New paper on advances in amino acid nitrogen isotopic analysis (Ohkouchi et al., Organic Geochemsitry)

Advances in the application of amino acid nitrogen isotopic analysis in ecological and biogeochemical studies

Naohiko Ohkouchi1,*, Yoshito Chikaraishi1,13, Hilary G. Close2, Brian Fry3, Thomas Larsen4, Daniel J. Madigan5, Matthew D. McCarthy6, Kelton W. McMahon7, Toshi Nagata8, Yuichi I. Naito1,14, Nanako O. Ogawa1, Brian N. Popp9, Shawn Steffan10,11, Yoshinori Takano1, Ichiro Tayasu12, Alex S.J. Wyatt8, Yasuhiko T. Yamaguchi8,15, Yusuke Yokoyama8

1 Department of Biogeochemistry, Japan Agency for Marine-Earth Science and Technology, Japan; 2 Rosenstiel School of Marine and Atmospheric Science, University of Miami, USA3 Australian Rivers Institute, Griffith University, Australia; 4 Leibniz-Laboratory, University of Kiel, Germany; 5 Harvard University Center for the Environment, USA; 6 Department of Ocean Sciences, University of California, Santa Cruz, USA; 7 Graduate School of Oceanography, University of Rhode Island, USA; 8 Atmosphere and Ocean Research Institute, The University of Tokyo, Japan; 9 Department of Geology and Geophysics, University of Hawaii, USA; 10 US Department of Agriculture, Agricultural Research Service, USA; 11 Department of Entomology, University of Wisconsin-Madison, USA; 12 Research Institute of Humanity and Nature, Japan; 13 Present address: Institute of Low Temperature Science, Hokkaido University, Japan; 14 Present address: Nagoya University Museum, Japan; 15 Present address: Lake Biwa Environmental Research Institute, Japan

Compound-specific isotopic analysis of amino acids (CSIA-AA) has emerged in the last decade as a powerful approach for tracing the origins and fate of nitrogen in ecological and biogeochemical studies. This approach is based on the empirical observation that source amino acids (AAs) (i.e., phenylalanine), fractionate 15N very little (< 0.5‰) during trophic transfer, whereas trophic AAs (i.e., glutamic acid), are greatly (∼6–8‰) enriched in 15N during each trophic step. The differential fractionation of these two AA groups can provide a valuable estimate of consumer trophic position that is internally indexed to the baseline δ15N value of the integrated food web. In this paper, we critically review the analytical methods for determining the nitrogen isotopic composition of AAs by gas chromatography–isotope-ratio mass spectrometry. We also discuss methodological considerations for accurate trophic position assessment of organisms using CSIA-AA. We then discuss the advantages and challenges of the CSIA-AA approach using published case studies across a range of topics, including trophic position assessment in various ecosystems, reconstruction of ancient human diets, reconstruction of animal migration and environmental variability, and assessment of marine organic matter dynamics with new classification of microbial fractionation patterns. It is clear that the CSIA-AA approach can provide unique insight into the sources, cycling, and trophic modification of organic nitrogen as it flows through systems. However, this approach will be greatly improved through continued exploration into how biochemical, physiological, and ecological mechanisms affect isotopic fractionation of individual AAs. We end this review with a perspective on future work that will promote the evolution of the rapidly growing field of CSIA-AA.

Bio-physical interactions on a coral reef island (Leichter et al., Oceanography)

Biological and Physical Interactions on a Tropical Island Coral Reef: Transport and Retention Processes on Moorea, French Polynesia

Leichter, J.J., Alldredge, A.L., Bernadi, G., Brooks, A.J., Carlson, C.A., Carpenter, R.C., Edmunds, P.J., Fewings, M.R., Hanson, K.M., Holbrook, S.J., Hench, J.L., Nelson, C.E., Schmitt, R.J., Toonen, R.J., Washburn, L. and Wyatt, A.S.J.

The Moorea Coral Reef Long Term Ecological Research project funded by the US National Science Foundation includes multidisciplinary studies of physical processes driving ecological dynamics across the fringing reef, back reef, and fore reef habitats of Moorea, French Polynesia. A network of oceanographic moorings and a variety of other approaches have been used to investigate the biological and biogeochemical aspects of water transport and retention processes in this system. There is evidence to support the hypothesis that a low-frequency counterclockwise flow around the island is superimposed on the relatively strong alongshore currents on each side of the island. Despite the rapid flow and flushing of the back reef, waters over the reef display chemical and biological characteristics distinct from those offshore. The patterns include higher nutrient and lower dissolved organic carbon concentrations, distinct microbial community compositions among habitats, and reef assemblages of zooplankton that exhibit migration behavior, suggesting multigenerational residence on the reef. Zooplankton consumption by planktivorous fish on the reef reflects both retention of reef-associated taxa and capture by the reef community of resources originating offshore. Coral recruitment and population genetics of reef fishes point to retention of larvae within the system and high recruitment levels from local adult populations. The combined results suggest that a broad suite of physical and biological processes contribute to high retention of externally derived and locally produced organic materials within this island coral reef system.

