The optical method based on gas injection overestimates leaf vulnerability to xylem embolism in three woody species

Plant hydraulic traits related to leaf drought tolerance like the water potential at turgor loss point (TLP) and the water potential inducing 50% loss of hydraulic conductance (P50), are extremely useful to predict potential impacts of drought on plants. While novel techniques allowed the inclusion of TLP in studies targeting a large group of species, fast and reliable protocols to measure leaf P50 are still lacking. Recently, the optical method coupled with the gas-injection (GI) technique has been proposed as a possibility to speed up P50 estimation. Here, we present a comparison of leaf optical vulnerability curves (OVc) measured in three woody species, namely F Acer campestre (Ac), Ostya carpinifolia (Oc) and Populus nigra (Pn), based on bench dehydration (BD) or gas-injection (GI) of detached branches. For Pn, we also compared optical data with direct micro-CT imaging in both intact saplings and cut shoots subjected to BD. Based on the BD e procedure, Ac, Oc and Pn had P50 values of -2.87, -2.47 and -2.11 MPa, respectively, while the GI procedure overestimated leaf vulnerability (2.68, 2.04 and 1.54 MPa for Ac, Oc and Pn, respectively). The overestimation was higher for Oc and Pn than for Ac, likely reflecting the species-specific e vessel lengths. According to micro-CT observations performed on Pn, the leaf midrib showed none or very few embolized conduits at -1.2 MPa, consistent with the OVc obtained with the BD procedure but at odds with that derived on the basis of GI. Overall, our data suggest that coupling the optical method with GI might not be a reliable technique to quantify leaf hydraulic vulnerability, since it could be affected by the ‘open-vessel’ artefact. Accurate detection of xylem embolism in the leaf vein network should be based on BD, preferably of intact up-rooted plants.