Characterizing the secret diets of siphonophores (Cnidaria: Hydrozoa) using DNA metabarcoding.

Siphonophores (Cnidaria: Hydrozoa) are abundant and diverse gelatinous predators in open-ocean ecosystems. Due to limited access to the midwater, little is known about the diets of most deep-dwelling gelatinous species, which constrains our understanding of food-web structure and nutrient flow in these vast ecosystems. Visual gut-content methods can rarely identify soft-bodied rapidly-digested prey, while observations from submersibles often overlook small prey items. These methods have been differentially applied to shallow and deep siphonophore taxa, confounding habitat and methodological biases. DNA metabarcoding can be used to assess both shallow and deep species' diets under a common methodological framework, since it can detect both small and gelatinous prey. We (1) further characterized the diets of open-ocean siphonophores using DNA metabarcoding, (2) compared the prey detected by visual and molecular methods to evaluate their technical biases, and (3) evaluated tentacle-based predictions of diet. To do this, we performed DNA metabarcoding analyses on the gut contents of 39 siphonophore species across depths to describe their diets, using six barcode regions along the 18S gene. Taxonomic identifications were assigned using public databases combined with local zooplankton sequences. We identified 55 unique prey items, including crustaceans, gelatinous animals, and fish across 47 siphonophore specimens in 24 species. We reported 29 novel predator-prey interactions, among them the first insights into the diets of nine siphonophore species, many of which were congruent with the dietary predictions based on tentilla morphology. Our analyses detected both small and gelatinous prey taxa underrepresented by visual methods in species from both shallow and deep habitats, indicating that siphonophores play similar trophic roles across depth habitats. We also reveal hidden links between siphonophores and filter-feeders near the base of the food web. This study expands our understanding of the ecological roles of siphonophores in the open ocean, their trophic roles within the 'jelly-web', and the importance of their diversity for nutrient flow and ecosystem functioning. Understanding these inconspicuous yet ubiquitous predator-prey interactions is critical to predict the impacts of climate change, overfishing, and conservation policies on oceanic ecosystems.

Effects of chemical preservation on bulk and amino acid isotope ratios of zooplankton, fish, and squid tissues.

RATIONALE: It is imperative to understand how chemical preservation alters tissue isotopic compositions before using historical samples in ecological studies. Specifically, although compound-specific isotope analysis of amino acids (CSIA-AA) is becoming a widely used tool, there is little information on how preservation techniques affect amino acid δ15 N values. METHODS: We evaluated the effects of chemical preservatives on bulk tissue δ13 C and δ15 N and amino acid δ15 N values, measured by gas chromatography/isotope ratio mass spectrometry (GC/IRMS), of (a) tuna (Thunnus albacares) and squid (Dosidicus gigas) muscle tissues that were fixed in formaldehyde and stored in ethanol for 2 years and (b) two copepod species, Calanus pacificus and Eucalanus californicus, which were preserved in formaldehyde for 24-25 years. RESULTS: Tissues in formaldehyde-ethanol had higher bulk δ15 N values (+1.4, D. gigas; +1.6‰, T. albacares), higher δ13 C values for D. gigas (+0.5‰), and lower δ13 C values for T. albacares (-0.8‰) than frozen samples. The bulk δ15 N values from copepods were not different those from frozen samples, although the δ13 C values from both species were lower (-1.0‰ for E. californicus and -2.2‰ for C. pacificus) than those from frozen samples. The mean amino acid δ15 N values from chemically preserved tissues were largely within 1‰ of those of frozen tissues, but the phenylalanine δ15 N values were altered to a larger extent (range: 0.5-4.5‰). CONCLUSIONS: The effects of preservation on bulk δ13 C values were variable, where the direction and magnitude of change varied among taxa. The changes in bulk δ15 N values associated with chemical preservation were mostly minimal, suggesting that storage in formaldehyde or ethanol will not affect the interpretation of δ15 N values used in ecological studies. The preservation effects on amino acid δ15 N values were also mostly minimal, mirroring bulk δ15 N trends, which is promising for future CSIA-AA studies of archived specimens. However, there were substantial differences in phenylalanine and valine δ15 N values, which we speculate resulted from interference in the chromatographic resolution of unknown compounds rather than alteration of tissue isotopic composition due to chemical preservation.

Long-term trends in the foraging ecology and habitat use of an endangered species: an isotopic perspective.

Evaluating long-term drivers of foraging ecology and population productivity is crucial for providing ecological baselines and forecasting species responses to future environmental conditions. Here, we examine the trophic ecology and habitat use of North Atlantic leatherback turtles (St. Croix nesting population) and investigate the effects of large-scale oceanographic conditions on leatherback foraging dynamics. We used bulk and compound-specific nitrogen isotope analysis of amino acids (CSIA-AA) to estimate leatherback trophic position (TP) over an 18-year period, compare these estimates with TP estimates from a Pacific leatherback population, and elucidate the pre-nesting habitat use patterns of leatherbacks. Our secondary objective was to use oceanographic indices and nesting information from St. Croix leatherbacks to evaluate relationships between trophic ecology, nesting parameters, and regional environmental conditions measured by the North Atlantic Oscillation (NAO) and Atlantic Multidecadal Oscillation. We found no change in leatherback TP over time and no difference in TP between Atlantic and Pacific leatherbacks, indicating that differences in trophic ecology between populations are an unlikely driver of the population dichotomy between Pacific and Atlantic leatherbacks. Isotope data suggested that St. Croix leatherbacks inhabit multiple oceanic regions prior to nesting, although, like their conspecifics in the Pacific, individuals exhibit fidelity to specific foraging regions. Leatherback nesting parameters were weakly related to the NAO, which may suggest that positive NAO phases benefit St. Croix leatherbacks, potentially through increases in resource availability in their foraging areas. Our data contribute to the understanding of leatherback turtle ecology and potential mechanistic drivers of the dichotomy between populations of this protected species.