Assessing accuracy of GAP and LANDFIRE land cover datasets in winter habitats used by greater sage-grouse in Idaho and Wyoming, USA.

Remotely sensed land cover datasets have been increasingly employed in studies of wildlife habitat use. However, meaningful interpretation of these datasets is dependent on how accurately they estimate habitat features that are important to wildlife. We evaluated the accuracy of the GAP dataset, which is commonly used to classify broad cover categories (e.g., vegetation communities) and LANDFIRE datasets, which classifies narrower cover categories (e.g., plant species) and structural features of vegetation. To evaluate accuracy, we compared classification of cover types and estimates of percent cover and height of sagebrush (Artemisia spp.) derived from GAP and LANDFIRE datasets to field-collected data in winter habitats used by greater sage-grouse (Centrocercus urophasianus). Accuracy was dependent on the type of dataset used as well as the spatial scale (point, 500-m, and 1-km) and biological level (community versus dominant species) investigated. GAP datasets had the highest overall classification accuracy of broad sagebrush cover types (49.8%) compared to LANDFIRE datasets for narrower cover types (39.1% community-level; 31.9% species-level). Percent cover and height were not accurately estimated in the LANDFIRE dataset. Our results suggest that researchers must be cautious when applying GAP or LANDFIRE datasets to classify narrow categories of land cover types or to predict percent cover or height of sagebrush within sagebrush-dominated landscapes. We conclude that ground-truthing is critical for successful application of land cover datasets in landscape-scale evaluations and management planning, particularly when wildlife use relatively rare habitat types compared to what is available.

Metagenomic sequencing provides insights into microbial detoxification in the guts of small mammalian herbivores (Neotoma spp.).

Microbial detoxification of plant toxins influences the use of plants as food sources by herbivores. Stephen's woodrats (Neotoma stephensi) specialize on juniper, which is defended by oxalate, phenolics and monoterpenes, while closely related N. albigula specialize on cactus, which only contains oxalate. Woodrats maintain two gut chambers harboring dense microbial communities: a foregut chamber proximal to the major site of toxin absorption, and a cecal chamber in their hindgut. We performed several experiments to investigate the location and nature of microbial detoxification in the woodrat gut. First, we measured toxin concentrations across gut chambers of N. stephensi. Compared to food material, oxalate concentrations were immediately lower in the foregut, while concentrations of terpenes remained high in the foregut, and were lowest in the cecal chamber. We conducted metagenomic sequencing of the foregut chambers of both woodrat species and cecal chambers of N. stephensi to compare microbial functions. We found that most genes associated with detoxification were more abundant in the cecal chambers of N. stephensi. However, some genes associated with degradation of oxalate and phenolic compounds were more abundant in the foregut chambers. Thus, microbial detoxification may take place in various chambers depending on the class of chemical compound.

Microbial detoxification in the gut of a specialist avian herbivore, the Greater Sage-Grouse.

One function of the gut microbiota gaining recent attention, especially in herbivorous mammals and insects, is the metabolism of plant secondary metabolites (PSMs). We investigated whether this function exists within the gut communities of a specialist avian herbivore. We sequenced the cecal metagenome of the Greater Sage-Grouse (Centrocercus urophasianus), which specializes on chemically defended sagebrush (Artemisia spp.). We predicted that the cecal metagenome of the sage-grouse would be enriched in genes associated with the metabolism of PSMs when compared to the metagenome of the domestic chicken. We found that representation of microbial genes associated with 'xenobiotic degradation and metabolism' was 3-fold higher in the sage-grouse cecal metagenomes when compared to that of the domestic chicken. Further, we identified a complete metabolic pathway for the degradation of phenol to pyruvate, which was not detected in the metagenomes of the domestic chicken, bovine rumen or 14 species of mammalian herbivores. Evidence of monoterpene degradation (a major class of PSMs in sagebrush) was less definitive, although we did detect genes for several enzymes associated with this process. Overall, our results suggest that the gut microbiota of specialist avian herbivores plays a similar role to the microbiota of mammalian and insect herbivores in degrading PSMs.

