Evaluation of the open-vessel artefact
As embolism cannot, theoretically, be induced via air seeding by exposing the xylem to less negative water potentials than the min- imum water potential previously experienced, the open-vessel artefact was evaluated by spinning xylem samples at a velocity that corresponded to a less negative pressure than the pressure reached by stem branches during a previous drought treatment. Two different rotor designs, the original rotor design (Alder et al., 1997) and the Cavitron (Cochard et al., 2005), were used to check whether the type of rotor could have an effect on the results. Several branches (≥ 2.0 m in length) of B. pendula and Q. palustris were collected in the early morning and transported to the laboratory, where they were allowed to dehydrate progres- sively (up to 5 h, depending on the species). For O. europaea (var. arbequina for the Cavitron design and manzanilla for the original design), saplings were exposed to increasing water stress by with- holding irrigation. The xylempressure of each branch and sapling was monitored regularly by measuring the leaf water potential with a Scholander-type pressure chamber (DGMeca, Gradignan, France) on two leaves per branch or sapling that had been previously bagged up (at least 1 h before taking the water potential measurements). Once the xylempressure reached values of 1.8 MPa or lower, a xylem sample was immediately collected for K s measurements. Samples were collected according to the
observe xylem reﬁlling. In this study, even xylem positive pres- sure did not lead successfully to xylem reﬁlling in all cases. Xylem pressures of 0.02 to 0.05 MPa magnitude were observed, which should correspond to a 2- to 5-m- high water column, while the apical portion remained at a slightly negative potential (20.02 to 20.1 MPa) without reﬁlling observed at the apex (Fig. 7). Xylempressure may have been dissipated along the plant stems and/or gas did not dissolve into xylem sap, delaying the occurrence of positive pressure at higher parts. Although xylem reﬁlling was not observed at the apex during our exper- iment, it may have occurred after a longer period. How- ever, the occurrence of negative water potential after more than 3 d without active transpiration suggests that this phenomenon is not routine for V. vinifera. It is important to consider that only bulk xylem pressures were assessed in this study. There is a possibility that pressure gradients are not homogenous across a portion of the stem or even between vessels that lie in close proximity to each other. Currently, experimental approaches do not exist for assessing in situ pressures at this scale, but this difﬁculty needs to be acknowledged. Given that reﬁlling is a phe- nomenon occurring at the level of an individual vessel, one would expect that it would be the local pressure
coordination between the water potential causing stomatal closure and the xylempressure that provokes cavitation, i.e. 35
xylem vulnerability to cavitation (VC) (Martin-St Paul et al. 2017). This VC is considered as a critical trait for drought 36
Vulnerability to drought-induced embolism was determined at the Caviplace (University of Bordeaux, Talence, France; http://sylvain-delzon.com/caviplace ) with the Cavitron technique (Cochard 2002; Cochard 2005). To prevent artefactual losses in hydraulic conductance due to the presence of open vessels in the samples (Pivovaroff et al., 2016; Torres-Ruiz et al 2017a), branches were recut under water to a 1m-long length, debarked at both ends and then installed in a large cavitron equipped with a 1m-diameter custom-built honeycomb rotor (DGMeca, Gradignan, France). Several samples per species were used to test the presence of open vessels by air injection at 2 bars and none of the studied species presented open vessels in 1 m long branches. Centrifugal force was used to establish negative pressure in the xylem (P i ) and to provoke water stress-induced cavitation. Samples were spun for two minutes at a given speed to decrease the xylempressure progressively at its center from -0.8 MPa to -10.5 MPa (those pressures correspond to centrifugation rotation from 764 rpm to 2768 rpm respectively). Samples were three to five years old with a diameter varying from 10.3 to 12.2 mm. As this technique enables measuring the hydraulic conductance of the samples under negative pressure, the vulnerability curves were generated by plotting the percentage loss of hydraulic conductivity (PLC) against the different target pressures applied. For each branch,
During the last two decades, research on xylem embolism has considerably improved our understanding of tree responses to drought. In trees, water is transported under tension (negative pressure) in the xylem vessels. Under drought conditions, the tension in the xylem conduits increases and cavitation can occur. Cavitation causes an embolism, which restricts the water supply to leaves and other organs. It occurs only when the xylempressure drops below a specific threshold value (ψ cav ) related to pit wall porosity (Tyree and Sperry 1988). Xylem vulnerability to cavitation is correlated with drought tolerance (Maherali et al. 2004, Pockman and Sperry 2000, Tyree et al. 2003), more xerophilous species being more resistant to xylem cavitation. This suggests that species able to keep xylem vessels func- tional under extreme droughts can more readily survive such events. Vulnerability