Formation and maintenance of high-nitrate, low pH layers (Waite et al., Biogeosciences)

Formation and maintenance of high-nitrate, low pH layers in the eastern Indian Ocean and the role of nitrogen fixation

A. M. Waite, V. Rossi, M. Roughan, B. Tilbrook, P. A. Thompson, M. Feng, A. S. J. Wyatt, and E. J. Raes

We investigated the biogeochemistry of low dissolved oxygen high-nitrate (LDOHN) layers forming against the backdrop of several interleaving regional water masses in the eastern Indian Ocean, off northwest Australia adjacent to Ningaloo Reef. These water masses, including the forming Leeuwin Current, have been shown directly to impact the ecological function of Ningaloo Reef and other iconic coastal habitats downstream. Our results indicate that LDOHN layers are formed from multiple subduction events of the Eastern Gyral Current beneath the Leeuwin Current (LC); the LC originates from both the Indonesian Throughflow and tropical Indian Ocean. Density differences of up to 0.025 kg m−3 between the Eastern Gyral Current and the Leeuwin Current produce sharp gradients that can trap high concentrations of particles (measured as low transmission) along the density interfaces. The oxidation of the trapped particulate matter results in local depletion of dissolved oxygen and regeneration of dissolved nitrate (nitrification). We document an associated increase in total dissolved carbon dioxide, which lowers the seawater pH by 0.04 units. Based on isotopic measurements (δ15N and δ18O) of dissolved nitrate, we determine that ~ 40–100% of the nitrate found in LDOHN layers is likely to originate from nitrogen fixation, and that, regionally, the importance of N-fixation in contributing to LDOHN layers is likely to be highest at the surface and offshore.

DON isotopes over a coral reef (Thibodeau et al., Coral Reefs)

Heterogeneous dissolved organic nitrogen supply over a coral reef: first evidence from nitrogen stable isotope ratios

B. Thibodeau, T. Miyajima, I. Tayasu, A. S. J. Wyatt, A. Watanabe, N. Morimoto, C. Yoshimizu, T. Nagata

Dissolved organic nitrogen (DON) potentially plays a major role in sustaining the high productivity and biological diversity of coral reefs. However, data are scarce regarding sources and sinks of DON. This study, for the first time, determined the 15N isotopic composition of total dissolved nitrogen (δ15NTDN), reflecting the isotopic signature of DON, in the water column over a coral reef. The uniformity in δ15NTDN during high tide (3.2 ± 0.3 ‰) indicated that the DON was mainly derived from offshore waters. In contrast, higher spatial heterogeneity of δ15NTDN (3.1 ± 0.9 ‰) and DON concentrations during low tide indicated the existence of local DON sources patchily distributed over the reef. Low δ15NTDN values located mid-reef were indicative of DON release from organisms that obtained their N via N2 fixation, whereas high δ15NTDN appeared to reflect localized release of DON by organisms exposed to dissolved inorganic nitrogen with elevated 15N, such as from terrestrial and offshore inputs. Collectively, the results highlight the importance of spatial patterns in DON release from reef communities in the N cycling of coral reefs.

Coral reef POM dynamics (Wyatt et al., L&O)

Particulate nutrient fluxes over a fringing coral reef: Source-sink dynamics inferred from carbon to nitrogen ratios and stable isotopes

Alex S. J. Wyatt, Ryan J. Lowe, Stuart Humphries and Anya M. Waite

We examined spatial and temporal variations in particulate organic matter (POM) dynamics over a fringing coral reef (Ningaloo Reef) in Western Australia during the austral autumn and spring. Total POM concentrations generally did not differ between seasons or reef zones, but the composition of POM, in terms of carbon isotope ratios (δ13C-POM), carbon to nitrogen ratios (C : N), and fatty acids, changed consistently in water flowing across the reef. Both δ13C-POM and C : N increased from the fore reef to the reef flat and lagoon, −23.0‰ to −20.1‰ and 7.31 to 8.34, respectively. Average rates of net POM uptake by the reef community were highest over the reef crest (4 to 30 mmol N m−2 d−1 and 6 to 130 mmol C m−2 d−1), with a Bayesian isotope model confirming independent measurements of high uptake rates of allochthonous POM (oceanic phyto- and zooplankton). In contrast, over the reef flat, net release of POM was observed (−4 to −5 mmol N m−2 d−1 and −50 mmol C m−2 d−1), with gross release rates (estimated as −6 to −8 mmol N m−2 d−1 and −30 to −90 mmol C m−2 d−1) indicating that the release of autochthonous POM may be of similar magnitude to allochthonous uptake. Examining POM dynamics in terms of gross fluxes reinforces the dependence of coral reef systems on oceanographic processes for allochthonous POM supply, as well as highlighting the potential for autochthonous POM production to supply nutrients to benthic and pelagic communities downstream.

