Physiological disorders related to water dynamics represent a significant limitation to fruit growth, particularly fruit cracking, which limits both production and quality, especially in berries and drupes, but also in citrus (Matthews et al., 2009; Agustí et al., 2014; Winkler et al., 2015, 2016; Fischer et al., 2021), and purple spot, which can affect up to 60% of the fruit in loquat (Gariglio et al., 2002). There are several other physiological disorders related to fruit water relations, but they are beyond the scope of this manuscript.

The aim of this review is to examine the water relations between internal and external tissues in growing fruit through the lens of the epidermal growth control hypothesis. We examine current knowledge on the influence of environmental, genetic, and physiological factors on water relations, focusing on skin tissue as a key indicator of growth stress associated with physiological disorders.

Fruit development involves processes of cell division and expansion after fertilization. It is regulated by distinct transcriptional patterns and complex regulatory systems involving genetic, epigenetic, and hormonal controls at all developmental stages (Teyssier et al., 2015). Traditional assessment of fruit growth is based on dry matter accumulation and changes in cell enlargement. Both cell division and cell enlargement exhibit a sigmoidal or double sigmoidal growth pattern, depending on the species, that develops over time, typically in fleshy fruits (Rodriguez et al., 2019).

Fruit dry matter accumulation is the result of photoassimilate transport through the phloem by mass flow, a phenomenon elucidated by Mason and Maskell (1928). The pressure gradient resulting from the altered sugar concentration between source and sink tissues facilitates phloem sap transport, allowing sink organs to modulate mass flow rates through various metabolic enzymes, sugar transporters, and transcriptional and post-translational regulation (Sun et al., 2022; Ren et al., 2023). In summary, it is possible to increase crop yield by increasing source capacity and carbohydrate production in leaves or by increasing the use of photoassimilates in sink tissues (Driesen et al., 2023). In light of this, a number of agronomic practices have been developed and adopted by farmers to improve assimilate allocation to fruit, thereby increasing both the quantity and quality of their crops. These practices include pruning (Araújo-Ferreira et al., 2016), irrigation, fertilization, fruit thinning (Singh-Sidhu et al., 2022), modification of flowering intensity (Agustí et al., 2022), and the use of growth regulators (Gill et al., 2023), among others.

A poorly explored facet of fruit growth is the relationship between internal and external tissues within a growing organ, mainly because of their different growth capacities. According to Kutschera and Niklas (2007), expansion of the internal tissues leads to elongation of the external tissues. Specifically, the inner tissues act as the primary force driving elongation, while the outer cell layers impose a mechanical constraint (Kraus, 1867). Consequently, this dynamic results in the generation of tension or stress within the outer cell layers due to the expansion of the internal tissues. The importance of the inner tissues in organ growth was highlighted by their active state of tension, which differed from that of the outer tissues. This led to a hypothesis that emphasized the role of the epidermis in growth regulation, the so-called “epidermal growth control hypothesis” (Kutschera and Niklas, 2007). This hypothesis focused the study on the architecture and properties of the epidermal cell walls, the structure and function of the cuticle, and the modification of these properties by hormonal actions, especially the changes that occur during fruit ripening (Kutschera and Niklas, 2007).

Parenchyma cells with a thin primary wall are the predominant cell type in fleshy fruits. In these cells, the turgor pressure (P) resulting from osmotic water uptake provides the necessary force to induce plastic deformation of the cell walls, allowing cell expansion and, consequently, the permanent increase in size or growth of different organs (Ortega, 2023). Plastic deformation occurs only when the turgor pressure exceeds a critical value (Pc), according to the following equations:

where ϕ = coefficient of irreversible extensibility of the cell wall (h⁻¹ MPa⁻¹), P = turgor pressure (MPa), and Pc = critical turgor pressure for plastic extension (MPa).

