M2 Internship

Supervisor : Nicolas Pollet in collaboration with François Brischoux (CEBC, Chizé)

Title : Effects of glyphosate exposure on the gut microbiota of common toad tadpoles

Keywords: Microbiome – Ecotoxicology – Amphibians – Glyphosate

Context : Biodiversity erosion is in part due to the massive use of plant protection products. Low doses of pesticides present in the environment can exert sublethal effects and profoundly modify the physiology, reproduction and survival of non-target organisms. The potential effects of environmental contamination on ponds and stagnant water that contain a very diverse and rich community of macro and microorganisms are almost non-existent.

We are involved in a project named AmphiTox in collaboration with the Centre d’Etudes Biologiques de Chizé (CEBC). One AmphiTox aim is to study the effects of environmental doses of glyphosate, a contaminant of agricultural origin, on the development of amphibians raised under controlled laboratory conditions. The amphibian species chosen is the spiny toad (Bufo spinosus) whose populations persist in heavily agricultural environments and which are confronted with environmental contamination.

Glyphosate is a specific inhibitor of the EPSP synthase, an enzyme part of the shikimate metabolic pathway found in archaea, bacteria, apicomplexa, algae, fungi and plants. This pathway is involved in the biosynthesis of aromatic amino acids and the vitamins folic acid and menaquinone. Thus, glyphosate inhibits the growth of plants and of various organisms including bacteria and various microorganisms.

Chronic exposure to low doses of glyphosate can lead to the appearance of resistance mechanisms in bacteria and modify their metabolism, and indirectly lead to their resistance to natural antibiotic agents. In turn, these changes can lead to dysbiosis in their hosts. Indeed different studies have shown that glyphosate exposure changes the gut microbial communities in various animals (daphnia, crabs, honey bees, chicken, cow, rat).

Objectives : The objective of this internship is to analyze the microbiota of tadpoles taken at different stages of their development and submitted to a chronic exposition of glyphosate and compare it to control tadpoles of the same age. The symbiotic microorganisms of the intestinal microbiome hosted by amphibians are important players in their immunity and their nutritional physiology. Several studies have shown that these amphibian microbiomes originate from the aquatic and terrestrial environments they frequent.

This internship project will aim  to compare the composition of bacteria, archaea, protists and fungal communities using a metatranscriptomic and metabarcoding strategy involving 16S, ITS and 18S rRNA gene sequencing using a long-read nanopore sequencing approach. This will require the use of a combination of wet lab and bioinformatic experiments and analysis. A pure bioinformatic and data analysis internship is also possible.

By measuring the impact of glyphosate and describing the sublethal effects of poorly studied molecules on amphibians, we will gain a better understanding on how these pesticides disrupt physiological mechanisms. This is crucial to predict the responses of organisms to current environmental upheavals.

References :

Ackermann W, Coenen M, Schrödl W, Shehata AA, Krüger M. 2015. The influence of glyphosate on the microbiota and production of botulinum neurotoxin during ruminal fermentation. Current Microbiology 70:374–382. DOI: 10.1007/s00284-014-0732-3.

Chiang Y-R, Wei ST-S, Wang P-H, Wu P-H, Yu C-P. 2020. Microbial degradation of steroid sex hormones: implications for environmental and ecological studies. Microbial Biotechnology 13:926–949. DOI: https://doi.org/10.1111/1751-7915.13504.

Colombo BM, Scalvenzi T, Benlamara S, Pollet N. 2015. Microbiota and mucosal immunity in amphibians. Frontiers in Immunology 6:111. DOI: 10.3389/fimmu.2015.00111.

Hertel R, Gibhardt J, Martienssen M, Kuhn R, Commichau FM. Molecular mechanisms underlying glyphosate resistance in bacteria. Environmental Microbiology n/a. DOI: https://doi.org/10.1111/1462-2920.15534.

Holert J, Cardenas E, Bergstrand LH, Zaikova E, Hahn AS, Hallam SJ, Mohn WW. 2018. Metagenomes Reveal Global Distribution of Bacterial Steroid Catabolism in Natural, Engineered, and Host Environments. mBio 9. DOI: 10.1128/mBio.02345-17.

Scalvenzi T, Clavereau I, Bourge M, Pollet N. 2021. Gut microbial ecology of Xenopus tadpoles across life stages. bioRxiv:2020.05.25.110734. DOI: 10.1101/2020.05.25.110734.

Shehata AA, Schrödl W, Aldin AA, Hafez HM, Krüger M. 2013. The effect of glyphosate on potential pathogens and beneficial members of poultry microbiota in vitro. Current Microbiology 66:350–358. DOI: 10.1007/s00284-012-0277-2.

Suppa A, Kvist J, Li X, Dhandapani V, Almulla H, Tian AY, Kissane S, Zhou J, Perotti A, Mangelson H, Langford K, Rossi V, Brown JB, Orsini L. 2020. Roundup causes embryonic development failure and alters metabolic pathways and gut microbiota functionality in non-target species. Microbiome 8:170. DOI: 10.1186/s40168-020-00943-5.

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