Biogeochemical cycling of N-osmolytes in the surface ocean

Lead Research Organisation: University of Oxford
Department Name: Engineering Science

Abstract

Nitrogen-containing compounds, including glycine betaine (GBT), choline and trimethylamine N-oxide (TMAO) are ubiquitous in marine organisms. They are used by marine organisms as compatible solute in response to changes in environmental conditions, such as increasing salinity, because they do not interfere with cell metabolisms. They also have beneficial effects in protecting proteins against denaturation due to chemical or physical damages.

In the marine environments, these compounds are frequently released into the sea water due to the change of environmental conditions, such as viral attack or grazing. The released nitrogenous osmolytes serve as important nutrients for marine microorganisms, which can use them as carbon, nitrogen and energy sources. It is well known that the degradation of these nitrogenous osmolytes contribute to the release of climate-active gases, including volatile methylated amines. Methylated amines are important sources of aerosols in the marine atmosphere, which help to reflect sunlight and cause a cooling effect of the climate. There is an urgent need to understand the microbial metabolism of these compounds and their seasonal cycles in the marine water column, in order to better understand their role in marine biogeochemical cycles and their role in future climate change.

Built on the recent progress on the discovery of the new pathway of TMAO degradation in marine organisms and the development of a powerful liquid chromatography with mass spectrometry (LC-ESI-MS) method for simultaneous quantification of these nitrogenous osmolytes from the applicants' laboratories, this timely proposal aims to determine the seasonal cycle of nitrogenous osmolytes in the surface seawater and to address how these environmentally-relevant compounds are degraded and what are the major microorganisms that are involved in the process. The data generated will fill in a major gap in our knowledge of marine carbon and nitrogen cycles and the contribution of these compounds in future climate change through the release of climate-active molecules.

Using newly developed analytic techniques, we aim to determine the seasonal cycle of standing concentrations of nitrogenous osmolytes in the surface seawater and microbial oxidation activities. These data will be incorporated to a biogeochemical model for future prediction of biogeochemical cycles of N-osmolytes under climate change.

Using cultivated model organisms, we aim to define the key genes, enzymes and the metabolic pathways in GBT and TMAO degradation by marine planktonic microbes.

Using molecular and single cell manipulation techniques, we aim to further determine the key microbial players involved in the metabolism of nitrogenous osmolytes in surface seawater from the English Channel.

This work will generate novel knowledge about our understanding of microbial transformation of these nitrogen containing compounds, and will fill in a serious gap in knowledge of marine carbon and nitrogen cycles. The project is expected to further strengthen the UK as a leading country not only in the research of marine biogeochemical cycles and marine microbiology, but also in the development of cutting edge technology in environmental science.

Planned Impact

This project addresses fundamental questions relating to the marine biogeochemical cycles and climate-active gas emissions from the natural environment, both of which are crucial to our understanding of global biogeochemical cycles and in the drive for a more sustainable future. The work is therefore of the utmost relevance to NERC's strategic aims, particularly Biodiversity Science and Climate Change themes.

The proposal work relies on the recent development of the two new techniques, i.e. Raman microspectroscopy in combination with stable isotope labelling and liquid chromatography (LC) with electrospray ionization mass spectrometry (LC-ESI-MS) for quantifying N-osmolytes. The proposed work is therefore of direct relevance to the immediate science community and NERC's Technologies theme.

The main beneficiary of the knowledge generated from this study is anticipated to be public sectors focusing on marine ecosystems services and the scientific community working on marine aerosol research and marine microbes. Marine aerosols play an important role in the Earth system, not only in climate, but also in atmospheric chemistry and human health problems. Until recently, sea-salt formation from bubble bursting and dimethylsulfide (DMS) oxidation were thought to be the main sources of marine aerosols. There is growing evidence in the past five years that methylated amines play an important role in marine aerosol formation, in addition to DMS. Methylated amine concentrations measured over the North Atlantic Ocean and the North Pacific Ocean have shown distinct seasonal variation and are directly linked to the primary productivity of phytoplankton. However, researchers working on atmospheric chemistry and modelling have not yet been able to pinpoint the origin of the methylated amines found in the marine atmosphere. There is a lack of communication to such a community which can be catalyzed through the proposed research and subsequent impact plan activities.