Consequently, the water potential (Ψ_a) of the parenchyma cells in fleshy fruits is controlled by two primary components. Firstly, the tissue pressure (P) within the fruit, due to the cell wall and the skin of the fruit, limits the expansion of both the cell and the flesh. Secondly, the osmotic potential (π) induced by the presence of soluble metabolites; hence, Ψ_a = P – π. As a result, the osmotic potential favors the influx of water into the cell, whereas the pressure potential exerts a restrictive influence. While elevated metabolite concentrations are uncommon in plant vegetative organs, they are a common phenomenon during the ripening of fleshy fruits (Jia et al., 2020).

According to the epidermal growth control hypothesis, the regulation of fruit growth is controlled by the epidermal tissue (Kutschera and Niklas, 2007) according to Equation 1. The cell wall strength and cell–cell adhesion of the fruit epidermis are not constant, but change over time (Canton et al., 2020; Shi et al., 2023). Therefore, the critical turgor for fruit growth and the coefficient of irreversible extensibility of the cell wall are subject to modification throughout the course of fruit development (Ortega, 2023), resulting in a corresponding change in fruit growth rate. However, the period of most pronounced change in cell wall structure, and consequently on fruit growth rate, is during fruit ripening. This involves a variety of physiological, structural, and metabolic changes that ultimately lead to the development of edible fruit. The regulation of this process occurs at different molecular levels, with phytohormones, transcription factors, and epigenetic modifications all playing a crucial role (Perotti et al., 2023).

Ethylene has been highlighted as a key phytohormone in the ripening of climacteric fruits, while abscisic acid also plays a central role in non-climacteric fruits (Li et al., 2021; Li et al., 2022). During the ripening process of climacteric fruits, changes in primary cell wall metabolism are mainly associated with the induction of several gene families, including expansins (EXPs), xyloglucan endotransglucosylase (XET)/hydrolases (XTH), and endo-1,4-β-glucanases (EGase or Cel) (Shi et al., 2023). While some changes in cell wall composition and structure are common to different species, others are species-specific (Canton et al., 2020). Interestingly, auxin and other agents can modulate cell wall growth rates in living plants very rapidly, within minutes or even seconds. Furthermore, this rapid response could be controlled by altering wall pH, which would activate or inactivate expansins, the proteins responsible for acid-induced wall extension in growing tissues (Cosgrove, 2000). In addition, cellulose can achieve a much higher degree of structural order, or crystallinity, which has a strong effect on the mechanical properties of cell walls, as the ratio of crystalline to amorphous cellulose is associated with the rate of extension (Bringmann et al., 2012).

During ripening, the metabolism of climacteric fruit also induces the degradation of polymers such as starch and cell wall polysaccharides, leading to changes in cellular water status due to catabolism. As a result, the cellular concentrations of metabolites increases, resulting in a lower cellular osmotic potential, which increases water uptake and turgor pressure, allowing the fruit to maintain a high growth rate even under adverse conditions (Blum, 2017; Miranda Fernandes et al., 2018). The contribution of metabolites to osmotic adjustment also includes acids, phenolics, amino acids, soluble pectins, and minerals, all measured as part of the soluble solids content (Sharma et al., 2019; Hou et al., 2020).

However, sugar metabolism and accumulation during ripening are also important in non-climacteric fruits, as observed in loquat, where almost 90% of the total sugar accumulation takes place within a 15-day period from color break (Gariglio et al., 2003a), and this period coincides with the phase of higher fruit growth rate (Gariglio et al., 2003b). Similar trends were observed in grapes (Matthews et al., 2009) and sweet cherries (Schumann et al., 2014).

As observed in fruits, the enzymatic modification of the cell wall of the epidermal tissue is also responsible for modifying of the growth rate of vegetative tissues, such as the leaves of Lolium temulentum (Bacon et al., 1997). In Cucurbita pepo, the periods of maximum growth rate in both fruit and leaves coincide with the highest levels of ascorbic acid oxidase (AAO) in the epidermis (Lin and Varner, 1991). This enzyme softens the cell walls, thereby facilitating an increase in growth rate. Furthermore, the epidermal growth control hypothesis has been proposed to explain the growth tension between internal and external tissues of the hypocotyl of sunflower seedlings (Kutschera and Niklas, 2007).

