Intrapopulation variability shaping isotope discrimination and turnover: experimental evidence in arctic foxes

Background

Tissue-specific stable isotope signatures can provide insights into the trophic ecology of consumers and their roles in food webs. Two parameters are central for making valid inferences based on stable isotopes, isotopic discrimination (difference in isotopic ratio between consumer and its diet) and turnover time (renewal process of molecules in a given tissue usually measured when half of the tissue composition has changed). We investigated simultaneously the effects of age, sex, and diet types on the variation of discrimination and half-life in nitrogen and carbon stable isotopes (δ¹⁵N and δ¹³C, respectively) in five tissues (blood cells, plasma, muscle, liver, nail, and hair) of a top predator, the arctic fox Vulpes lagopus.

Methodology/Principal Findings

We fed 40 farmed foxes (equal numbers of adults and yearlings of both sexes) with diet capturing the range of resources used by their wild counterparts. We found that, for a single species, six tissues, and three diet types, the range of discrimination values can be almost as large as what is known at the scale of the whole mammalian or avian class. Discrimination varied depending on sex, age, tissue, and diet types, ranging from 0.3‰ to 5.3‰ (mean  = 2.6‰) for δ¹⁵N and from 0.2‰ to 2.9‰ (mean  = 0.9‰) for δ¹³C. We also found an impact of population structure on δ¹⁵N half-life in blood cells. Varying across individuals, δ¹⁵N half-life in plasma (6 to 10 days) was also shorter than for δ¹³C (14 to 22 days), though δ¹⁵N and δ¹³C half-lives are usually considered as equal.

Conclusion/Significance

Overall, our multi-factorial experiment revealed that at least six levels of isotopic variations could co-occur in the same population. Our experimental analysis provides a framework for quantifying multiple sources of variation in isotopic discrimination and half-life that needs to be taken into account when designing and analysing ecological field studies.     

Effects of trophic level and metamorphosis on discrimination of hydrogen isotopes in a plant-herbivore system

The use of stable isotopes in ecological studies requires that we know the magnitude of discrimination factors between consumer and element sources. The causes of variation in discrimination factors for carbon and nitrogen have been relatively well studied. In contrast, the discrimination factors for hydrogen have rarely been measured. We grew cabbage looper caterpillars (Trichoplusia ni) on cabbage (Brassica oleracea) plants irrigated with four treatments of deuterium-enriched water (δD = −131, −88, −48, and −2‰, respectively), allowing some of them to reach adulthood as moths. Tissue δD values of plants, caterpillars, and moths were linearly correlated with the isotopic composition of irrigation water. However, the slope of these relationships was less than 1, and hence, discrimination factors depended on the δD value of irrigation water. We hypothesize that this dependence is an artifact of growing plants in an environment with a common atmospheric δD value. Both caterpillars and moths were significantly enriched in deuterium relative to plants by ∼45‰ and 23‰ respectively, but the moths had lower tissue to plant discrimination factors than did the caterpillars. If the trophic enrichment documented here is universal, δD values must be accounted for in geographic assignment studies. The isotopic value of carbon was transferred more or less faithfully across trophic levels, but δ¹⁵N values increased from plants to insects and we observed significant non-trophic ¹⁵N enrichment in the metamorphosis from larvae to adult.     

Tissue Turnover Rates and Isotopic Trophic Discrimination Factors in the Endothermic Teleost,Pacific Bluefin Tuna (Thunnus orientalis)