Our topic also has immediate general interest for the wider public, including general public, school pupils. With its relevance to climate and marine biology, this type of work is guaranteed to attract considerable attention in the media.

A variety of methods will be used to engage with the end-users, including workshops, a detailed project website, regular updates in social media (such as PML and Warwick's twitter accounts https://twitter.com/PMLGroup; https://twitter.com/WarwickLifeSci), publications in popular magazines (e.g. Planet Earth), and visits and exhibitions at local schools. One of the things that we are keen to do is to have end user opinion at very early stages of our research. We consider that engagement of end users from the very early stages of the project will also benefit our research and help to formulate new impact dissemination activities.

Publications

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Berry D (2015) Tracking heavy water (D2O) incorporation for identifying and sorting active microbial cells. in Proceedings of the National Academy of Sciences of the United States of America

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Fan C (2020) Chromosome-free bacterial cells are safe and programmable platforms for synthetic biology. in Proceedings of the National Academy of Sciences of the United States of America

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Hsu CC (2020) A single-cell Raman-based platform to identify developmental stages of human pluripotent stem cell-derived neurons. in Proceedings of the National Academy of Sciences of the United States of America

 
Description We have developed Raman activated cell ejection which is able to sort cells based on label-free biochemical fingerprint of cells- single cell Raman spectra.
To our knowledge, this is the first report of the use of Raman activated cell sorting coupled to single cell genomics. Single cell genomics based on Raman sorting can not only sort cells with specific compounds but also isolate cells with carbon, nitrogen and general metabolic activity when it is coupled with stable isotope probing, which would help link the specific metabolism of single cells (e.g. carbon, nitrogen substrate metabolism or general metabolic activity) and assist in defining the ecological functions of uncultivated bacteria in the environment. This technology will open a new frontier to understand previously uncultured bacteria in nature which account for 99% species.
Exploitation Route 1. We have filed a patent on 24th Feb 2016, which has been granted (Cell sorting WO 2017/144886).
2. The patent has been licensed to Horiba Scientific Ltd which is the largest manufacturer of Raman instrument in the global market.
3. The project PI has organised an international conference - Ramafest 24-25 June, 2019 at Oxford University to disseminate Raman sorting technology developed in the project.
Sectors Aerospace, Defence and Marine,Agriculture, Food and Drink,Chemicals,Creative Economy,Energy,Environment,Healthcare,Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology

 
Description 1. We have filed a patent on 24th Feb 2016, which has been granted (Cell sorting WO 2017/144886). 2. The patent has been licensed to Horiba Scientific Ltd which is the largest manufacturer of Raman instrument in the global market. 3. The project PI has organised an international conference - Ramafest 24-25 June, 2019 at Oxford University to disseminate Raman sorting technology developed in the project. 4. The prototype of Raman activated cell sorter has been made. At least 5 academic research groups want to copy the system in their lab. They are: three institutes of Chinese Academy of Sciences,Qingdao Institute of Bioenergy and Bioprocess Technology, Institute of Urban Environment, Suzhou Institute of Biomedical Engineering and Technology, as well as Shanghai Jiaotong University and Oklahoma University.
First Year Of Impact 2015
Sector Aerospace, Defence and Marine,Agriculture, Food and Drink,Chemicals,Energy,Environment,Healthcare,Pharmaceuticals and Medical Biotechnology
Impact Types Economic

 
Description Chair an international meeting Ramanfest 2019
Geographic Reach Multiple continents/international 
Policy Influence Type Influenced training of practitioners or researchers
URL https://www.horiba.com/en_en/company/news/detail/news/4/2019/horiba-uk-hosts-ramanfest-2019/
 
