Isotope Analysis Of Amino Acids Research Articles

Unveiling the importance of heterotrophy for coral symbiosis under heat stress.

Global warming endangers reef-building corals as they lose their photosynthetic symbionts, which limits their ability to feed autotrophically. Consequently, heterotrophy, the capture of zooplankton, can become crucial for the energy budget of heat-stressed corals. However, it is difficult to assess the extent of the heterotrophic contribution in corals, as well as the dynamics of nutrient exchange between the host and its symbionts. In this pioneering study, we employed a suite of isotopic markers, including 13C- and 15N bulk tissue isotope measurements, compound-specific isotope analysis of amino acids (CSIA-AAs), and 13C- and 15N-labeled food incubations, to investigate nutrient acquisition and allocation in the coral Stylophora pistillata under controlled and heat-induced bleaching conditions. Bulk isotope values and inorganic carbon assimilation remained unchanged in the bleached corals compared to the control corals, overall indicating undisturbed autotrophic activity of the symbionts under heat stress. However, CSIA-AAs showed an increased dependence on heterotrophy for amino acid synthesis in both the host and the symbionts despite reduced assimilation of 15N-labeled food. Overall, these results suggest that although S. pistillata reduces its assimilation of heterotrophic food under heat stress, the acquisition of amino acids by the coral host and symbionts still relies on heterotrophy. This study emphasizes the importance of using multiple indicators to gain a comprehensive understanding of coral nutrition. It shows that coral dependence on heterotrophy is not only associated with a decline in autotrophic availability. Rather, it demonstrates the ability of S. pistillata to adapt its utilization of food sources to the prevailing environmental conditions.IMPORTANCEThis work highlights that every isotopic marker displays a piece of different information concerning the diet of the model coral S. pistillata. By combining all markers, we observed that although S. pistillata exhibited reduced heterotrophic assimilation under heat stress, amino acid acquisition and synthesis remained dependent on heterotrophy. The findings emphasize the adaptability of corals in utilizing different food sources, which is vital for their resilience and recovery in changing environmental conditions. This research underscores the complexity of coral symbiosis and highlights the need for multiple indicators to understand dietary dynamics comprehensively.

Dietary partitioning in sympatric Paradoxurinae civets in Borneo suggested by compound-specific nitrogen isotope analysis of amino acids

We applied stable carbon and nitrogen isotopic analyses to understand the faunivory of the four sympatric wild Paradoxurinae civet species in Borneo, which share similar ecological characteristics. We also employed compound-specific nitrogen isotope analysis of amino acids to estimate these species’ trophic positions (TPs). The bulk stable isotope analysis revealed distinctly lower nitrogen isotope ratios in binturongs than in the other three species. This suggests that binturongs exhibit the lowest degree of faunivory among the four species. Binturongs had the lowest TP estimated from the nitrogen isotope ratios of amino acids (2.0–2.1), followed by small-toothed palm civets (2.4–2.5), masked palm civets (2.7), and common palm civets (2.9). These results suggest that there is little faunivory in binturong and variations in faunivory in the other species. Although the number of samples measured for the nitrogen isotope ratios of amino acids is small (n = 2 for each species), our results suggest that the varying degree of consumption of animal food sources, such as insects, is the key mechanism of niche partitioning in these four Paradoxurinae civet species in Borneo. Such subtle but essential differences in closely related sympatric species would maintain high biodiversity in tropical regions.

A long-term study of stable isotope ratios of fingernail keratin and amino acids in a mother-infant dyad.

To evaluate the potential of compound-specific isotope analysis of amino acids (CSIA-AA) for investigating infant feeding practices, we conducted a long-term study that compared infant and maternal amino acid (AA) nitrogen isotope ratios. Fingernail samples were collected from a single mother-infant dyad over 19 months postpartum. Carbon and nitrogen stable isotope ratios were measured in the bulk keratin of the fingernail samples. Selected samples were then hydrolyzed and derivatized for compound-specific nitrogen isotope analysis of keratin AAs. As in previous studies, infant bulk keratin nitrogen isotope values increased during exclusive breastfeeding and fell with the introduction of complementary foods and eventual cessation of breastfeeding. Infant trophic AAs had elevated nitrogen isotope values relative to the mother, while the source AAs were similar between the mother and infant. Proline and threonine appeared to track the presence of human milk in the infant's diet as the isotopic composition of these AAs remained offset from maternal isotope values until the cessation of breastfeeding. Although CSIA-AA is costly and labor intensive, it appears to hold potential for estimating the duration of breastfeeding, even after the introduction of complementary foods. Through the analysis of a full suite of AAs, it may also yield insights into infant physiology and AA synthesis.

