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Article

Stem Photosynthesis Affects Hydraulic Resilience in the Deciduous Populusalba but Not in the Evergreen Laurus nobilis

by
Patrizia Trifilò
1,*,
Sara Natale
2,
Sara Gargiulo
3,
Elisa Abate
1,
Valentino Casolo
3 and
Andrea Nardini
2
1
Dipartimento di Scienze Chimiche, Biologiche, Farmaceutiche ed Ambientali, Università di Messina, Salita Stagno D, Alcontres, 31, 98166 Messina, Italy
2
Dipartimento di Scienze della Vita, Università di Trieste, Via L. Giorgieri, 10, 34127 Trieste, Italy
3
Dipartimento di Scienze AgroAlimentari, Ambientali e Animali, Università di Udine, Via delle Scienze, 91, 33100 Udine, Italy
*
Author to whom correspondence should be addressed.
Water 2021, 13(20), 2911; https://doi.org/10.3390/w13202911
Submission received: 27 September 2021 / Revised: 8 October 2021 / Accepted: 14 October 2021 / Published: 16 October 2021
(This article belongs to the Special Issue Nonstructural Carbohydrates, Water Status, and Hydraulic Dynamics in Plants)
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Abstract

:
Stem photosynthesis has been suggested to play relevant roles to cope with different biotic and abiotic stress factors, including drought. In the present study, we performed measurements of stem hydraulic conductance and non-structural carbohydrate content in the evergreen Laurus nobilis L. and the deciduous Populus alba L., subjected to inhibition of stem photosynthesis and successive exposure to a drought-recovery cycle in order to check if stem photosynthesis may be involved in allowing hydraulic recovery after drought stress relief. Stem shading affected the growth of L. nobilis but not of P. alba saplings. By contrast, inhibition of stem photosynthesis was coupled to inhibition of hydraulic recovery following embolism build-up under drought in P. alba but not in L. nobilis. The two study species showed a different content and behavior of nonstructural carbohydrates (NSCs). The differences in NSCs’ trend and embolism reversal ability led to a significant relationship between starch content and the corresponding hydraulic conductance values in L. nobilis but not in P. alba. Our findings suggest that stem photosynthesis plays a key role in the maintenance of hydraulic functioning during drought especially in the deciduous species. This, in turn, may increase their vulnerability under current global climate change scenarios.

1. Introduction

Most terrestrial plants perform photosynthesis at the leaf level, but some of them can only use stems to this purpose, as leaves have been eliminated or transformed into thorns or other organs serving different functions in peculiar habitats. However, even several leaf-bearing plants, including both herbaceous and woody species, do assimilate CO2 at stem level, as suggested by the presence of chloroplasts in the bark and even in wood parenchyma and pith [1,2,3,4]. Photosynthetic activity of stems has been reported in many species and families, encompassing their biogeographic and phylogenetic features [5,6,7].
In trees and other woody plants, carbon assimilation is typically higher in current-year stems compared to older ones [8]. The actual contribution of stem photosynthesis to plant carbon balance, compared to carbon uptake at leaf level, remains debated [7,9,10,11,12,13]. Different studies have suggested that carbon assimilation by tree stems can provide different functional advantages for plants. The increased carbon gain would offer additional carbohydrate availability for the production of new leaves, flowers and fruits under both optimal and stressful environmental conditions [3,14,15,16,17,18,19]. In accordance, different studies have shown that stem photosynthesis contributes to stem growth and bud development, especially in young woody plants and in deciduous species [20,21,22,23,24]. Photosynthetic activity of stems also plays a relevant role to cope with defoliation caused by biotic or abiotic agents [25,26,27]. Finally, it contributes to face thermal stress [8,28,29,30,31,32] and drought, as reported in different desert and semi-desert non-succulent species, as well as in Mediterranean plants (i.e., [4,5,8,10,11,12,33,34]).
Research on stem photosynthesis has recently received renewed interest because of its potential role in resistance and resilience of plants to severe global change-type droughts [35]. Hydraulic failure caused by massive xylem embolism is a major and recurrent cause of drought-induced tree mortality [36,37,38], with an additional role suggested for carbon starvation. Interestingly, carbohydrate content decline, even when non-lethal, has been reported to impact plant hydraulic efficiency [39,40,41,42,43]. Non-structural carbohydrates (NSCs) are apparently involved in the maintenance of plant hydraulic function via different processes, which are only partially understood. NSCs are involved in xylogenesis, thus providing plants with the possibility to grow new xylem to increase or maintain their hydraulic capacity, despite possible occurrence of xylem embolism or mechanical damage impairing older conduits [44,45]. Moreover, NSCs are also involved in osmoregulation processes that, at the level of wood parenchyma, might protect cells from damage caused by drought- or freeze-induced protoplast dehydration [46,47,48].
Besides the relatively well-known role of NSCs in xylem growth and symplast osmoregulation, NSCs stored in wood parenchyma have also been reported to be involved in lowering the osmotic potential of xylem sap during and after stressful events. Xylem sap osmoregulation by soluble NSCs might lead to lowering its freezing point, thus limiting formation of ice in xylem conduits during freeze stress [49,50]. Accumulation of soluble NSCs in the xylem sap during and/or after freeze or drought stress might also provide the necessary driving force to refill embolized conduits with water once stress is relieved and the xylem pressure rises to near-atmospheric values [41,42,51,52,53,54,55]. An additional intriguing role of NSCs might be the direct or indirect contribution to the production of surfactants that, by modifying xylem sap surface tension or stabilizing gas nanobubbles, would modify the xylem vulnerability to embolism formation on diurnal or seasonal time scales [55,56,57,58].
Based on the above, stem-level photosynthesis might play a primary role not only in the production of NSCs contributing to the whole plant carbon gain but also in the modulation and regulation of xylem hydraulic function. Indeed, some recent studies have reported experimental evidence for a role of stem photosynthesis in reducing xylem vulnerability to embolism and improving the capacity for xylem embolism reversal [59,60,61,62,63].
In the present study, we report measurements of stem hydraulic conductance in response to drought stress and re-irrigation in plants subjected to inhibition of stem photosynthesis via localized shading. We aimed at testing the hypotheses that the carbon gain obtained by stem photosynthesis is involved in allowing hydraulic recovery after drought stress relief. We focused on two woody species, namely the evergreen Laurus nobilis L. and the deciduous Populus alba L, because, for both of them, there is available experimental evidence suggesting their capacity to reverse xylem embolism following drought stress, with an involvement of NSC stores [55,64].