ANTIOXIDANT CAPACITY OF WYOMING BIG SAGEBRUSH (ARTEMISIA TRIDENTATA SSP. WYOMINGENSIS) VARIES SPATIALLY AND IS NOT RELATED TO THE PRESENCE OF A SAGEBRUSH DIETARY SPECIALIST.

Sagebrush (Artemisia spp.) in North America is an abundant native plant species that is ecologically and evolutionarily adapted to have a diverse array of biologically active chemicals. Several of these chemicals, specifically polyphenols, have antioxidant activity that may act as biomarkers of biotic or abiotic stress. This study investigated the spatial variation of antioxidant capacity, as well as the relationship between a mammalian herbivore and antioxidant capacity in Wyoming big sagebrush (Artemisia tridentata wyomingensis). We quantified and compared total polyphenols and antioxidant capacity of leaf extracts from sagebrush plants from different spatial scales and at different levels of browsing by a specialist mammalian herbivore, the pygmy rabbit (Brachylagus idahoensis). We found that antioxidant capacity of sagebrush extracts was positively correlated with total polyphenol content. Antioxidant capacity varied spatially within and among plants. Antioxidant capacity in sagebrush was not related to either browsing intensity or duration of association with rabbits. We propose that the patterns of antioxidant capacity observed in sagebrush may be a result of spatial variation in abiotic stress experienced by sagebrush. Antioxidants could therefore provide a biomarker of environmental stress for sagebrush that could aid in management and conservation of this plant in the threatened sagebrush steppe.

Monoterpenes as inhibitors of digestive enzymes and counter-adaptations in a specialist avian herbivore.

Many plants produce plant secondary metabolites (PSM) that inhibit digestive enzymes of herbivores, thus limiting nutrient availability. In response, some specialist herbivores have evolved digestive enzymes that are resistant to inhibition. Monoterpenes, a class of PSMs, have not been investigated with respect to the interference of specific digestive enzymes, nor have such interactions been studied in avian herbivores. We investigated this interaction in the Greater Sage-Grouse (Phasianidae: Centrocercus urophasianus), which specializes on monoterpene-rich sagebrush species (Artemisia spp.). We first measured the monoterpene concentrations in gut contents of free-ranging sage-grouse. Next, we compared the ability of seven individual monoterpenes present in sagebrush to inhibit a protein-digesting enzyme, aminopeptidase-N. We also measured the inhibitory effects of PSM extracts from two sagebrush species. Inhibition of aminopeptidase-N in sage-grouse was compared to inhibition in chickens (Gallus gallus). We predicted that sage-grouse enzymes would retain higher activity when incubated with isolated monoterpenes or sagebrush extracts than chicken enzymes. We detected unchanged monoterpenes in the gut contents of free-ranging sage-grouse. We found that three isolated oxygenated monoterpenes (borneol, camphor, and 1,8-cineole) inhibited digestive enzymes of both bird species. Camphor and 1,8-cineole inhibited enzymes from chickens more than from sage-grouse. Extracts from both species of sagebrush had similar inhibition of chicken enzymes, but did not inhibit sage-grouse enzymes. These results suggest that specific monoterpenes may limit the protein digestibility of plant material by avian herbivores. Further, this work presents additional evidence that adaptations of digestive enzymes to plant defensive compounds may be a trait of specialist herbivores.

Modeling trade-offs between plant fiber and toxins: a framework for quantifying risks perceived by foraging herbivores.

