Within the framework of the Master research programs funded by the Fondation pour la Recherche sur la Biodiversité (FRB), the members of the GloMEc (Global Change in Mountain Ecosystems) project and the BIOREF team (Biodiversity, trophic networks and flows in aquatic ecosystems) of the Functional Ecology and Environment Laboratory (UMR 5245) are looking for a candidate for a project at the nexus of biodiversity (microbial ecology) and health (epidemiology of amphibian infectious diseases).
Background :
Mountain ecosystems are of paramount importance for human societies (provision of drinking water, among others) but very sensitive to global change (climate change, chemical and nutrient pollution, habitat degradation, introduction of invasive species) caused by human activities (pastoralism, tourism, dam construction, fish-stocking of high-altitude lakes). Two indicators of mountain freshwater ecosystem health are considered here: biofilms and amphibians. The former are very productive and diverse communities in freshwater ecosystems. At high elevation, they form the basis of food webs: the tadpoles of certain amphibians feed on them. Amphibians play an active role in water purification and form the link between the aquatic and terrestrial worlds. Their productivity (high biomass) also makes them keystone species of mountain ecosystems. Amphibian chytridiomycosis, a panzootic disease caused by the aquatic fungus Batrachochytrium dendrobatidis (Bd), spread worldwide by humans, is a serious threat to amphibian biodiversity, having caused hundreds of declines and extinctions worldwide. This disease is observed in the Pyrenees where it causes declines in the populations of the common-midwife toad (Alytes obstetricans), which our team has been studying for over 10 years.
However, the epidemiology of amphibian chytridiomycosis is not yet elucidated. There is evidence to suggest that amphibians may be protected by biotic environmental factors, such as aquatic microfauna (Schmeller et al. 2014). Our aim is to assess whether benthic biofilms in Pyrenean lakes can reduce the infectious risk posed by Bd. We also want to determine whether different types of biofilms have different protective capacities, and whether they can help limit infection. Indeed, Bd has an aquatic, motile life stage, the zoospore (the infective stage), which is likely to come into contact with benthic biofilms. The latter are literally mini-ecosystems known either to be able to harbour and protect certain human pathogens causing waterborne diseases (when the latter manage to colonise a biofilm), or to eliminate them (biofilms can harbour micropredators, or contain microorganisms antagonistic to Bd, for example by secreting antifungal substances). Our hypothesis is that the greater the biodiversity in the biofilms, the more they will be able to reduce the number of zoospores in the water column, thus decreasing the infective pressure. The alteration of biofilms by human activities would then lead to a higher risk of infection and/or parasite load for amphibians.
To test this hypothesis, we have adopted a transdisciplinary approach, linking in- and ex-situ investigations (study of biofilm’s taxonomic composition by metabarcoding, correlation with risk of infection by Bd). We would like to complete this approach first with an initial laboratory experiment to find out whether these biofilms alone inhibit Bd (experiment 1). Infections of tadpoles in the laboratory are the next step (experiment 2). Using the results of experiment 1 (if promising), we will select two types of biofilms for this project that are most likely to answer the research question. We will breed tadpoles on a good quality biofilm (diatom-dominated), another group on a poor quality biofilm (rich in cyanobacteria, from a « degraded » lake) and a final group without biofilm (fed with industrial food tablets). We will experimentally infect the tadpoles with a known load of Bd. We will evaluate, by qPCR, the prevalence and average infection intensity of Bd in each treatment group over time. The presence of ‘negative’ controls (not exposed to Bd) for each group will also provide information on the nutritional quality of the two biofilms, by measuring tadpole weight.
The approach is multidisciplinary, involving veterinary (nutrition, epidemiology, diagnosis), ecological (limnology) and microbiological sciences. The scale is regional (Pyrenees). We have already obtained a favourable opinion from the ethics committee n°73 and the Ministry of Higher Education and Research (ref APAFIS#25575-2020052812331078 v4) for the experimental protocol of this project, and have the necessary infrastructures to carry it out.
Keywords: Biofilms, parasite, ecosystem health, microbial ecology, host-pathogen-environment-microbiome interactions, disease pyramid
Skills required:
-Good level of English and inferential statistics required (GLM level if possible, advantage if basic knowledge of GLMM). Mastery of R software required.
-Excellent synthesis, writing and oral presentation skills (FR or EN)
-The student will have to demonstrate absolute meticulousness and assiduity, as well as self-denial. He/she should have a strong sense of responsibility. The care of animals is necessary to guarantee their well-being, but sometimes requires flexibility in terms of time.
-Strong organisational skills are essential.
-Ability to work in a team, but also autonomously and reliably (ability to follow and adapt to instructions)
-The problematic is complex and new, which means that the protocols are not perfected. The student must be able to manage the unexpected by solving minor technical problems. He/she must therefore be astute and ingenious, but also know how to call for help when necessary (major problems).
-Basic knowledge of the epidemiology of infectious diseases is desirable.
-Previous experience of laboratory work (e.g. qPCR) will be a considerable advantage. Experience with amphibians or other laboratory animals is a plus.
-The work will involve trips to the Pyrenees to collect water samples, biofilms, etc. A good level of hiking is desirable.
Email to hugo.sentenac@toulouse-inp.fr

