Dissecting the evolutionary ecology of a unique secondary endosymbiosis.


Summary: Endosymbiosis is recognised as a fundamental evolutionary innovation that underpins the origins of many unicellular and all multicellular lifeforms (Chomicki et al., 2019). Understanding the biology of such phenomena can shed light on key drivers of inter-species cooperation and provide an important window into early origins of cellular life on the planet. The PhD student will link to a recently funded project to deploy new tools: single cell genomics and transcriptomics, as well as metabolomics; to explore a unique and poorly understood endosymbiosis involving an emergent disease agent of major economic importance. Supported by a world class supervisory team (Prof. Mike Barrett, Dr. Martin Llewellyn, Wellcome Trust Centre for Integrative Parasitology, Glasgow; Dr. Guillame Chimocki, Life Sciences, Durham) and a range in international collaborators (Prof. John Archibald, Dalhousie University, Canada; Dr. Neil Ruane, Marine Institute, Ireland), the student will have the opportunity to develop skills at the cutting edge of genomics and molecular biology, undertake training at international centres of excellence in parasitology and evolutionary biology in the UK and North America, and engage in marine biological fieldwork on the west coast of Ireland and Scotland. Finally, this project has strong links with aquaculture industry via project partners Scottish Sea Farms (SSF, Dr. Ralph Bickerdike) and the student will also get valuable experience working alongside industry.

Secondary endosymbiosis: The phenomenon in which eukaryotic organisms engulf other eukaryotes is termed ‘secondary endosymbiosis’. Secondary endosymbiosis underpins the evolution of many eukaryotic phototrophs and is thought to have involved the engulfment of an ancestral eukaryotic rhodophyte (Oborník 2018). The number of times this has occurred in evolutionary history is a moot point. However, it is clear that rhodophyte-origin plastids play a key role in their host cell’s biology. In some cases, the symbiont has lost the ability to photosynthesize, which leaves them a relic non-photosynthetic plastid in a secondarily heterotrophic cell. This is the case for the apicomplexans, which include the causative agents of toxoplasmosis and malaria. The basis of ongoing metabolic dependency is not always clear, however some conserved functions across plastids belonging to different apicomplexan lineages include isoprenoid (IPP and DMAPP), tetrapyrrole, and fatty acid biosynthesis (Janouškovec et al. 2015).

The study system: Paramoeba perurans causes amoebic gill disease (AGD) and is a major pathogen in salmonid aquaculture, causing > £400 million in losses per annum world-wide. There are currently no drugs available to treat AGD. P. perurans has a unique cellular biology that can be readily exploited given the right tools. Enclosed within its cytoplasm is a bizarre endosymbiont – Perkinsela. Genomic sequence data suggest that the basic physiology of this endosymbiont has many of the same biochemical features as found in kinetoplastid pathogens of man and domestic livestock (e.g. Sleeping sickness, Leishmaniasis and Chagas disease).

The endosymbiosis between P. perurans and Perkinsela is unique among eukaryotes because it does not involve an originally photosynthetic symbiont. Prior investigations have established interdependence between the kinetoplasitid and amoeba based on predicted gene content and ontogeny in the related Parameoba pemaquidensis (Tanifuji et al. 2017).

This studentship has three major aims:
Aim 1: Understand the molecular basis of the obligate dependence between P. perurans and Perkinsela. The student will use genome sequencing (long read technologies jointly with illumina short reads for polishing), single-cell transcriptomics as well as metabolomics to dissect the molecular basis of the symbiosis. Specific drug knock outs jointly with transcriptomic analysis will allow to functionally test metabolic dependences.

Aim 2: Undertake rational Amoebic gill disease (AGD) drug discovery. A detailed understanding of dependences in the between P. perurans and Perkinsela symbiosis will provide a window to test drugs efficient on AGD. In collaboration with the Wellcome Trust Centre for Intergrative Parasitology, the student will test drugs targeting metabolic dependences of P. perurans. Ultimately trials will be performed in fish farms with SSF.

