Phenotyping xylem connections in grafted plants using X-ray micro-computed tomography

Plants are able to naturally graft or inosculate their trunks, branches and roots together, this mechanism is used by humans to graft together different genotypes for a range of purposes. Grafts are considered successful if functional vascular connections between the two genotypes occur. Various techniques can evaluate xylem connections across the graft interface. However, these methods are generally unable to assess the heterogeneity and three-dimensional (3D) structure of xylem vessel connections. Here we present the use of X-ray micro-computed tomography to characterize the 3D morphology of grafts of grapevine. We show that xylem vessels form between the two plants of natural root and human-made stem grafts. The main novelty of this methodology is that we were able to visualize the 3D network of functional xylem vessels connecting the scion and rootstock in human-made stem grafts thanks to the addition of a contrast agent to the roots and improved image analysis pipelines. In addition, we reveal the presence of extensive diagonal xylem connections between the main axial xylem vessels in 2-year old grapevine stems. In conclusion, we present a method that has the potential to provide new insights into the structure and function of xylem vessels in large tissue samples.

This workwas funded with the project EFA 324/19 - VITES QUALITAS, 65% co-funded by the European Regional Development Fund (FEDER)through the Interreg Programme V-A Spain-France-Andorra (POC-TEFA 2014-2020) and VitiGraft funded by the Plant2Pro ® CarnotInstitute in the frame of its 2021 call, Plant2Pro® is supportedby ANR (agreement #21-CARN-024-01 - 2021). This study alsoreceived financial support from the French government in theframework of the University of Bordeaux's France 2030 pro-gramme/GPR Bordeaux Plant Sciences. Micro-CT acquisitionswere performed in Montpellier, France, at the MRI platform memberof the national infrastructure France-BioImaging and supported bythe French National Research Agency (ANR-10-INBS-04, « Invest-ments for the Future »), and by the Labex CEMEB (ANR-10-LABX-0004) and NUMEV (ANR-10-LABX-0020).