Stable isotope analysis (SIA) of highly migratory marine pelagic animals can improve understanding of their migratory patterns and trophic ecology. However, accurate interpretation of isotopic analyses relies on knowledge of isotope turnover rates and tissue-diet isotope discrimination factors. Laboratory-derived turnover rates and discrimination factors have been difficult to obtain due to the challenges of maintaining these species in captivity. We conducted a study to determine tissue- (white muscle and liver) and isotope- (nitrogen and carbon) specific turnover rates and trophic discrimination factors (TDFs) using archived tissues from captive Pacific bluefin tuna (PBFT), Thunnus orientalis, 1–2914 days after a diet shift in captivity. Half-life values for ¹⁵N turnover in white muscle and liver were 167 and 86 days, and for ¹³C were 255 and 162 days, respectively. TDFs for white muscle and liver were 1.9 and 1.1‰ for δ ¹⁵N and 1.8 and 1.2‰ for δ ¹³C, respectively. Our results demonstrate that turnover of ¹⁵N and ¹³C in bluefin tuna tissues is well described by a single compartment first-order kinetics model. We report variability in turnover rates between tissue types and their isotope dynamics, and hypothesize that metabolic processes play a large role in turnover of nitrogen and carbon in PBFT white muscle and liver tissues. ¹⁵N in white muscle tissue showed the most predictable change with diet over time, suggesting that white muscle δ ¹⁵N data may provide the most reliable inferences for diet and migration studies using stable isotopes in wild fish. These results allow more accurate interpretation of field data and dramatically improve our ability to use stable isotope data from wild tunas to better understand their migration patterns and trophic ecology.     

Two categories of c/c ratios for higher plants

¹³C/¹²C ratios have been determined for plant tissue from 104 species representing 60 families. Higher plants fall into two categories, those with low δ_PDBI¹³C values (—24 to —34‰) and those with high δ ¹³C values (—6 to —19‰). Algae have δ ¹³C values of —12 to —23‰. Photosynthetic fractionation leading to such values is discussed.     

Sequential isotopic signature along gladius highlights contrasted individual foraging strategies of jumbo squid (Dosidicus gigas)

Background

Cephalopods play a major role in marine ecosystems, but knowledge of their feeding ecology is limited. In particular, intra- and inter-individual variations in their use of resources has not been adequatly explored, although there is growing evidence that individual organisms can vary considerably in the way they use their habitats and resources.

Methodology/Principal Findings

Using δ¹³C and δ¹⁵N values of serially sampled gladius (an archival tissue), we examined high resolution variations in the trophic niche of five large (>60 cm mantle length) jumbo squids (Dosidicus gigas) that were collected off the coast of Peru. We report the first evidence of large inter-individual differences in jumbo squid foraging strategies with no systematic increase of trophic level with size. Overall, gladius δ¹³C values indicated one or several migrations through the squid''s lifetime (∼8–9 months), during which δ¹⁵N values also fluctuated (range: 1 to 5‰). One individual showed an unexpected terminal 4.6‰ δ¹⁵N decrease (more than one trophic level), thus indicating a shift from higher- to lower-trophic level prey at that time. The data illustrate the high diversity of prey types and foraging histories of this species at the individual level.

Conclusions/Significance

The isotopic signature of gladii proved to be a powerful tool to depict high resolution and ontogenic variations in individual foraging strategies of squids, thus complementing traditional information offered by stomach content analysis and stable isotopes on metabolically active tissues. The observed differences in life history strategies highlight the high degree of plasticity of the jumbo squid and its high potential to adapt to environmental changes.     

Chaetognatha of the Namibian Upwelling Region: Taxonomy,Distribution and Trophic Position