In summary, specific phytohormones and enzymes play a crucial role in modulating the water relations and, consequently, the growth rate of growing tissues by altering the structure of cell walls. These modifications impact pivotal parameters of the water relations (see Equation 1), including the coefficient of irreversible extensibility of the cell wall and the critical turgor pressure required for plastic expansion. Furthermore, they contribute to the catabolism of reserve substances, thereby triggering osmotic adjustment processes. By influencing these factors, phytohormones and enzymes contribute to the dynamic regulation of growth processes in response to environmental and developmental cues.

The epidermal growth control hypothesis establishes the occurrence of stress between internal and external tissues during growth; however, the water potentials and their components in both internal and external tissues have not been thoroughly studied. Nevertheless, some studies have indirectly investigated the evolution of water potentials in flesh tissues, and their analysis is crucial for understanding this physiological process in fruit. Rain-induced cracking of sweet cherry fruit significantly limits its global production and has been the subject of extensive research (Winkler et al., 2016). An accepted conceptual framework to explain this phenomenon is the critical turgor model, originally proposed by Considine and Kriedemann (1972), with a specific focus on grape (Vitis vinifera L.) berries. According to this theoretical model, the flesh of the berry is compressed by an elastically stretched skin. As the fruit absorbs water, the internal pressure within the fruit increases. Once the critical threshold is reached, the fruit skin is overstressed beyond its elastic limit, resulting in cracking (Considine and Kriedemann, 1972).

However, a study in which fruits of 19 sweet cherry cultivars were incubated in water to analyze water uptake and cracking contradicted the notion that cracking is a simple function of the amount of water absorbed, as proposed by the critical turgor model (Winkler et al., 2016). In the following years, extensive research was carried out to improve the understanding of water relations in cherries in order to explain the cracking process (Schumann et al., 2014). Tissue water potential and fruit turgor showed a decrease 55 days after full bloom and remained low until maturity. In contrast to the critical turgor model, cell turgor decreased from the beginning of stage III of sweet cherry fruit growth, dropping from 350 to 25 kPa within the same period. Consequently, in the absence of cell turgor, both water potential and osmotic potential tended to converge to similar values. Surprisingly, fruit growth continued at high rates despite the significant reduction in cell turgor (Schumann et al., 2014). Based on these results, the hypothesis that cracking is induced by an increase in tissue turgor was rejected (Schumann et al., 2014). In another study of 17 European plum cultivars, cell turgor pressure decreased significantly from 0.33 to 0.35 MPa 78 days after full bloom to 0.02 MPa at harvest. This final measurement was remarkably low compared with the markedly negative osmotic potential (<−3.00 MPa) and water potential recorded at maturity, both of which reached similar values at this time. Furthermore, cell turgor in European plum was found to be independent of osmotic potential (Knoche and Grimm, 2022). These results showed a parallel trend to those mentioned for sweet cherry (Schumann et al., 2014).

In Cabernet Sauvignon berries grown in a greenhouse, cell turgor declined significantly, from a peak of 0.30 MPa at 48 days after anthesis to approximately 0.05 MPa at 60 days after anthesis, and this level was maintained throughout ripening (Matthews et al., 2009). During the initial phase of turgor decline, from 0.30 to 0.16 MPa, there was only a marginal increase in soluble solids, but cell turgor continued to decline, increasing to 3.6°Brix within 3 days. This trend continued, indicating that significant sugar accumulation did not begin until cell turgor reached about 0.10 MPa or less. In the case of Pinot Noir, the pattern of cell turgor is similar to that described for Cabernet Sauvignon (Matthews et al., 2009).

In loquat fruit, the flesh turgor is consistently low, remaining below 0.15 MPa throughout the growth period. Interestingly, in a similar context, turgor becomes almost non-existent around the time of color break, a phase when the fruit reaches its highest growth rate (Gariglio et al., 2008a; Reig et al., 2016). In contrast to observations in sweet cherry (Schumann et al., 2014), European plum (Knoche and Grimm, 2022), and grape (Matthews et al., 2009), there is no significant decrease in turgor, osmotic potential, and water potential in loquat at the beginning of phase III of fruit growth (Gariglio et al., 2008a).

