Using Nuclear Magnetic Resonance Spectroscopy to Develop Physiological Profiles for Bighorn Sheep (Poster)

This study employs new techniques using nuclear magnetic resonance (NMR) to assess the relative health, physiological condition, and reproductive function of wild bighorn sheep (Ovis canadensis) in Montana and Wyoming. Ongoing bighorn studies in Montana and the Greater Yellowstone Ecosystem are focused on herd attributes and the population dynamics which are affected by disease, climate, habitat and physiology. Indices of herd health and physiological status are typically obtained through expensive and time consuming lab assays and field measurements. Recently, NMR spectroscopy has been used to revolutionize the assessment of human metabolic health, and we expect that there is similar potential for studies of wildlife populations. Using NMR spectroscopy to assess metabolites associated with disease, nutrition and stress may eliminate the need for many traditional assays and techniques used today. NMR can be used to evaluate a large suite of metabolites associated with a variety of physiological functions from as little as 500 uL of serum or plasma. Blood samples from 242 sheep from 13 different herds were collected during the winters of 2013-14 and 2014-15 to develop a comprehensive metabolite panel for bighorn sheep. We have used a recently developed statistical program known as MetaboAnalyst™ to begin to analyze and evaluate differences in NMR metabolic profiles among herds and across the fall-winter season when nutritional and physiological stress is expected to be acute. We will be presenting the results of this preliminary study and discussing the potential for application in wildlife management

Nuclear Magnetic Resonance Spectroscopy Metabolic Profiles to Distinguish Geographically Isolated Populations of Mountain Goats

Basic physiological studies on mountain goats (Oreamnos americanus) are conspicuously lacking in the literature, and the physiology of this species is perhaps the least known of the high mountain ungulates. The objective of this study was to evaluate metabolic profiles of female mountain goats from five geographically distinct populations using Nuclear Magnetic Resonance (NMR) spectroscopy. Serum samples were collected from nannies located in Alaska in September (AK) from Glacier in August (GMT), from the Grand Tetons in November-December (GT), from NE Yellowstone in December (NEY) and from Absaroka in March (AB). Serum was extracted with acetone, dried and re-suspended in a standard NMR buffer. NMR spectra were analyzed with Chenomix™ software. Metabolites were identified and concentrations determined using the Chenomix™ database and the Human Metabolome Database. We identified 55 metabolites in the serum of mountain goats using this emerging technology. Of these 42 metabolites differed among the herds (P < 0.05). Of these 42 metabolites; creatinine, lactate and pyruvate distinguished (P < 0.05) each herd from another. Furthermore, using Principal Component Analyses of these metabolites allowed us to clearly differentiate metabolic profiles in carbohydrate, protein and lipid metabolism in nannies from these five populations. This study has the potential to enhance our understanding of how changes in nutrition, reproduction, susceptibility to disease, and survival rates drive population dynamics

Using Nuclear Magnetic Resonance (NMR) Metabolic Profiling to Distinguish Herds of Bighorn Sheep

The objective of this study was to determine if nuclear magnetic resonance (NMR) metabolic profiling has the potential to serve as a management tool for evaluating herds of bighorn (Ovis canadensis) sheep. Two-hundred and forty bighorn sheep serum samples from 13 herds located in Montana and Wyoming were processed for NMR spectra, profiled for small molecule metabolites using Chenomx®, and then analyzed with MetaboAnalyst (v3.0). Fifty-six small molecule metabolites were identified in ungulate serum.  To determine if NMR metabolic profiles can distinguish herds that are geographically distinct with access to different nutritional resources, herds collected in December were compared to herds collected in March. Partial least square discriminant analysis (PLS-DA) indicated a clear, majority separation of metabolic shifts with minor overlaps. Biomarker analysis identified 15 potential biomarkers from the compounds with variables of importance (VIP) scores greater than 1.0. These molecules enabled us to identify ‘significantly’ important metabolic pathways that discriminate herds sampled in December and herds sampled in March. Key biomarkers resulting from the pathway analysis, included: 2-oxoisocaproate, choline, tyrosine, creatinine, and trimethylamine n-oxide. To determine if metabolic profiling can distinguish individual herds within a month, herds in December, January and March were compared to a domestic, Rambouillet ewes (control) sampled during the sample months.  PLS-DA of all herds showed clear metabolic shifts and complete separation between each individual herd and the control ewes for each month. Potential biomarkers for herds within a season that were found to be good discriminants for the December herds included: trimethylamine n-oxide and sarcosine; for January herds included: creatinine and asparagine; and, for March herd included, creatinine. Through identification of small molecule metabolites, it is possible to discriminate herds from each other within and between seasons. These biomarkers represent a potential panel of metabolites that may be used for assessing nutritional status, environmental stress, and herd health through the identification of significantly important metabolic pathways related to energy and protein balance

Characterization of Fatty Acids in Crenarchaeota by GC-MS and NMR

Lipids composed of condensed isoprenyl units connected to glycerol backbones by ether linkages are a distinguishing feature of Archaea. Data suggesting that fatty acids with linear hydrocarbon chains are present in some Archaea have been available for decades. However, lack of genomic and biochemical evidence for the metabolic machinery required to synthesize and degrade fatty acids has left the field unclear on this potentially significant biochemical aspect. Because lipids are energy currency and cell signaling molecules, their presence in Archaea is significant for understanding archaeal biology. A recent large-scale bioinformatics analysis reignited the debate as to the importance of fatty acids in Archaea by presenting genetic evidence for the presence of enzymes required for anabolic and catabolic fatty acid metabolism across the archaeal domain. Here, we present direct biochemical evidence from gas chromatography-mass spectrometry (GC-MS) and nuclear magnetic resonance (NMR) spectroscopy for the presence of fatty acids in two members of the Crenarchaeota, Sulfolobus solfataricus and Ignicoccus hospitalis. This is the first report providing biochemical data for the existence of fatty acids in these Crenarchaeota, opening new discussions on energy balance and the potential for the discovery of new thermostable enzymes for industry