In October 2010, the vertical distribution, biodiversity and maturity stages of Chaetognatha species were investigated at four stations located off Walvis Bay, Namibia. Seventeen species were detected and classified as pelagic, shallow-mesopelagic, deep-mesopelagic and bathypelagic species based upon the weighted mean depth derived from their average vertical distribution. High abundances of Chaetognatha were found in the upper 100 m at all stations of the Walvis Bay transect with a maximum value of 20837 ind. 1000 m⁻³ at the outer shelf station near the surface. The community was dominated by species of the Serratodentata group. Furthermore, the distribution of Chaetognatha did not seem to be influenced by low oxygen concentrations. Stable isotope ratios of carbon and nitrogen in Chaetognatha were determined for seven different areas located off northern Namibia. The values of δ¹⁵N ranged from 6.05 ‰ to 11.39 ‰, while the δ¹³C values varied between −23.89 ‰ and −17.03 ‰. The highest values for δ¹⁵N were observed at the Walvis Bay shelf break station. The lowest δ¹³C values were found at the Rocky Point offshore station, which was statistically different from all other areas. Stable isotopes of carbon and nitrogen were determined for four taxa (Sagitta minima, Planctonis group, Sagitta enflata, Sagitta decipiens). In this case, the δ¹⁵N values ranged from 6.17 ‰ to 10.38 ‰, whereas the δ¹³C values varied from −22.70 ‰ to −21.56 ‰. The lowest δ¹⁵N values were found for S. minima. The C- and N-content revealed maximum C-values for S. decipiens and maximum N-values for the Planctonis group. The C:N ratio of Chaetognatha ranged between 5.25 and 6.20. Overall, Chaetognatha are a diverse group in the pelagic food web of the Benguela Upwelling System and act as competitors of fish larvae and jelly fish by preying on copepods.     

Preservation Methods Alter Carbon and Nitrogen Stable Isotope Values in Crickets (Orthoptera: Grylloidea)

Stable isotope analysis (SIA) is an important tool for investigation of animal dietary habits for determination of feeding niche. Ideally, fresh samples should be used for isotopic analysis, but logistics frequently demands preservation of organisms for analysis at a later time. The goal of this study was to establish the best methodology for preserving forest litter-dwelling crickets for later SIA analysis without altering results. We collected two cricket species, Phoremia sp. and Mellopsis doucasae, from which we prepared 70 samples per species, divided among seven treatments: (i) freshly processed (control); preserved in fuel ethanol for (ii) 15 and (iii) 60 days; preserved in commercial ethanol for (iv) 15 and (v) 60 days; fresh material frozen for (vi) 15 and (vii) 60 days. After oven drying, samples were analyzed for δ ¹⁵N, δ ¹³C values, N(%), C(%) and C/N atomic values using continuous flow isotope ratio mass spectrometry. All preservation methods tested, significantly impacted δ ¹³C and δ ¹⁵N and C/N atomic values. Chemical preservatives caused δ ¹³C enrichment as great as 1.5‰, and δ ¹⁵N enrichment as great as 0.9‰; the one exception was M. doucasae stored in ethanol for 15 days, which had δ ¹⁵N depletion up to 1.8‰. Freezing depleted δ ¹³C and δ ¹⁵N by up to 0.7 and 2.2‰, respectively. C/N atomic values decreased when stored in ethanol, and increased when frozen for 60 days for both cricket species. Our results indicate that all preservation methods tested in this study altered at least one of the tested isotope values when compared to fresh material (controls). We conclude that only freshly processed material provides adequate SIA results for litter-dwelling crickets.     

Methane-Derived Carbon in the Benthic Food Web in Stream Impoundments

Methane gas (CH₄) has been identified as an important alternative source of carbon and energy in some freshwater food webs. CH₄ is oxidized by methane oxidizing bacteria (MOB), and subsequently utilized by chironomid larvae, which may exhibit low δ¹³C values. This has been shown for chironomid larvae collected from lakes, streams and backwater pools. However, the relationship between CH₄ concentrations and δ¹³C values of chironomid larvae for in-stream impoundments is unknown. CH₄ concentrations were measured in eleven in-stream impoundments located in the Queich River catchment area, South-western Germany. Furthermore, the δ¹³C values of two subfamilies of chironomid larvae (i.e. Chironomini and Tanypodinae) were determined and correlated with CH₄ concentrations. Chironomini larvae had lower mean δ¹³C values (−29.2 to −25.5 ‰), than Tanypodinae larvae (−26.9 to −25.3 ‰). No significant relationships were established between CH₄ concentrations and δ¹³C values of chironomids (p>0.05). Mean δ¹³C values of chironomid larvae (mean: −26.8‰, range: −29.2‰ to −25.3‰) were similar to those of sedimentary organic matter (SOM) (mean: −28.4‰, range: −29.3‰ to −27.1‰) and tree leaf litter (mean: −29.8 ‰, range: −30.5‰ to −29.1‰). We suggest that CH₄ concentration has limited influence on the benthic food web in stream impoundments.     