Reef fish trophodynamics (Wyatt et al., Coral Reefs)

Stable isotope analysis reveals community-level variation in fish trophodynamics across a fringing coral reef

A. S. J. Wyatt, A. M. Waite, S. Humphries

In contrast to trophodynamic variations, the marked zonation in physical and biological processes across coral reefs and the concomitant changes in habitat and community structure are well documented. In this study, we demonstrate consistent spatial changes in the community-level trophodynamics of 46 species of fish across the fringing Ningaloo Reef, Western Australia, using tissue stable isotope and fatty acid analyses. Increasing nitrogen (δ15N) and decreasing carbon (δ13C) isotope ratios in the tissues of herbivores, planktivores and carnivores with increasing proximity to the ocean were indicative of increased reliance on oceanic productivity. In contrast, detritivores and corallivores displayed no spatial change in δ15N or δ13C, indicative of the dependence on reef-derived material across the reef. Higher δ13C, as well as increased benthic- and bacterial-specific fatty acids, suggested reliance on reef-derived production increased in back-reef habitats. Genus-level analyses supported community- and trophic group-level trends, with isotope modelling of species from five genera (Abudefduf sexfasciatus, Chromis viridis, Dascyllus spp.,Pomacentrus spp. and Stegastes spp.), demonstrating declining access to oceanic zooplankton and, in the case of Pomacentrus spp. and Stegastes spp., a switch to herbivory in the back-reef. The spatial changes in fish trophodynamics suggest that the relative roles of oceanic and reef-derived nutrients warrant more detailed consideration in reef-level community ecology.

Coral reef nutrient dynamics (Wyatt et al., L&O)

Oceanographic forcing of nutrient uptake and release over a fringing coral reef

Alex S. J. Wyatt, James L. Falter, Ryan J. Lowe, Stuart Humphries and Anya M. Waite

Nitrate and nitrite (NOx) and phosphate (PO4) dynamics over Ningaloo Reef, Western Australia, are shown to depend on oceanographic forcing of coupled mass transfer limited (MTL) gross uptake and gross release from remineralized oceanic particulate organic matter (POM). Estimates of gross release rates increased significantly with increasing POM uptake and were of the same order as gross uptake rates. Gross uptake rates increased significantly with increasing oceanic concentrations and wave energy dissipation, were 35–80% higher over the reef crest (7–9 mmol NOx m−2 d−1 and 4–5 mmol PO4 m−2 d−1), and were significantly correlated with independent estimates of POM-mediated gross NOx uptake, supporting both MTL uptake and the strong role of oceanic POM supply. The relative supply of NOx and POM was linked to the seasonal dynamics of a regional current system. In late spring, upwelling associated with seasonally strong equator-ward winds led to increased NOx concentrations (0.71 ± 0.2 µmol L−1), POM < NOx and the reef was a net nutrient sink (5390 mmol NOxm−1 d−1 and 270 mmol PO4 m−1 d−1). In contrast, during the autumn, NOx was low (0.16 ± 0.06 µmol L−1), but POM > NOx and the reef was a net nutrient source (−7060 mmol NOx m−1 d−1 and −730 mmol PO4 m−1 d−1). The autumn enhancement of oceanic POM supply to the reef can be attributed to a regional phytoplankton bloom associated with acceleration of the oligotrophic Leeuwin Current, which may result in a significant supply of dissolved nutrients to downstream communities.

Organic Matter Release by Benthic Primary Producers (Haas et al., PLoS One)

Effects of Coral Reef Benthic Primary Producers on Dissolved Organic Carbon and Microbial Activity

Andreas F. Haas, Craig E. Nelson, Linda Wegley Kelly, Craig A. Carlson, Forest Rohwer, James J. Leichter, Alex Wyatt, Jennifer E. Smith

