Discussion
Rootstocks that strongly differed in their dwarfing capacity of the scion also affected stem water potential, leaf gas exchange, and carbon isotope composition in both leaves and stems. Rootstocks with lower stem water potential also showed more strongly restricted growth and trunk diameter. The influence of rootstocks on scion growth and canopy development is an important component in the understanding of rootstock-scion water use (Higgs and Jones 1990). In this experiment, rootstocks with a high dwarfing capacity such as B.9. had lower stem water potential, net CO2 assimilation, stomatal conductance, and transpiration. These rootstock-induced changes in leaf function translated to differences in carbon isotope composition in both leaves and stems.
Changes to stem water potential are normally induced via changes in soil water availability (Suter et al., 2019). Stem water potential contributes to the control of stomatal conductance (Buckley, 2019). Stomatal behavior depends on the physiological traits of the plant and the host environment. At night, water is taken up and stored within the plant (Knipfer et al., 2019). In the morning, stomata open and transpirations rates increase, reducing leaf water potential and turgor pressure (Rodriguez-Dominguez et al., 2019). Drought stress forces earlier stomatal closure due to loss of leaf turgor and, consequently, a decline in leaf xylem hydraulic conductance to avoid excessive negative pressure in xylem (Knipfer et al., 2020). Stomatal closure is an effect of a decline in root hydraulic conductance and changes within the root-to-soil hydraulic continuum (Knipfer et al., 2020).
In apple, stem water potential has been reported to vary between – 0.8 and – 1.0 MPa in fully irrigated trees (Naor and Cohen. 2003). Here, stem water potential measurements corresponded with those previously reported. Valverdi et al. (2021) reported that rootstocks with a higher dwarfing capacity like B.9 did not vary as much in shoot growth between well-watered conditions compared to semi-dwarfing rootstocks like G.890. Similar to Valverdi et al. (2019), more dwarfing rootstocks increased inherent water limitations indicated by lower stem water potential, stomatal conductance, and transpiration. This was especially true in this study as trees matured from 2018 to 2019. Rooting volume may, in part, contribute to some of these responses (Costes and Garcia-Villanueva, 2007; Ma et al., 2013; Harrison et al., 2016; Foster et al., 2017). However, in the semi-arid environment where this study occurred, more than 95% of the water was supplied through drip irrigation and with a young orchard, irrigated volume was similar for all rootstocks. Therefore, differences observed among rootstocks were more likely a result of elevated limitations in water uptake and transport to the scion rather than simply access to water within the soil profile.
In this study, rootstocks modulated scion leaf water relations by affecting stomatal closure and leaf gas exchange which had downstream effects on δ13C composition. There have been previous studies which have reported differences in water relations when different rootstocks were used in woody plants (Bongi et al., 1994; Padgett-Johnson et al., 2000; Gibberd et al., 2001; Cohen and Naor, 2002; Ma et al., 2010; Liu et al., 2012a; Galbignani et al., 2016; Peccoux et al., 2018; Villalobos-Gonzales et al., 2019; Frioni et al., 2020). However, these studies did not show the linear relationship between these traits among many phenotypically diverse rootstocks like this study. More often, differences have been observed among scion cultivars or among treatments that affect water supply to the roots. However, Liu et al (2012b) described how rootstocks manipulated stomatal conductance and photosynthetic capacity in ‘Gale Gala’. Here, carbon isotope composition was different for Honeycrisp when grafted to apple rootstocks with differences in dwarfing capacities. A reduction in discrimination against 13CO2 may indicate an increase in water use efficiency (Jones; 1984; Farquhar et al. 1989). In apples, carbon isotope composition was reported to be lower for Fuji compared to Braeburn apples because of limitations in stomatal and leaf respiration rates (Massonnet et al., 2007). In this study, elevated carbon isotope composition in more dwarfing rootstocks like B.9 indicated increased conservation of water through reduced transpiration rates and lower stomatal conductance in ‘Honeycrisp’ leaves. These effects on water relations have downward effects on scion vigor and were consistent with our knowledge of dwarfing capacity of the rootstocks tested in this study (Seleznyova et al.,2008; Kosina, 2010; Tworkoski and Fazio, 2015; Foster et al., 2017). The most obvious influence of dwarfing rootstock is the ability to produce lower vegetative biomass relative to more vigorous rootstocks. Trunk diameter and shoot extension measurements are two of the main traits used to characterize tree growth (Lauri et al; 2006). When we measured trunk diameter at rootstock above and below the graft union, we notice differences in growth rates. In both years, B.9 rootstock was smaller than all other rootstocks and our observations were similar to Gjamovski and Kiprijanovski (2011). Corresponding reductions in stem water potential, an accurate measure of plant water status (Shackel et al., 1997) were observed among rootstock cultivars.
