Indexing melon physiological decline to fruit quality and vine morphometric parameters

Scientia Horticulturae 203 (2016) 207–215

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Indexing melon physiological decline to fruit quality and vine morphometric parameters Georgios A. Soteriou a , Lambros C. Papayiannis b , Marios C. Kyriacou c,∗ a b c

Vegetable Science Laboratory Plant Pathology Laboratory Postharvest Technology Laboratory, Agricultural Research Institute, Nicosia, Cyprus

a r t i c l e

i n f o

Article history: Received 30 November 2015 Received in revised form 22 March 2016 Accepted 23 March 2016 Available online 31 March 2016 Keywords: Cucumis melo L. Melon decline Carbohydrates Internode Intespecific hybrid rootstocks Hypocotyl Texture

a b s t r a c t While grafting cucurbits has become essential in the management of soil borne diseases and for improving performance under conditions of abiotic stress, commercial melon grafting has been curbed by the incidence of non-pathological decline, usually expressed right before harvest and attributed to physiological scion-rootstock incompatibility. The current study investigated the potential indexing of physiological incompatibility to plant performance, vine morphometric parameters and fruit physicochemical characteristics. Graft combinations sensitive to physiological incompatibility (scions: melon cvs. Elario, Polynica and Raymond; rootstocks: interspecific hybrids ‘TZ148’, ‘N101’, ‘Carnivore’ and ‘30900’) were grown between February and May in a disease-free soil environment. Results indicate that plant collapse shortly before harvest is a onetime event that does not necessarily reflect on the performance of the asymptomatic, surviving plants. However, a negative rootstock effect on scion dry weight was indicative of rootstock-scion combinations subject to incompatibility and prone to decline. The attenuation of the 1st internode’s diameter relative to the hypocotyl’s (‘Elario’ 29.1%; ‘Raymond’ 41.5%; ‘Polynica’ 44.0%) and the loss of mesocarp firmness reflected the categorical sensitivity of the scions to physiological decline. No systematic pattern was identified connecting fruit soluble carbohydrate (fructose, glucose and sucrose) content to physiological incompatibility and plant decline. However, earliness of harvest maturity, pronounced in sensitive climacteric scions ‘Polynica’ and ‘Raymond’, may relate to ethylene-mediated comprehensive acceleration of ripening stressing rootstock-scion synergy to collapse. © 2016 Elsevier B.V. All rights reserved.

1. Introduction Owing to the environmental restrictions imposed on the use of chlorofluorocarbon-based soil fumigants, grafting cucurbits has become an essential tool in the management of soil borne disease, employing most notably the resistance of inter-specific Cucurbita maxima × C. moschata rootstocks. Further to conferring phytoprotective resistance, a resilient rootstock system may augment crop yield by enhancing nutrient uptake (Colla et al., 2010a) and water use efficiency (Rouphael et al., 2008). Increase in yield has also been attributed to stimulation of scion growth (San Bautista et al., 2011) mediated by increased synthesis and translocation of endogenous hormones from the acquired root (Dong et al., 2008). Under conditions of edaphoclimatic stress the exploitation of resilient rootstocks may sustain high yields by exploiting traits such as

∗ Corresponding author. E-mail address: [email protected] (M.C. Kyriacou). http://dx.doi.org/10.1016/j.scienta.2016.03.032 0304-4238/© 2016 Elsevier B.V. All rights reserved.

increased tolerance to salinity (Colla et al., 2006; He et al., 2009; Romero et al., 1997), to temperature extremes (Rivero et al., 2003; Schwartz et al., 2010) and soil alkalinity (Colla et al., 2010b), and heavy metal contaminations (Kumar et al., 2015a,b). The use however of grafted plants in commercial melon production is not the one anticipated. This can be attributed partly to the higher cost incurred but also to frequent problems of incompatibility involving inter-specific Cucurbita rootstocks, which may eventually lead to plant decline (Aloni et al., 2010). The rather inconsistent information available on the performance of melon scions grafted onto Cucurbita rootstocks constitutes an additional disincentive for grafting. In fact, the ultimate conclusion of several reports is that incompatibility in melon is foremost subject to the dynamics of rootstock/scion interaction (Aloni et al., 2010; Cohen et al., 2002; Davis et al., 2008; Rouphael et al., 2010), in contrast to watermelon where compatibility of squash inter-specific hybrid rootstocks is usually unequivocal across commercial scions (Kyriacou and Soteriou, 2015; Soteriou and Kyriacou, 2015).