Aim 3: Trace the evolution of this unique endosymbiosis. Using a recent approach (Kwong et al., 2019), we will reconstruct the phylogenetic histories of both the host P. perurans and the Perkinsela symbiont clades, relying on a range of archival environmental samples as well as new marine collections. Using targeted sequence enrichment, we will sequence the genomic regions of the host identified as driving the obligate dependence (Aim 1), and analyses of substitution rates (dn/ds) will inform of their functionality. When possible, close relatives will be cultured to assay the presence of the symbiont using microscopy and FISH. This will allow t evolutionary history of this unique endosymbiosis, specifically testing (i) the number of origins of this symbiosis across the clade encompassing P. perurans, (ii) whether the obligate dependence has been lost or is retained throughout the clade, and (iii) whether all P. perurans strains evolved the same or distinct dependences on Perkinsela.

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Via genome sequencing, standard and single-cell transcriptomics, as well as metabolomic analyses to validate predicted pathways, the student will establish the role of Perkinsela in P. perurans biology. At the University of Glasgow, the student will undergo training in genome sequencing and annotation (Aim 1 and 2). At Durham University, Biosciences, the student will receive training in key ecological and evolutionary theory around symbioses as well as in-depth phylogenetic comparative methods including phylogenetic inference, ancestral state estimation and gene substitution rate with the aim of reconstructing the origins of the symbiosis (Aim 3). At the University of Glasgow, Wellcome Trust Centre for Integrative Parasitology the student will learn how to exploit plastid-targeted drug knock-outs and single cell sequencing to unpick the metabolic interactions between host and symbiont. During a secondment to John Archibald’s laboratory at Dalhousie University, Canada, the student will receive further training in amoebozoan genome assembly and annotation with particular reference to secondary endosymbiosis.

Project Timeline

Year 1

Student, assisted by PIs and dedicated post-docs, sequences P. perurans genome and transcriptome. Visits Canada to undertake training in genome assembly

Year 2

Student assists with targeted drug knock-outs of P. perurans organelles alongside metabolomics and transcriptomics to unpick ecological/biochemical basis of symbiosis

Year 3

Student will undertake sample collection and sequencing of P. perurans and symbiont clades to establish the evolution of the symbiosis

Year 3.5

Data analysis

& Skills

The student will receive training in genomics, transcriptomics (inc. single cell), phylogenetic, metabolomics, molecular biology, microscopy and more. The student will have access to world class supervision and benefit from links to international research networks as well as to industry.

References & further reading

Rodger HD (2013.) Amoebic gill disease (AGD) in farmed salmon (Salmo salar) in Europe. . Fish Veterinary Journal 16.

Harmer J, Yurchenko V, Nenarokova A, Lukeš J, and Ginger ML 2018 Farming, slaving and enslavement: histories of endosymbioses during kinetoplastid evolution
Parasitolgy 145, pp. 1311-1323

Creek DJ, Barrett MP (2013) Determination of antiprotozoal drug mechanisms by metabolomics approaches. Parasitology 141: 83-92.

Schwabl, P., Imamura, H., Van den Broeck, F. … & Llewellyn, MS Meiotic sex in Chagas disease parasite Trypanosoma cruzi. Nat Commun 10, 3972 (2019)

Räz B, Iten M, Grether-Bühler Y, Kaminsky R, Brun R (1997) The Alamar Blue® assay to determine drug sensitivity of African trypanosomes (T.b. rhodesiense and T.b. gambiense) in vitro. Acta Tropica 68: 139-147.

Chomicki, G., Weber, M., Antonelli, A., Bascompte, J. and Kiers, E.T., 2019. The impact of mutualisms on species richness. Trends in ecology & evolution, 34(8), pp.698-711.

Chomicki, G., Kiers, E.T. and Renner, S.S., 2020. The evolution of mutualistic dependence. Annual Review of Ecology, Evolution, and Systematics, 51. (in press)

Kwong, W.K., Del Campo, J., Mathur, V., Vermeij, M.J. and Keeling, P.J., 2019. A widespread coral-infecting apicomplexan with chlorophyll biosynthesis genes. Nature, 568(7750), pp.103-107.

Further Information

Application procedure:  For IAPETUS2 applications to the University of Glasgow please use the dedicated application portal: www.gla.ac.uk/ScholarshipApp  (you will still need to submit your administrative details to the IAPETUS2 website as well).

All enquiries please email martin.llewellyn@glasgow.ac.uk


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