Improving trophic position estimates from amino acid stable isotopes by accounting for physiology and environment

AbstractNitrogen isotope analyses of amino acids (δ15N‐AA) are being increasingly used to decipher trophic dynamics. Interpretation of δ15N‐AA in consumers relies on the assumption that consumer physiological status and nutritional status of prey have negligible influences on the trophic discrimination factor (TDF), hence a constant TDF value is used in trophic position (TP) equations. Recent experiments have shown that this is not always the case and there is also a need to validate derived TP estimates in the field. We take advantage of the uniquely long time series of environmental monitoring data and archived (frozen) samples from the species‐poor Baltic Sea. We analyzed δ15N‐AA in similar sized individuals of cod and in its prey herring from four decades, 1980–2018; including time periods where dramatic reduction in condition status of cod has occurred. We expected that TDF in trophic AAs would increase during periods of poor cod condition, resulting in inflated TP estimates. We found that calculated TP and empirical estimates of TDF (difference in δ15N in trophic AAs between cod and herring) for cod increased in recent decades and that this was linked to condition status, herring (prey) lipid content and the hypoxic state of the ecosystem. Statistically adjusting TP for condition and prey lipid content as well as environmental stress (hypoxia) resulted in lower cod TP which better resembled the observed decrease in herring TP in recent decades. TP calculated from stomach analysis data in cod individuals over the same period showed no trend over time and confirmed that adjusted TP estimates mirror the real dietary TP better than unadjusted. By simultaneously measuring condition/nutritional status in both predator and prey it is possible to adjust for them as confounding variables and decipher actual consumer TP, partly overcoming the issues of unknown and variable TDF‐values. Our study also highlights the importance of including environmental stressors (here hypoxia) when interpreting TP and reconstructing food webs.

Lipid extraction alters amino acid composition and bulk, but not amino acid, carbon and nitrogen isotope values.

Concerns exist over observed shifts in value and variance of nitrogen isotopes following physicochemical extraction of lipids from organic matter. The mechanisms behind these apparent changes in bulk tissue δ15N values are not fully understood yet have major implications for analytical costs and integrity of data interpretations. Changes in proximate analysis, amino acid composition, C:N ratios, bulk tissue and amino acid δ13C and δ15N values, and resulting isotope-based food web metrics were compared between lipid-intact and lipid-extracted muscle tissue of fishes spanning <1% to >20% muscle fat content to identify mechanisms of nitrogen isotope fractionation associated with physicochemical lipid extraction. Bulk δ13C and δ15N values increased and %N, C:N ratios and crude protein content decreased following lipid extraction. Resulting bulk isotope niche spacing and overlap varied significantly between lipid-intact and lipid-extracted tissues. While amino acid composition significantly changed during lipid extraction, particularly for lipid-associated amino acids (e.g., Glu, Lys, Ser), individual amino acid δ13C and δ15N values, and their associated compound-specific isotope analysis of amino acids (CSIA-AA)-based food web metrics, did not. Physicochemical lipid extraction caused significant tissue composition changes (e.g., leaching of amino acids and 15N-deplete nitrogenous waste) that affected δ13C and δ15N values and tissue %C and %N beyond simply removing lipids. However, lipid extraction did not alter individual amino acid δ13C or δ15N values or their associated CSIA-AA-based food web metrics.

Enhanced dietary reconstruction of Korean prehistoric populations by combining δ13C and δ15N amino acids of bone collagen.

