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The chirality of amino acids in extraterrestrial materials may provide an insight into the origin of the essential l-enantiopure amino acids in the terrestrial biosphere. In 2020, the Hayabusa2 mission succeeded in bringing back surface materials from the C-type asteroid (162173) Ryugu to the Earth. Amino acids were one of the targeted organic molecules to be studied in the Ryugu samples. To analyze the various structural isomers of amino acids, which were expected to be present, from the limited amount of the returned samples, the development of a highly-sensitive and selective analytical method was necessary. In the present study, a three-dimensional high-performance liquid chromatography (3D-HPLC) system has been developed for the enantioselective determination of five proteinogenic and three non-proteinogenic amino acids in the Ryugu samples, in which amino acids in the sample were separated by reversed-phase, anion-exchange and enantioselective columns after the fluorescence derivatization with 4-fluoro-7-nitro-2,1,3-benzoxadiozole. The applicability of the analytical system to the extraterrestrial samples was evaluated by analyzing several types of carbonaceous meteorites before applying the system to the Ryugu samples. In the analysis of the Ryugu samples, all of the target amino acids were successfully determined quantitatively. Non-proteinogenic amino acids including 2-amino-n-butyric acid, isovaline and norvaline, rarely present in the terrestrial environment, were found as almost racemic mixtures with 47.1 to 55.2%l.

Abstract Based on laboratory observations, planktonic foraminifers are omnivorous, feeding zooplankton and phytoplankton. Spinose species tend toward greater dependence on zooplankton prey than on phytoplankton prey, while non-spinose species are more adapted to herbivorous diets. However, the trophic activity of planktonic foraminifers in the natural environment and their trophic positions in the marine food web have not yet been fully understood. The trophic position (TP) of two non-spinose species, Neogloboquadrina dutertrei and Pulleniatina obliquiloculata, was determined by differences in the nitrogen isotopic composition between two amino acids (glutamic acid and phenylalanine). Results show that TP values of N. dutertrei were ~ 2.4, indicating dependence on omnivorous (mixed herbivorous and carnivorous) diets, while those of P. obliquiloculata were ~ 2.1, indicating dependence on herbivorous diets. Together with previous laboratory observations, these TP values suggest that N. dutertrei is a detritivore or scavenger, while P. obliquiloculata is generally a herbivore. This trophic niche separation likely allows these two planktonic foraminiferal species to live within a similar depth zone in the open water column and provides a clue for understanding causes of spatial and temporal changes in their relative abundances in living and sediment assemblages.

Tracking migration of highly migratory marine fish using isotope analysis (iso-logging) has become a promising tool in recent years. However, application of this method is often hampered by the lack of essential information such as spatial variations in isotope ratios across habitats (isoscapes) and ontogenetic shifts of isotope ratios of target animals. Here, we test the utility of geostatistical analysis to generate isoscapes of δ13C and δ15N in the western Pacific and estimate the ontogenetic shifts in δ13C and δ15N values of a target species. We first measured δ13C and δ15N in the white muscle of juvenile (n = 210) and adult (n = 884) skipjack tuna Katsuwonus pelamis sampled across the northwest Pacific. Next we fitted a geostatistical model to account for the observed spatial variations in δ13C and δ15N of skipjack by fork length and other environmental variables with spatial random effects. We then used the best-fit models to predict the isoscapes of δ13C and δ15N in 2021. Furthermore, we measured δ15N of amino acids (δ15NAAs) of skipjack (n = 5) to determine whether the observed spatial variation of isotope ratios resulted from baseline shifts or differences in trophic position. The geostatistical model reasonably estimated both isoscapes and ontogenetic shifts from isotope ratios of skipjack, and the isoscapes showed that δ13C and δ15N can clearly distinguish the latitudinal migration of skipjack in the western Pacific. The δ15NAAs supported the results of the geostatistical model, that is, observed variations in skipjack δ15N were largely derived from a baseline shift rather than regional differences in trophic position. Thus, we showed that geostatistical analysis can provide essential basic information required for iso-logging without compound-specific isotope analysis.