2. Materials and Methods

2.1. Plant Material and Growth Condition

Measurements were performed during spring and summer 2019 on P. alba and L. nobilis saplings (50 samples per species). In April, one-year-old cuttings of P. alba and L. nobilis growing in 4-liter pots were transferred to a greenhouse at University of Messina, Italy. Samples were randomly divided in two experimental groups: control (C) and shaded (S) samples. In S samples, the main stem and any lateral branch were wrapped with aluminum foil, similarly to the experimental procedure described by Saveyn et al. [20]. This treatment was expected to significantly reduce the presence of differentiated chloroplasts and, in any case, to inhibit stem photosynthesis, without blocking gas exchange between stem tissues and atmosphere [20,59].
In the greenhouse, plants received only natural light, with maximum photosynthetic photon flux density (PPFD) daily values averaging 1050 ± 180 μmol s−1 m−2. Air temperature ranged from 22 ± 2 °C to 31 ± 3 °C (night/day) and the mean value of air humidity was 80 ± 3%. All plants were regularly irrigated to field capacity (FC), every second day from April until early June, when the drought treatment was imposed (Figure 1). The drought treatment started after stem and leaf production and growth had ceased. Water stress was imposed by irrigating plants to 30% FC once per week, for 4 weeks (drought period). This irrigation regime induced a strong reduction of gas exchange and xylem water potential (Ψx), thus inducing substantial loss of stem hydraulic conductivity (see below). After the drought period, C and S samples were further divided in two groups. A first set of plants was used to immediately measure different physiological parameters (see below) at the end of drought. A second group of samples was re-irrigated to field capacity and measured 7 days later.
Moreover, five additional plants per species were maintained fully irrigated during the whole experimental period and grew under the same environmental conditions experienced by C and S samples. These plants were used to collect samples for estimating NSC content (see below) in plants not experiencing drought.

2.2. Morphological Measurements

To estimate the possible effect of stem shading on plant growth, the number (Ncy) and the length (Lcy, as recorded by a ruler) of current-year stems as well as the total number of leaves on current-year branches (NL) were measured weekly during the whole experimental period (i.e., from April to July) in 3 C and 3 S samples per species.
Leaf area (AL) and leaf mass per unit area (LMA) were estimated on 40 leaves randomly collected at the end of hydraulic measurements (see below). AL was recorded by a scanner (HP Scanjet G4050, Palo Alto, CA, USA) and image analysis based on the software ImageJ (http://imagej.nih.gov/ij/, accessed on 8 October 2021). Leaves were oven-dried for 3 days at 70 °C to obtain their dry weight (DW) and the leaf mass per unit area (LMA) was calculated as DW/AL.

2.3. Leaf Water Potential Isotherms

At the beginning of June (i.e., before starting the drought treatment) and in July (i.e., after one month of water stress), leaf water potential isotherms (PV-curves) were performed on 5 C and 5 S leaves per species and per treatment, sampled from different plants. PV-curves were elaborated to obtain the leaf water potential at the turgor loss point (Ψtlp), the osmotic potential at full turgor (π0), and the bulk modulus of elasticity (εmax). In particular, Ψtlp was estimated as the flex point of the relationship between 1/ΨL and water loss, π0 was calculated by the y-intercept of the linear region of this relationship, and εmax was calculated by the ratio between the change of turgor pressure and the relative change of the leaf water content [65].