Using a miniaturized pressure probe, Kutschera and Niklas (2007) observed that cell turgor in the outer tissues of sunflower hypocotyls of 4-day-old dark-grown seedlings is comparable to that of the inner tissues (0.48–0.49 MPa). However, there is a marked difference in osmotic potential between the epidermal layer (0.63 MPa) and the internal tissues (0.56 MPa). Consequently, the water potential is more negative in the outer tissues (−0.14 MPa) compared to the inner tissues (−0.08 MPa), meaning that the water potential gradient drives the movement of water from the inner tissues into the epidermis (Kutschera and Niklas, 2007).

A limited number of studies have been identified that specifically investigate water potentials and their components within the internal and external tissues of growing fruits. One such investigation, which sheds light on the physiological origins of loquat purple spot, is reported in the work of Gariglio et al. (2008a). This study focuses on the analysis of water potential, providing an important perspective for understanding the underlying physiology associated with water relations between internal and external tissues.

Purple spot appears as an extensive area with a slightly depressed surface, characterized by a purple color and irregular shape, affecting only the epidermal tissue of the fruit. The affected areas begin in the deepest layers of the skin cells and progress to the more superficial cell layers. The cuticle shows no signs of damage and its water permeability is unaffected. Therefore, water loss from the fruit to the atmosphere cannot be considered as the cause of the skin dehydration that characterizes purple spot (Gariglio et al., 2002). Subsequent research has shown that epidermal dehydration results from a change in the water balance between the flesh and the rind, which occurs at the same time as fruit color breakdown. This period is characterized by both a significant increase in sugar accumulation and a high fruit growth rate. The dehydration process is also influenced by cultural practices, such as the intensity of fruit thinning, and environmental factors, such as low temperature and exposure to sunlight. These factors affect the assimilation and partitioning of sugars and minerals, favoring the flesh and increasing the solute concentration gradient between the two tissues (Gariglio et al., 2008b). In conclusion, the purple spot of the loquat fruit is a physiological disorder explained by the tension between internal and external tissues, as postulated by the epidermal growth control hypothesis. Therefore, their results have the potential to shed light on this hypothesis and the water relations established in this context.

As the incidence of purple spot increases with the intensity of fruit thinning, a comparative analysis was carried out between plants thinned at different rates of fruit per panicle. In particular, unthinned trees showed no incidence of purple spot, whereas plants thinned at one fruit per panicle showed a significant fruit damage rate of 34%. Intermediate incidence levels were observed in plants thinned at five and three fruits per panicle (Gariglio et al., 2003a). Comparing the extreme treatments of fruit thinning, the average water potential of both flesh and skin tissues did not show significant differences, but varied with the sampling date. This is true for both unthinned plants ( Figure 1A ) and plants thinned to one fruit per panicle ( Figure 2A ). In the absence of purple spot risk (unthinned plants), the osmotic potential of the skin tends to be slightly lower than that of the flesh tissue ( Figure 1B ), while the turgor remains slightly higher throughout the fruit growth period.

In plants thinned to one fruit per panicle, where the susceptibility to purple spot is maximal, the evolution of water potential in both flesh and skin tissues does not show differences ( Figure 2A ), as observed in unthinned plants. However, significant changes are observed in the components, osmotic potential and turgor pressure. The osmotic potential of the skin tends to decrease compared to the flesh tissue from the end of February, coinciding with the appearance of purple spot symptoms. At this time, a sudden drop in skin osmotic potential is observed ( Figure 2B ), corresponding to an abrupt increase in turgor pressure ( Figure 2C ) within the skin tissue. The drop in osmotic potential observed at color break in fruit from thinned trees ( Figure 2B ) suggests that either water uptake lags behind sugar accumulation in the skin or there is a loss of water from the skin to the flesh in fruit from thinned trees. This dehydration is a characteristic symptom of purple spot (Gariglio et al., 2002).