Compound specific stable isotope analysis of amino acids (CSIA-AA) is a powerful tool for determining dietary behaviors in complex environments and improving dietary reconstructions. Here, we conducted CSIA-AA on human (n = 32) and animal (n = 13) remains from two prehistoric archaeological sites (Mumun, Imdang) to assess in more detail the dietary sources consumed by prehistoric Korean populations. Results of estimated trophic position (TP) using Δ15NGlx-Phe show that the Imdang individuals consumed aquatic resources, as well as terrestrial resources. Principal component analysis (PCA) using δ13C and δ15N essential amino acid (EAA) values show that the Imdang humans closely cluster with game birds and terrestrial herbivores, whilst the Mumun humans closely cluster with C4 plants. Quantitative estimation by a Bayesian mixing model (MixSIAR) indicates that the Imdang humans derived a large proportion of their proteins from terrestrial animals and marine fish, whereas the main protein sources for the Mumun humans were C4 plants and terrestrial animals. Additionally, the comparison between the EAA and bulk isotope models shows that there is a tendency to overestimate the consumption of plant proteins when using bulk isotopic data. Our CSIA-AA approach reveals that in prehistoric Korea there were clear differences in human diets through time. This study adds to a growing body of literature that demonstrates the potential of CSIA-AA to provide more accurate estimations of protein consumption in mixed diets than previous bulk isotopic studies.

Omnivorous summer feeding by juvenile Antarctic krill in coastal waters

AbstractThe Antarctic krill Euphausia superba is often considered an herbivore but is notable for its trophic flexibility, which includes feeding on protistan and metazoan zooplankton. Characterizing krill trophic position (TP) is important for understanding carbon and energy flow from phytoplankton to vertebrate predators and to the deep ocean, especially as plankton composition is sensitive to changing climate. We used repeated field sampling and experiments to study feeding by juvenile krill during three austral summers in waters near Palmer Station, Antarctica. Our approach was to combine seasonal carbon budgets, gut fluorescence measurements, imaging flow cytometry, and compound‐specific isotope analysis of amino acids. Field measurements coupled to experimentally derived grazing functional response curves suggest that phytoplankton grazing alone was insufficient to support the growth and basal metabolism of juvenile krill. Phytoplankton consumption by juvenile krill was limited due to inefficient feeding on nanoplankton (2–20 μm), which constituted the majority of autotrophic prey. Mean krill TP and the metazoan dietary fraction increased in years with higher mesozooplankton biomass, which was not coupled to phytoplankton biomass. Comparing TP estimates using δ15N of different amino acids indicated a substantial and consistent food‐web contribution from heterotrophic protists. Phytoplankton, metazoans, and heterotrophic protists all were important contributors to a diverse krill diet that changed substantially among years. Juvenile krill fed mostly on heterotrophic prey during summer near Palmer Station, and this food web complexity should be considered more broadly throughout the changing Southern Ocean.

Trophic positions of soil microarthropods in forests increase with elevation, but energy channels remain unchanged

AbstractMountain forests are at risk as the consequences of climate change will likely lead to altered tree species boundaries. Characterizing food webs along elevation gradients in primary forests may help to predict the potential consequences of such changes, for example with regard to the decomposition of dead organic matter. Here, for the first time, we studied trophic variations in two species‐rich microarthropod taxa, Collembola and Oribatida, along an elevation gradient of primary forest at Changbai Mountain, China. Samples were taken at seven elevations of 150‐m elevational difference between 800 and 1700 m. At each elevation, Collembola and Oribatida were extracted from litter samples of eight subplots. We applied three state‐of‐the‐art methods to elucidate trophic positions and basal resource use at community level: Bulk stable isotope analysis of nitrogen (Δ15Nbulk) and carbon (Δ13Cbulk), compound‐specific stable isotope analysis of amino acids (CSIA‐AA), and dietary routing of neutral lipid fatty acids (NLFAs). Trophic positions calculated using Δ15Nbulk and CSIA‐AA (TPCSIA) in both taxa increased similarly with elevation by about half and one third of a trophic position, respectively. Stable isotope mixing models and linear discriminant analysis bootstrapping using δ13C of essential amino acids indicated fungi as the most important resource at all elevations for both taxa. Also, proportions of marker NLFAs changed little across elevations in both taxa; overall high proportions of linoleic acid indicated high fungal contributions, but in Collembola the contribution of bacterial markers was generally higher than in Oribatida. Δ13Cbulk did not respond linearly to the elevation gradient; however, changes in elevation differed between Collembola and Oribatida. A strong linear relationship between δ15N of phenylalanine and δ15N of litter indicated litter as the basis of energy channels in both taxa. Overall, food web functioning likely changes with changing forest types along elevation gradients, with microarthropods switching from feeding closer to the base of the food web at lower elevations to feeding at higher trophic levels at higher elevations, potentially compromising their role in litter decomposition and nutrient cycling.