2.4. Gas Exchange and Water Potential Measurements

In order to check gas exchange rates and water status in C versus S samples under drought and recovery, leaf conductance to water vapor (gL) and photosynthetic rate (An) were measured from the beginning of May (i.e., when fully expanded leaves of poplar samples were observed) to the end of the experiment (Figure 1). gL and An were measured once per week until the beginning of the drought treatment and, afterwards, twice per week. Measurements were performed at midday on at least 4 leaves per group and per treatment, selected from different plants, using a portable LCi Analyzer System (ADC Bioscientific Ltd., Herts, UK). To avoid excessive defoliation, minimum diurnal xylem water potential (Ψx) was measured at midday only the week before starting the drought treatment, and then on the same days of gas exchange measurements. This experimental procedure allowed us to check that the applied water stress led plants to experience Ψx values of about –1.5 MPa (i.e., Ψx inducing significant loss of hydraulic conductivity for both species [66,67]). Ψx was also measured at midday and in the morning (at about 6:00 a.m.) on the same samples used for hydraulic measurements. Measurements of Ψx were performed on 3 leaves from different plants per species and per treatment, using a portable pressure chamber (3005 Plant Water Status Console, Soilmoisture Equipment Corp., Goleta, CA, USA). Leaves were wrapped in cling film and aluminum foil 2–3 h before sampling.

2.5. Hydraulic Conductivity Measurements

Xylem-specific hydraulic conductivity (Kx) was measured in C and S samples after the drought period and upon recovery. Measurements were performed at midday (i.e., 12.00) and on the following morning (i.e., 6:00 a.m.) on about 10 cm-long current-year samples using a hydraulic apparatus [68,69]. Perfusion solution was a commercial mineral water in which were added 15 mM KCl [70]. Samples were perfused at a pressure (P) of 8 kPa. When flow rate (F) became stable, samples were flushed at P = 0.2 MPa for 15 min to remove embolism and F was re-measured at 8 kPa. Kx was computed as (F/P) × (L/Ax), where L is sample length and Ax is the xylem cross-sectional area. The initial branch hydraulic conductivity value (K) and the K value measured after embolism removal (Kmax) were used to estimate the PLC as PLC = 1 − (K/Kmax) × 100.
To avoid possible cutting artifacts [71], 1 h before their collection, branches were girdled for their entire length at about 3 cm intervals, by removing about 3 mm wide bark rings. The exposed wood was immediately covered with a thin layer of silicone grease to avoid desiccation. After girdling, at least six samples per treatment per species were cut under water and maintained with their cut end immersed into the water for 1 h in order to relax xylem tension [72]. Then, the current-year segment was collected and measured. All hydraulic measurements were performed at a temperature of 20 °C.

2.6. NSC Content

At least 5 samples from those used for hydraulic measurements were collected to estimate NSC content (sugars and starch). Moreover, at the end of drought treatment, five additional samples were collected from samples maintained fully irrigated during the whole experimental period and growing under the same environmental conditions experienced by C and S samples. After hydraulic measurements, stem samples were immediately microwaved at 700 W for 3 min to stop enzymatic activities, oven-dried at 70 °C for 24 h, and finally grounded to fine powder (particle size < 0.15 mm). NSC extraction followed the standardized method proposed by Quentin et al. [73] and Landhausser et al. [74], with minor modifications to account for small amount of material. An aliquot of 15 ± 1 mg of dry sample was suspended in 0.5 mL of ethanol 80% (v/v), incubated at 80 °C for 30 min and centrifuged (with Mikro 120, Hettuch zentrifugen, Tuttlingen, Germany) at 14.000 RPM for three minutes. This step was repeated, suspending the pellet with 0.3 mL of ethanol 80%. Once mixed in the same Eppendorf tube, the extracts were dried till complete evaporation. The resulting crystalized carbohydrates were re-suspended with 0.5 mL of 50 mM Tris-HCl pH 7.5 and centrifuged at 14.000 RPM for three minutes. Sugars contained in supernatants were measured with Anthrone assay [75], using spectrophotometer at wavelength of 620 nm. The absorbance values were converted to mg glucose g−1 DW using a calibration curve prepared with known amounts of glucose. Pellets were resuspended in 1 mL of 0.2 M Sodium Acetate Trihydrate pH 4.6 and incubated at 100 °C for 1 h to allow starch gelatinization. The enzymatic hydrolysis—using 100 U of α-amylase and 25 U of amyloglucosidase per sample—was performed at 55 °C. To stop enzymatic activity, samples were boiled for 5 min. Glucose obtained from starch digestion was evaluated as NADPH (absorbance at 340 nm) by action of 0.3 U per sample of hexokinase and 0.5 U per sample of glucose-6-phosphate-deihydrogenase in 0.1 mL of buffer solution (50 mM Tris-HCl, 2 M MgCl2, 50 mM NADP+ and 0.4 M ATP) at 32 °C for 20 min. Starch content (mg g−1 DW) was calculated comparing NADPH obtained with known amounts of commercial amylose from potato (Sigma-Aldrich, Milan, Italy) that followed the same hydrolysis protocol of samples. Spectrophotometer analysis was performed in a VICTOR3 Multilabel Counter Plate Reader (Perkin Elmer, Boston, MA, USA), for both sugars and starch, using 300 µL per sample.

