In situ ESEM using 3-D printed and adapted accessories to observe living plantlets and their interaction with enzyme and fungus

Romain Roulard (1, 2) , Michel Trentin , Valerie Lefebvre (1, 3) , Francoise Fournet (1, 3) , Ludivine Hocq (1, 3) , Jerome Pelloux (1, 3) , Eric Husson (4) , Christophe Pineau (5) , Loic Dupont (6, 2) , Arash Jamali (2)

1 BioEcoAgro BIOPI-UPJV
2 PME - Plateforme Microscopie Électronique - UPJV
3 BioEcoAgro - Equipe 1 - Functioning and adaptation of the plant in interaction with its environment
4 GEC UPJV - Génie Enzymatique et Cellulaire. Reconnaissance Moléculaire et Catalyse - UMR CNRS 7025
5 Linéa Semences de Lin
6 LRCS - Laboratoire réactivité et chimie des solides - UMR CNRS 7314

This paper describes an innovative way of using environmental scanning electron microscopy (ESEM) and the development of a suitable accessory to perform in situ observation of living seedlings in the ESEM. We provide details on fabrication of an accessory that proved to be essential for such experiments but inexpensive and easy to build in the laboratory, and present our in situ observations of the tissue and cell surfaces. Sample-specific configurations and optimized tuning of the ESEM were defined to maintain Arabidopsis and flax seedlings viable throughout repetitive exposure to the imaging conditions in the microscope chamber. This method permitted us to identify cells and tissues of the live plantlets and characterize their surface morphology during their early stage of growth and development. We could extend the application of this technique, to visualize the response of living cells and tissues to exogenous enzymatic treatments with polygalacturonase in Arabidopsis, and their interaction with hyphae of the wilt fungus Verticillium dahliae during artificial infection in flax plantlets. Our results provide an incentive to the use of the ESEM for in situ studies in plant science and a guide for researchers to optimize their electron microscopy observation in the relevant fields.

Domaines: Sciences du Vivant [q-bio]

Romain Roulard, Michel Trentin, Valerie Lefebvre, Francoise Fournet, Ludivine Hocq, et al.. In situ ESEM using 3-D printed and adapted accessories to observe living plantlets and their interaction with enzyme and fungus. Micron, 2022, 153, ⟨10.1016/j.micron.2021.103185⟩. ⟨hal-03610097⟩