Rootstocks affect the vulnerability to embolism and pit membrane thickness in Citrus scions.

Embolism resistance of xylem tissue varies among species and is an important trait related to drought resistance, with anatomical attributes like pit membrane thickness playing an important role in avoiding embolism spread. Grafted Citrus trees are commonly grown in orchards, with the rootstock being able to affect the drought resistance of the whole plant. Here, we evaluated how rootstocks affect the vulnerability to embolism resistance of the scion using several rootstock/scion combinations. Scions of 'Tahiti' acid lime, 'Hamlin', 'Pera' and 'Valencia' oranges grafted on a 'Rangpur' lime rootstock exhibit similar vulnerability to embolism. In field-grown trees, measurements of leaf water potential did not suggest significant embolism formation during the dry season, while stomata of Citrus trees presented an isohydric response to declining water availability. When 'Valencia' orange scions were grafted on 'Rangpur' lime, 'IAC 1710' citrandarin, 'Sunki Tropical' mandarin or 'Swingle' citrumelo rootstocks, variation in intervessel pit membrane thickness of the scion was found. The 'Rangpur' lime rootstock, which is known for its drought resistance, induced thicker pit membranes in the scion, resulting in higher embolism resistance than the other rootstocks. Similarly, the rootstock 'IAC 1710' citrandarin generated increased embolism resistance of the scion, which is highly relevant for citriculture.

Angiosperms follow a convex trade-off to optimize hydraulic safety and efficiency.

Intervessel pits are considered to function as valves that avoid embolism spreading and optimize efficient transport of xylem sap across neighbouring vessels. Hydraulic transport between vessels would therefore follow a safety-efficiency trade-off, which is directly related to the total intervessel pit area (Ap ), inversely related to the pit membrane thickness (TPM ) and driven by a pressure difference. To test this hypothesis, we modelled the relative transport rate of gas (ka ) and water (Q) at the intervessel pit level for 23 angiosperm species and correlated these parameters with the water potential at which 50% of embolism occurs (Ψ50 ). We also measured ka for 10 species using pneumatic measurements. The pressure difference across adjacent vessels and estimated values of ka and Q were related to Ψ50 , following a convex safety-efficiency trade-off based on modelled and experimental data. Minor changes in TPM and Ap exponentially affected the pressure difference and flow, respectively. Our results provide clear evidence that a xylem safety-efficiency trade-off is not linear, but convex due to flow across intervessel pit membranes, which represent mesoporous media within microporous conduits. Moreover, the convex nature of long-distance xylem transport may contribute to an adjustable fluid balance of plants, depending on environmental conditions.

Water stress signaling and hydraulic traits in three congeneric citrus species under water deficit.

Morpho-physiological strategies to deal with water deficit vary among citrus species and the chemical signaling through ABA and anatomical, hydraulic, and physiological traits were evaluated in saplings of Rangpur lime, Swingle citrumelo and Valencia sweet orange. Trunk and roots of Swingle citrumelo presented lower vessel diameter and higher vessel frequency as compared to the other species. However, relative water content at the turgor loss point (RWCTLP), the osmotic potential at full turgor (Ψ0), the osmotic potential at the turgor loss point (ΨTLP), bulk modulus of elasticity (ε) and the xylem water potential when hydraulic conductivity is reduced by 50% (Ψ50) and 88% (Ψ88) indicated similar hydraulic traits among citrus species, with Rangpur lime showing the highest hydraulic safety margin. Roots of Rangpur lime and Swingle citrumelo were more water conductive than ones of Valencia sweet orange, which was linked to higher stomatal conductance. Chemical signaling through ABA prevented shoot dehydration in Rangpur lime under water deficit, with this species showing a more conservative stomatal behavior, sensing, and responding rapidly to low soil moisture. Taken together, our results suggest that Rangpur lime - the drought tolerant species - has an improved control of leaf water status due to chemical signaling and effective stomatal regulation for reducing water loss as well as decreased root hydraulic conductivity for saving water resources under limiting conditions.

A User Manual to Measure Gas Diffusion Kinetics in Plants: Pneumatron Construction, Operation, and Data Analysis.

The Pneumatron device measures gas diffusion kinetics in the xylem of plants. The device provides an easy, low-cost, and powerful tool for research on plant water relations and gas exchange. Here, we describe in detail how to construct and operate this device to estimate embolism resistance of angiosperm xylem, and how to analyse pneumatic data. Simple and more elaborated ways of constructing a Pneumatron are shown, either using wires, a breadboard, or a printed circuit board. The instrument is based on an open-source hardware and software system, which allows users to operate it in an automated or semi-automated way. A step-by-step manual and a troubleshooting section are provided. An excel spreadsheet and an R-script are also presented for fast and easy data analysis. This manual aims at helping users to avoid common mistakes, such as unstable measurements of the minimum and maximum amount of gas discharged from xylem tissue, which has major consequences for estimating embolism resistance. Major advantages of the Pneumatron device include its automated and accurate measurements of gas diffusion rates, including highly precise measurements of the gas volume in intact, embolised conduits. It is currently unclear if the method can also be applied to woody monocots, gymnosperm species that possess torus-margo pit membranes, or to herbaceous species.