When selecting habitats, herbivores must weigh multiple risks, such as predation, starvation, toxicity, and thermal stress, forcing them to make fitness trade-offs. Here, we applied the method of paired comparisons (PC) to investigate how herbivores make trade-offs between habitat features that influence selection of food patches. The method of PC measures utility and the inverse of utility, relative risk, and makes trade-offs and indifferences explicit by forcing animals to make choices between two patches with different types of risks. Using a series of paired-choice experiments to titrate the equivalence curve and find the marginal rate of substitution for one risk over the other, we evaluated how toxin-tolerant (pygmy rabbit Brachylagus idahoensis) and fiber-tolerant (mountain cottontail rabbit Sylviagus nuttallii) herbivores differed in their hypothesized perceived risk of fiber and toxins in food. Pygmy rabbits were willing to consume nearly five times more of the toxin 1,8-cineole in their diets to avoid consuming higher levels of fiber than were mountain cottontails. Fiber posed a greater relative risk for pygmy rabbits than cottontails and cineole a greater risk for cottontails than pygmy rabbits. Our flexible modeling approach can be used to (1) quantify how animals evaluate and trade off multiple habitat attributes when the benefits and risks are difficult to quantify, and (2) integrate diverse risks that influence fitness and habitat selection into a single index of habitat value. This index potentially could be applied to landscapes to predict habitat selection across several scales.

The dilemma of foraging herbivores: dealing with food and fear.

For foraging herbivores, both food quality and predation risk vary across the landscape. Animals should avoid low-quality food patches in favour of high-quality ones, and seek safe patches while avoiding risky ones. Herbivores often face the foraging dilemma, however, of choosing between high-quality food in risky places or low-quality food in safe places. Here, we explore how and why the interaction between food quality and predation risk affects foraging decisions of mammalian herbivores, focusing on browsers confronting plant toxins in a landscape of fear. We draw together themes of plant-herbivore and predator-prey interactions, and the roles of animal ecophysiology, behaviour and personality. The response of herbivores to the dual costs of food and fear depends on the interplay of physiology and behaviour. We discuss detoxification physiology in dealing with plant toxins, and stress physiology associated with perceived predation risk. We argue that behaviour is the interface enabling herbivores to stay or quit food patches in response to their physiological tolerance to these risks. We hypothesise that generalist and specialist herbivores perceive the relative costs of plant defence and predation risk differently and intra-specifically, individuals with different personalities and physiologies should do so too, creating individualised landscapes of food and fear. We explore the ecological significance and emergent impacts of these individual-based foraging outcomes on populations and communities, and offer predictions that can be clearly tested. In doing so, we provide an integrated platform advancing herbivore foraging theory with food quality and predation risk at its core.

Plant protein and secondary metabolites influence diet selection in a mammalian specialist herbivore.

For herbivores, nutrient intake is limited by the relatively low nutritional quality of plants and high concentrations of potentially toxic defensive compounds (plant secondary metabolites, PSMs) produced by many plants. In response to phytochemical challenges, some herbivores selectively forage on plants with higher nutrient and lower PSM concentrations relative to other plants. Pygmy rabbits (Brachylagus idahoensis) are dietary specialists that feed on sagebrush (Artemisia spp.) and forage on specific plants more than others within a foraging patch. We predicted that the plants with evidence of heavy foraging (browsed plants) would be of higher dietary quality than plants that were not browsed (unbrowsed). We used model selection to determine which phytochemical variables best explained the difference between browsed and unbrowsed plants. Higher crude protein increased the odds that plants would be browsed by pygmy rabbits and the opposite was the case for certain PSMs. Additionally, because pygmy rabbits can occupy foraging patches (burrows) for consecutive years, their browsing may influence the nutritional and PSM constituents of plants at the burrows. In a post hoc analysis, we did not find a significant relationship between phytochemical concentrations, browse status and burrow occupancy length. We concluded that pygmy rabbits use nutritional and chemical cues while making foraging decisions.

A pharm-ecological perspective of terrestrial and aquatic plant-herbivore interactions.