Spatial and Temporal Variations in Stable Carbon (δ13C) and Nitrogen (δ15N) Isotopic Composition of Symbiotic Scleractinian Corals

Tropical scleractinian corals are considered autotrophic as they rely mainly on photosynthesis-derived nutrients transferred from their photosymbionts. Corals are also able to capture and ingest suspended particulate organic matter, so heterotrophy can be an important supplementary trophic pathway to optimize coral fitness. The aim of this in situ study was to elucidate the trophic status of 10 coral species under contrasted environmental conditions in a French Polynesian lagoon. Carbon (δ¹³C) and nitrogen (δ¹⁵N) isotopic compositions of coral host tissues and photosymbionts were determined at 3 different fringing reefs during wet and dry seasons. Our results highlighted spatial variability in stable isotopic compositions of both coral host tissues and photosymbionts. Samples from the site with higher level of suspended particulate matter were ¹³C-depleted and ¹⁵N-enriched relative to corals and photosymbionts from less turbid sites. However, differences in both δ¹³C and δ¹⁵N between coral host tissues and their photosymbionts (Δ^{host-photosymbionts 13}C and Δ^{host-photosymbionts 15}N) were small (0.27 ± 0.76‰ and 1.40 ± 0.90‰, respectively) and similar at all sites, thus indicating no general increases in the heterotrophic pathway. Depleted δ¹³C and enriched δ¹⁵N values of coral host tissues measured at the most turbid site were explained by changes in isotopic composition of the inorganic nutrients taken up by photosymbionts and also by changes in rate of isotopic fractionation with environmental conditions. Our results also highlighted a lack of significant temporal variations in δ¹³C and δ¹⁵N values of coral host and photosymbiont tissues and in Δ^{host-photosymbionts 13}C and Δ^{host-photosymbionts 15}N values. This temporal stability indicated that corals remained principally autotrophic even during the wet season when photosymbiont densities were lower and the concentrations of phytoplankton were higher. Increased coral heterotrophy with higher food availability thus appears to be species-specific.     

Spatial patterns of leaf δ13C and δ15N of aquatic macrophytes in the arid zone of northwestern China

Analysis of stable isotope composition is an important tool in research on plant physiological ecology. However, large‐scale patterns of leaf‐stable isotopes for aquatic macrophytes have received considerably less attention. In this study, we examined the spatial pattern of stable isotopes of carbon (δ¹³C) and nitrogen (δ¹⁵N) of macrophytes leaves collected across the arid zone of northwestern China (approximately 2.4 × 10⁶ km²) and attempted to illustrate its relationship with environmental factors (i.e., temperature, precipitation, potential evapotranspiration, sediment total carbon and nitrogen). Our results showed that the mean values of the leaf δ¹³C and δ¹⁵N in the macrophytes sampled from the arid zone were −24.49‰ and 6.82‰, respectively, which were far less depleted than those measured of terrestrial plants. The order of averaged leaf δ¹³C from different life forms was as follows: submerged > floating‐leaved > emergent. Additionally, our studies indicated that the values of foliar δ¹³C values of all the aquatic macrophytes were only negatively associated with precipitation, but the foliar δ¹⁵N values were mainly associated with temperature, precipitation, and potential evapotranspiration. Therefore, we speculated that water‐relation factors are the leaf δ¹³C determinant of macrophytes in the arid zone of northwestern China, and the main factors affecting leaf δ¹⁵N values are the complex combination of water and energy factors.     