Compound-Specific Isotope Analysis of Amino Acids for Aquatic Systems -Problems, Challenges, Solutions: A Review

Compound-Specific Isotope Analysis of Amino Acids for Aquatic Systems -Problems, Challenges, Solutions: A Review

Origin and alteration of sinking and resuspended organic matter on a benthic nepheloid layer influenced continental shelf

Understanding the biogeochemical transformations that particulate organic matter (POM) undergoes from production in surface waters to deposition on the seafloor is fundamental for interpreting the paleoceanographic record preserved in sediments. Here we use compound-specific N isotope analysis of amino acids (δ15NAA) from two sediment traps (deployed at 150 m and 250 m water depth) and underlying push-core sediments to investigate temporal and depth related changes in the export flux on a continental shelf basin influenced by a prominent benthic nepheloid layer (BNL). Despite apparent isotopic similarities between traps, molecular data show distinct patterns between the shallower (150 m) and deeper (250 m) sediment traps – specifically, depth-related differences in the relative contributions of particles with different processing histories. The export flux in the shallower trap is dominated by animal sources rather than phytodetritus. Significant microbial degradation in the deeper trap and sediments emphasizes the key roles of resuspension-deposition cycles that are mainly (but not exclusively) facilitated by a thick year-around BNL in the region. The BNL acts as a ‘microbial reactor’ reworking sinking POM before its ultimate deposition, and also diluting exported material with previously deposited surface sediments. This heavy modification of POM during its passage through and residence in the BNL opens potentially important questions for our current understanding of OC preservation and interpretations of proxy δ15N records in BNL-influenced areas.

Integrated trophic position as a proxy for food‐web complexity

Abstract There are two distinct approaches to describing the distributions of biomass and species in food webs: one to consider them as discrete trophic levels (TLs); and the other to consider them as continuous trophic positions (TPs). Bridging the gap between these two perspectives presents a nontrivial challenge in integrating biodiversity and food‐web structure. Food network unfolding (FNU) is a technique used to bridge this gap by partitioning the biomass of species into integer TLs to compute three complexity indices, namely vertical (DV), horizontal (DH) and range (DR) diversity (D indices), through decomposition of Shannon's index H′. Using FNU, the food web (a network of species with unique TPs) is converted to a linear food chain (a biomass distribution at discrete TLs). This enables us to expect that the unfolded biomass within species decreases exponentially as the TL increases. Under this condition, the mean TL value in unfolded food chains is hypothesized to have an exponential relationship with the vertical diversity, DV. To explore this, we implemented FNU and calculated D indices for food webs publicly available at EcoBase (n = 158) and calculated the integrated TP (iTP), defined as the biomass‐weighted average TP of a given food web. The iTP corresponds to the mean TL in unfolded food chains and can be empirically measured through compound‐specific isotope analysis of amino acids (CSIA‐AA). Although our analysis is biased towards marine ecosystems, we revealed an exponential relationship between iTP and DV, suggesting that iTP can serve as a measurable proxy for DV. Furthermore, we found a positive correlation between the iTP observed in the total communities (total iTP) and the iTPs of partial communities consisting only of species with 2.0 ≤ TP &lt; 3.0 (partial iTP; r2 = 0.48), suggesting that DV can be predicted using partial iTP. Our findings suggest that the net effect of species diversity, excluding the effect of biomass (corresponding to H′ − DV), on food‐web complexity can be revealed by combining CSIA‐AA with biodiversity analysis (e.g. environmental DNA).

