic and non-destructive method, to assess the diet composition of the Pyrenean chamois Rupicapra pyrenaica pyrenaica. Fecal samples (n = 192) were collected from two chamois populations in the French and Spanish Pyrenees. Diet composition was initially assessed by fecal cuticle microhistological analysis (CMA) and categorized into four functional groups, namely: woody, herbaceous, graminoid and Fabaceae plants. Regressions of modified partial least squares and several combinations of scattering correction and derivative treatments were tested. The results showed that models based on the second derivative processing obtained the higher determination coefficient for woody, herbaceous and graminoid plants (R2CAL, coefficient of determination in calibration, ranged from 0.86 to 0.91). The Fabaceae group, however, was predicted with lower accuracy (R2CAL = 0.71). Even though an agreement between NIRS and CMA methods was confirmed by a Bland-Altman analysis, confidence limits of agreement differed by up to 25%. Our results support the viability of fecal NIRS analysis to study spatial and temporal variations of the Pyrenean chamois' diets in summer and winter when differences in the consumption of woody and annual plants are the greatest. This new use for the NIRS technique would be useful to assess the consequences of global change on the feeding behavior of this mountain ungulate and also in other ungulate counterparts.

Mildly toxic shrubs as indicators of goats herbivory give information for the management of natural landscapes on Mediterranean islands.

Herbivory of insular plant communities by introduced animals has been widely studied for decades. Though their diet mainly includes palatable and highly nutritive species, goats will also eat plants that are toxic to other animals. Thus, severe affection of toxic species may indicate high herbivore pressure or a low quality of vegetative food. To evaluate whether herbivory damage to low-palatability shrubs could give us information about feral goat pressure on vegetation, we assessed the predation impact of feral goats on the shrub Euphorbia dendroides (Euphorbiaceae) on Mallorca Island (Spain). We aimed to investigate whether goats consume juvenile E. dendroides and affect their population structure and determine if the plants increase the concentrations of toxic compounds as an adaptation to herbivory. Overall, two experimental plots and analysis of eleven natural populations indicated E. dendroides is affected by ungulates and that the population structure change with the presence of feral goats. Euphorbia dendroides could be used as an ecological indicator to determine the extent of ungulate damage to vegetation or indicate poor food availability, and thus inform the maintenance of optimal animal populations. Depending on the management objective for the territory, E. dendroides could be used as an ecological indicator to determine the extent of ungulate damage to vegetation or indicate poor food availability for animals, and thus inform the maintenance of optimal animal populations.

Historic exposure to herbivores, not constitutive traits, explains plant tolerance to herbivory in the case of two Medicago species (Fabaceae).

Mechanisms that allow plants to survive and reproduce after herbivory are considered to play a key role in plant evolution. In this study, we evaluated how tolerance varies in species with different historic exposure to herbivores considering ontogeny. We exposed the range-restricted species Medicago citrina and its closely related and widespread species M. arborea to one and two herbivory simulations (80 % aerial biomass loss). Physiological and growth parameters related to tolerance capacity were assessed to evaluate constitutive values (without herbivory) and induced tolerance after damage. Constitutive traits were not always related to greater tolerance, and each species compensated for herbivory through different traits. Herbivory damage only led to mortality in M. citrina; adults exhibited root biomass loss and increased oxidative stress after damage, but also compensated aerial biomass. Despite seedlings showed a lower death percentage than adults after herbivory in M. citrina, they showed less capacity to recover control values than adults. Moderate tolerance to M. arborea herbivory and low tolerance to M. citrina is found. Thus, although the constitutive characteristics are maintained in the lineage, the tolerance of plants decreases in M. citrina. That represents how plants respond to the lack of pressure from herbivores in their habitat.

Preference by Donkeys and Goats among Five Mediterranean Forest Species: Implications for Reducing Fire Hazard.

