Meals, Metabolism, Microbes, and Management: Linking Foraging Behavior and Physiology to Predict Demographics in Wildlife (hosted by TWS)

At every meal, herbivores face the challenge of balancing their nutritional needs with potential exposure to toxic plant secondary compounds. New genetic and chemistry techniques are providing unprecedented views into the nutritional and chemical opportunities and challenges that plants present herbivores, and likewise, how herbivores and their gut microbial communities respond to the nutritional and chemical profiles of their food plants. In this symposium, we use wild herbivore systems including the sage-grouse, pygmy rabbits, woodrats and mule deer as case studies of how plant chemicals challenge herbivores, and the genetic, behavioral and physiological mechanisms that mammalian and avian herbivores have evolved to overcome these challenges. We explore the role of gut microbial communities in herbivore foraging ecology to understand how microbes aid herbivores in converting plant matter into biomass while also limiting exposure to toxins. Moreover, we demonstrate how the molecular interactions between plants, herbivores and their gut microbes could alter the nutritional condition of wild herbivores. Using integrated models we predict individual fitness and population trajectories resulting from changes in plant quality and the foraging behavior and physiology of herbivores and their gut microbes. These new technologies and approaches provide wildlife managers new insight into how to monitor and manipulate the plant and herbivores traits that allow herbivores to grow, survive and reproduce, and provide new opportunities to manage landscapes and populations of herbivores.

1:10PM Plant Secondary Compounds; Why Animals Hate Them but Love the Plants They’Re in.
  Lora Richards, Danny Nielsen, Kaitlin Ochsenrider, Chris Jeffrey, Marjorie Matocq
Divergence in ecologically adaptive traits can be maintained over small spatial scales, even in the presence of gene flow. Strong selective gradients such as those that exist at ecological boundaries often present animals with a sharp transition in various resources, including the availability of food. Mammalian herbivores meet the demands of nutrient acquisition and plant detoxification through their own metabolic mechanisms, as well as those of their gut microbiome. In order to begin investigating how mammalian herbivores respond to spatial and temporal variation in plant availability, we use DNA sequencing and chemical profiling of fecal samples to: Quantify seasonal shifts in foraging ecology, gut microbiome, and metabolomics across a sharp ecotone where two closely related small mammal species (Neotoma bryanti and N. lepida) come into contact and hybridize. We found sharp transitions in diet, microbial community and metabolomic signatures that coincide with the rapid habitat transition. Woodrats maintained habitat-specific diets that contain distinctly different phytochemistry that would likely require different metabolic processing. Diet diversity increased in the spring for both species. This included taking advantage of ephemerally-available annual forbs, like Phacelia, and their gut microbiome may shift with these seasonal changes in diet. Fecal metabolomic profiles are also habitat specific, but converge in spring when diets diversify and widespread annuals become available. Through fecal analyses we were gained insight in the feeding ecology of two woodrat species that come in close contact and generate hypotheses on the mechanisms maintaining the separation of these species.
1:30PM Natural Diets Promote Retention of Native Microflora of Captive Wildlife
  Rodolfo Martinez-Mota, Denise Dearing, Teri Orr, Kevin Kohl
Wildlife entering into captivity experience radical lifestyle changes resulting in microbiome alterations. However, it is unclear which factors drive microbial community shifts in captivity, and what actions could mitigate microbial changes. Using the white-throated woodrat (Neotoma albigula), we tested whether offering natural diets in captivity facilitates retention of native microbial communities of captive animals. Wild-caught woodrats were brought to laboratory conditions. Woodrats received either a natural diet of cactus or an artificial diet of commercial chow over three weeks. Microbial inventories from woodrat feces at the time of capture and in captivity were generated using Illumina 16S rRNA sequencing. We found that providing woodrats with wild natural diets significantly mitigated alterations in their microbiota, promoting a 90% retention of native microbial communities across the experiment. In contrast, the artificial diet significantly impacted microbial structure to the extent that 38% of the natural microflora was lost. Core bacteria including Bifidobacterium and Allobaculum were lost, and abundances of microbes related to oxalate degradation decreased in individuals fed the artificial but not the natural diet. These results highlight the importance of supplementing captive diets with natural foods to maintain native microbiomes of wildlife kept in artificial conditions for scientific or conservation purposes.