The proportion of amino acids (AAs) in sediment can be used as an indicator of microbial degradation, which is primarily the product of benthic prokaryote activity. The microbial activity would be reflected with the stable isotope ratio of nitrogen (δ15N) at the time of mineralization and resynthesis of AAs. In this study, the compound-specific isotope analysis of individual AAs was used to investigate δ15N variation associated with microbial processes in marine sediment samples. Our results showed a decrease in AA concentrations in core-top sediment was accompanied by an increase in δ15N values, suggesting large 15N enrichment in buried AAs. Phenylalanine displayed an increase in δ15N from the surface to depths greater than 2 cm, whereas relatively constant δ15N values at depths below 2 cm, suggesting that microbial utilization of phenylalanine varies with depth. Glycine showed the highest relative molar contribution (from 12.1 to 36.4%), with the largest δ15N increase (from 0.8 to 8.7) in deep sediment, implying that such information can serve as a measure of AA diagenesis in sedimentary environments. Our results also indicated that the δ15N values of individual AAs in sediment reflected the microbial alteration of organic matter at water-sediment interfaces and in sub-surface environments. These findings form an important basis for interpreting the δ15N values of AAs in sediment.

Abstract In the high‐productivity subarctic ocean, phytoplankton growth is assumed to rely mainly on nitrate, based on snapshot observations conducted in spring or summer. However, the year‐round utilization of nitrate and ammonium by phytoplankton remains unexplored. Here, we show the seasonal transition of the dominant nitrogen source for phytoplankton by using the nitrogen isotopic composition (δ15N) of nitrate, ammonium, and sinking particles in the northwestern North Pacific. To decouple the combined effects of nitrogen sources for phytoplankton and subsequent trophic transfers in δ15N of sinking particles, we used compound‐specific δ15N analysis of amino acids (δ15NAA) in sinking particles. The seasonal change in δ15NAA revealed that most of the seasonal variation in δ15N of sinking particles arose from changes in the availability of nitrogenous nutrients in the euphotic layer. The δ15N of sinking particles suggested that the dominant nitrogen source for phytoplankton switches from nitrate to ammonium in autumn.

ABSTRACT Microbial symbiosis drives physiological processes of higher-order systems, including the acquisition and consumption of nutrients that support symbiotic partner reproduction. Metabolic analytics provide new avenues to examine how chemical ecology, or the conversion of existing biomass to new forms, changes over a symbiotic life cycle. We applied these approaches to the nematode Steinernema carpocapsae, its mutualist bacterium, Xenorhabdus nematophila, and the insects they infect. The nematode-bacterium pair infects, kills, and reproduces in an insect until nutrients are depleted. To understand the conversion of insect biomass over time into either nematode or bacterium biomass, we integrated information from trophic, metabolomic, and gene regulation analyses. Trophic analysis established bacteria as meso-predators and primary insect consumers. Nematodes hold a trophic position of 4.6, indicative of an apex predator, consuming bacteria and likely other nematodes. Metabolic changes associated with Galleria mellonella insect bioconversion were assessed using multivariate statistical analyses of metabolomics data sets derived from sampling over an infection time course. Statistically significant, discrete phases were detected, indicating the insect chemical environment changes reproducibly during bioconversion. A novel hierarchical clustering method was designed to probe molecular abundance fluctuation patterns over time, revealing distinct metabolite clusters that exhibit similar abundance shifts across the time course. Composite data suggest bacterial tryptophan and nematode kynurenine pathways are coordinated for reciprocal exchange of tryptophan and NAD+ and for synthesis of intermediates that can have complex effects on bacterial phenotypes and nematode behaviors. Our analysis of pathways and metabolites reveals the chemistry underlying the recycling of organic material during carnivory. IMPORTANCE The processes by which organic life is consumed and reborn in a complex ecosystem were investigated through a multiomics approach applied to the tripartite Xenorhabdus bacterium-Steinernema nematode-Galleria insect symbiosis. Trophic analyses demonstrate the primary consumers of the insect are the bacteria, and the nematode in turn consumes the bacteria. This suggests the Steinernema-Xenorhabdus mutualism is a form of agriculture in which the nematode cultivates the bacterial food sources by inoculating them into insect hosts. Metabolomics analysis revealed a shift in biological material throughout progression of the life cycle: active infection, insect death, and conversion of cadaver tissues into bacterial biomass and nematode tissue. We show that each phase of the life cycle is metabolically distinct, with significant differences including those in the tricarboxylic acid cycle and amino acid pathways. Our findings demonstrate that symbiotic life cycles can be defined by reproducible stage-specific chemical signatures, enhancing our broad understanding of metabolic processes that underpin a three-way symbiosis.