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Incompatibility expressed as graft failure and premature seedling death may be caused by unfavorable conditions (e.g. temperature) prevailing during the connection of vascular bundles between scion and rootstock at the nursery (Davis et al., 2008). Andrews and Marquez (1993) referred discretely to physiological incompatibility which results in plant decline either at the early stages after transplanting a healthy grafted seedling, or as late as the beginning of harvest. Such incompatibility has been related to oxidative stress triggered by hormonal signaling from the scion to the rootstock (Aloni et al., 2008a).Moreover, the effect of factors (e.g. high air temperature) suspected of triggering physiological incompatibility is random, causing decline only to a variable fraction of the presumably incompatible scion-rootstock population. Limited evidence exists linking the effect of grafting on plant morphological characteristics to the incidence of plant decline (Edelstein et al., 2004; Traka-Mavrona et al., 2000). Hence, extensive field trials are still essential before recommending specific scion × rootstock combinations. Conflicting reports regarding the effect of grafting on melon fruit quality characteristics (Colla et al., 2006, 2010a; Crinò et al., 2007; Trionfetti-Nisini et al., 2002), can be explained partially on the basis of variation in the prevailing production practices (Rouphael et al., 2010). It is possible however that latent, non-pathological incompatibility elicited by rootstock-scion interaction under triggering stress conditions may impair certain fruit quality traits or plant morphometric characteristics in the surviving population. This hypothesis is underpinned by the association of incompatibility with changes in water and nutrient flow through the graft union potentially causing plant wilting (Davis et al., 2008; Rouphael et al., 2010). An etiology for melon physiological incompatibility and plant decline proposed by Aloni et al. (2008a) is the occurrence of hormonal imbalance in the rootstock after the establishment of vascular connections. Any potential stress imposed by self-grafting has never been found deleterious on melon plants, provided a healthy transplant stock; neither any positive or negative effects of self-grafting on yield and quality traits have been reported (Guan et al., 2014; Guan et al., 2015; Schultheis et al., 2015; Zhao et al., 2011). Indexing physiological incompatibility to fruit quality and vegetative morphometric parameters in the surviving plant population, may provide useful tools for diagnosing latent incompatibility and identifying rootstock-scion combinations prone to decline. It may also enhance our understanding of the physiological mechanisms behind grafted melon plant decline. Accordingly, the objective of the current work has been to investigate the possible expression of physiological incompatibility in plant performance and morphometric parameters, and in fruit physicochemical characteristics. For the purposes of this study scion-rootstock combinations sensitive to physiological incompatibility were selected on the basis of preliminary experimental results and extension service reports on grafted melon decline incidence. Scion selection in particular was based on the relative frequency of physiological incompatibility incidence prior to harvest as follows: (a) infrequent plant mortality – cv. Elario; (b) frequent plant mortality – cv. Raymond; (c) ubiquitous symptomatic incompatibility and persistent plant mortality – cv. Polynica on inter-specific rootstock ‘N101’.

2. Materials and methods 2.1. Plant material Transplants of melon (Cucumis melo L.) Galia type cv. Elario and Ananas type cv. Raymond were grafted onto four inter-specific (Cucurbita maxima Duchesne × C. moschata Duchesne) hybrid rootstocks: ‘TZ148’, ‘N101’, ‘Carnivore’ and ‘30900’. Moreover, Galia

Table 1 Incidence of plant decline in grafted and non-grafted melon plants of cultivars Elario, Polynica and Raymond (N = 30). Cultivar

Rootstock

Plant Decline (%)

Elario

TZ148 N101 Carnivore 30900 Non-grafted TZ148 N101 Carnivore 30900 Non-grafted

3.3 0.0 3.3 3.3 0.0 n.s. 23.3 a 36.7 a 16.7 a 30.0 a 0.0 b

N101 Non-grafted

20.0 b 0.0 a

Raymond

Polynica

Pearson Prob > ChiSq

0.3839

0.0069*

0.0112*

* Signicant rootstock effect according to Pearson ChiSquare test (P ≤ 0.05). Scionspecific values within columns followed by the same letter are not significantly different according to Pearson ChiSquare test (P ≤ 0.05).