Compound-specific stable nitrogen isotope analysis of amino acids shows that bulk methods provide higher estimates of mercury biomagnification in the Gulf of St. Lawrence

Compound-specific stable isotope analysis of amino acids (CSIA-AA) provides a method to estimate baseline δ15N values of food chains, allowing less biased estimates of trophic positions for organisms. Greater accuracy in trophic positions can improve estimates of contaminant biomagnification. We calculated trophic positions with various CSIA-AA equations for four species of fish and northern gannets (Morus bassanus) from the Gulf of St. Lawrence. We examined the effect of CSIA-AA-derived trophic positions on mercury biomagnification metrics (trophic magnification factors (TMF) and biomagnification factors) and compared these with trophic position estimates and metrics obtained from traditional bulk stable isotope analysis. The TMFs for the CSIA-AA equations ranged from 10 to 19, and bulk stable isotope analysis produced TMFs of 43, one of the highest TMFs recorded yet in the literature. Biomagnification factors between prey and northern gannets ranged from 20 to 42 using dietary observations and stable isotope mixing models. Our study demonstrates that discrepancies in biomagnification assessed using different approaches may go undetected when using a single approach.

Loss of sea ice and intermittent winds alter distributions and diet resources of young forage fish in the Chukchi sea

Loss of sea ice alters habitat for organisms that function as energetic links in Arctic food webs. To address the impacts of such changes on lower trophic levels we focus on forage fish during the early life stages (polar cod: Boreogadus saida and saffron cod: Eleginus gracilis) and copepods (Calanus spp. and Pseudocalanus spp.) in the Chukchi Sea with differing reliance on sea ice environments. We assess distributions throughout years of varying ice extent (2010–2013, 2015, 2017–2018) and diet resources during unprecedented warm years (2017, 2018) using stable isotope analyses (bulk δ13C and δ15N and δ13C compound-specific isotope analysis of amino acids). Calanus spp. and polar cod were found at relatively higher latitudes in closer proximity to recent sea ice, whereas saffron cod were rare and Pseudocalanus spp. were ubiquitous. Polar cod juveniles, but not larvae, were more common in open water, suggesting that low ice and northward ocean currents interact to influence juvenile distributions. Low summer sea ice also coincided with northward expansion of warm Pacific-origin water across the shelf. Stable isotope δ13C values reflected this latitudinal variation in water masses as well as inshore-offshore gradients due to convergence of ocean currents. When summer sea ice was absent during 2017, juvenile polar cod were dispersed across the shelf, abundances were low for both copepod taxa, isotopic niche was reduced for polar cod, and carbon sources for some larval polar cod reflected boreal-associated phytoplankton. Distributions and isotopic patterns suggest that under low ice conditions, along-shelf wind reversals and weak currents can lead to dominance of Pacific-origin water, mixing of organisms from various source locations, and reductions in sea ice-influenced environments. Northward expansion of subarctic water masses and organisms across the Chukchi shelf alters basal resources for lower trophic levels and modifies pelagic habitats to more uniformly resemble lower latitude marine ecosystems.

Small particles and heterotrophic protists support the mesopelagic zooplankton food web in the subarctic northeast Pacific Ocean

AbstractA quantitative understanding of the mesopelagic zooplankton food web is key to development of accurate carbon budgets and geochemical models in marine systems. Here we use compound specific nitrogen stable isotope analysis of amino acids to quantify the trophic structure of the microzooplankton and mesozooplankton community during summer in the subarctic northeast Pacific Ocean during the EXport Processes in the Ocean from Remote Sensing (EXPORTS) field campaign. Source amino acid values in particles and zooplankton provide strong evidence that basal resources for the mesopelagic zooplankton food web were primarily small (), suspended or slow‐sinking particles, but that surface organic matter delivered by vertically migrating zooplankton may have also been important. Comparisons of values of source and trophic amino acids provide estimates of food web length, which decrease significantly with depth and suggest that protistan microzooplankton are key components of the food web from the surface to at least 500. These results emphasize the importance of small particles as a source of carbon and nitrogen to mesopelagic communities in this region, support observations of an inverse relationship between zooplankton vertical migration and small particles as sources of carbon to deep‐sea food webs in low productivity environments, and document the role of heterotrophic protists as key trophic intermediaries in the mesopelagic zone at this location.