Abstract A novel species of the family Alepocephalidae (slickheads), Narcetes shonanmaruae, is described based on four specimens collected at depths greater than 2171 m in Suruga Bay, Japan. Compared to other alepocephalids, this species is colossal (reaching ca. 140 cm in total length and 25 kg in body weight) and possesses a unique combination of morphological characters comprising anal fin entirely behind the dorsal fin, multiserial teeth on jaws, more scale rows than congeners, precaudal vertebrae less than 30, seven branchiostegal rays, two epurals, and head smaller than those of relatives. Mitogenomic analyses also support the novelty of this large deep-sea slickhead. Although most slickheads are benthopelagic or mesopelagic feeders of gelatinous zooplankton, behavioural observations and dietary analyses indicate that the new species is piscivorous. In addition, a stable nitrogen isotope analysis of specific amino acids showed that N. shonanmaruae occupies one of the highest trophic positions reported from marine environments to date. Video footage recorded using a baited camera deployed at a depth of 2572 m in Suruga Bay revealed the active swimming behaviour of this slickhead. The scavenging ability and broad gape of N. shonanmaruae might be correlated with its colossal body size and relatively high trophic position.

Abstract Stable nitrogen (15N/14N) and carbon (13C/12C) isotopic compositions of amino acids in organisms have widely been employed as a powerful tool to evaluate resource utilization and trophic connection among organisms in diverse ecosystems. However, little is known about the physiological factors or mechanisms responsible for determining the isotopic discrimination (particularly for carbon) within amino acids of organisms. In the present study, we investigated the inter-trophic discrimination of nitrogen and carbon isotopes within amino acids (Δδ 15NAA and Δδ 13CAA, respectively) using four consumer–diet pairs. Each pairing illustrates a metabolic perspective of isotopic fractionation of amino acids. The Δδ 15NAA values in these combinations reveal a trend consistent with those observed in many other combinations in previous studies. This further validates a standard scenario: the deamination preferentially removes 14N amino group from diet-derived amino acids, leaving behind the 15N-enriched amino acids in consumer biomass. The Δδ 15NAA values thus mirror the activity of amino acid deamination in consumers. In contrast, the trends in the Δδ 13CAA value suggest a different metabolic fate for the amino acid carbon isotope. Based on our results, we predict the following scenario: decarboxylation preferentially removes 12C α-carbon (i.e., carbonyl-carbon) from pyruvic acid in glycolysis, and from α-ketoglutaric acid in the tricarboxylic acid cycle, leaving behind the 13C-enriched both pyruvic and α-ketoglutaric acids. The 13C is then transferred to amino acids that are synthesized from the 13C-enriched precursor molecules within consumers. The Δδ 13CAA values therefore mirror the pathways of de novo amino acid synthesis in consumers. The proposed link between nitrogen and carbon isotopes can refine our knowledge of the potential processes affecting the isotopic fractionation within diet and consumer compartments, as well as environmental samples. Graphical abstract