type cv. Polynica was grafted solely onto ‘N101 since preliminary work and commercial experience showed that, under stressful temperature conditions, simulated in the controlled environment of the current study, cv. Polynica grafted on common commercial C. maxima × C. moschata rootstocks, such as TZ148, N101, Carnivore and 30900, demonstrates a persistent morphological incompatibility that results in significant plant decline. Grafts were made after the appearance of the first true leaf on seedlings using the approach method (Lee et al., 2010). The root system of the scion was removed about 15 days after grafting. Non-grafted (self-rooted) plants were used as the control. All plants were produced by Solomou Nurseries (Nicosia, Cyprus). Potted plants were spaced 0.5 m apart within rows and 1.0 m apart between rows amounting to a plant density of 20,000 plants ha−1 . 2.2. Experimental conditions The study was conducted at the Zygi Experimental Station (34◦ 00 N; 33◦ 20 15 E) of the Agricultural Research Institute of Cyprus between February and May 2014 in a 400 m2 greenhouse covered with double-layer polyethylene sheet and insect proof windows. A climate controller system (Galileo, Galcon, Kfar Blum, Israel) regulated maximum air temperature set at 28 ◦ C during the day and night air temperature was kept above 15 ◦ C, until the oldest fruits (first to harvest) reached a diameter of about 8 cm. Subsequently, maximum day and minimum night greenhouse temperatures were raised to 32 and 19 ◦ C, respectively, for better simulation of summer field conditions. A mixture of perlite and peat moss (in 1:2 vol ratio) was used as the growing medium in 30.0 l black plastic pots (height = 0.30 m; diameter = 0.40 m). Soil moisture sensors (Mas-1, Decagon Devices, Pullman, USA), calibrated for the specific medium and connected to the climate control system, were established for monitoring pot volumetric water content. Daily irrigation was regulated to allow 20–30% leaching of the total irrigation volume per plant per day. Pressure-compensated drip irrigation at a rate of 4 l h−1 per plant was used in combination with constant nutrient feeding. The nutrient solution formula, based on variable N levels at different stages of plant growth, was adopted from Rodriguez et al. (2006). All treatments were irrigated and fertilized uniformly. Plants were trellised in a single-truss system and allowed to reach the next plant within row on the horizontal trellis cable before they had their tops pruned. All lateral shoots below the 6th node were removed. Lateral shoots from the 7th node and above were allowed to set fruit on the 1st or 2nd node and pruned past two nodes. When fruits reached approximately the size of a lemon, 44

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Table 2 Mean comparison for fresh weight, dry weight, rootstock and scion diameter of melon cultivars Elario and Raymond grown non-grafted or grafted on rootstocks TZ148, N101, Carnivore and 30900, and cultivar Polynica grown non-grafted or grafted on rootstock N101. Source of variance

Fresh weight (g)

Dry weight (g)

Hypocotyl diameter (cm)

1st Internode diameter (cm)

1st Internode/hypocotyl diameter

Elario TZ148 N101 Carnivore 30900 Non-grafted

800.0 786.7 946.7 813.3 913.3 n.s.

149.7 143.3 168.0 149.0 168.3 n.s.

1.8 1.7 1.9 1.7 1.5 n.s.

1.5 ab 1.3 b 1.5 ab 1.4 ab 1.7 a

0.82 b 0.79 b 0.84 b 0.81 b 1.15 a

*

**

600.0 553.3 586.7 700.0 766.7

125.0 117.7 123.7 139.0 151.0

1.9 a 1.9 a 1.9 a 1.9 a 1.1 b

0.81 b 0.78 b 0.71 b 0.72 b 1.29 a

*

**

***

1.5 1.4 1.4 1.4 1.4 n.s.

453.3 b 1046.7 a

109.3 b 190.0 a

1.0 b 1.8 a

0.65 b 1.16 a

***

**

1.5 1.5 n.s.

***

***

Raymond TZ148 N101 Carnivore 30900 Non-grafted Polynica N101 Non-grafted

***

n.s. non-significant effect. Scion-specific values within columns followed by the same letter are not significantly different according to Tukey-Kramer HSD Test (P ≤ 0.05). * Significant effect at the 0.05 level. ** Significant effect at the 0.01 level. *** Significant effect at the 0.001 level.

they were enclosed in a plastic net supported by lateral strings. Standard pest and disease control practices were applied. Pollination was facilitated by placing a hive of 35–40 bumble bees in the centre of the greenhouse three weeks after planting. Experimental treatments were arranged in a completely random design with 30 plants per treatmentHarvest indices applied included clear abscission of the fruit from the plant by applying light pressure (3/4 slip), yellowing of the skin, raised netting and aroma development. Three fruits from each surviving plant were sampled for quality assessment, beginning at 14 weeks after planting. 2.3. Fruit quality and vine morphometric analyses Fruits were weighed and equatorial and meridian diameters determined at harvest and shape index was calculated. Marketable yield and number of fruits per surviving plant were also determined. Assessment of quality was performed on the day of harvest. Fruit rind and flesh thickness were measured at two representative points on each fruit cross-section using an electronic calliper. Flesh firmness and fruit hardness were measured using a TA·XT plus Texture Analyser (Stable Micro Systems, Surrey, UK) equipped with a 50 kg load cell. Flesh firmness was recorded as the maximum resistance force to penetration performed at two points per sectioned fruit in the direction from the locular interior toward the periphery; penetration was performed with a flat, 6-mm diameter probe mechanically operated at a speed of 2 mm s−1 in a displacement depth of 10 mm. Fruit hardness was determined by a compression test, corresponding to 3% strain, applied once on each fruit using a 75-mm flat probe at a test speed of 2 mm s−1 . Flesh color was measured at two intermediate loci on the mesocarp of each cross-sectioned fruit using an 8 mm-aperture Minolta CR-400Chroma Meter (Minolta, Osaka, Japan). Measurements were performed in the CIELAB color space (McGuire, 1992), and recorded parameters were lightness (L*), color components a* (+red/−green) and b*(+yellow/−blue), chroma [C* = (a*2 + b*2 )1/2 ] and hue angle [h◦ = arctan (b/a)]. Sensory analysis for overall organoleptic acceptability was performed by a trained panel based on 1–5 hedonic scale (1 = dislike, 3 = neither like nor dislike, 5 = like). From each cross-sectioned fruit mesocarp tissue samples of about 100 g were obtained using a core borer and were lyophilised to constant weight on a Christ, Alpha 1–4 (Osterode, Germany) freeze drier to determine dry matter content. About 50 g were similarly obtained and homogenized under low speed. Part of