The rootstock-modulated effect on scion water relations represents an important component in the understanding of rootstock-scion water use (Higgs and Jones., 1990). When net carbon assimilation was measured among rootstocks, we observed that less conservative rootstocks like semi-dwarfing G.890 had higher photosynthetic rates. These results follow similar patterns with those reported by Fallahi et al (2001), who reported lower net photosynthetic rates for B.9 compared to more vigorous rootstocks like Ottawa 3 and M.7. Clearly, the capacity of the tree to improve net carbon assimilation and thus, increase carbon fixation is related to the ability of the hydraulic system to supply water to the leaves (Koepke and Dhingra. 2013). Therefore, the hypothesis that dwarfing in rootstocks is associated with rootstock-mediated water restrictions at a particular point in the tree (Cohen and Naor, 2002; Atkinson et al., 2003; Cohen et al; 2007; Cohen et al, 2003; Atkinson et al, 2001; Tworkoski and Fazio, 2015)..These restrictions could either occur at the root, the graft union, or in the impact of rootstocks on stem and leaf hydraulic traits. We acknowledge that this isn’t fully answered by this study and more work is needed to better discern how rootstocks might affect each restriction point.
The positive relationship between carbon isotope composition between leaf and stem demonstrates short-term and long-term impacts of rootstocks on scion water relations and carbon assimilation. We observed enrichment of stems compared to leaves and this is well aligned with previous studies (Arndt and Wanek, 2002; Cernusak et al., 2005). Specifically, B.9 showed a clear divergence in carbon isotope composition between stem and leaves where leaves were depleted relative to stems that could be explained by several phenomenon. Cernusak et al., 2009 presented six hypotheses for this effect. One of the outlined hypotheses to explain enrichment of non-photosynthetic tissues compared with their leaves rely on post-photosynthetic carbon fractionation due to seasonal separation of growth (Cernusak et al., 2009). Development and growth of leaves occur in spring with plentiful soil water content. Since variability in carbon discrimination is also affected by intercellular to ambient CO2 partial pressure, the photosynthetic carbon assimilation is heavily discriminating against13C at this time (Werner and Gessler, 2011). Wood growth occurs later in the season, when temperatures and VPD are elevated which can reduce stomatal conductance producing less discrimination against 13C at the time carbon is used for synthesis of non-photosynthetic tissues (Cernusak et al., 2009). Differences in δ13C between leaves and stems were less pronounced when discrimination and vegetative vigor were also greater. Since more vigorous rootstocks extend growth into later parts of the season, we would expect a greater agreement in δ13C between leaves and stems than for less vigorous rootstocks like B.9.
In conclusion, we report the effect of rootstocks on carbon isotope composition in leaves and stems and show the close association between leaf gas exchange and stem water potential. Rootstocks were able to strongly affect water-use efficiency and carbon assimilation for the same apple scion. Furthermore, there was a close association between rootstock-influenced gas exchange and overall shoot vigor when soil water content was uniform. Rootstocks that are more vigorous have more negative carbon isotope composition indicating lower water restrictions in transport to above ground parts compared to rootstocks with lower vigor. These results increase our understanding of the physiological mechanisms underlying dwarfing in composite woody plants like apple and show how below-ground traits imparted from rootstocks can affect water-use efficiency and carbon isotope composition in leaves and stems.