Abstract To adapt to ecological and environmental conditions, species can change their ecological niche (e.g., interactions among species) and function (e.g., prey‐predation, diet competition, and habitat segregation) at the species and guild levels. Stable isotope analysis of bulk carbon and nitrogen of organisms has conventionally been used to evaluate such adaptabilities in the scenopoetic and bionomic views as the isotopic niche width. Compound‐specific stable isotope analysis (CSIA) of nitrogen within amino acids provides trophic information without any disruption of scenopoetic views in the isotope ratios, unlike conventional bulk isotope analysis provides both information and therefore frequently hinders its usefulness for trophic information. We performed CSIA of amino acids to understand the trophic variability of the pike gudgeon Pseudogobio esocinus and largemouth bass Micropterus salmoides as representative specialist and generalist fish species, respectively, from 16 ecologically variable habitats in the four major rivers of Korea. There was little variation (1σ) in the trophic position (TP) among habitats for P. esocinus (± 0.2); however, there was considerably large variation for M. salmoides (± 0.6). The TP of M. salmoides was negatively correlated with the benthic invertebrate indices of the habitats, whereas the TP of P. esocinus showed no significant correlation with any indices. Thus, these two representative fish species have different trophic responses to ecological conditions, which is related to known differences in the trophic niche between specialists (i.e., small niche width) and generalists (i.e., large niche width). Over the past four decades, the conventional bulk isotope analysis has not been capable of deconvoluting “scenopoetic” and “bionomic” information. However, in the present study, we demonstrated that the CSIA of amino acids could isolate trophic niches from the traditional ecological niche composed of trophic and habitat information and evaluated how biological and ecological indices influence the trophic response of specialists and generalists.

As a key nutrient, nitrogen can limit primary productivity and carbon cycle dynamics, but also evolutionary progress. Given strong redox-dependency of its molecular speciation, environmental conditions can control nitrogen localization and bioavailability. This particularly applies to periods in Earth history with strong and frequent redox fluctuations, such as the Neoproterozoic. We here report on chlorophyll-derived porphyrins and maleimides in Ediacaran sediments from Oman. Exceptionally light δ15N values (< –10‰) in maleimides derived from anoxygenic phototrophs point towards ammonium assimilation at the chemocline, whereas the isotopic offset between kerogens and chlorophyll-derivatives indicates a variable regime of cyanobacterial and eukaryotic primary production in surface waters. Biomarker and maleimide mass balance considerations imply shallow euxinia during the terminal Ediacaran and a stronger contribution of anoxygenic phototrophs to primary productivity, possibly as a consequence of nutrient ‘lockup’ in a large anoxic ammonium reservoir. Synchronous δ13C and δ15N anomalies at the Ediacaran–Cambrian boundary may reflect one in a series of overturn events, mixing ammonium and isotopically-light DIC into oxic surface waters. By modulating access to nitrogen, environmental redox conditions may have periodically affected Ediacaran primary productivity, carbon cycle perturbations, and possibly played a role in the timing of the metazoan radiation across the terminal Ediacaran and early Cambrian.

To study the influence of different ocean currents on the trophic spectra found in a sympatric fish community, we analyzed the radiocarbon contents (Δ14C) and stable carbon (δ13C) and nitrogen (δ15N) isotopic compositions in their bulk muscle tissues and the δ15N of individual amino acids in 26 species of demersal fish collected from off Tohoku (Pacific coast), northeastern Japan. The Δ14C values of the fish varied from –42 to +41‰, consistent with the Δ14C of the dissolved inorganic carbon in the cold Oyashio Current (typically about –50‰) and the warm Kuroshio Current (typically about +50‰). The trophic positions (TPs) of the fish estimated from the δ15N values of amino acids increased from 3.1 to 4.5 with increasing snout length. A negative correlation was observed between Δ14C and TP in gadiform fish and flatfish, suggesting that the Oyashio Current delivers a high TP diet to these fish groups. These results suggest that the trophic ecology of marine fish in the coastal western North Pacific is primarily controlled by the two major surface water currents, but is also significantly influenced by a combination of species-specific feeding and migration strategies.