to xylem cavitation has also been found to vary substantially within species (Lopez et al. 2005). For some species, these variations have been associated with a genotypic variability (Cochard et al. 2007, Ennajeh et al. 2008). For others, xylem vul- nerability to cavitation has also been shown to acclimate to environmental conditions such as irradiance (Barigah et al. 2006, Cochard et al. 1999) or soil nutrient avail- ability (Harvey and Van Den Driessche 1999). However, ability to acclimate to drier soil conditions remains largely unexplored and has been restricted to drought- tolerant species (Beikircher and Mayr 2009, Ladjal et al. 2005). Only a slight increase in xylem vulnerability to cavitation was observed in response to drought for a drought-sensitive willow cultivar (Wikberg and ¨ Ogren 2007). Responses to drought are species-specific and depend on the plant’s hydraulic strategy (Br´eda et al. 2006). For example, only one of three Mediterranean cedar species tested showed an acclimation of its vul- nerability to cavitation to drier soil conditions (Ladjal et al. 2005).
In our study, the increased embolism resistance observed with increasing aridity could be explained by changes in pit membrane properties. Whereas several studies found that pit anatomical traits such as the torus overlap explains variability in conifers P 50 between and within conifer species ( Delzon et al. 2010 , Bouche et al. 2014 ), measurements that link the interves- sel pit membrane thickness with P 50 remain scarce in angios- perms. However, recent studies showed that pit membrane thickness is potentially the hydraulically most relevant anatomical feature that explains variation in embolism resistance across woody angiosperms ( Jansen et al. 2009 , Li et al. 2016 ). This pit anatomical feature and its variability need to be assessed at the Figure 4. Mean P 50 (in MPa, xylempressure inducing a 50% loss of hydraulic conductance; top panels) and P 88 (in MPa, xylempressure inducing an 88% loss of hydraulic conductance; bottom panels) values for the various European beech populations studied, according to latitude, mean annual air temperature (MAT) and aridity index (AI). Black lines correspond to linear regressions. Solid grey and dashed grey lines indicate 95% conﬁdence and prediction intervals, respectively. The error bars represent the standard errors.
The similarities between instruments used to measure water tension in soils and plants suggest that High- Capacity Tensiometers, which are widely used to monitor (negative) pore-water pressure in soils, could also be used to measure xylem water pressure in plants. Indeed,  have successfully attempted to measure xylem water pressure directly using a pressure probe made of a capillary tube filled with water and silicon oil. When the tip of the probe was inserted into a xylem channel, the xylem water pressure was transmitted through the liquid in the probe and measured by a pressure transducer. However, this pressure probe has never registered xylem water pressures below -0.65 MPa  and for no more than a few hours due to cavitation occurring in the probe . This was probably due to the absence of a high-air-entry porous interface, which is actually incorporated into high-capacity tensiometer used to measure pore-water tension in soil . This paper presents the use of High-Capacity Tensiometer (HCT) to monitor the negative xylem water pressure in plants. The measurement of the HCT was validated against the measurement of leaf water potential via the Pressure Chamber and the Transistor Psychrometer over a relatively wide range of xylem water pressures via field and laboratory experiments . However, this paper only focuses on one single experiment carried out on a cherry tree in the laboratory.
This study investigates the effects of a pressure gradient on the wall pressure beneath equilibrium turbulent boundary layers. Excitation of the walls of a vehicle by turbulent boundary layers indeed constitutes a major source of interior noise and it is necessary to take into account the presence of a pressure gradient to represent the effect of the curvature of the walls. With this aim, large-eddy simulations of turbulent boundary layers in the presence of both mild adverse and mild favourable pressure gradients are carried out by solving the compressible Navier–Stokes equations. This method provides both the aeroacoustic contribution and the hydrodynamic wall-pressure fluctuations. A critical comparison with existing databases, including recent measurements, is conducted to assess the influence of a free stream pressure gradient. The analyses of wall-pressure spectral densities show an increase in the low-frequency content from adverse to favourable conditions, yielding higher integrated levels of pressure fluctuations scaled by the wall shear stress. This is accompanied by a steeper decay rate in the medium-frequency portion for adverse pressure gradients. No significant difference is found for the mean convection velocity. Frequency–wavenumber spectra including the subconvective region are presented for the first time in the presence of a pressure gradient. A scaling law for the convective ridge is proposed, and the acoustic domain is captured by the simulations. Direct acoustic emissions have similar features in all gradient cases, even if slightly higher levels are noted for boundary layers subjected to an adverse gradient.