We describe some recent themes in the nutritional and chemical ecology of herbivores and the importance of a broad pharmacological view of plant nutrients and chemical defenses that we integrate as "Pharm-ecology". The central role that dose, concentration, and response to plant components (nutrients and secondary metabolites) play in herbivore foraging behavior argues for broader application of approaches derived from pharmacology to both terrestrial and aquatic plant-herbivore systems. We describe how concepts of pharmacokinetics and pharmacodynamics are used to better understand the foraging phenotype of herbivores relative to nutrient and secondary metabolites in food. Implementing these concepts into the field remains a challenge, but new modeling approaches that emphasize tradeoffs and the properties of individual animals show promise. Throughout, we highlight similarities and differences between the historic and future applications of pharm-ecological concepts in understanding the ecology and evolution of terrestrial and aquatic interactions between herbivores and plants. We offer several pharm-ecology related questions and hypotheses that could strengthen our understanding of the nutritional and chemical factors that modulate foraging behavior of herbivores across terrestrial and aquatic systems.

Mechanisms for eliminating monoterpenes of sagebrush by specialist and generalist rabbits.

Pygmy rabbits (Brachylagus idahoensis) are one of only three vertebrates that subsist virtually exclusively on sagebrush (Artemisia spp.), which contains high levels of monoterpenes that can be toxic. We examined the mechanisms used by specialist pygmy rabbits to eliminate 1,8-cineole, a monoterpene of sagebrush, and compared them with those of cottontail rabbits (Sylvilagus nuttalli), a generalist herbivore. Rabbits were offered food pellets with increasing concentrations of cineole, and we measured voluntary intake and excretion of cineole metabolites in feces and urine. We expected pygmy rabbits to consume more, but excrete cineole more rapidly by using less-energetically expensive methods of detoxification than cottontails. Pygmy rabbits consumed 3-5 times more cineole than cottontails relative to their metabolic body mass, and excreted up to 2 times more cineole metabolites in their urine than did cottontails. Urinary metabolites excreted by pygmy rabbits were 20 % more highly-oxidized and 6 times less-conjugated than those of cottontails. Twenty percent of all cineole metabolites recovered from pygmy rabbits were in feces, whereas cottontails did not excrete fecal metabolites. When compared to other mammals that consume cineole, pygmy rabbits voluntarily consumed more, and excreted more cineole metabolites in feces, but they excreted less oxidized and more conjugated cineole metabolites in urine. Pygmy rabbits seem to have a greater capacity to minimize systemic exposure to cineole than do cottontails, and other cineole-consumers, by minimizing absorption and maximizing detoxification of ingested cineole. However, mechanisms that lower systemic exposure to cineole may come with a higher energetic cost in pygmy rabbits than in other mammalian herbivores.

Inhibition of snowshoe hare succinate dehydrogenase activity as a mechanism of deterrence for papyriferic acid in birch.

The plant secondary metabolite papyriferic acid (PA) deters browsing by snowshoe hares (Lepus americanus) on the juvenile developmental stage of the Alaska paper birch (Betula neoalaskana). However, the physiological mechanism that reduces browsing remains unknown. We used pharmacological assays and molecular modeling to test the hypothesis that inhibition of succinate dehydrogenase (SDH) is a mode of action (MOA) of toxicity of PA in snowshoe hares. We tested this hypothesis by measuring the effect of PA on the activity of SDH in liver mitochondria isolated from wild hares. In addition, we used molecular modeling to determine the specific binding site of PA on SDH. We found that PA inhibits SDH from hares by an uncompetitive mechanism in a dose-dependent manner. Molecular modeling suggests that inhibition of SDH is a result of binding of PA at the ubiquinone binding sites in complex II. Our results provide a MOA for toxicity that may be responsible for the concentration-dependent anti-feedant effects of PA. We propose that snowshoe hares reduce the dose-dependent toxic consequences of PA by relying on efflux transporters and metabolizing enzymes that lower systemic exposure to dietary PA.