During the second half of the 20th century, European countries experienced an increase in their forest area due to the global change. Consequently, there has been an increase in large forest fires, mainly in the Mediterranean basin, and this has forced the development of several types of prevention programs. One of them is the control of the understory by livestock. In this sense, browsing with a combination of donkeys and goats could be a good option, as both animals usually feed on forest species. However, little is known about their preferences for the key species of the Mediterranean forest. Using a cafeteria test, the preferences and consumption of both animals have been determined for five typical species of the Mediterranean forest, such as Quercus ilex, Pinus halepensis, Phillyrea latifolia, Rubus ulmifolius, and Brachypodium retusum. Results showed that donkeys and goats could act complementarily in the reduction of the fuel biomass of forests. Donkeys appear to act more on fine fuel, such as B. retusum, and goats on the more pyrophyte species, in this case P. halepensis. In addition, given that donkeys are at severe risk of extinction in Europe, this role of providing ecosystem services could contribute to their conservation. Despite this study only showing that goats and donkeys would consume all five presented plant species and that there are some differences in consumption during a short-term test, it constitutes a useful first step for conservation and fire prevention in the Mediterranean forests.

Remote mapping of foodscapes using sUAS and a low cost BG-NIR sensor.

The assessment of landscape condition for large herbivores, also known as foodscapes, is fast gaining interest in conservation and landscape management programs worldwide. Although traditional approaches are now being replaced by satellite imagery, several technical issues still need to be addressed before full standardization of remote sensing methods for these purposes. We present a low-cost method, based on the use of a modified blue/green/near-infrared (BG-NIR) camera housed on a small-Unmanned Aircraft System (sUAS), to create foodscapes for a generalist Mediterranean ungulate: the Iberian Ibex (Capra pyrenaica) in Northeast Spain. Faecal cuticle micro-histological analyses were used to assess the dietary preferences of ibexes and then individuals of the most common plant species (n = 19) were georeferenced to use as test samples. Because of the seasonal pattern in vegetation activity, based on the NDVI (Smooth term Month = 21.5, p-value < .01, R2 = 43%, from a GAM), images were recorded in winter and spring to represent contrasting vegetation phenology using two flight heights above ground level (30 and 60 m). Additionally, the range of image pixel sizes was 3.5-30 cm with the smallest pixel size representing the highest resolution. Boosted Trees were used to classify plant taxa based on spectral reflectance and create a foodscape of the study area. The number of target species, the sampling season, the height of flight and the image resolution were analysed to determine the accuracy of mapping the foodscape. The highest classification error (70.66%) was present when classifying all plant species using a 30 cm pixel size from acquisitions at 30 m height. The lowest error (18.7%), however, was present when predicting plants preferred by ibexes, at 3.5 cm pixel size acquired at 60 m height. This methodology can help to successfully monitor food availability and seasonality and to identify individual species.

EasyModel: user-friendly tool for building and analysis of simple mathematical models in systems biology.

SUMMARY: EasyModel is a new user-friendly web application that contains ready-for-simulation versions of the BioModels Database, and allows for the intuitive creation of new models. Its main target audience is the experimental biologist and students of bioinformatics or systems biology without programming skills. Expert users can also benefit from it by implementing basic models quickly and downloading the code for further tailoring. AVAILABILITY AND IMPLEMENTATION: Freely available on the web at https://easymodel.udl.cat. Implementation is described in its own section.

Comparing the accuracy of PCR-capillary electrophoresis and cuticle microhistological analysis for assessing diet composition in ungulates: A case study with Pyrenean chamois.

The study of diet composition is required to understand the interactions between animal and plant ecosystems. Different non-invasive techniques applied on faecal samples have commonly been used for such purposes, with cuticle microhistological analysis (CMA) and emerging DNA-based methods being the most relevant. In this work, we refined and optimized a qualitative DNA-based approach combining PCR amplification of long trnL(UAA) and ITS2 fragments and capillary electrophoresis (PCR-CE), instead of short trnL(UAA) fragments and massive sequencing technologies commonly reported. To do so, we developed a controlled diet assay using a stabled Pyrenean chamois specimen (Rupicapra pyrenaica pyrenaica), which included representative herbaceous and shrubby plant species. We also assessed the impact of sample freshness on the diet determination of this mountain caprinae by exposing faecal samples to the outdoor environment for three weeks. Faecal samples from both experiments were analysed by qualitative PCR-CE and semi-quantitative CMA in order to compare the pros and cons of both approaches. Our results show that all of the offered plant species were detected by both methodologies although CMA over-detected shrubs compared to herbaceous species. At the same time, sample degradation due to sustained climate exposure is a limiting factor for molecular analysis, but not for CMA. Taken all together, our results suggest that the qualitative information obtained by CMA and PCR-CE can be interchangeable when faecal samples are fresh (less than one week after deposition) but, afterwards, molecular analysis underestimates diet composition probably due to DNA degradation. CMA, however, can accurately be used at least three weeks after defecation. Moreover, by combining the results of simultaneous PCR amplification of two complementary genes, this optimized PCR-CE methodology provides a reliable, feasible and more affordable alternative for multiple and routine analyses of complex samples. Neither CMA nor PCR-CE seems to solve comprehensively the quatification of herbivore diets and thus further research needs to be done.