Abstract The abundance and biomass of benthic foraminifera are high in intertidal rocky‐shore habitats. However, the availability of food to support their high biomass has been poorly studied in these habitats compared to those at seafloor covered by sediments. Previous field and laboratory observations have suggested that there is diversity in the food preferences and modes of life among rocky‐shore benthic foraminifera. In this study, we used the stable nitrogen isotopic composition of amino acids to estimate the trophic position, trophic niche, and feeding strategy of individual foraminifera species. We also characterized the configuration and structure of the endobiotic microalgae in foraminifera using transmission electron microscopy, and we identified the origin of endobionts based on nucleotide sequences. Our results demonstrated a large variation in the trophic positions of different foraminifera from the same habitat, a reflection of endobiotic features and the different modes of life and food preferences of the foraminifera. Foraminifera did not rely solely on exogenous food sources. Some species effectively used organic matter derived from endobionts in the cell cytoplasm. The high biomass and species density of benthic foraminifera found in intertidal rocky‐shore habitats are thus probably maintained by the use of multiple nitrogen resources and by microhabitat segregation among species as a consequence.

Abstract Relatively high 15N abundances in bone collagen of early anatomically modern humans in Europe have often been interpreted as a specific consumption of freshwater resources, even if mammoth is an alternative high 15N prey. At Buran-Kaya III, access to associated fauna in a secured archaeological context and application of recently developed isotopic analyses of individuals amino acids offer the opportunity to further examine this hypothesis. The site of Buran-Kaya III is located in south Crimea and has provided a rich archaeological sequence including two Upper Palaeolithic layers, from which human fossils were retrieved and directly dated as from 37.8 to 33.1 ka cal BP. Results from bulk collagen of three human remains suggests the consumption of a high 15N prey besides the contribution of saiga, red deer, horse and hare, whose butchered remains were present at the site. In contrast to bulk collagen, phenylalanine and glutamic acid 15N abundances reflect not only animal but also plant protein contributions to omnivorous diet, and allow disentangling aquatic from terrestrial resource consumption. The inferred human trophic position values point to terrestrial-based diet, meaning a significant contribution of mammoth meat, in addition to a clear intake of plant protein.

Abstract The relationship between biodiversity and ecosystem functioning is an important theme in environmental sciences. We propose a new index for configuration of the biomass pyramid in an ecosystem, named integrated trophic position (iTP). The iTP is defined as a sum of trophic positions (i.e. the average number of steps involved in biomass transfer) of all the animals in a food web integrated by their individual biomass. The observed iTP for stream macroinvertebrates ranged from 2.39 to 2.79 and was negatively correlated with the species density and the Shannon–Wiener diversity index of the local community. The results indicate a lower efficiency of biomass transfer in more diverse communities, which may be explained by the variance in edibility hypothesis and/or the trophic omnivory hypothesis. We found a negative effect of biodiversity on ecosystem functioning.

Abstract Over the last 100 yr, anthropogenic stressors have decimated the assemblage of deepwater coregonines that once underpinned the food webs of the Laurentian Great Lakes. As a part of ongoing restoration efforts, fisheries managers are interested in reintroducing deepwater coregonines from remnant populations to reestablish historical food web connections. However, little is known about historical trophic position and niche partitioning among deepwater coregonines in the Great Lakes. We used nitrogen stable isotope analysis of amino acids to compare trophic position of museum‐preserved (1920s) and present‐day forage fishes in Lakes Michigan and Superior. In the 1920s, deepwater coregonines exhibited clear trophic niche partitioning, with trophic positions spanning a full trophic level. Additionally, species trophic positions were tightly conserved between lakes. In Lake Superior, trophic niche partitioning has been maintained over the last 100 yr, but trophic position has shifted downward by ~0.5 trophic level. The more dramatic species loss in Lake Michigan corresponds with a sharp reduction in trophic niche breadth over time. Our study reveals remarkable trophic niche breadth among deepwater coregonines prior to the major anthropogenic impacts on the Laurentian Great Lakes and provides a food web benchmark for restoring the historical trophic diversity of this iconic species flock.