2.7. Statistical Analysis

All statistical analyses were performed with SigmaStat v. 12.0 (SPSS, Inc., Chicago, IL, USA). To test for differences between C and S samples in terms of Ncy, Lcy, AL and LMA for each species, a Student’s t-test (α = 0.05) was performed after checking for normality assumption. Effects of stem shading and drought on NL, gas exchange, water status, PV-curve data, PLC and NSC were assessed by two-way ANOVA. For statistically significant tests (p < 0.05), a Tukey’s post hoc test using Holm-Sidak p-values correction was carried out to perform pairwise comparisons.

3. Results

3.1. Stem Shading Effects on Plant Growth

Plant growth ceased by the end of May, and no further production of branches and leaves was observed until the end of the experiment (July) in both L. nobilis and P. alba. Table 1 reports data of Ncy, Lcy, AL and LMA as recorded at the end of the experimental period. Stem shading did not affect the growth of P. alba. In fact, the number and length of current-year branches were similar in C and S samples (Table 1). Similarly, no differences were recorded in terms of mean AL (~ 17 cm2) and LMA (~42 mg cm−2) in this species. By contrast, S samples of L. nobilis showed significantly lower values of AL compared to C plants (10.6 ± 3 versus 12.2 ± 3.2 cm2, respectively), but similar values of Ncy, Lcy and LMA. The drought treatment affected the number of leaves per plant in P. alba but not in L. nobilis, and NL decreased more in C than in S poplar (Table 2). This decrease was due to leaf shedding in response to water stress. No leaf fall was observed over the same time interval in well-watered poplar samples used for NSC estimation (see above).

3.2. Plant Water Relations

Stem shading did not affect plant water relations of the two study species. In accordance, no differences in Ψtlp, π0 and εmax values were recorded in C versus S samples (Table 3 and Table 4). L. nobilis plants lowered their Ψtlp value by decreasing π0 in response to drought. By contrast, no changes in these parameters were recorded in P. alba.
Midday values of gL, An and Ψx during the experimental period were statistically similar in C versus S samples (Figure 2). In response to drought, reductions of stomatal aperture and photosynthetic rates were recorded. Stomatal closure did not fully prevent the drop of xylem water potential to about −1.3 MPa, that, in turn, induced a significant loss of stem hydraulic conductivity (see below). In response to re-irrigation, gL and An partially recovered but remained below pre-drought values. However, it can be noted that we recorded values only until 7 days after re-irrigation. Therefore, it cannot be excluded that a full recovery of gas exchange may have occurred after a longer time interval.

3.3. Hydraulic Recovery and NSCs Content

Stem shading did not affect the ability to recover xylem hydraulic function upon re-irrigation in the evergreen species, but influenced this process in the deciduous one (Figure 3). In fact, in poplar, a significant interaction between time and treatment (i.e., C versus S) was recorded. By contrast, changes in PLC were affected only by time in laurel samples. In accordance, C and S samples of L. nobilis had PLC of about 40% at midday and about 20% on the next morning (i.e., 6:00 a.m.) while under drought stress, and of about 10% in response to re-irrigation. By contrast, despite a similar value of PLC recorded at midday in C and S samples in P. alba, only controls showed hydraulic recovery after re-irrigation.
Laurel and poplar samples not experiencing drought (i.e., samples collected, at the end of drought treatment, by plants maintained fully irrigated during all experimental period) showed a soluble sugar content of about 14 mg g−1 DW−1, and no differences in midday versus morning values were recorded (Table 5 and Table 6). Starch content was about 20 mg g−1 DW−1 in L. nobilis, but only up to 10 mg g−1 DW−1 in P. alba (Table 5 and Table 6).
C and S samples of both species showed similar trends of soluble NSCs’ content (Figure 4a,b). Higher values of soluble NSCs were recorded at midday compared to morning and no significant differences were recorded in response to re-irrigation compared to values measured in the early morning (i.e., 6:00 am). However, in laurel, a significant interaction between time and treatment (i.e., C versus S) was recorded, due to a higher decrease of soluble NSCs in control in respect to shaded samples. A statistically significant increase of starch content was recorded in C and S stems of laurel in response to re-irrigation respect values recorded at midday. A similar trend was recorded in S samples of P. alba. By contrast, in C poplar stems, a significant decrease in starch content was recorded in response to re-irrigation in respect to value recorded at midday.
The differences in NSC and PLC trends led to a significant relationship between starch content and the corresponding PLC values in L. nobilis but not in P. alba (Figure 5c,d). Moreover, a linear correlation near to be significant (p = 0.06) was also recorded between soluble NSCs and PLC values in laurel samples but not in poplar plants.