Diet Reconstruction and Resource Partitioning of a Caribbean Marine Mesopredator Using Stable Isotope Bayesian Modelling

The trophic ecology of epibenthic mesopredators is not well understood in terms of prey partitioning with sympatric elasmobranchs or their effects on prey communities, yet the importance of omnivores in community trophic dynamics is being increasingly realised. This study used stable isotope analysis of ¹⁵N and ¹³C to model diet composition of wild southern stingrays Dasyatis americana and compare trophic niche space to nurse sharks Ginglymostoma cirratum and Caribbean reef sharks Carcharhinus perezi on Glovers Reef Atoll, Belize. Bayesian stable isotope mixing models were used to investigate prey choice as well as viable Diet-Tissue Discrimination Factors for use with stingrays. Stingray δ¹⁵N values showed the greatest variation and a positive relationship with size, with an isotopic niche width approximately twice that of sympatric species. Shark species exhibited comparatively restricted δ¹⁵N values and greater δ¹³C variation, with very little overlap of stingray niche space. Mixing models suggest bivalves and annelids are proportionally more important prey in the stingray diet than crustaceans and teleosts at Glovers Reef, in contrast to all but one published diet study using stomach contents from other locations. Incorporating gut contents information from the literature, we suggest diet-tissue discrimination factors values of Δ¹⁵N ≊ 2.7‰ and Δ¹³C ≊ 0.9‰ for stingrays in the absence of validation experiments. The wide trophic niche and lower trophic level exhibited by stingrays compared to sympatric sharks supports their putative role as important base stabilisers in benthic systems, with the potential to absorb trophic perturbations through numerous opportunistic prey interactions.     

Photosynthetic carbon metabolism of a marine grass

The δ¹³C value of a tropical marine grass Thalassia testudinum is −9.04‰. This value is similar to the δ¹³C value of terrestrial tropical grasses. The δ¹³C values of the organic acid fraction, the amino acid fraction, the sugar fraction, malic acid, and glucose are: −11.2‰, −13.1‰, −10.1‰, −11.1‰, and −11.5‰, respectively. The δ¹³C values of malic acid and glucose of Thalassia are similar to the δ¹³C values of these intermediates in sorghum leaves and attest to the presence of the photosynthetic C₄-dicarboxylic acid pathway in this marine grass. The inorganic HCO₃⁻ for the growth of the grass fluctuates between −6.7 to −2.7‰ during the day. If CO₂ fixation in Thalassia is catalyzed by phosphoenolpyruvate carboxylase (which would result in a −3‰ fractionation between HCO₃⁻ and malic acid), the predicted δ¹³C value for Thalassia would be −9.7 to −5.7‰. This range is close to the observed range of −12.6 to −7.8‰ for Thalassia and agree with the operation of the C₄-dicarboxylic acid pathway in this plant. The early products of the fixation of HCO₃⁻ in the leaf sections are malic acid and aspartic acid which are similar to the early products of CO₂ fixation in C₄ terrestrial plants.     

C/C ratio changes in crassulacean Acid metabolism plants

¹³C/¹²C ratios have been found in totally combusted leaves of Crassulacean acid metabolism plants to range from −14 to −33 δ ¹³C‰ compared with a limestone standard. Crassulacean acid metabolism plants apparently utilize both ribulose-1, 5-diphosphate carboxylase and phosphoenolpyruvate carboxylase to assimilate atmospheric CO₂ and, depending on environmental conditions, have ¹³C/¹²C ratios indicative of either carboxylase or to any intermediate value. The degree of discrimination against ¹³C and the resultant ¹³C/¹²C ratio from the photosynthetically fixed CO₂ is influenced by environmental conditions and is not a specific and fixed characteristic of a Crassulacean acid metabolism plant. Certain Crassulacean acid metabolism plants may shift their ratios as much as 17 δ ¹³C‰ in specific environments.     