Workplace: Functional Ecology and Environment Laboratory – UMR 5245 -INP-ENSAT, Avenue de l’Agrobiopole, BP32607 Auzeville-Tolosane 31126 Castanet-Tolosan Cedex, France
Supervisors: Hugo Sentenac, Adeline Loyau, Dirk Schmeller
Examples of skills and experiences potentially acquired at the end of the course:
– In-depth introduction and familiarisation with the world of scientific research through collaboration with a team of researchers. The student will see how an international scientific project (GloMEc project Global Change in Mountain Ecosystems, https://www.p3mountains.org/) is run, and will significantly help its members with the implementation of this project.
– Literature review / Scientific reading / Critical thinking / Scientific writing. All these elements will help to improve the level of English language skills throughout the project
– Introduction to project design, carrying out fieldwork, introduction to laboratory work (microscope counting, handling microbial culture, etc.)
– Statistical analysis: generalized linear mixed models (GLMM)
– Theoretical and practical knowledge of ecology, microbial ecology (including biofilm communities), disease ecology and epidemiology. The student will become familiar with important concepts such as One Health/EcoHealth in the context of sustainability, the concept of the disease pyramid (host-pathogen-microbiota-environment) and the phenomenon of the emergence of infectious diseases, all of which are particularly relevant today. Best of all, he will be able to apply these concepts while focusing on the health of freshwater ecosystems, studying biofilm communities and the most aggressive pathogen ever known (which, by the way, is fortunately not zoonotic). This knowledge will be transferable to other systems and as such will most likely prove useful to the student.
Funding: stipend of approximately €600 per month.
Period: Flexible, but April-September 2022 preferred. Deadline October 2022: submission of the report to the FRB.
Note: This master project will be part of a larger project. This means that the Master student will benefit from significant advantages throughout the internship, namely
– continuous support provided throughout the project
– the possibility of being included in scientific publications if this Master project leads to new discoveries useful for the GloMEc project.
– if interested and available, the student could help the supervisors during the next fieldwork in the mountains in late spring and summer 2022 (we have many study sites in the Pyrenees) and learn more about the ecosystems and fauna of the Pyrenees, in particular amphibians (good level of hiking required)
– Potential for future collaboration, provided that this master project runs smoothly

For more information (GloMEc website + prior reading advice*):
– BIOREF team website: https://www.eco.omp.eu/research-team/biodiversite-reseaux-trophiques-et-flux-dans-les-ecosystemes-aquatiques-bioref/
– GloMEc project website: https://www.p3mountains.org/
(1) Fisher, M.C., Garner, T.W.J. (2020) « Chytrid fungi and global amphibian declines », Nature Reviews Microbiology, 1-12. Il existe des tonnes d’articles sur la chytridiomycose et la Bd des amphibiens, et dans une moindre mesure sur la Bsal. Celui-ci est récent (et donc à jour) et donne une bonne vue d’ensemble
(2) Flemming, H.-C., Wuertz, S. (2019) « Bacteria and archaea on Earth and their abundance in biofilms », Nature Reviews Microbiology, 17(4), 247-260. Bien que l’objectif principal du document ne nous concerne pas trop, il est bien écrit et magnifiquement illustré, et il fournit des définitions merveilleuses et claires. Là encore, il est récent, de sorte que vous pouvez y trouver de nombreuses bonnes références. Si vous devez en choisir une, je vous le recommande vivement :
(3) Battin, T.J., Besemer, K., Bengtsson, M.M., Romani, A.M., Packmann, A.I. (2016) « The ecology and biogeochemistry of stream biofilms », Nature Reviews Microbiology, 14(4), 251-263. Le titre parle de lui-même. Encore une fois, de superbes illustrations
(4) Schmeller, D.S., Blooi, M., Martel, A., Garner, T.W.J., Fisher, M.C., Azemar, F., Clare, F.C., Leclerc, C., Jäger, L., Guevara-Nieto, M., Loyau, A., Pasmans, F. (2014) « Microscopic Aquatic Predators Strongly Affect Infection Dynamics of a Globally Emerged Pathogen », Current Biology, 24(2), 176-180. Il établit un lien entre la composition des espèces du « plancton » des lacs pyrénéens et la dynamique de l’infection par le Bd. Car dans certains lacs, il y a des prédateurs compétents du Bd, et dans d’autres, il n’y en a pas. Fondamentalement, ici, nous voulons voir si c’est la même chose avec les communautés de biofilms.
(5) Schmeller, D.S., Loyau, A., Bao, K., Brack, W., Chatzinotas, A., De Vleeschouwer, F., Friesen, J., Gandois, L., Hansson, S.V., Haver, M., Le Roux, G., Shen, J., Teisserenc, R, Vredenburg, V.T. (2018) « People, pollution and pathogens – Global change impacts in mountain freshwater ecosystems », Science of The Total Environment, 622-623, 756-763. Pour avoir une idée plus précise du contexte de notre projet
(6) Bernardo-Cravo, A.P., Schmeller, D.S., Chatzinotas, A., Vredenburg, V.T., Loyau, A. (2020) « Environmental Factors and Host Microbiomes Shape Host-Pathogen Dynamics », Trends in Parasitology. Introduction au concept de pyramide des maladies avec, entre autres, le système d’amphibiens-Bd comme exemple
(7) Rapport, D.J., Costanza, R., McMichael, A.J. (1998) « Assessing ecosystem health », Trends in Ecology & Evolution, 13(10), 397-402. La base
If problems to get full version of articles, contact:
hugo.sentenac@toulouse-inp.fr

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