4. Discussion

Drought-induced hydraulic failure (i.e., whole blockage of the long-distance water transport system) poses a major threat to plant survival [76,77], but even carbon starvation can represent a significant challenge to plants facing water limitations [43,78]. The maintenance of basal metabolism under drought, when leaf gas exchange is strongly reduced by stomatal closure, relies on the consumption of stored NSCs [39]. However, NSCs might play even more important indirect roles under drought, e.g., being involved in the maintenance of hydraulic conductance [42]. Hence, the eventual extra carbon gain assured by stem photosynthesis under drought may be involved in the modulation of plant hydraulic functions [7,35,62,63]. Our results support the hypothesis that stem-level carbon gain plays a species-dependent role in the post-drought hydraulic recovery [21,35,79]. However, only in the deciduous poplar stem shading did affect the hydraulic recovery ability, while this was not the case for the evergreen laurel.
Inhibition of stem photosynthesis did not produce effects on the growth of poplar saplings. By contrast, smaller leaves were produced by S samples of L. nobilis compared to controls. Moreover, 5 weeks after the end of foliar expansion, lower starch content was recorded in well-watered samples of poplar compared to laurel (i.e., about 10 versus 20 mg g−1 DW−1) but soluble sugars’ content were similar (i.e., about 14 mg g−1 DW−1). Overall, these results strongly suggest that in P. alba, the decrease in stored NSCs, required to maintain similar annual growth in control and stem-shaded samples, affected the residual availability of NSCs to be used for the maintenance of hydraulic function under water stress. Sprouting is mainly supported by stored NSCs produced over the previous growing seasons, especially in deciduous species [80,81,82,83,84]. Woody tissue carbon storage plays a key role in annual growth, as reported in a large number of species [85,86]. However, the impact of the remobilization of stored NSCs on the magnitude of growth is still unclear [87,88,89]. Recently, Klein et al. [84] reported that more than 95% of branch starch content was consumed for production of new leaves in saplings of three deciduous species (including species showing stem photosynthesis), and the recovery of carbon reserves occurred only 2–6 weeks after complete leaf expansion. On this basis, it is reasonable to suppose that, in deciduous species, even when performing stem photosynthesis, annual growth is mainly sustained by already available wood NSCs’ stores. This, in turn, is expected to lead to relevant depletion of NSC reserves that may be recovered over weeks by mature leaf and, at least in some species, by stem photosynthesis. In accordance, in well- watered samples, where no stem photosynthesis inhibition occurred, lower NSCs’ content was recorded in the deciduous compared to evergreen species. Therefore, the inability to recover hydraulic function following embolism build-up under drought as recorded in S poplar samples was likely the consequence of inhibition of stem photosynthesis coupled to lower NSC reserves for the maintenance of hydraulic function during water stress, compared to laurel plants subjected to the same treatment. In accordance, xylem recovery occurred in control P. alba samples that, with equal NSC reserves of S samples, can rely on stem photosynthesis as well as in C and stem-shaded plants of L. nobilis.
In response to experimental drought, we observed a significant reduction in leaf gas exchange in the study species, leading to a severe reduction of leaf-level carbon assimilation. Under such conditions, plant metabolism of C plants was probably maintained by stored carbon compounds as well as by stem photosynthesis products. By contrast, in S samples, only stored carbon could be used. Soluble NSCs derived from photosynthesis or by starch hydrolysis are also necessary for osmoregulation processes and/or ROS scavenging [46,47,48,90]. Sugars lead to generation of local positive pressures to drive water inflow into embolized conduits when water potential rises upon re-irrigation, leading to hydraulic recovery, i.e., [61,91,92,93]. In accordance, L. nobilis plants under drought showed a significant increase in soluble NSC content, and a parallel decrease in starch concentration in both C and S samples, compared to well-watered plants. Moreover, in this species, changes in NSC content were clearly related to changes in PLC. A different trend was recorded in P. alba. In fact, despite (i) the significant increase in soluble NSCs’ content as recorded in C and S poplar samples in respect to well-watered plants and (ii) soluble sugars’ content showing similar trend of changes in C and S poplar and laurel plants in response to drought-recovery cycle, no correlation between NSCs and PLC values was recorded in P. alba. These results clearly suggest that, in poplar stems, modulation of soluble NSCs and starch content was not involved in restoring hydraulic function, at least in S samples, where no refilling occurred even after re-irrigation.
Our data invite to depict the following scenario, that might orient future studies on the role of stem photosynthesis in modulation of plant hydraulic functioning. In evergreens, the maintenance of leaf photosynthesis over the whole year may assure sufficient carbon uptake, so that carbon stores would be normally available and usable under stress conditions. Hence, for these species, stem photosynthesis could represent an extra carbon gain to be used only when leaf photosynthesis is severely inhibited. By contrast, in deciduous species, the extra carbon gain derived by stem photosynthesis might play a key role in whole plant metabolism, especially under drought. In these species, stem photosynthesis would not provide an “extra” carbon gain but would contribute to whole plant carbon assimilation.

5. Conclusions

In conclusion, our experiment has tried to disentangle the impact of leaf and stem photosynthesis on post-drought hydraulic recovery, showing that carbon assimilation at stem level differentially affects the maintenance of hydraulic functions in an evergreen versus a deciduous species. Our results, if confirmed in a wider number of deciduous versus evergreen species, will provide key information to predict the vulnerability of tree species under current global climate changes. Nevertheless, further studies investigating the impact of long- and short-term stem shading on species-specific drought resilience are needed.