1:50PM Nutrient Manipulation Differentially Affects Gut Anatomy and Microbiome Structure in Rodents with Distinct Dietary Niches
  Brian Trevelline, Rodolfo Martinez-Mota, Terry Derting, Andrea Darracq, Bret Pasch, Denise Dearing, Kevin Kohl
Mammals must extract sufficient energy and nutrients from their diets for survival and reproduction. The digestive system and its resident gut microbiota are highly dynamic and responsive to diet, likely aiding in the maintenance of optimal digestion. However, studies investigating microbial and physiological responses to diet are typically conducted on a single species. Therefore, we have poor understanding of how the flexibility of the digestive system and gut microbiome structure varies across species. We conducted feeding trials with three species of rodents with distinct dietary niches: montane voles (Microtus montanus, herbivorous), white-footed mice (Peromyscus leucopus, omnivorous), and southern grasshopper mice (Onychomys torridus, insectivorous). Rodents were fed four different diets varying in their concentrations of fiber and protein for a period of five weeks. Rodents were dissected for measurements of gut morphology, and gut content samples were collected to inventory microbial communities via 16S rRNA sequencing. We found that several aspects of gut anatomy exhibited species-specific responses to diet. For example, in voles small intestinal length showed no changes in voles, while in white-footed mice it increased in length in response to high fiber diets, and in grasshopper mice it increased in length in response to low protein diets. Similarly, the gut microbiota exhibited species-specific responses to diet. These data suggest that the flexibility of the digestive system and gut microbiota may be adapted to species-specific dietary niches.
2:10PM How Plants Outwit Their Consumers. What Plant Chemicals Do to Defeat Specialist Herbivores.
  Carolyn Dadabay, Jennifer Forbey
The sagebrush ecosystems of the western United States provide a model of complex interactions at the chemical level between co-evolving plants and herbivores. Plants of the sagebrush family (Artemisia tridentata) are under selective pressure to synthesize a large number and variety of toxic compounds to deter attacking herbivores. Despite this diversity of defensive phytochemicals, greater sage-grouse (Centrocercus urophasianus) is a dietary specialist herbivore, which has evolved a high toxin tolerance phenotype defined as maintaining physiological function even with relatively high toxin intake. Two major groups of genes contribute to metabolic mechanisms of toxin tolerance: metabolizing enzymes and efflux transporters. Metabolizing enzymes (e.g., cytochrome P450s/CYP enzymes) function to speed the metabolism of toxins and efflux transporters (e.g., ABC type such as P-gp or MDR1) function to limit intracellular concentrations of toxins. While these mechanisms enable the greater sage-grouse to utilize dietary sagebrush, particularly in winter months, sage grouse are highly selective toward the species, patch, and even individual sagebrush plants, which they consume. We hypothesize that these avoided species of sagebrush produce defensive chemical panels that act upon the toxin metabolizing pathways of sage-grouse, knocking out the herbivore’s metabolic defenses. We have characterized two classes of sagebrush defensive compounds, polyphenols and sesquiterpene lactones. We utilize grouse microsomal CYP enzymes in assays with sagebrush compounds as both substrates and inhibitors to characterize the metabolism of these compounds as well as to ascertain the ability of plant compounds to interfere with herbivore chemical defenses. Understanding the metabolic mechanism of toxin tolerance in herbivores is one component of predicting demographic consequences of toxin exposure in wildlife species.
2:30PM How Herbivores Outwit Their Plants over Evolutionary Time; Sage-Grouse Versus Sagebrush.
  Sara Oyler-McCance
Herbivores rely on plants for food and, in turn, many of those plants have evolved defense mechanisms against such herbivory. Herbivores must then develop counter-adaptations and co-evolve in order to survive. This evolutionary arms race between plants and the herbivores that eat them results in a series of adaptations aimed at continually outwitting the other player in the plant-herbivore system. Here, I discuss current efforts to investigate the link between plants and herbivores in the sagebrush ecosystem by examining local adaptation in Greater and Gunnison Sage-grouse, both sagebrush-obligate species. Substantial geographic variation exists in both the composition and concentration of plant secondary metabolites in sagebrush, indicating that sage-grouse populations throughout their range may be exposed to distinct natural selection imposed by the chemistry of local sagebrush varieties. Research efforts are ongoing examining whether Greater and Gunnison Sage-grouse populations are locally adapted to the specific varieties of sagebrush growing in those populations. Independent studies in both species of sage-grouse using different genomic approaches revealed similar patterns of selection in single nucleotide polymorphisms linked to genes in the cytochrome P450 gene family, which could indicate adaptive divergence for genes involved in the metabolism of plant secondary metabolites in sagebrush. Such information is important for management in that it could guide efforts in sagebrush restoration and the translocation of individuals of both species of sage-grouse.