Compared to mature trees, the unfolding of spring leaves started 10–40 d earlier in seedlings ( Vitasse, 2013 ). Vieira et al. (2013) investigated trees of the same age and size under identical localized conditions. Their results demonstrated that the timings of xylem phenology changed with cell production, but not with age or size per se. Rathgeber et al. (2011) found that trees of different social status and sizes, growing in even- aged stands, exhibited different xylem phenology. Compared to intermediate and suppressed trees, the cambial reactivation of dominant trees exhibited an earlier onset and a later termination and, therefore, a longer duration of xylem growth. De Luis et al. (2009) investigated an even-aged conifer plantation (Pinus halepenisi and Pinus pinea) under semi-arid Mediterranean climate conditions. The faster-growing pines were able to resume cambial activity in response to early autumn precipitation ( De Luis et al., 2007 ); thus, they speculated that both size and growth rate affected their sensitivity to rain events. Some studies applied grafting techniques to separate the effect of size from the effect of age ( Mencuccini et al., 2005, 2007; Abdul- Hamid and Mencuccini, 2009 ). According to this method, it has been suggested that photosynthesis and tree growth decline with increasing tree size ( Abdul-Hamid and Mencuccini, 2009 ). Similarly, Mencuccini et al. (2005) concluded that tree size, not age, mediated changes in growth, carbon assimilation, and leaf biochemistry. Based on available literature, it seems that changes in phenology and the growth-climate relationship may mostly be associated with the size rather than the age of trees.
Figure 4-5 Graph showing the particle size distribution for the different feed solutions to compare directionality
The results from these tests suggest that the permeability of the xylem may depend weakly on the direction that the feed solution is flown through it. Excluding the final data point for each test (under the assumption that an anomaly occurred and the membrane may have ruptured), both samples showed higher flow rates at all points along the experiment in the reverse configuration. Additionally, the standard sample had a student t-test value of 0.026 for these results, which is below the threshold value of 0.05 for it to be statistically significant. Additionally, the rejection rate of the samples was higher when they were oriented in the reverse configuration. For the xylem to be an effective filter, we want both a high permeability and high rejection rate, to produce the maximum amount of clean and potable water. These results suggest that, for final implementation, the xylem should be loaded into the hosing in the reverse orientation. Finally, it appears that fouling is a viable method by which, the cutoff particle size for the xylem can be decreased, allowing the membranes to remove more particles from the feed solution.
Materials and methods Plant material
The perfusion experiments were carried out on stems from an old beech tree (Fagus sylvatica) from Allagnat forest in central France (45 ◦ 45 23 N, 2 ◦ 56 26 E, 1000 m a.s.l.) and on stems from an adult poplar tree (Populus tremula) from Bort-l’Etang in central France (45 ◦ 47 02 N, 3 ◦ 25 43 E, 333 m a.s.l.). Stems were sampled on the same tree to avoid intraspecific variation. Branches were harvested from October to December, i.e. after the vegetative season and before the temperature decrease drastically below 0 ◦ C. We analyzed sunlit shoots of comparable age and growth. Selected stems were 0.5–1 cm in diameter and 0.5 m long. The freshly sampled stems were sealed in airtight black plastic bags to reduce water loss through transpiration and taken straight to the lab to measure xylem hydraulic conductance on the same day or the day after. Samples taken to study VC were wrapped in moist paper, bagged and stored at 4 ◦ C until analysis.
conduits near the pith (Cochard & Tyree, 1990; Brodersen et al., 2013). Collapse of leaf vein xylem conduits has been reported in some (Cochard et al., 2004a) but not all gymnosperms (Brodribb & Holbrook, 2005; Zhang et al., 2014), but has yet to be observed in angiosperms.