Significantly, the stress associated with growth according to the epidermal growth control hypothesis is evident in the solute potential and turgor pressure of the skin tissue rather than in the flesh. This distinction is noteworthy as most studies of fruit growth tend to focus on the water potential and its component in the flesh tissue.

Similar to the modulation observed in other fleshy fruits, the primary water import pathway of grape (Vitis sp.) berries undergoes a transition from xylem to phloem at the onset of veraison. Consequently, berry water uptake through the xylem is reduced, although the structural integrity of the berry xylem remains unaffected and unhindered throughout the ripening process (Zhang and Keller, 2016).

Sucrose is unloaded from the phloem in sink tissues either apoplasmically or symplasmically (Ruan, 2014). Apparently, there is a change in the primary water transport pathway in grape berries with a transition from symplastic to apoplastic phloem unloading at the onset of ripening (Zhang et al., 2006). This hypothesis was confirmed by studying the movement of xylem mobile dyes in grape berries and root pressure treatments. Modeling studies have shown that some phloem-derived water contributes to both berry growth and transpiration, with the excess being recirculated through the xylem (xylem reflux) ( Figure 3 ). Restricting the release of water through the xylem and/or the skin limits the accumulation of solutes in the berry and its color change. This mechanism can be viewed as a strategy to increase the berry sink strength and promotes normal grape ripening and may be particularly important during periods of rapid sugar accumulation and under environmental conditions that limit berry transpiration (Zhang and Keller, 2016).

At the color break stage, loquat fruits exhibit their highest growth rate, which is 2.5 times faster in fruits from plants thinned at one fruit per panicle compared to unthinned plants (Gariglio et al., 2003a). The concentration of total soluble carbohydrates in flesh tissue (over dry mass) at color break was also 3.52 times higher in plants thinned at one fruit per panicle compared to unthinned plants (Gariglio et al., 2003a). In peel tissue, the concentration of total soluble carbohydrates at color break is less than a quarter of that in flesh tissue. In grapes, the ratio of sugar concentration between flesh and skin tissue is in the range of 2 to 3 during fruit development (Zhu et al., 2017).

In this context, the progressive weakening of the peel tissue observed in loquat fruits ( Figures 1 , 2 ) can be attributed to the complex biochemical changes associated with the onset of ripening. This phenomenon facilitates the influx of water with minimal restriction, given the low turgor and relatively constant osmotic potential of the flesh tissue at color break ( Figure 4 ), despite the increased sugar accumulation (expressed as dry weight). This phenomenon explains the exponential growth of loquat fruit from the onset of color break to harvest.

As a result of the expansion of the flesh, the cell volume of the external tissues is compressed, leading to an increase in their turgor and, consequently, their water potential ( Figure 4 ). The equilibrium is restored by the migration of water from the outer tissues (higher water potential) to the inner tissues (lower water potential). As a result, there is a gradual decrease in osmotic potential (due to water loss) and a simultaneous increase in turgor in the skin of loquat fruits. In some cases, the intensity and duration of this growth stress can lead to irreversible cell dehydration and collapse of the skin cells, resulting in symptoms of purple spot ( Figure 4B ). The extreme values of the osmotic potential recorded in the fruit skin during pre-down measurements are −2.5 MPa (Gariglio, 2001; Gariglio et al., 2008a).

In low crop load conditions, the osmotic potential of the peel tissue is dependent on the rate of sugar accumulation, the sugar concentration gradient (dry basis) between the flesh and the peel, and the intensity of the fruit growth rate. The combined effect of these factors determines the risk of irreversible peel dehydration (Gariglio et al., 2008a, b). Conversely, in situations of high crop load, there is no correlation between the peel osmotic potential and any of the aforementioned factors. This indicates that the intensity of these factors is insufficient to influence the peel’s hydration status (Gariglio, 2001; Gariglio et al., 2008a). In contrast to the previous result, a more negative osmotic potential of the flesh compared to the skin tissue was observed in sweet cherry (Grimm and Knoche, 2015). In the case of tomatoes, the osmotic potential of the flesh was found to be slightly lower than that of the skin tissue. Additionally, a low cell turgor (0.04-0.08 MPa) was observed in the skin (Ikeda et al., 1999; van Ieperen et al., 2005).