2:50PM Refreshment Break
3:20PM Towards Functional Community Ecology in the Metagenomes of Herbivore Specialists
  Eric J. Hayden, Jennifer Forbey
Recent research is beginning to reveal the importance of the microbiome for herbivore specialization. Sagebrush herbivory is an excellent model system for studying the role of the microbiome in herbivore specialization. Sagebrush and its herbivores have been locked in a co-evolutionary arms race for at least 12 million years. Sagebrush synthesizes hundreds of monoterpenes, polyphenolics and sesquiterpene lactones that have known cytotoxic, anti-herbivore and anti-microbial properties. Moreover, ancestral clades of Artemisia are well known to possess chemicals with diverse, toxic, mechanisms of action. Despite this toxicity, there are a number of invertebrate and vertebrate herbivores that specialize on sagebrush for food. Because specialist herbivores often have mechanisms that limit the absorption of ingested lipophilic monoterpenes, microbes must be able to thrive within a gutt environment that contains concentrated and unaltered plant defense chemicals. Here, we describe our recent efforts to understand changes in microbial communities associated with site-specific and temporal shifts in sagebrush chemicals. We are focusing on shotgun metagenomics data to identify changes in the abundance of specific taxonomic groups and to identify important gene functions underlying these changes. In addition, we are developing analysis approaches to understand the assembly and stability of microbial community ecology within a given host. Our long-term goals include studying multiple species of herbivores and coupling microbiome data to metabolomic data in order to enable more general conclusions about the role of the microbiome in the plant herbivore arms race.
3:40PM Stealing Good Ideas from Cows: In-Vitro Gut Systems to Test Microbial Manipulations and Their Potential Use in “Soft Releases” of Wildlife.
  Mozart Fonseca
Gut fermentation studies and their role in animal nutritional requirements have been quantitatively studied since the 1940s. A combination of in vitro and in situ techniques are used in order to isolate gut microorganisms, quantify metabolites, and compare efficiency of gut microbial populations upon degradation of specific substrates in various compartments of the gastrointestinal tract. Other instrumental techniques such as the solubility test, the two-stage technique, the enzymatic-free cells methods, the ruminal simulation technique, gas production technique are also used in order to explore our understanding of how animals would benefit from by-products produced by gut microflora or even adapted to other compartments where microbial fermentation occurs (e.g.: large intestine). In continuous culture systems, because there is a regular addition of buffer and nutrients and a continuous withdrawal of fermentation products, microbial populations tend to stabilize and simulation of field conditions would no longer rely on harvesting animals for inoculum sampling every so often. These systems allow us to identify and quantify microbial population dynamics as well as digestion of nutrients under controlled conditions for long periods of time. Simulating in vivo and in loco dietary characteristics draw the baseline for comparisons between nutrient utilization under similar conditions for microbial populations of interest. The construction of anaerobic fermentation chambers that mimic the digestive processes are feasible for various animal species with adjustments performed according to anatomy, chemical and physiological characteristics of species of study, pH, redox potential, metabolites present, substrate supply characteristics, temperature, microbial site of attachment, particle size, nature of chemical compounds present, among many other factors. There meaningful biological interpretations help in the understanding of the dynamics of passage and degradation as regulators of intake, motility, gastric emptying, and overall requirements that dictate the capability of animals to survive and reproduce in specific ecological sites.