Mapping the Interacting Regions between Troponins T and C. Binding of TnT and TnI peptides to TnC and NMR mapping of the TnT-binding site on TnC

Muscular contraction is triggered by an increase in calcium concentration, which is transmitted to the contractile proteins by the troponin complex. The interactions among the components of the troponin complex (troponins T, C, and I) are essential to understanding the regulation of muscle contraction. While the structure of TnC is well known, and a model for the binary TnC·TnI complex has been recently published (Tung, C.-S., Wall, M. E., Gallagher, S. C., and Trewhella, J. (2000)Protein Sci. 9, 1312–1326), very little is known about TnT. Using non-denaturing gels and NMR spectroscopy, we have analyzed the interactions between TnC and five peptides from TnT as well as how three TnI peptides affect these interactions. Rabbit fast skeletal muscle peptide TnT-(160–193) binds to TnC with a dissociation constant of 30 ± 6 µm. This binding still occurs in the presence of TnI-(1–40) but is prevented by the presence of TnI-(56–115) or TnI-(96–139), both containing the primary inhibitory region of TnI. TnT-(228–260) also binds TnC. The binding site for TnT-(160–193) is located on the C-terminal domain of TnC and was mapped to the surface of TnC using NMR chemical shift mapping techniques. In the context of the model for the TnC·TnI complex, we discuss the interactions between TnT and the other troponin subunits

Developing Physiological Profiles using Nuclear Magnetic Resonance Spectroscopy to Inform Bighorn Sheep Management

This study employs new techniques using nuclear magnetic resonance (NMR) to assess the relative health, physiological condition, and reproductive function of wild bighorn sheep (Ovis canadensis)  in Montana and Wyoming. Ongoing bighorn studies in Montana and the Greater Yellowstone Ecosystem are focused on herd attributes and the population dynamics which are affected by disease, climate, habitat and physiology. Indices of herd health and physiological status are typically obtained through expensive and time consuming lab assays and field measurements. Recently, NMR spectroscopy has been used to revolutionize the assessment of human metabolic health, and we expect that there is similar potential for studies of wildlife populations. Using NMR spectroscopy to assess metabolites associated with disease, nutrition and stress may eliminate the need for many traditional assays and techniques used today. NMR can be used to evaluate a large suite of metabolites associated with a variety of physiological functions from as little as 500 ?L of serum or plasma. Blood samples from 242 sheep from 13 different herds were collected during the winters of 2013-14 and 2014-15 to develop a comprehensive metabolite panel for bighorn sheep. We have used a recently developed statistical program known as MetaboAnalyst™ to begin to analyze and evaluate differences in NMR metabolic profiles among herds and across the fall-winter season when nutritional and physiological stress is expected to be acute. We will be presenting the results of this preliminary study and discussing the potential for application in wildlife management

Group demography affects ant colony performance and individual speed of queen and worker aging

Background: The performance and fitness of social societies mainly depends on the efficiency of interactions between reproductive individuals and helpers. Helpers need to react to the group's requirements and to adjust their tasks accordingly, while the reproductive individual has to adjust its reproductive rate. Social insects provide a good system to study the interrelations between individual and group characteristics. In general, sterile workers focus on brood care and foraging while the queen lays eggs. Reproductive division of labor is determined by caste and not interchangeable as, e.g., in social mammals or birds. Hence, changing social and environmental conditions require a flexible response by each caste. In the ant Cardiocondyla obscurior, worker task allocation is based on age polyethism, with young workers focusing on brood care and old workers on foraging. Here, we examine how group age demography affects colony performance and fitness in colonies consisting of only old or young workers and a single old or young queen. We hypothesized that both groups will be fully functional, but that the forced task shift affects the individuals' performance. Moreover, we expected reduced worker longevity in groups with only young workers due to precocious foraging but no effect on queen longevity depending on group composition. Results: Neither the performance of queens nor that of workers declined strongly with time per se, but offspring number and weight were influenced by queen age and the interaction between queen and worker age. Individual residual life expectancy strongly depended on colony demography instead of physiological age. While worker age affected queen longevity only slightly, exposing old workers to the conditions of colony founding increased their life spans by up to 50% relative to workers that had emerged shortly before colony set-up. Conclusions: The social environment strongly affected the tempo of aging and senescence in C. obscurior, highlighting the plasticity of life expectancy in social insects. Furthermore, colonies obtained the highest reproductive output when consisting of same-aged queens and workers independent of their physiological age. However, workers appeared to be able to adjust their behavior to the colony's needs and not to suffer from age-dependent restrictions

Cocolysis: Stable-Coil Based Coiled Coil Protein Database

Abstract—We discuss the development of a database called Cocolysis that stores all coiled coil domains found in all the proteins stored in the ExPaSy database. This database is hosted on a Linux box, and is automatically updated on a regular basis. It has been already been used to make some interesting observations about coiled coils. Index Terms—protein database, coiled coils, protein structure, protein structure predictio