Deconvolving mechanisms of particle flux attenuation using nitrogen isotope analyses of amino acids

AbstractParticulate organic matter settling out of the euphotic zone is a major sink for atmospheric carbon dioxide and serves as a primary food source to mesopelagic food webs. Degradation of this organic matter encompasses a suite of mechanisms that attenuate flux, including heterotrophic metabolic processes of microbes and metazoans. The relative contributions of microbial and metazoan heterotrophy to flux attenuation, however, have been difficult to determine. We present results of compound specific nitrogen isotope analysis of amino acids of sinking particles from sediment traps and size‐fractionated particles from in situ filtration between the surface and 500 m at Ocean Station Papa, collected during NASA EXPORTS (EXport Processes in the Ocean from RemoTe Sensing). With increasing depth, we observe: (1) that, based on the δ15N values of threonine, fecal pellets dominate the sinking particle flux and that attenuation of downward particle flux occurs largely via disaggregation in the upper mesopelagic; (2) an increasing trophic position of particles in the upper water column, reflecting increasing heterotrophic contributions to the nitrogen pool and the loss of particles via remineralization; and (3) increasing δ15N values of source amino acids in submicron and small (1–6 μm) particles, reflecting microbial particle solubilization. We further employ a Bayesian mixing model to estimate the relative proportions of fecal pellets, phytodetritus, and microbially degraded material in particles and compare these results and our interpretations of flux attenuation mechanisms to other, independent methods used during EXPORTS.

Use of amino acid isotope analysis to investigate capital versus income breeding strategies in migratory avian species

Abstract Income and capital breeding represent opposing ends of a continuum of reproductive strategies. Quantifying nutrient allocation to reproduction is challenging, but recent advances in compound‐specific stable isotope analysis hold promise for tracing the source of individual compounds allocated to reproduction. Here, we describe a novel approach of using measured carbon (δ13C) and nitrogen (δ15N) isotope values of individual amino acids (AAs) in pectoral muscle of egg‐laying females and egg yolk as a useful tool to quantify the reliance on income versus capital breeding in migrating species. We used white‐fronted (Anser albifrons frontalis), lesser snow (A. caerulescens caerulescens) and black brant (Branta bernicla nigricans) geese breeding in tundra ecosystems of northern Alaska as model organisms. All three species relied on mixed capital–income breeding strategies, but models based on AA isotope data estimated higher proportional contributions of endogenous resources to yolk synthesis compared to results based on bulk tissue isotope analyses. Tracing income versus capital nutrient allocation in migratory species at the compound level is a major advance from the current ‘elemental’ perspective obtained from bulk tissue stable isotope analyses. Our framework is applicable to all taxonomic groups, as long as there is a sufficient spatial or temporal isotopic gradient between resources obtained during the breeding and non‐breeding periods.

Amino acid carbon isotope fingerprints are unique among eukaryotic microalgal taxonomic groups

AbstractEukaryotic microalgae play critical roles in the structure and function of marine food webs. The contribution of microalgae to food webs can be tracked using compound‐specific isotope analysis of amino acids (CSIA‐AA). Previous CSIA‐AA studies have defined eukaryotic microalgae as a single functional group in food web mixing models, despite their vast taxonomic and ecological diversity. Using controlled cultures, this work characterizes the amino acid δ13C (δ13CAA) fingerprints—a multivariate metric of amino acid carbon isotope values—of four major groups of eukaryotic microalgae: diatoms, dinoflagellates, raphidophytes, and prasinophytes. We found excellent separation of essential amino acid δ13C (δ13CEAA) fingerprints among four microalgal groups (mean posterior probability reclassification of 99.2 ± 2.9%). We also quantified temperature effects, a primary driver of microalgal bulk carbon isotope variability, on the fidelity of δ13CAA fingerprints. A 10°C range in temperature conditions did not have significant impacts on variance in δ13CAA values or the diagnostic microalgal δ13CEAA fingerprints. These δ13CEAA fingerprints were used to identify primary producers at the base of food webs supporting consumers in two contrasting systems: (1) penguins feeding in a diatom‐based food web and (2) mixotrophic corals receiving amino acids directly from autotrophic endosymbiotic dinoflagellates and indirectly from water column diatoms, prasinophytes, and cyanobacteria, likely via heterotrophic feeding on zooplankton. The increased taxonomic specificity of CSIA‐AA fingerprints developed here will greatly improve future efforts to reconstruct the contribution of diverse eukaryotic microalgae to the sources and cycling of organic matter in food web dynamics and biogeochemical cycling studies.