Author Contributions

Conceptualization, P.T., A.N. and V.C.; investigation, E.A., S.G., S.N., P.T. and V.C.; resources, P.T. and V.C.; data curation, P.T.; writing—original draft preparation, P.T; writing—review and editing, P.T., A.N. and V.C. All authors have read and agreed to the published version of the manuscript.

Funding

This research received no external funding.

Data Availability Statement

The data presented in this study are available on request from the corresponding author.

Acknowledgments

We are very grateful to Dipartimento Regionale Azienda Foreste Demaniali, Messina, Sicily, Italy, for kindly providing plant material.

Conflicts of Interest

The authors declare no conflict of interest.

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Figure 1. (A) Experimental design and (B) measurements performed on control and stem-shaded samples (C and S, respectively) of L. nobilis and P. alba plants; (A) At the beginning of June, C and S samples were subjected to a water stress treatment. One month later, a set of samples was measured for different physiological parameters. Another set of plants was re-irrigated at field capacity and measured after 7 days to evaluate the capacity for recovery; (B) Leaf gas exchange rates were measured from May to the end of the experiment; xylem water potential (Ψx) values were recorded from June to the end of experiments; hydraulic measurements and NSC content were performed after the drought treatment and 7 days after re-irrigation. Moreover, PV-curves were measured before and at the end of the drought-recovery treatments. Leaf area and leaf mass per area were measured at the end of the experiments (for details, see the text). PV: pressure volume curves; NL: number of leaves of the current-year branches; Ncy: number of current-year branches; Lcy: length of the current-year branches; AL: leaf area; LMA: leaf mass area.
Figure 1. (A) Experimental design and (B) measurements performed on control and stem-shaded samples (C and S, respectively) of L. nobilis and P. alba plants; (A) At the beginning of June, C and S samples were subjected to a water stress treatment. One month later, a set of samples was measured for different physiological parameters. Another set of plants was re-irrigated at field capacity and measured after 7 days to evaluate the capacity for recovery; (B) Leaf gas exchange rates were measured from May to the end of the experiment; xylem water potential (Ψx) values were recorded from June to the end of experiments; hydraulic measurements and NSC content were performed after the drought treatment and 7 days after re-irrigation. Moreover, PV-curves were measured before and at the end of the drought-recovery treatments. Leaf area and leaf mass per area were measured at the end of the experiments (for details, see the text). PV: pressure volume curves; NL: number of leaves of the current-year branches; Ncy: number of current-year branches; Lcy: length of the current-year branches; AL: leaf area; LMA: leaf mass area.
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Figure 2. Mean values ± SEM of (a,b) stomatal conductance to water vapor, gL; (c,d) photosynthetic rate (An) and (e,f) xylem water potential (ΨX) as recorded in May (M), June (J) and July (Jl) in control (C, light green circle) and shaded samples (S, dark green circle) of L. nobilis and P. alba plants. Arrows indicate the day starting drought treatment (D) and re-irrigation (I). F values, as obtained by the two-way ANOVA analysis, are reported. Time of measurement, T, and light treatment, L (i.e., control and shaded samples) are the explanatory variables ** = p < 0.01; *** = p < 0.001.
Figure 2. Mean values ± SEM of (a,b) stomatal conductance to water vapor, gL; (c,d) photosynthetic rate (An) and (e,f) xylem water potential (ΨX) as recorded in May (M), June (J) and July (Jl) in control (C, light green circle) and shaded samples (S, dark green circle) of L. nobilis and P. alba plants. Arrows indicate the day starting drought treatment (D) and re-irrigation (I). F values, as obtained by the two-way ANOVA analysis, are reported. Time of measurement, T, and light treatment, L (i.e., control and shaded samples) are the explanatory variables ** = p < 0.01; *** = p < 0.001.
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Figure 3. Mean values ± SEM of percentage loss of hydraulic conductivity (PLC) as recorded in control (C) and shaded stem (S) samples of L. nobilis (a) and P. alba (b) as measured at midday, on the following morning (Morning) as well as in samples re-irrigated at field capacity and measured 7 days after re-irrigation (Recovery). Light green and dark green columns refer to C and S samples, respectively. F values, as obtained by the two-way ANOVA analysis, are reported. Time of measurement, T, and light treatment, L (i.e., control and shaded samples) are the explanatory variables. * = p < 0.05; ** = p < 0.01; *** = p < 0.001.