Spatial Differences in East Scotia Ridge Hydrothermal Vent Food Webs: Influences of Chemistry,Microbiology and Predation on Trophodynamics

The hydrothermal vents on the East Scotia Ridge are the first to be explored in the Antarctic and are dominated by large peltospiroid gastropods, stalked barnacles (Vulcanolepas sp.) and anomuran crabs (Kiwa sp.) but their food webs are unknown. Vent fluid and macroconsumer samples were collected at three vent sites (E2, E9N and E9S) at distances of tens of metres to hundreds of kilometres apart with contrasting vent fluid chemistries to describe trophic interactions and identify potential carbon fixation pathways using stable isotopes. δ¹³C of dissolved inorganic carbon from vent fluids ranged from −4.6‰ to 0.8‰ at E2 and from −4.4‰ to 1.5‰ at E9. The lowest macroconsumer δ¹³C was observed in peltospiroid gastropods (−30.0‰ to −31.1‰) and indicated carbon fixation via the Calvin-Benson-Bassham (CBB) cycle by endosymbiotic gamma-Proteobacteria. Highest δ¹³C occurred in Kiwa sp. (−19.0‰ to −10.5‰), similar to that of the epibionts sampled from their ventral setae. Kiwa sp. δ¹³C differed among sites, which were attributed to spatial differences in the epibiont community and the relative contribution of carbon fixed via the reductive tricarboxylic acid (rTCA) and CBB cycles assimilated by Kiwa sp. Site differences in carbon fixation pathways were traced into higher trophic levels e.g. a stichasterid asteroid that predates on Kiwa sp. Sponges and anemones at the periphery of E2 assimilated a proportion of epipelagic photosynthetic primary production but this was not observed at E9N. Differences in the δ¹³C and δ³⁴S values of vent macroconsumers between E2 and E9 sites suggest the relative contributions of photosynthetic and chemoautotrophic carbon fixation (rTCA v CBB) entering the hydrothermal vent food webs vary between the sites.     

Stable isotopes document the trophic structure of a deep-sea cephalopod assemblage including giant octopod and giant squid

Although deep-sea cephalopods are key marine organims, their feeding ecology remains essentially unknown. Here, we report for the first time the trophic structure of an assemblage of these animals (19 species) by measuring the isotopic signature of wings of their lower beaks, which accumulated in stomachs of stranded sperm whales. Overall, the species encompassed a narrow range in δ¹³C values (1.7‰), indicating that they lived in closely related and overlapping habitats. δ¹³C values can be interpreted in terms of distribution with the more ¹³C-depleted species (e.g. Stigmatoteuthis arcturi, Vampyroteuthis infernalis) having a more pelagic habitat than the more ¹³C-enriched, bathyal species (e.g. Todarodes sagittatus and the giant squid Architeuthis dux). The cephalopods sampled had δ¹⁵N values ranging 4.6‰, which is consistent with the species spanning approximately 1.5 trophic levels. Neither the giant octopod (Haliphron atlanticus) nor the giant squid reached the highest trophic position. Species δ¹⁵N was independent of body size, with large squids having both the highest (Taningia danae) and lowest (Lepidoteuthis grimaldii) δ¹⁵N values. Their trophic position indicates that some species share the top of the food web, together with other megacarnivores such as the sperm whale.     

Carbon and Nitrogen Isotopes from Top Predator Amino Acids Reveal Rapidly Shifting Ocean Biochemistry in the Outer California Current