Reconstructions of individual fish trophic geographies using isotopic analysis of eye-lens amino acids.

Fish eye lenses are a proteinaceous structure that grows by accumulating layers in a chronological manner. Each layer becomes metabolically inert, capturing the ratio of heavy/light carbon and nitrogen isotopes at time of formation. Therefore, eye lenses contain chronological isotopic records and can be used to create a temporal isotopic history throughout an individual's lifetime. We analyzed eye lens amino-acid δ15N to address spatio-temporal baseline variability and to reconstruct trophic histories of 10 individual Red Snapper. Proteins from sequential eye lens laminae were derivatized to measure 10 amino acids, from which glutamic acid (trophic) and phenylalanine (source) were used to estimate trophic positions at different points in life. Best-fitting regressions were generated to represent individual (R2 ≥ 0.89) and generalized (R2 = 0.77) trophic trajectory for Red Snapper. The resulting trophic trajectories indicated an increase in trophic position with increasing length. Until recently, there has not been a lifetime isotopic structure with enough organic nitrogen to recreate geographic histories using compound-specific stable isotope analysis of amino acids (CSIA-AA). This study confirms that eye-lens laminae can be used to reconstruct trophogeographic histories via CSIA-AA.

Trophic ontogeny of a generalist predator is conserved across space.

Consumers can influence ecological patterns and processes through their trophic roles and contributions to the flow of energy through ecosystems. However, the diet and associated trophic roles of consumers commonly change during ontogeny. Despite the prevalence of ontogenetic variation in trophic roles of most animals, we lack an understanding of whether they change consistently across local populations and broad geographic gradients. We examined how the diet and trophic position of a generalist marine predator varied with ontogeny across seven broadly separated locations (~ 750km). We observed a high degree of heterogeneity in prey consumed without evidence of spatial structuring in this variability. However, compound-specific isotope analysis of amino acids revealed remarkably consistent patterns of increasing trophic position through ontogeny across local populations, suggesting that the roles of this generalist predator scaled with its body size across space. Given the high degree of diet heterogeneity we observed, this finding suggests that even though the dietary patterns differed, the underlying food web architecture transcended variation in prey species across locations for this generalist consumer. Our research addresses a gap in empirical field work regarding the interplay between stage-structured populations and food webs, and suggests ontogenetic changes in trophic position can be consistent in generalist consumers.

Stable isotope analyses of amino acids reveal the importance of aquatic resources to Mediterranean coastal hunter-gatherers.

Determining the degree to which humans relied on coastal resources in the past is key for understanding long-term social and economic development, as well as for assessing human health and anthropogenic impacts on the environment. Prehistoric hunter-gatherers are often assumed to have heavily exploited aquatic resources, especially those living in regions of high marine productivity. For the Mediterranean, this view has been challenged, partly by the application of stable isotope analysis of skeletal remains which has shown more varied coastal hunter-gatherer diets than in other regions, perhaps due to its lower productivity. By undertaking a more specific analysis of amino acids from bone collagen of 11 individuals from one of the oldest and best-known Mesolithic cemeteries in the Mediterranean, at El Collado, Valencia, we show that high levels of aquatic protein consumption were achieved. By measuring both carbon and nitrogen in amino acids, we conclude that some of the El Collado humans relied heavily on local lagoonal fish and possibly shellfish, rather than open marine species. By contrast to previous suggestions, this study demonstrates that the north-western coast of the Mediterranean basin could support maritime-oriented economies during the Early Holocene.