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EQUIP Report Summary

Project ID: 661063
Funded under: H2020-EU.1.3.2.

Periodic Reporting for period 1 - EQUIP (Elemental quota in marine phytoplankton for effective carbon sequestration, clean energy and biogeochemical modelling)

Reporting period: 2015-05-07 to 2017-05-06

Summary of the context and overall objectives of the project

The EQUIP Project seeks to address the use of phytoplankton for the development of effective phytoplankton-based biotechnology strategies. In the oceans, phytoplankton are responsible for ~50% of the atmospheric carbon fixed on earth, and they have contributed substantially to the mitigation of the negative effects caused by the human-driven increase in atmospheric CO2. This natural phenomenon can be engineered for carbon sequestration, and to produce 3rd generation biofuels.
The aim of the EQUIP Project is to exhaustively describe the element composition of representatives from 3 key phytoplankton groups, and relate it to phytoplankton biochemical composition, to understand how the organisms behave in environmental conditions with respect to resource availability and element assimilation and allocation. This project focuses on diatoms, dinoflagellates and coccolitophores because they are key biogeochemical players in the ocean, in terms of primary production and carbon export, as well as because of their ability to impact the pelagic system through blooms.
This project also aims to give visibility to a novel single-cell methodology, X-Ray Microanalysis (XRMA), for the analysis of elemental composition at a single-cell level. In order to predict climate scenarios using biogeochemical models, accurate conversion factors (e.g. element/volume ratios) are needed. Biogeochemical models enable the study of exchanges among C pools or reservoirs (e.g. between hydrosphere and atmosphere) in order to know whether a system is a source or a sink of CO2, and to predict primary production and carbon export. Up to now, the study of conversion factors has been addressed using bulk analysis methods that overestimate C and N, mostly, because debris, dead cells and other organic excretion products are included with the plankton samples. The use of single-cell XRMA to determine the elemental composition of plankton will overcome this drawback.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

Experimental work:
We compared growth curves and nutrient utilization (nitrate and phosphate) of fast-growing and bloom-forming species of 3 key plankton groups: diatoms (Phaeodactylum tricornutum, Cylindrotecha fusiformis and Thalassiosira pseudonana), dinoflagellates (Alexandrium minutum and Prorocentrum micans) and coccolitophore (Emiliania huxleyi). We focused our studies on nutrient (C, N, P, S) allocation in diatoms as they have the highest potential for obtaining products of interest, especially P. tricornutum, which have low silicon requirement.
We cultivated monocultures of P. tricornutum in triplicate and sampled for elemental and biochemical analysis, focusing on the first 12 h, to follow the draw-down of phosphate in the media and its conversion to different phosphate molecules within the cell. We optimized sample preparation for XRMA to avoid the interference of the nutrient-rich media. We also combined the method for biochemical analysis used in Dr. Vaidyanathan's lab for proteins, carbohydrates and pigments determination, with a method to analyse phospholipids, and set up a method for the analysis of intracellular dissolved nutrients and dissolved silicate analysis. Finally, we established collaborations with other researchers for the analysis of polyphosphate (Dr. S. Duhamel, LDEO-Columbia University, USA) and DNA/RNA (Dr. Elisa Berdalet, Institute of Marine Sciences, ICM-CSIC, Spain). Part of this samples are going to be used to describe a new model to convert from elemental to biochemical composition, since the models tried so far are not valid for diatoms.
We cultivated all the species described above, plus a phosphate-limited P. tricornutum, and collected samples during the exponential and stationary phases of the growth curve. We used this data to compare the elemental compositions obtained with both the novel single-cell method and the bulk analysis method. To do so, we set up new methods for the analysis of particulate organic phosphorus and carbon, nitrogen and sulphur elemental analysis using filters, a procedure not used routinely in Dr. Vaydianathan's lab. Dinoflagellate samples were also cultivated for training in ToF-SIMS analysis.
Field work:
With the help of Dr. Claire Widdicombe, Plymouth Marine Laboratory, we obtained samples from the Western Channel Observatory to determine the elemental composition of single diatom cells by XRMA from four different months with contrasting environmental regimes. We are analysing this data to build new regressions element vs. volume that will be used to develop more accurate biogeochemical models of the C-cycle. We will use our model obtained with cultures to convert from elemental to biochemical composition.
Part of the results have already been disseminated through conferences: 2016 Challenger Society - Oceans and Climate, 5-9 September 2016, Liverpool, UK, and 6th UK Algae Conference, Sheffield, UK.

Professional development training:
The University of Sheffield is an ideal place for the professional development of early-career researchers. During her time at the University of Sheffield, the fellow successfully completed the Sheffield Teaching Assistant program, leading to a certification as an Associate Fellow of the UK Higher Education Academy (AFHEA). The fellow also had the opportunity to join leadership and writing in the sciences workshops, as well as to participate in several outreach events organized by The University of Sheffield.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

The conclusions of this work advance our understanding of element allocation in diatoms. This knowledge would be useful for the optimization of processes that use diatoms as a source to sustainable manufacture products of interest from CO2. Also, our study provides more accurate values of elemental composition of key phytoplankton species, in terms of impact on the pelagic oceanic system and global C-cycle. This knowledge is relevant for the determination of the error introduced to models when data obtained using bulk analysis is used.
During this project we obtained for the first time the single-cell elemental composition of Emiliania huxleyi, a well-known and bloom-forming coccolitophore, increasing the number of key plankton groups that have been studied with the single-cell X-Ray Microanalysis method. Moreover, we singnificantly increased the knowledge about the elemental composition of diatoms from the field, in terms of single-cell analysis and number of species, which was, so far, limited to few species from the NW Mediterranean Sea.
We would also like to take this opportunity to emphasize the importance of Time-Series Programs, like the Western Channel Observatory, for the advancement of science. Apart from providing the full infrastructure needed for sample collection and process (research vessels, laboratories, staff expertise), we were able to firstly, put our elemental composition data in context with the historical average climatology of the Western Channel Observatory, and secondly, complement our results with additional biogeochemical data collected at the same time, like nutrients, dissolved organic and inorganic carbon, and physical characteristics of the water column. Since phytoplankton can adapt their biochemical composition, and hence, their elemental quota (carbon, oxygen, nitrogen, phosphor and sulphur) depending on the environmental conditions, the complementary variables provided by Plymouth Marine Laboratory are key to understand and extrapolate our results.

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