Overview

Project Highlights:

  • Develop mass spectrometry imaging and high-throughput toxicity screening to study a key model organism in the environmental sciences.
  • Discover how pollutants affect the biochemistry of Daphnia.
  • Join a dynamic team of 25 metabolomics scientists and work alongside chemists, biologists and computer scientists.

 

Desorption electrospray ionisation (DESI) mass spectrometry imaging is an analytical tool for biomolecular research that can accurately locate and identify chemicals within a tissue section. While increasingly applied in biomedical research to study the metabolism of naturally occurring low molecular weight chemicals (metabolites), drugs and drug metabolites, its application to the environmental sciences, and in particular environmental (nano)toxicology, has yet to be realised. Rapid evaporative ionisation mass spectrometry (REIMS) is a novel technique that allows near real-time characterisation of tissue samples through analysis of the vapours released during sample analysis. Waters Corporation is at the forefront of the development of DESI and REIMS technologies.

The freshwater crustacean Daphnia is a model organism in biology, with immediate relevance for ecology, human and environmental health.

Their short generation time, large clutch sizes and ease of laboratory and field manipulation have assured their importance for setting environmental health standards, for testing chemical safety, for monitoring water quality, and as a model for ecological and evolutionary research. Recently, Daphnia has been included in the list of thirteen model organisms by the US National Institutes of Health. The importance of this organism has inspired the development of international consortia including the Daphnia Genomics Consortium and Daphnia Metabolome Project. However, to date, there have been no studies mapping the spatial localisation of metabolites and pollutants in Daphnia species. Such information is critical for mechanistically understanding the toxicity caused by environmental pollutants as well as, more broadly, biochemical changes induced by environmental stressors. Specifically, this knowledge is essential for the construction of Adverse Outcome Pathways (AOPs) that provide a knowledge management framework for the effects of pollutants on organisms.

The student will develop and optimise DESI and REIMS mass spectrometry approaches to study Daphnia. Subsequently they will apply DESI to characterise the spatial localisation of the baseline metabolome of this important species. Next they will investigate how exposure to pollutants can modulate the naturally occurring metabolites, as well as co-mapping the spatial location of the pollutants themselves. In addition, the student will explore the application of REIMS as a novel approach for high-throughput toxicity testing.

Desorption Electrospray Ionization (DESI) image of a tissue section, courtesy of Waters.

Methodology

The student will benefit from Waters technologies available at both the University of Birmingham and the new Waters Mass Spectrometry Headquarters in Wilmslow, less than 2 hrs from Birmingham. Specifically, the laboratory at Birmingham includes six new Waters UPLC-MS systems with one instrument configured for DESI research. The Waters Mass Spectrometry HQ houses a large suite of DESI and REIMS instrumentation. All of the biological investigations will be conducted at Birmingham utilising the recently rebuilt Daphnia culturing and exposure facility. Daphnia strains will be experimentally manipulated to perturb the baseline metabolome. This will be accomplished using environmentally relevant pollutants, including chemicals and/or nanomaterials. Histology will be conducted locally at Birmingham. Large molecular datasets will then be generated using mass spectrometry imaging and analysed using the newly expanded data storage and computational facilities at the University of Birmingham.

Training and Skills

CENTA students are required to complete 45 days of training throughout their PhD including a 10-day placement. In the first year, students will be trained as a single cohort on environmental science, research methods and core skills. Throughout the PhD, training will progress from core skills sets to master classes specific to the student's projects and themes. 

The student will be embedded within a large multi-disciplinary team of ca. 25 PhD students and postdocs all developing and applying metabolomics technologies. Consequently they will acquire a broad set of interdisciplinary skills, encompassing both analytical chemistry and computational research. Extensive training will be provided in mass spectrometry imaging, and more broadly in mass spectrometry based metabolomics. Wet lab training will also include the use of the Daphnia experimental facility in Birmingham. Bioinformatics training will enable the student to process, mine and visualise the molecular datasets generated in their research.

Timeline

Year 1:  The student will design, develop, test and optimise experimental approaches for studying the metabolites in Daphnia using both Desorption Electrospray Ionization (DESI) and Rapid Evaporative Ionization Mass Spectrometry (REIMS) technologies. The imaging studies will include the optimisation of cryopreservation, histological sectioning and DESI measurements. The REIMS experiments will investigate the feasibility of measuring chemical fingerprints of Daphnia in very high throughput within the context of screening for environmental (in particular pollutant) stress. During Year 1 the student will also begin acquiring bioinformatics skills for the handling and analysis of ‘omics Big Data.

Year 2:  Optimisation of the mass spectrometry technologies will continue alongside their application to conduct the first ever spatial mapping of the Daphnia metabolome. This will complement on-going research at Birmingham in the Daphnia Metabolome Project. The student will aim to localise multiple classes of metabolites in these animals – for example those involved in the endocrine axis, phospholipids, and important classes of sulfated lipids that are known to play an important role in cross species signalling from Daphnia (primary consumer) to unicellular algae (primary producer). A primary objective of year 2 will be the creation and publication of the Daphnia Metabolome Atlas.

Year 3:  The effects of pollutants (chemicals and/or nanomaterials) on the Daphnia metabolome will be investigated in the third year. Specifically to localise sites of uptake of the pollutants (when those chemicals can be imaged), and sites of biochemical disruption. These studies will help to inform on the mechanisms of toxicity of chemicals and/or nanomaterials at the organ level, an integral piece of knowledge for the construction of Adverse Outcome Pathways. As such this project will feed important new information into the Birmingham research team’s efforts to build a deeper mechanistic understanding of toxicology. The student will complete their data analysis and transition to thesis writing at the end of year 3.

Partners and collaboration (including CASE)

This is a collaborative project between the University of Birmingham and Waters Corporation, one of the leading developers and manufacturers of LC-MS instrumentation worldwide. Waters will provide an attractive CASE studentship package together with the opportunity for the student to learn mass spectrometry imaging approaches in their new Mass Spectrometry facility in Wilmslow. The School of Biosciences at Birmingham is a world leader in metabolomics and more broadly environmental ‘omics research, achieving an impressive performance in the Research Excellence Framework 2014, rising up to 6th place in the elite, research-focused Russell Group of UK universities.

Further Details

Prof Mark Viant, University of Birmingham – m.viant@bham.ac.uk

Metabolomics at Birmingham:

http://www.birmingham.ac.uk/research/activity/metabolomics/index.aspx

Training in metabolomics at Birmingham

http://www.birmingham.ac.uk/bmtc

School of Biosciences

http://www.birmingham.ac.uk/schools/biosciences/news/2014/18Dec-biosciences-top-six-for-research.aspx

Waters Mass Spectrometry HQ, Wilmslow

http://www.waters.com/waters/en_GB/New-Mass-Spectrometry-Facility/nav.htm?cid=134653003&locale=en_GB