DINOTROPHY: Deuterium in Organic Biomarkers: A new tool to investigate the role of Marine Mixotrophy in the Global Carbon Cycle

Marine microscopic organisms drifting in the oceans (plankton) produce 50% of the oxygen we breath. Their activity helps maintain the balance of oxygen and carbon dioxide in the atmosphere. Much of this production is undertaken by eukaryote (non-bacterial) microbes called protists. Science has traditionally separated these marine protists between those akin to microscopic plants (autotrophs), getting their energy directly from the sunlight, or akin to microscopic animals (heterotrophs), getting their energy by eating other microbes. Recently, however, scientists have come to appreciate that many marine protists combine both autotrophic and heterotrophic (feeding) activities in the same single cell (mixotrophy). Mixotrophy alters marine food web functioning and affects the capacity of the oceans to remove carbon dioxide (counter climate change) from the atmosphere by increasing marine photosynthesis.

This revelation overturns a century of understand in marine science, but also presents a severe challenge; traditional methods are targeted at autotrophy or heterotrophy, in separate organisms and we lack methods to determine mixotrophic activity. This project will develop a new tool to tackle this critical challenge. The high interest met in developing this project has already created a wide interdisciplinary network across the United Kingdom, France and Switzerland.

Our team have previously shown that hydrogen isotopic signature of compounds produced in plants and bacteria (for instance into lipids) is uniquely sensitive to the balance of heterotrophy and autotrophy. This project will extend this investigation for compounds produced by marine microorganisms using similar approaches. We will target a particularly important group of mixotrophic protist plankton, the dinoflagellates, by measuring the hydrogen isotopic composition of specific compounds produced by dinoflagellates. Specifically, we will investigate the hydrogen isotope fractionation mechanisms (at molecular level, i.e. "site-specific") during biosynthesis of organic compounds and seek to establish the hydrogen isotope signature of marine lipids biomarkers (also used as molecular fossils) as a novel tool to investigate the behaviours of dinoflagellates in modern and past oceans and the impact of mixotrophy on the global carbon cycle.

The core of the project will also produce of a numerical biochemical model uniquely possible by combining microorganism cultures and cutting-edge chemical analyses (including Mass Spectrometry and Nuclear Magnetic Resonance Spectroscopy) that will identify in detail the chemical steps responsible for the isotopic signature. By doing so, the project is also fostering a new cross-disciplinary group uniquely skilled in chemistry, isotope biogeochemistry and marine ecology dedicated to extend our knowledge of the marine microorganisms and their role on the Earth System.

New conceptual understanding sees the traditional dichotomy between producers and consumers in the marine food web replaced by one that recognises that mixotrophy is widespread. Many "phytoplankton" eat, while 50% of "microzooplankton" perform photosynthesis. This mixotrophic behaviour enhances primary production, biomass transfer to higher trophic levels, and affects the sequestration of atmospheric CO2. Science requires a tool to measure the contributions of phototrophy & heterotrophy in plankton to aid in climate change modelling. We have shown that hydrogen (H) isotopic signature of lipids is uniquely sensitive to the expression of heterotrophy relative to photosynthesis. This project will investigate the H isotope fractionation mechanisms (at molecular level, i.e. "site-specific") during biosynthesis of organic compounds in mixotrophic protists. It will establish the H isotope signature of lipid biomarkers as a novel tool to investigate the metabolic changes in these oceanic protists and thus provide a tool to determine the impact of mixotrophy on the global carbon cycle. This project will develop a numerical biochemical model based on a combination of chemostat experiments and cutting-edge mass spectrometry (MS) and nuclear magnetic resonance (NMR) isotopic ratio (ir) measurements. This include using irMS coupled to Gas Chromatography to measure the isotopic composition of lipids and to a high Temperature Conversion Elemental Analyzer after isotope equilibrations for measurement of non-exchangeable H in carbohydrates. NMR spectrometry adapted for the emerging technique of irm-NMR (ir measured by NMR) that can perform position-specific isotope analysis will elucidate the non-statistical distribution of 2H in different sites of a given molecule. The project will foster a new cross-disciplinary group uniquely skilled in isotope biogeochemistry and dedicated to extend our knowledge of the Plankton Physiological Ecology as well as its role on the Earth System.