According to the hydraulic vulnerability segmentation hypothesis (Tyree & Ewers, 1991), the most distal parts of the xylem pathways should be more vulnerable to hydraulic failure than basal portions. This would allow distal portions to buffer more basal parts from cavitation events, as these supply water to the entire lamina and are therefore less expendable. This hypothesis has been supported by the finding that leaves are more vulnerable to hydraulic decline than stems (Bucci et al., 2012; Pivovaroff et al., 2014). If this hypothesis applies within leaves, the minor veins should be more vulnerable to embolism than the petiole and lower-order veins. Greater susceptibility of higher-order veins would also be consistent with these experiencing the strongest tensions in the plant xylem system. The relative vulnerability of leaf vein orders to embolism formation will influence their contribution to the leaf xylem hydraulic decline. Models and recent imaging techniques have suggested that embolism in the midrib would have a stronger impact on hydraulic conductance than in the smaller diameter minor veins, given the high density of minor veins (McKown et al., 2010; Brodribb et al., 2016a).
In this work, more than 75% of the total starch measured in cambial region and wood 250
ray parenchyma was detected during dormancy (DOY 33) and after the end of xylem differentiation (DOY 236). These results support the hypothesis that starch is directly used in differentiating tissues during wood formation as source of glucose for cellulose biosynthesis. Elle and Sauter (2000) found that the increased endoamylase activity, related with the starch degradation, leads to higher concentrations of malto-
The Suppression of Dwarﬁsm Is Associated with the Recovery of Xylem Morphology
esk1-5 dwarﬁsm is associated with an impairment of water transport due to the deformation of xylem vessels (Lefebvre et al., 2011). Observation with a light micro- scope of Toluidine Blue Orange-stained cross sections of 10-week-old, fully developed stem bases showed a clear suppression of the irx phenotype in beem396b (Fig. 5). Based on the measurement of 10 to 20 xylem vessel surface areas in 19 to 21 different poles from six different plants (2,299 data points; Supplemental Fig. S8), we observed a bimodal distribution (Supplemental Fig. S9). Consequently, the data were analyzed for two popula- tions: those with large vessels and those with medium- sized vessels. Overall, we observed a pole effect in individual plants and a plant effect in genotypes. Be- cause vessel area was heteroscedastic, we compared vessel area between genotypes with nonparametric tests (permutation-based pairwise Student’s t tests). beem396b presents wild-type-like round-shaped vessels of area intermediate between esk1-5 and the wild type in the large vessel as well as the medium vessel populations (Fig. 5; Supplemental Fig. S9). The double mutants esk1-5 kak-7 and esk1-5 kak-8 also showed suppression of the irx phenotype (Fig. 5), with round-shaped vessels showing an area similar to the wild type, in the large vessel as well as the medium vessel populations (Fig. 5; Supplemental Fig. S9). For kak-7 and kak-8 mutants, the vessel area distribution was large, heteroscedastic, and not signiﬁcantly different from the wild type, but there was a trend toward higher vessel area, as observed with the wide range of values from the median to the third quartile. These results suggest that rosette enlargement in beem396b and in esk1-5 kak double mutants is associ- ated with the recovery of xylem function. However, the proportion of xylem relative to phloem in the vascular pole area was not signiﬁcantly different within geno- types (Fig. 5). This recovery of xylem morphology can be viewed as an increase in water transport capacity, which may account for the suppression of dwarﬁsm in the beem line.
Moreover, the number of people dependent upon the borrower increases the likelihood of transition from the status of a poor borrower to that of a good one, from the second loan onwards. This is an interesting result since it points to the fact that another form of social pressure can complete the guarantor mechanism which only seems efficient for the first loan. In fact, we can imagine that dependents with access to the financing via the borrower (head of the family) may put pressure on him to keep more closely to the repayment schedule. A social pressure mechanism within the family (in the broadest sense of the term) would then complete the pressure exerted by guarantors for a first loan. However, there is an alternative explanation which, to some extent, is the opposite of the one just proposed. In fact, it would rather consist of social pressure exerted by the head of the family over his dependents to force them to participate in the microcredit repayment schedule. We would then find some of the drift brought to light in the group lending system where the head of the group can potentially misuse his dominant position to get others to make repayments instead of him.