Sink strength of citrus rootstocks under water deficit.

Carbon allocation between source and sink organs determines plant growth and is influenced by environmental conditions. Under water deficit (WD), plant growth is inhibited before photosynthesis and shoot growth tends to be more sensitive than root growth. However, the modulation of the source-sink relationship by rootstocks remains unsolved in citrus trees under WD. Citrus plants grafted on Rangpur lime are drought tolerant, which may be related to a fine coordination of the source-sink relationship for maintaining root growth. Here, we followed 13C allocation and evaluated physiological responses and growth of Valencia orange trees grafted on three citrus rootstocks (Rangpur lime, Swingle citrumelo and Sunki mandarin) under WD. As compared with plants on Swingle and Sunki rootstocks, ones grafted on Rangpur lime showed higher stomatal sensitivity to the initial variation of water availability and less accumulation of non-structural carbohydrates in roots under WD. High 13C allocation found in Rangpur lime roots indicates this rootstock has high sink demand associated with high root growth under WD. Our data suggest that Rangpur lime rootstock used photoassimilates as sources of energy and carbon skeletons for growing under drought, which is likely related to increases in root respiration. Taken together, our data revealed that carbon supply by leaves and delivery to roots are critical for maintaining root growth and improving drought tolerance, with citrus rootstocks showing differential sink strength under WD.

The Pneumatron: An automated pneumatic apparatus for estimating xylem vulnerability to embolism at high temporal resolution.

Xylem vulnerability to embolism represents an important trait to determine species distribution patterns and drought resistance. However, estimating embolism resistance frequently requires time-consuming and ambiguous hydraulic lab measurements. Based on a recently developed pneumatic method, we present and test the "Pneumatron", a device that generates high time-resolution and fully automated vulnerability curves. Embolism resistance is estimated by applying a partial vacuum to extract air from an excised xylem sample, while monitoring the pressure change over time. Although the amount of gas extracted is strongly correlated with the percentage loss of xylem conductivity, validation of the Pneumatron was performed by comparison with the optical method for Eucalyptus camaldulensis leaves. The Pneumatron improved the precision of the pneumatic method considerably, facilitating the detection of small differences in the (percentage of air discharged [PAD] < 0.47%). Hence, the Pneumatron can directly measure the 50% PAD without any fitting of vulnerability curves. PAD and embolism frequency based on the optical method were strongly correlated (r2 = 0.93) for E. camaldulensis. By providing an open source platform, the Pneumatron represents an easy, low-cost, and powerful tool for field measurements, which can significantly improve our understanding of plant-water relations and the mechanisms behind embolism.

Exogenous nitric oxide improves sugarcane growth and photosynthesis under water deficit.

MAIN CONCLUSION: Nitric oxide (NO)-mediated redox signaling plays a role in alleviating the negative impact of water stress in sugarcane plants by improving root growth and photosynthesis. Drought is an environmental limitation affecting sugarcane growth and yield. The redox-active molecule nitric oxide (NO) is known to modulate plant responses to stressful conditions. NO may react with glutathione (GSH) to form S-nitrosoglutathione (GSNO), which is considered the main reservoir of NO in cells. Here, we investigate the role of NO in alleviating the effects of water deficit on growth and photosynthesis of sugarcane plants. Well-hydrated plants were compared to plants under drought and sprayed with mock (water) or GSNO at concentrations ranging from 10 to 1000 µM. Leaf GSNO sprayed plants showed significant improvement of relative water content and leaf and root dry matter under drought compared to mock-sprayed plants. Additionally, plants sprayed with GSNO (≥ 100 µM) showed higher leaf gas exchange and photochemical activity as compared to mock-sprayed plants under water deficit and after rehydration. Surprisingly, a raise in the total S-nitrosothiols content was observed in leaves sprayed with GSH or GSNO, suggesting a long-term role of NO-mediated responses to water deficit. Experiments with leaf discs fumigated with NO gas also suggested a role of NO in drought tolerance of sugarcane plants. Overall, our data indicate that the NO-mediated redox signaling plays a role in alleviating the negative effects of water stress in sugarcane plants by protecting the photosynthetic apparatus and improving shoot and root growth.