Figure 3. Mean values ± SEM of percentage loss of hydraulic conductivity (PLC) as recorded in control (C) and shaded stem (S) samples of L. nobilis (a) and P. alba (b) as measured at midday, on the following morning (Morning) as well as in samples re-irrigated at field capacity and measured 7 days after re-irrigation (Recovery). Light green and dark green columns refer to C and S samples, respectively. F values, as obtained by the two-way ANOVA analysis, are reported. Time of measurement, T, and light treatment, L (i.e., control and shaded samples) are the explanatory variables. * = p < 0.05; ** = p < 0.01; *** = p < 0.001.
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Figure 4. Mean values ± SEM of (a,b) soluble NSCs and (c,d) starch content as recorded in control (C) and shaded stem (S) samples of L. nobilis and P. alba at midday, on the following morning (Morning) and at morning after re-irrigation at field capacity (Recovery). Light and dark green columns refer to C and S samples, respectively. F values, as obtained by the two-way ANOVA analysis, are reported. Time of measurement, T, and light treatment, L (i.e., control and shaded samples) are the explanatory variables. * = p < 0.05; ** = p < 0.01; *** = p < 0.001.
Figure 4. Mean values ± SEM of (a,b) soluble NSCs and (c,d) starch content as recorded in control (C) and shaded stem (S) samples of L. nobilis and P. alba at midday, on the following morning (Morning) and at morning after re-irrigation at field capacity (Recovery). Light and dark green columns refer to C and S samples, respectively. F values, as obtained by the two-way ANOVA analysis, are reported. Time of measurement, T, and light treatment, L (i.e., control and shaded samples) are the explanatory variables. * = p < 0.05; ** = p < 0.01; *** = p < 0.001.
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Figure 5. Relationships between percentage loss of hydraulic conductivity (PLC) and (a,c) soluble NSCs and (b,d) starch content as recorded in control (light green) and woody tissue-shaded (dark green) stem of L. nobilis and P. alba stem samples. Regression equation (dotted and solid lines for correlation with p > 0.05 and p < 0.05, respectively), coefficient values, correlation coefficients (r2) and p-values are also reported.
Figure 5. Relationships between percentage loss of hydraulic conductivity (PLC) and (a,c) soluble NSCs and (b,d) starch content as recorded in control (light green) and woody tissue-shaded (dark green) stem of L. nobilis and P. alba stem samples. Regression equation (dotted and solid lines for correlation with p > 0.05 and p < 0.05, respectively), coefficient values, correlation coefficients (r2) and p-values are also reported.
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Table
Table 1. Mean ± SEM of the total number (Ncy) and length (Lcy) of current-year branches, mean leaf surface area (AL) and leaf mass per area (LMA) as recorded in control (C) and shaded (S) samples of L. nobilis and P. alba. Differences between C and S samples were checked by a Student’s t-test. p-Values are reported between brackets; n = 3 plants for Ncy and Lcy; n = 40 leaf samples for AL and LMA.
Table 1. Mean ± SEM of the total number (Ncy) and length (Lcy) of current-year branches, mean leaf surface area (AL) and leaf mass per area (LMA) as recorded in control (C) and shaded (S) samples of L. nobilis and P. alba. Differences between C and S samples were checked by a Student’s t-test. p-Values are reported between brackets; n = 3 plants for Ncy and Lcy; n = 40 leaf samples for AL and LMA.
ParameterL. nobilisP. alba
CSCS
Ncy2.7 ± 0.32.7 ± 0.37.7 ± 1.19.3 ± 1.9
(p-value = 1.0)(p-value = 0.292)
Lcy (cm)13.3 ± 0.913.4 ± 2.317.6 ± 3.113.7 ± 1.7
(p-value = 1.0)(p-value = 0.201)
AL (cm2)12.2 ± 0.510.6 ± 0.617.4 ± 0.917.1 ± 1.1
(p-value = 0.022)(p-value = 0.414)
LMA (g m−2)104.0 ± 2.6111.5 ± 2.941.1 ± 1.541.6 ± 1.3
(p-value = 0.06)(p-value = 0.797)
Table
Table 2. Mean ± SEM (n = 3 plants) of total number of leaves (NL) as recorded in June (before starting drought treatment) and in July (at the end of the experimental period, for details, see the text). Different letters indicate statistically significant differences based on two-way ANOVA. F values are reported as resulted by statistical analysis of NL measurements by time, T (i.e., June, before drought, and July, at the end of experimental treatment) and light treatment, L (i.e., control and shaded samples).
Table 2. Mean ± SEM (n = 3 plants) of total number of leaves (NL) as recorded in June (before starting drought treatment) and in July (at the end of the experimental period, for details, see the text). Different letters indicate statistically significant differences based on two-way ANOVA. F values are reported as resulted by statistical analysis of NL measurements by time, T (i.e., June, before drought, and July, at the end of experimental treatment) and light treatment, L (i.e., control and shaded samples).