The physiological significance of cell turgor pressure differs between flesh and skin tissue. In flesh, turgor pressure is generated by osmotic water uptake ( Figure 4 ), which is a consequence of sugar accumulation (Ortega, 2023). Additionally, since the internal tissue is limited in its expansion, it is in a state of active tension in accordance with the growth control hypothesis (Vines, 1886; Kutschera and Niklas, 2007).

In the outer tissues, the pressure exerted by the expanding flesh tends to reduce the skin cell volume ( Figure 4 ), which indirectly increases turgor. Consequently, the skin tissue is in a state of passive tension (Vines, 1886; Kutschera and Niklas, 2007). In light of these concepts, it can be argued that in tissues that are in a state of active tension, the flesh, the turgor pressure should be referred to as “active turgor pressure” or “true turgor pressure”. Conversely, in external tissues, turgor results from a decrease in cell volume due to the driving forces of the internally expanding tissue. In this case, turgor should be referred to as “passive turgor pressure” or “false turgor pressure” ( Figure 4 ). In the case of loquat fruit, passive turgor pressure is responsible for maintaining a high water potential of the skin cells even as the tissue dehydrates under conditions predisposing to purple spot (Gariglio et al., 2008a).

In sweet cherry, an osmotic potential gradient of 1.1 MPa was observed between the fruit skin and its flesh, with the osmotic potential being more negative in the flesh (Grimm and Knoche, 2015). The low turgor pressure measured in the flesh (Schumann et al., 2014) establishes a significant water potential gradient from skin to flesh (Grimm and Knoche, 2015). However, the process by which water potential equilibrium is achieved between the two tissues remains unclear. There is currently a lack of data available on apoplastic phloem unloading (Zhang et al., 2006) and the xylem reflux mechanisms (Zhang and Keller, 2016) in different species. It is therefore possible that a significant proportion of the solutes measured in the extracted juice from sweet cherry pulp (Grimm and Knoche, 2015) are apoplastic and therefore may not accurately represent the osmotic potential of the pulp tissue. In tomato, the study by Ikeda et al. (1999) showed no water potential gradient between flesh and skin tissue during the pre-dawn period. Nevertheless, an osmotic gradient was observed between the apoplastic and symplastic spaces, which could facilitate substantial water import into the cells. In practice, however, this import did not occur. This may be attributed to the presence of matrix potentials in the apoplastic space, resulting from capillary forces within the cell walls (van Ieperen et al., 2005).

The manuscript by Gariglio et al. (2008a) presents an original hypothesis that sheds light on the changes in water relations between the flesh and skin tissues of developing loquat fruits, and the prospective emergence of physiological disorders. In the present manuscript, the explanation and definition of the presence of passive turgor in the peel tissue is also and original physiological concept. The hypothesis and the introduction of the concept of passive turgor are of great importance as they provide a novel contribution to the understanding of the regulation of water dynamics in fruit under different crop load conditions.

This review puts forth a hypothesis regarding the alterations in water relations between internal and external tissues in developing fruit, situated within the context of the epidermal growth control hypothesis. The pressure potential of the outer tissues is a consequence of the expansion of the inner tissues, rather than osmotic water uptake, as observed in the flesh. This results in a false turgor, which, when considered with the osmotic potential of the skin, can be used as an indicator of the level of growth stress and the risk of skin cell dehydration. The primary factors identified as potential determinants of the water balance between internal and external tissues include the growth rate of the fruit, the gradient of sugar concentration (on a dry weight basis) between the flesh and skin tissues, the proportion of apoplastic and symplastic solutes, and the strength of the skin. These factors render the fruit susceptible to physiological disorders associated with water relations. While the relevance of skin water potential and its components to the hypothesis presented may be validated in future research, it is recommended that they be included in studies of physiological disorders under different predisposing conditions to improve our understanding of the physiological process involved.