Benthic primary producers in marine ecosystems may significantly alter biogeochemical cycling and microbial processes in their surrounding environment. To examine these interactions, we studied dissolved organic matter release by dominant benthic taxa and subsequent microbial remineralization in the lagoonal reefs of Moorea, French Polynesia. Rates of photosynthesis, respiration, and dissolved organic carbon (DOC) release were assessed for several common benthic reef organisms from the backreef habitat. We assessed microbial community response to dissolved exudates of each benthic producer by measuring bacterioplankton growth, respiration, and DOC drawdown in two-day dark dilution culture incubations. Experiments were conducted for six benthic producers: three species of macroalgae (each representing a different algal phylum: Turbinaria ornata – Ochrophyta; Amansia rhodantha – Rhodophyta; Halimeda opuntia – Chlorophyta), a mixed assemblage of turf algae, a species of crustose coralline algae (Hydrolithon reinboldii) and a dominant hermatypic coral (Porites lobata). Our results show that all five types of algae, but not the coral, exuded significant amounts of labile DOC into their surrounding environment. In general, primary producers with the highest rates of photosynthesis released the most DOC and yielded the greatest bacterioplankton growth; turf algae produced nearly twice as much DOC per unit surface area than the other benthic producers (14.0±2.8 µmol h−1 dm−2), stimulating rapid bacterioplankton growth (0.044±0.002 log10 cells h−1) and concomitant oxygen drawdown (0.16±0.05 µmol L−1 h−1 dm−2). Our results demonstrate that benthic reef algae can release a significant fraction of their photosynthetically-fixed carbon as DOC, these release rates vary by species, and this DOC is available to and consumed by reef associated microbes. These data provide compelling evidence that benthic primary producers differentially influence reef microbial dynamics and biogeochemical parameters (i.e., DOC and oxygen availability, bacterial abundance and metabolism) in coral reef communities.


Picoplankton and virus uptake by a coral reef (Patten et al., Coral Reefs)

Uptake of picophytoplankton, bacterioplankton and virioplankton by a fringing coral reef community (Ningaloo Reef, Australia)

Nicole L. Patten, Alex S.J. Wyatt, Ryan J. Lowe, Anya M. Waite

We examined the importance of picoplankton and virioplankton to reef trophodynamics at Ningaloo Reef, (north-western Australia), in May and November 2008. Picophytoplankton (Prochlorococcus,Synechococcus and picoeukaryotes), bacterioplankton (inclusive of bacteria and Archaea), virioplankton and chlorophyll a (Chl a) were measured at five stations following the consistent wave-driven unidirectional mean flow path of seawater across the reef and into the lagoon.Prochlorococcus, Synechococcus, picoeukaryotes and bacterioplankton were depleted to similar levels (~40% on average) over the fore reef, reef crest and reef flat (=‘active reef’), with negligible uptake occurring over the sandy bottom lagoon. Depletion of virioplankton also occurred but to more variable levels. Highest uptake rates, m, of picoplankton occurred over the reef crest, while uptake coefficients, S (independent of cell concentration), were similarly scaled over the reef zones, indicating no preferential uptake of any one group. Collectively, picophytoplankton, bacterioplankton and virioplankton accounted for the uptake of 29 mmol C m−2 day−1, with Synechococcuscontributing the highest proportion of the removed C. Picoplankton and virioplankton accounted for 1–5 mmol N m−2 day−1 of the removed N, with bacterioplankton estimated to be a highly rich source of N. Results indicate the importance of ocean–reef interactions and the dependence of certain reef organisms on picoplanktonic supply for reef-level biogeochemistry processes.

Keywords: Coral reef, Picoplankton, Virus, Uptake, Ningaloo Reef, Indian Ocean

Reef fish discrimination (Wyatt et al., PLoS One)

Variability in Isotope Discrimination Factors in Coral Reef Fishes: Implications for Diet and Food Web Reconstruction

Alex S.J. Wyatt, Stuart Humphries, Anya M. Waite

Interpretation of stable isotope ratios of carbon and nitrogen (δ13C and δ15N) is generally based on the assumption that with each trophic level there is a constant enrichment in the heavier isotope, leading to diet-tissue discrimination factors of 3.4‰ for 15N (ΔN) and ~0.5‰ for13C (ΔC). Diet-tissue discrimination factors determined from paired tissue and gut samples taken from 152 individuals from 26 fish species at Ningaloo Reef, Western Australia demonstrate a large amount of variability around constant values. While caution is necessary in using gut contents to represent diet due to the potential for high temporal variability, there were significant effects of trophic position and season that may also lead to variability in ΔN under natural conditions. Nitrogen enrichment increased significantly at higher trophic levels (higher tissue δ15N), with significantly higher ΔN in carnivorous species. Changes in diet led to significant changes in ΔN, but not tissue δ15N, between seasons for several species:Acanthurus triostegus, Chromis viridis, Parupeneus signatus and Pomacentrus moluccensis. These results confirm that the use of meta-analysis averages for ΔN is likely to be inappropriate for accurately determining diets and trophic relationships using tissue stable isotope ratios. Where feasible, discrimination factors should be directly quantified for each species and trophic link in question, acknowledging the potential for significant variation away from meta-analysis averages and, perhaps, controlled laboratory diets and conditions.

Keywords: diet, food web, gut contents, mixing models, Ningaloo Reef, tissue isotopes