Climatic variation alters biochemical and ecological processes, but it is difficult both to quantify the magnitude of such changes, and to differentiate long-term shifts from inter-annual variability. Here, we simultaneously quantify decade-scale isotopic variability at the lowest and highest trophic positions in the offshore California Current System (CCS) by measuring δ¹⁵N and δ¹³C values of amino acids in a top predator, the sperm whale (Physeter macrocephalus). Using a time series of skin tissue samples as a biological archive, isotopic records from individual amino acids (AAs) can reveal the proximate factors driving a temporal decline we observed in bulk isotope values (a decline of ≥1 ‰) by decoupling changes in primary producer isotope values from those linked to the trophic position of this toothed whale. A continuous decline in baseline (i.e., primary producer) δ¹⁵N and δ¹³C values was observed from 1993 to 2005 (a decrease of ∼4‰ for δ¹⁵N source-AAs and 3‰ for δ¹³C essential-AAs), while the trophic position of whales was variable over time and it did not exhibit directional trends. The baseline δ¹⁵N and δ¹³C shifts suggest rapid ongoing changes in the carbon and nitrogen biogeochemical cycling in the offshore CCS, potentially occurring at faster rates than long-term shifts observed elsewhere in the Pacific. While the mechanisms forcing these biogeochemical shifts remain to be determined, our data suggest possible links to natural climate variability, and also corresponding shifts in surface nutrient availability. Our study demonstrates that isotopic analysis of individual amino acids from a top marine mammal predator can be a powerful new approach to reconstructing temporal variation in both biochemical cycling and trophic structure.     

Stable isotope biogeochemistry of seabird guano fertilization: results from growth chamber studies with maize (Zea mays)

Background

Stable isotope analysis is being utilized with increasing regularity to examine a wide range of issues (diet, habitat use, migration) in ecology, geology, archaeology, and related disciplines. A crucial component to these studies is a thorough understanding of the range and causes of baseline isotopic variation, which is relatively poorly understood for nitrogen (δ¹⁵N). Animal excrement is known to impact plant δ¹⁵N values, but the effects of seabird guano have not been systematically studied from an agricultural or horticultural standpoint.

Methodology/Principal Findings

This paper presents isotopic (δ¹³C and δ¹⁵N) and vital data for maize (Zea mays) fertilized with Peruvian seabird guano under controlled conditions. The level of ¹⁵N enrichment in fertilized plants is very large, with δ¹⁵N values ranging between 25.5 and 44.7‰ depending on the tissue and amount of fertilizer applied; comparatively, control plant δ¹⁵N values ranged between −0.3 and 5.7‰. Intraplant and temporal variability in δ¹⁵N values were large, particularly for the guano-fertilized plants, which can be attributed to changes in the availability of guano-derived N over time, and the reliance of stored vs. absorbed N. Plant δ¹³C values were not significantly impacted by guano fertilization. High concentrations of seabird guano inhibited maize germination and maize growth. Moreover, high levels of seabird guano greatly impacted the N metabolism of the plants, resulting in significantly higher tissue N content, particularly in the stalk.

Conclusions/Significance

The results presented in this study demonstrate the very large impact of seabird guano on maize δ¹⁵N values. The use of seabird guano as a fertilizer can thus be traced using stable isotope analysis in food chemistry applications (certification of organic inputs). Furthermore, the fertilization of maize with seabird guano creates an isotopic signature very similar to a high-trophic level marine resource, which must be considered when interpreting isotopic data from archaeological material.     

In Vitro Study of Lipid Biosynthesis in an Anaerobically Methane-Oxidizing Microbial Mat

