4.4 CSIA
What is the difference between CSIA & SIA and how does CSIA work?
Compound specific stable isotope analysis (CSIA) elevates traditional bulk stable isotope analysis (SIA) by analyzing isotope ratios of multiple compounds.
Bulk SIA
Entire sample is homogenized & freeze dried
Tin capsules containing the sample are fed into an elemental analyzer
Samples are combusted
Gas Chromatography separates heavier from lighter elements
The flow of gasses is fed into the mass spectrometer where they are pushed around a large magnet, separating
Results show peaks for each element
CSIA
Initial processing (lipid extraction)
Samples are homogenized & freeze dried & fed into gas chromatograph combustion.
Gas Chromatography separates compounds
Combustion breaks down compounds
The flow of gasses is fed into the mass spectrometer where they are pushed around a large magnet, separating
Results show peaks for each compound
Repeat steps 3-6 for each element
Why use CSIA on corals
As mixotrophs, it is difficult to disentangle the energetic inputs
CSIA allows us to ask very specific questions about food source and trophic level that we haven't been able to ask before.
References
Examples of CSIA on corals
Fujii 2020
The authors investigated CSIA (N only) and SIA at multiple sites, multiple species, and multiple seasons. Their conclusions were:
C:N ratios are species specific and not sensitive to environmental change
δ13C driven by isotope fractionation during DIC uptake and are likely related to morphology, season and habitat
δ15N reflect the DIN values and varied based on reef-scale polluion gradients
heterotrophy of corals causes shifts in C and N, but we need more info to make appropriate models.
Treignier 2009
Quantified the δ13C in fatty acids and sterols of both host and symbionts. Found significant effects of both light availability and food availability. Made some conclusions about de novo synthesis of lipids and how lipids are transferred within the holobiont.
McMahon 2018
Proteinaceous deep sea corals deposit metabolically inert layers of skeleton (not CaCO3) that reflect the δ13C and δ15N values of the metabolically active polyps. these can be used as a record of historical climate flux because the corals grow for hundreds to thousands of years.
Martinez 2020
Transplanted corals from mesophotic to shallow (60m - 5m) to quantify CSIA and other photo-physiological metrics. Found that some traits easily acclimated to the shallow environment, but a few remained more similar to mesophotic counterparts. Also made some interesting hypotheses of their CSIA results (no fingerprinting) because the trophic position of both shallow and mesophotic was the same, indicating that light availability does not shift to heterotrophy. Also, the TP of the symbionts was higher than 1, indicating a large amount of sharing/recycling between sym and host.
Martinez 2022
Feeding experiments 1. hetero-, auto- and mixotrophy treatments 2. labeled 15N food with daily collections to see how quickly the host transfers metabolites to symbionts. They found that heterotrophic acquisition of nutrients (N) was quickly transmitted to the symbionts, making nutrient recycling and transfer within the holobiont more complex than traditionally understood. Didn't find huge differences in C or N sources between symbiont and host, indicating that they share a lot.
Fox et al. 2019
Ran CSIA for C only on corals around an island. Patterns of δ13C values of individual AAs provided separation of autotrophic & heterotrophic nutritional inputs. Traditional bulk measurements were a good proxy for heterotrophy vs autotrophy. No spacial patterns found.
Wall et al. 2021
Ran CISA for N and C on corals under light & feeding conditions. Had very small sample sizes so very few results were significant.
Lesser, Slattery & Macartney 2022
Rebuttal to the criticism against using stable isotopes to analyze the transition from autotrophy to heterotrophy. Re-analysis of SIA and CSIA from previous data sets along a depth gradient. AAs valine and isoleucine showed a significant effect of depth, with shallow corals having less negative 13C values than their mesophotic counterparts. CSIA on for trophic position on glu and phe identified differences between host and sym but not along depth gradients.
Ferrier-Pages et al. 2021
CSIA on coral and symbiont for both N and C. They found expected results in Glu and Phe N. Did not use fingerprinting methods, and instead just looked for trends in EAAs and did not find any (duh).
Examples of CSIA on other mixotrophs
Macartney, Slattery & Lesser 2021
CSIA on mixotrophic sponges. Used ΣV value which measures the bacterial resynthesis & translocation of organic matter. More focus on microbes than symbionts. No significant change in C but yes in N based on particulate availability across depth gradients & changes in trophic level.
CSIA on non-mixotrophs
Larsen, Hansen & Dierking 2020
Conducted CSIA on fish to determine feeding differences in marine habitats. Found significant differences between six functional groups of fish and some species exhibited site dependent values.
McMahon et al. 2016
CSIA on 7 reef fishes with four C source end members including coral. Determined that CSIA was effective at predicting the C source of the fishes, which often followed expected sources, but often showed that in oceanic sites, there was a shift to planktonic sources.
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