SpeciesTimeNLp-Value
CS
L. nobilisJune32.3 ± 3.533.7 ± 5.5T: 0.819
L: 1
July32.3 ± 3.5 33.7 ± 5.5T × L: 1
P. albaJune177.3 ± 15.5a168.7 ± 9.6aT: <0.001
L: 0.1
July38.7 ± 10.11b95.7 ± 3.6cT × L: <0.05
Table
Table 3. Mean ± SEM (n = 5) of leaf water potential at turgor loss point (−Ψtlp), osmotic potential at full turgor (−πo) and bulk modulus of elasticity (εmax) as recorded in June (before starting drought treatment), and in July (at the end of the experimental period) in control (C) and shaded (S) samples of L. nobilis and P. alba plants. Different letters indicate statistically significant differences based on two-way ANOVA.
Table 3. Mean ± SEM (n = 5) of leaf water potential at turgor loss point (−Ψtlp), osmotic potential at full turgor (−πo) and bulk modulus of elasticity (εmax) as recorded in June (before starting drought treatment), and in July (at the end of the experimental period) in control (C) and shaded (S) samples of L. nobilis and P. alba plants. Different letters indicate statistically significant differences based on two-way ANOVA.
ParameterTimeL. nobilisP. alba
CSCS
−Ψtlp (MPa)June1.99 ± 0.05a1.97 ± 0.06a2.27 ± 0.032.21 ± 0.10
July2.64 ± 0.11b2.62 ± 0.10b2.19 ± 0.012.19 ± 0.09
−πo (MPa)June1.71 ± 0.04a1.72 ± 0.04a1.73 ± 0.031.72 ± 0.05
July2.07 ± 0.0.3b2.08 ± 0.09b1.53 ± 0.111.72 ± 0.09
εmax (MPa)June27.4 ± 3.631.2 ± 4.418.2 ± 4.525.7 ± 3.5
July35.7 ± 2.537.9 ± 3.130.1 ± 3.927.6 ± 4.1
Table
Table 4. Results of the two-way ANOVA analysis of parameters determined from PV-curves’ analysis by time of measurement, T (i.e., Before drought and at the end of experiment) and light treatment, L (i.e., control and shaded samples). Numbers represent F values; *** = p < 0.001.
Table 4. Results of the two-way ANOVA analysis of parameters determined from PV-curves’ analysis by time of measurement, T (i.e., Before drought and at the end of experiment) and light treatment, L (i.e., control and shaded samples). Numbers represent F values; *** = p < 0.001.
ParameterL. nobilisP. alba
TLT × LTLT × L
Ψtlp (MPa)43.58 ***0.0250.00060.180.080.04
πo (MPa)33.91 ***0.050.011.230.911.11
εmax (MPa)3.760.580.052.361.301.24
Table
Table 5. Mean values ± SEM of soluble NSCs and starch content as recorded in well-watered samples of L. nobilis and P. alba at midday and on the following morning on the same days of hydraulic measurements in water-stressed samples. Different letters indicate statistically significant differences based on two-way ANOVA.
Table 5. Mean values ± SEM of soluble NSCs and starch content as recorded in well-watered samples of L. nobilis and P. alba at midday and on the following morning on the same days of hydraulic measurements in water-stressed samples. Different letters indicate statistically significant differences based on two-way ANOVA.
SpeciesSoluble Sugars
(mg g−1 DW−1)		Starch
(mg g−1 DW−1)
MiddayMorningMiddayMorning
L. nobilis14.8 ± 7.313.2 ± 9.719.2 ± 1.8b17.9 ± 2.5b
P. alba13.1 ± 1.313.2 ± 1.26.9 ± 2.3a610.5 ± 2.8ab
Table
Table 6. Results of the two-way ANOVA analysis of parameters by species, Sp (i.e., laurel versus poplar) and time, T (i.e., Midday versus Morning). Numbers represent F values; *** = p < 0.001.
Table 6. Results of the two-way ANOVA analysis of parameters by species, Sp (i.e., laurel versus poplar) and time, T (i.e., Midday versus Morning). Numbers represent F values; *** = p < 0.001.
ParametersSpTSp × T
soluble NSCs0.530.410.62
starch16.71 ***0.2151.01
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Trifilò, P.; Natale, S.; Gargiulo, S.; Abate, E.; Casolo, V.; Nardini, A. Stem Photosynthesis Affects Hydraulic Resilience in the Deciduous Populusalba but Not in the Evergreen Laurus nobilis. Water 2021, 13, 2911. https://doi.org/10.3390/w13202911

AMA Style

Trifilò P, Natale S, Gargiulo S, Abate E, Casolo V, Nardini A. Stem Photosynthesis Affects Hydraulic Resilience in the Deciduous Populusalba but Not in the Evergreen Laurus nobilis. Water. 2021; 13(20):2911. https://doi.org/10.3390/w13202911

Chicago/Turabian Style

Trifilò, Patrizia, Sara Natale, Sara Gargiulo, Elisa Abate, Valentino Casolo, and Andrea Nardini. 2021. "Stem Photosynthesis Affects Hydraulic Resilience in the Deciduous Populusalba but Not in the Evergreen Laurus nobilis" Water 13, no. 20: 2911. https://doi.org/10.3390/w13202911

APA Style

Trifilò, P., Natale, S., Gargiulo, S., Abate, E., Casolo, V., & Nardini, A. (2021). Stem Photosynthesis Affects Hydraulic Resilience in the Deciduous Populusalba but Not in the Evergreen Laurus nobilis. Water, 13(20), 2911. https://doi.org/10.3390/w13202911

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