Description An integrated microfluidic - single cell Raman technology for rapid diagnosis of pathogens and their antibiotic resistance
Amount £748,508 (GBP)
Funding ID 104984 
Organisation Innovate UK 
Sector Public
Country United Kingdom
Start 01/2019 
End 12/2021
 
Description Effector gene persistence in bacterial plant pathogens
Amount £430,764 (GBP)
Funding ID BB/R009236/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 04/2018 
End 04/2021
 
Description LinkPI: Linking Phenotype function with Identity: a novel integrated single-cell technology and metagenomics approach
Amount £83,396 (GBP)
Funding ID NE/S008721/1 
Organisation Natural Environment Research Council 
Sector Public
Country United Kingdom
Start 12/2018 
End 11/2020
 
Description CAS Bioenergy collaboration 
Organisation Chinese Academy of Sciences
Country China 
Sector Public 
PI Contribution Prof Jian Xu is Director of BioEnergy Directorate, Qingdao Institute of BioEnergy and Bioprocess Technology (QIBEBT), Chinese Academy of Sciences, has established a formal collaboration with me on bioenergy research. They will characterise novel gene from algae, which will be introduced into our SimCells to biofuel production.
Collaborator Contribution He has visited Oxford in Dec 2015. He has paid my travelling, accommodation cost (about CYN20K) to visit China and give a talk in his institute. He would like to pay and get a licence for this technology.
Impact We are in the process of commercialization collaboration.
Start Year 2007
 
Description Collaboration project with Chinese Academy of Sciences 
Organisation Chinese Academy of Sciences
Country China 
Sector Public 
PI Contribution Oxford University and Chinese Academy of Sciences have signed a research agreement. Suzhou Institute of Biomedical Engineering and Technology (SIBET), CAS will support £450,000 in Oxford and more than £1M in SIBET to establish a Raman research centre.
Collaborator Contribution SIBET will contribute financial support and development of intrumentation.
Impact 1. Oxford will get 3 year PDRA working on Raman technology. 2. SIBET will set up a research centre for Raman technology.
Start Year 2019
 
Title CELL SORTING 
Description The present invention relates to a screening chip for cell sorting, said screening chip comprising a substrate having opposing first and second surfaces, wherein at least a portion of said first surface is coated with a Raman-inactive coating material which can be vaporised by laser irradiation at a wavelength and wherein said substrate is transparent to laser radiation at wavelength In further aspects of the invention, a cell sorting method employing the screening chip and a cell sorting apparatus employing the screening chip are provided. 
IP Reference WO2017144886 
Protection Patent application published
Year Protection Granted 2017
Licensed Yes
Impact It has been licensed to Horiba Scientific Ltd, the largest Raman manufacturer in the world.
 
Company Name Oxford Molecular Biosensors 
Description OMB is offering a range of products and services, including several biosensors with differing functionalities, greater robustness and sensitivities than those currently on market. The company has also developed a platform technology which enables rapid design and construction of bespoke biosensors tailored to the specific needs of the customer. A further development is a portable device which will enable biosensor assessment to be made on site and in remote locations. License agreements to employ our biosensors, training courses for their application and data analysis as well as consultation on best measures for remediation of contamination are also available. 
Year Established 2017 
Impact Provide service to Primark, Bangladesh government water project.
Website https://www.omb.co.uk/
 
Description Research Expo St Edmund Hall 2017 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact The second St Edmund Hall Research Expo will take place on Saturday 25 February 2017 (6th Week), 12:30-5:00pm. Once again, the College will showcase the wide diversity of research being undertaken by its students and academics - from undergraduates to Fellows - via sessions of short 'Teddy Talks' (all aimed at a non-specialist audience) and three rooms of interactive displays and exhibits.
Year(s) Of Engagement Activity 2017
URL https://www.seh.ox.ac.uk/expo