Different effects of alpine woody plant expansion on domestic and wild ungulates.

Changes in land-use and climate affect the distribution and diversity of plant and animal species at different spatiotemporal scales. The extent to which species-specific phenotypic plasticity and biotic interactions mediate organismal adaptation to changing environments, however, remains poorly understood. Woody plant expansion is threatening the extent of alpine grasslands worldwide, and evaluating and predicting its effects on herbivores is of crucial importance. Here, we explore the impact of shrubification on the feeding efficiency of Pyrenean chamois (Rupicapra p. pyrenaica), as well as on the three most abundant coexisting domestic ungulate species: cattle, sheep and horses. We use observational diet composition from May to October and model different scenarios of vegetation availability where shrubland and woodland proliferate at the expense of grassland. We then predicted if the four ungulate species could efficiently utilize their food landscapes with their current dietary specificities measuring their niche breath in each scenario. We observed that the wild counterpart, due to a higher trophic plasticity, is less disturbed by shrubification compared to livestock, which rely primarily on herbaceous plants and will be affected 3.6 times more. Our results suggest that mixed feeders, such as chamois, could benefit from fallow landscapes, and that mountain farmers are at a growing economic risk worldwide due to changing land-use practices and climate conditions.

Effects of boom and bust grazing management on vegetation and health of beef cattle used for wildfire prevention in a Mediterranean forest.

Humans and wildfires have historically driven landscape structure in the Mediterranean basin. The Iberian Peninsula is not an exception to that rule, and therefore, farmers, researchers, and governments seek alternative tools to minimize the loss of biodiversity and wildfire risks. Extensive livestock including beef cattle is currently promoted as a suitable management tool by European agro-environmental policies yet pieces of evidence exist regarding the reciprocal effects between cows and Mediterranean woody vegetation. In this work, we performed a field manipulation to evaluate whether free-ranging beef cattle without supplementary feeding, at high density (2 livestock units (LU)/ha) for a short period of time i.e. "boom and bust grazing" management, are able to adapt their grazing preferences to the Mediterranean woody vegetation without health impairment, and prevent from bush encroachment and wildfires. For our purposes, a native herd of 14 adult cows was kept captive without supplementary feeding in a 14 ha enclosure covered by Mediterranean vegetation for two months (April-June 2016). Plant and cattle fecal and blood samples were collected to assess diet composition (plant cuticle microhistological analysis), fecal nitrogen and protein contents of consumed plants, and the nutritional status (non-esterified fatty acids) of cattle. Our results showed that cattle adapted their feeding habits toward a more woody diet including potentially flammable taxa but with some detrimental effects on health status. Hence, cattle cannot control woody vegetation for long periods of time without supplementary feeding. Further research should be oriented to explore other alternative approaches to minimize the health impairment of cattle used for control flammable vegetation in Mediterranean regions.

Seasonal diet composition of Pyrenean chamois is mainly shaped by primary production waves.