The anaerobic oxidation of methane (AOM) is a key process in the global methane cycle, and the majority of methane formed in marine sediments is oxidized in this way. Here we present results of an in vitro ¹³CH₄ labeling study (δ¹³CH₄, ~5,400‰) in which microorganisms that perform AOM in a microbial mat from the Black Sea were used. During 316 days of incubation, the ¹³C uptake into the mat biomass increased steadily, and there were remarkable differences for individual bacterial and archaeal lipid compounds. The greatest shifts were observed for bacterial fatty acids (e.g., hexadec-11-enoic acid [16:1Δ11]; difference between the δ¹³C at the start and the end of the experiment [Δδ¹³C_start-end], ~160‰). In contrast, bacterial glycerol diethers exhibited only slight changes in δ¹³C (Δδ¹³C_start-end, ~10‰). Differences were also found for individual archaeal lipids. Relatively high uptake of methane-derived carbon was observed for archaeol (Δδ¹³C_start-end, ~25‰), a monounsaturated archaeol, and biphytanes, whereas for sn-2-hydroxyarchaeol there was considerably less change in the δ¹³C (Δδ¹³C_start-end, ~2‰). Moreover, an increase in the uptake of ¹³C for compounds with a higher number of double bonds within a suite of polyunsaturated 2,6,10,15,19-pentamethyleicosenes indicated that in methanotrophic archaea there is a biosynthetic pathway similar to that proposed for methanogenic archaea. The presence of group-specific biomarkers (for ANME-1 and ANME-2 associations) and the observation that there were differences in ¹³C uptake into specific lipid compounds confirmed that multiple phylogenetically distinct microorganisms participate to various extents in biomass formation linked to AOM. However, the greater ¹³C uptake into the lipids of the sulfate-reducing bacteria (SRB) than into the lipids of archaea supports the hypothesis that there is autotrophic growth of SRB on small methane-derived carbon compounds supplied by the methane oxidizers.     

Experimentally Derived δ13C and δ15N Discrimination Factors for Gray Wolves and the Impact of Prior Information in Bayesian Mixing Models

Stable isotope analysis of diet has become a common tool in conservation research. However, the multiple sources of uncertainty inherent in this analysis framework involve consequences that have not been thoroughly addressed. Uncertainty arises from the choice of trophic discrimination factors, and for Bayesian stable isotope mixing models (SIMMs), the specification of prior information; the combined effect of these aspects has not been explicitly tested. We used a captive feeding study of gray wolves (Canis lupus) to determine the first experimentally-derived trophic discrimination factors of C and N for this large carnivore of broad conservation interest. Using the estimated diet in our controlled system and data from a published study on wild wolves and their prey in Montana, USA, we then investigated the simultaneous effect of discrimination factors and prior information on diet reconstruction with Bayesian SIMMs. Discrimination factors for gray wolves and their prey were 1.97‰ for δ¹³C and 3.04‰ for δ¹⁵N. Specifying wolf discrimination factors, as opposed to the commonly used red fox (Vulpes vulpes) factors, made little practical difference to estimates of wolf diet, but prior information had a strong effect on bias, precision, and accuracy of posterior estimates. Without specifying prior information in our Bayesian SIMM, it was not possible to produce SIMM posteriors statistically similar to the estimated diet in our controlled study or the diet of wild wolves. Our study demonstrates the critical effect of prior information on estimates of animal diets using Bayesian SIMMs, and suggests species-specific trophic discrimination factors are of secondary importance. When using stable isotope analysis to inform conservation decisions researchers should understand the limits of their data. It may be difficult to obtain useful information from SIMMs if informative priors are omitted and species-specific discrimination factors are unavailable.     

Isotope Fractionation in Photosynthetic Bacteria during Carbon Dioxide Assimilation

The δ _PDB¹³C values have been determined for the cellular constituents and metabolic intermediates of autotrophically grown Chromatium vinosum. The isotopic composition of the HCO₃^- in the medium and the carbon isotopic composition of the bacterial cells change with the growth of the culture. The δ _PDB¹³C value of the HCO₃^- in the media changes from an initial value of −6.6‰ to +8.1‰ after 10 days of bacterial growth and the δ _PDB¹³C value of the bacterial cells change from −37.5‰ to −29.2‰ in the same period. The amount of carbon isotope fractionation during the synthesis of hexoses by the photoassimilation of CO₂ has a range of −15.5‰ at time zero to −22.0‰ after 10 days. This range of fractionation compares to the range of carbon isotope fractionation for the synthesis of sugars from CO₂ by ribulose 1,5-diphosphate carboxylase and the Calvin cycle.