4:00PM Tradeoffs on the Landscape for Deer: How Fearscapes Interact with Foodscapes
  Kevin Monteith, Hall Sawyer, Jill Randall, Jeff Beck, Jennifer Forbey, Gary Fralick, Samantha Dwinnell
The availability and quality of forage on the landscape constitute the foodscape within which animals make behavioral decisions to acquire food. Novel changes to the foodscape, such as human disturbance, can alter behavioral decisions that favor avoidance of perceived risk over food acquisition. Although behavioral changes and population declines often coincide with the introduction of human disturbance, the link(s) between behavior and population trajectory are difficult to elucidate. To identify a pathway by which human disturbance may affect ungulate populations, we tested the Behaviorally Mediated Forage-Loss Hypothesis, wherein behavioral avoidance is predicted to reduce use in regions of the foodscape adjacent to disturbance. We used GPS-collar data collected from migratory mule deer (Odocoileus hemionus) to evaluate habitat selection, movement patterns, and time-budgeting behavior in response to varying levels of forage availability and human disturbance in three different populations exposed to a gradient of energy development. Subsequently, we linked animal behavior with measured use of forage relative to human disturbance, forage availability, and quality. Mule deer avoided human disturbance at both home range and winter range scales, but showed negligible differences in vigilance rates at the site level. Use of the primary winter forage, sagebrush (Artemisia tridentata), increased as production of new annual growth increased and was not related strongly to forage quality, but use of available forage decreased with proximity to disturbance. Consequently, avoidance of human disturbance prompted loss of otherwise available forage, resulting in indirect habitat loss that was 4.6-times greater than direct habitat loss from roads, well pads, and other infrastructure. The multiplicative effects of indirect habitat loss, as mediated by behavior, functionally reduced the amount of available forage for mule deer. Reductions in nutritional carrying capacity are implicit with loss of food and habitat—an outcome that would be expected to have population-level consequences.
4:20PM Modeling the Impact of Age-Dependent Toxicity Defense of Woody Plant to the Demographic Consequences and Population Persistence of Herbivores.
  Rongsong Liu
we study the effects that woody plant chemical defenses may have on interactions between boreal hares that in winter feed almost entirely on twigs. We focus particularly on the fact that toxin concentration often varies with the age of twig segments. The model incorporates the fact that the woody internodes of the youngest segments of the twigs of the deciduous angiosperm species that these hares prefer to eat are more defended by toxins than the woody internodes of the older segments that subtend and support the younger segments. Thus, the per capita daily intake of the biomass of the older segments of twigs by hares is much higher than their intake of the biomass of the younger segments of twigs. This age-dependent toxicity of twig segments is modeled using age-structured model equations which are reduced to a system of delay differential equations involving multiple delays in the woody plant–hare dynamics. A novel aspect of the modeling was that it had to account for mortality of non-consumed younger twig segment biomass when older twig biomass was bitten off and consumed. Basic mathematical properties of the model are established together with upper and lower bounds on the solutions. Necessary and sufficient conditions are found for the linear stability of the equilibrium in which the hare is extinct, and sufficient conditions are found for the global stability of this equilibrium. Numerical simulations confirmed the analytical results and demonstrated the existence of limit cycles over ranges of parameters reasonable for hares browsing on woody vegetation in boreal ecosystems. This showed that age dependence in plant chemical defenses has the capacity to cause hare–plant population cycles, a new result.
4:40PM Manipulating Molecular and Microbial Mechanisms of Toxin Tolerance to Manage Wild Herbivores
  Jennifer Forbey
A great interest in the conservation and management of wild vertebrate herbivores is to predict how various processes influence the health of individuals and stability of populations. In that context, much is known about how predators influence stress physiology, foraging and migratory behavior, and population dynamics of vertebrate herbivores. As a result, management of vertebrate herbivores often includes manipulation of predator risks. There is increasing evidence that diet quality, specifically plant toxins, can influence habitat selection, foraging behavior, nutritional condition, and reproductive success of long-lived avian and mammalian herbivores. We propose that management of vertebrate herbivores, particularly those undergoing translocations and reintroductions, could be enhanced through improved capacity to monitor and mitigate the risks of toxic foodscapes. We demonstrate how manipulating plant toxins across landscapes and the genetic mechanisms of toxin tolerance within herbivores and their gut microbes could contribute to the conservation and management of wild herbivore populations.
5:00PM Panel Discussion

Organizers: Marjorie Matocq

Location: Reno-Sparks CC Date: September 30, 2019 Time: 1:10 pm - 5:00 pm