In alpine habitats, the seasonally marked climatic conditions generate seasonal and spatial differences in forage availability for herbivores. Vegetation availability and quality during the growing season are known to drive life history traits of mountain ungulates. However, little effort has been made to understand the association between plant phenology and changes in the foraging strategies of these mountain dwellers. Furthermore, this link can be affected by the seasonal presence of livestock in the same meadows. The objective of this work was to study the seasonal changes in diet composition of Pyrenean chamois (Rupicapra p. pyrenaica) and its relationship to primary production trends in a Mediterranean alpine environment. Moreover, diet composition in two populations with contrasting livestock pressure was compared in order to study the effect of sheep flocks on the feeding behaviour of chamois. From 2009 to 2012, monthly diet composition was estimated by cuticle microhistological analysis of chamois faeces collected in the eastern Pyrenees. The primary production cycle was assessed by remote sensing, using the Normalized Difference Vegetation Index. Additionally, the diet of sheep sharing seasonally the subalpine and alpine meadows with chamois was analysed. Diet selection of chamois and sheep and their overlap was also assessed. Our results show an intra-annual variation in the diet composition of Pyrenean chamois and demonstrate a strong relationship between plant consumption dynamics and phenology in alpine areas. In addition, Calluna vulgaris, Cytisus spp. and Festuca spp., as well as forbs in the summer, are found to be key forage species for Pyrenean chamois. Furthermore, this study couldn't detect differences between both chamois populations despite the presence of sheep flocks in only one area. However, the detection of a shift in the diet of chamois in both areas after the arrival of high densities of multi-specific livestock suggest a general livestock effect. In conclusion, Pyrenean chamois are well adapted to the variations in the seasonal availability of plants in alpine habitats but could be disturbed by the seasonal presence of livestock. Due to the key plants in their diet, we suggest that population management programmes should focus on the preservation of mixed grasslands composed of patches of shrubs and herbs. The effects of climate change and shrub expansion should be studied as they may potentially affect chamois population dynamics through changes in habitat composition and temporal shifts in forage availability.

Predicting herbivore faecal nitrogen using a multispecies near-infrared reflectance spectroscopy calibration.

Optimal management of free-ranging herbivores requires the accurate assessment of an animal's nutritional status. For this purpose 'near-infrared reflectance spectroscopy' (NIRS) is very useful, especially when nutritional assessment is done through faecal indicators such as faecal nitrogen (FN). In order to perform an NIRS calibration, the default protocol recommends starting by generating an initial equation based on at least 50-75 samples from the given species. Although this protocol optimises prediction accuracy, it limits the use of NIRS with rare or endangered species where sample sizes are often small. To overcome this limitation we tested a single NIRS equation (i.e., multispecies calibration) to predict FN in herbivores. Firstly, we used five herbivore species with highly contrasting digestive physiologies to build monospecies and multispecies calibrations, namely horse, sheep, Pyrenean chamois, red deer and European rabbit. Secondly, the equation accuracy was evaluated by two procedures using: (1) an external validation with samples from the same species, which were not used in the calibration process; and (2) samples from different ungulate species, specifically Alpine ibex, domestic goat, European mouflon, roe deer and cattle. The multispecies equation was highly accurate in terms of the coefficient of determination for calibration R2 = 0.98, standard error of validation SECV = 0.10, standard error of external validation SEP = 0.12, ratio of performance to deviation RPD = 5.3, and range error of prediction RER = 28.4. The accuracy of the multispecies equation to predict other herbivore species was also satisfactory (R2 > 0.86, SEP < 0.27, RPD > 2.6, and RER > 8.1). Lastly, the agreement between multi- and monospecies calibrations was also confirmed by the Bland-Altman method. In conclusion, our single multispecies equation can be used as a reliable, cost-effective, easy and powerful analytical method to assess FN in a wide range of herbivore species.

Mass scans from a proton transfer mass spectrometry analysis of air over Mediterranean shrubland browsed by horses.

Plants usually emit large amount and varieties of volatiles after being damaged by herbivores. However, analytical methods for measuring herbivore-induced volatiles do not normally monitor the whole range of volatiles and the response to large herbivores such as large mammals is much less studied than the response to other herbivores such as insects. In this paper we present the results of using a highly sensitive proton transfer reaction-mass spectrometry (PTR-MS) technique that allows simultaneous monitoring of leaf volatiles in the pptv range. The resulting mass scans in air over Mediterranean shrubland browsed by horses show 70 to 100% higher concentrations of the masses corresponding to mass fragments 57, 43 and 41 (mostly hexenals, acetone and acetic acid) than scans over control non-browsed shrubland. These compounds are biogeochemically active and they are significant components of the volatile organic carbon found in the atmosphere. They influence the performance of living organisms and, the chemical and physical processes of Earth's atmosphere.