Shallow-water hydrothermal vent ecosystems are distinct from the deep-sea counterparts, because they are in receipt of sustenance from both chemosynthetic and photosynthetic production and have a lack of symbiosis. The trophic linkage and energy flow in these ecosystems, however remain elusive, which allows us poor understanding of the whole spectrum of biological components distributed across such environmental gradients. In this study, a thorough isotopic survey was conducted on various biological specimens and suspended particulates collected along four transects across the venting features of a shallow-water hydrothermal field off Kueishan Island, Taiwan. The isotope data combined with a Bayesian-based mixing model indicate that the vent-associated particulate organic matter (vent POM), as primary contribution of chemoautotrophic populations, has a high δ13C value (-18.2 ± 1.1‰) and a low δ15N value (-1.7 ± 0.4‰). Zooplankton and epibenthic crustaceans, as the fundamental consumers, exhibit δ13C and δ15N values ranging from -21.3 to -19.8‰ and +5.1 to +7.5‰, respectively, and can utilize the vent POM for 38-53% of their diets. The vent-obligate crab Xenograpsus testudinatus shows a large variation in δ13C (from -18.8 to -13.9‰) and δ15N values (from 1.1 to 9.8‰), although an omnivorous trophic level (2.5) is identified for it using δ15N values of amino acids, and it can utilize the vent POM for 6-87% of its diet. The consistently low (< 10.0‰) and overlapping δ15N values for most of the analyzed macroinvertebrates suggest extensive ingestion of chemosynthetic production complementing the photosynthetic production, a weak prey-predator relationship and low trophic complexity possibly imposed by the extreme environmental contexts of shallow-water hydrothermal ecosystems.

Food web ecologists have long sought to characterize the trophic niches of animals using stable isotopic analysis. However, distilling trophic position from isotopic composition has been difficult, largely because of the variability associated with trophic discrimination factors (inter-trophic isotopic fractionation and routing). We circumvented much of this variability using compound-specific isotopic analysis (CSIA). We examined the (15)N signatures of amino acids extracted from organisms reared in pure culture at four discrete trophic levels, across two model communities. We calculated the degree of enrichment at each trophic level and found there was a consistent trophic discrimination factor (~7.6‰). The constancy of the CSIA-derived discrimination factor permitted unprecedented accuracy in the measurement of animal trophic position. Conversely, trophic position estimates generated via bulk-(15)N analysis significantly underestimated trophic position, particularly among higher-order consumers. We then examined the trophic hierarchy of a free-roaming arthropod community, revealing the highest trophic position (5.07) and longest food chain ever reported using CSIA. High accuracy in trophic position estimation brings trophic function into sharper focus, providing greater resolution to the analysis of food webs.

The sea slug Plakobranchus ocellatus (Sacoglossa, Gastropoda) retains photosynthetically active chloroplasts from ingested algae (functional kleptoplasts) in the epithelial cells of its digestive gland for up to 10 months. While its feeding behavior has not been observed in natural habitats, two hypotheses have been proposed: 1) adult P. ocellatus uses kleptoplasts to obtain photosynthates and nutritionally behaves as a photoautotroph without replenishing the kleptoplasts; or 2) it behaves as a mixotroph (photoautotroph and herbivorous consumer) and replenishes kleptoplasts continually or periodically. To address the question of which hypothesis is more likely, we examined the source algae for kleptoplasts and temporal changes in kleptoplast composition and nutritional contribution. By characterizing the temporal diversity of P. ocellatus kleptoplasts using rbcL sequences, we found that P. ocellatus harvests kleptoplasts from at least 8 different siphonous green algal species, that kleptoplasts from more than one species are present in each individual sea slug, and that the kleptoplast composition differs temporally. These results suggest that wild P. ocellatus often feed on multiple species of siphonous algae from which they continually obtain fresh chloroplasts. By estimating the trophic position of wild and starved P. ocellatus using the stable nitrogen isotopic composition of amino acids, we showed that despite the abundance of kleptoplasts, their photosynthates do not contribute greatly to the nutrition of wild P. ocellatus, but that kleptoplast photosynthates form a significant source of nutrition for starved sea slugs. The herbivorous nature of wild P. ocellatus is consistent with insights from molecular analyses indicating that kleptoplasts are frequently replenished from ingested algae, leading to the conclusion that natural populations of P. ocellatus do not rely on photosynthesis but mainly on the digestion of ingested algae.