the homogenate was filtered through double cheesecloth and the soluble solids content (SSC) at 20 ◦ C of the filtered juice was measured on a digital refractometer (RFM870; Bellingham-Stanley Ltd., Kent, UK). Part of the homogenate was transferred to 50 ml falcon tubes, instantly frozen in liquid nitrogen, and then stored at −80 ◦ C for phytochemical analyses. Analysis of non-structural carbohydrates (glucose, fructose, and sucrose) in juice was performed by liquid chromatography on an Agilent HPLC system (Agilent Technologies, Santa Clara, CA, USA) equipped with 1200 Series quaternary pump and a 1260 Series refractive index detector commanded by ChemStation software. Separation was performed on a 4.6 × 250 mm carbohydrate column at 35 ◦ C (Waters, Milford, MA, USA) using an acetonitrile:water (75:25) mobile phase at a flow rate of 1.4 ml min−1 . Quantification was performed against fructose, glucose and sucrose external standard calibrating curves with a coefficient of determination (R2 ) > 0.9999. Recovery trials performed under the same operating conditions were in all cases on the order of 100%. Plant hypocotyl and 1st internode diameters were measured with an electronic vernier calliper at about ten days before the end of the fruiting cycle. Measurements for plant fresh and dry weights were recorded for all non-declined plants at the end of the experiment. Plant dry weight was determined by drying plants in an oven at 65 ◦ C for 72 h, whereby the dry weight had stabilised. 2.4. Detection of plant pathogens All plants were visually inspected at weekly intervals for the presence of symptoms that could be attributed to plant pathogens. Identification of fungal infections in symptomatic plants was performed either directly by microscopic examination or indirectly by isolation onto potato dextrose agar (PDA) culture. For that purpose, plant tissues were excised and surface-sterilized in 1% sodium hypochlorite, followed by two rinses, 1 min each, in sterile water. Small pieces of tissue were plated directly onto potato dextrose agar and incubated at 20 ◦ C for up to 10 days or until sufficient growth or sporulation enabled identification of fungal infection. Previously reported molecular based assays and markers were used for the detection of soilborne vascular wilt diseases (Haegi et al., 2013; Luongo et al., 2012). For the detection of plant viruses, a young fully developed leaf from each plant was collected and tested by the standard double-antibody sandwich (DAS), Enzyme linked immuno-sorbent assay (ELISA) using polyclonal antibodies against

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Table 3 Mean comparison for plant yield, fruit number, fruit mean weight, fruit shape (meridian/equatorial diameter) and period to 1st harvest (harvest maturity) of melon cultivars Elario and Raymond grown non-grafted or grafted on rootstocks TZ148, N101, Carnivore and 30900, and cultivar Polynica grown non-grafted or grafted on rootstock N101. Source of variance

Yield (kg/ plant)

Fruits per plant

Fruit weight (kg)

Fruit Shape (M/E)

Harvest maturity (days)

Elario TZ148 N101 Carnivore 30900 Non-grafted

5.6 5.7 5.6 6.0 6.7 n.s. 6.6 6.5 6.6 6.2 6.8 n.s. 6.0b 7.3a

5.2 5.6 5.4 5.8 5.9 n.s. 3.1 3.0 3.3 2.9 3.5 n.s. 5.8 6.5 n.s.

1.118 1.068 0.979 1.017 1.003 n.s. 2.151 2.195 2.016 2.194 2.098 n.s. 0.912b 1.084a

1.0 1.1 1.1 1.1 1.1 n.s. 1.5 1.5 1.4 1.5 1.5 n.s. 1.0 1.0 n.s.

103 a 101 ab 97 b 97 b 99 ab

Raymond TZ148 N101 Carnivore 30900 Non-grafted Polynica N101 Non-grafted

**

*

**

104b 105b 104b 104b 112a ***

94b 101a ***

n.s. non-significant effect. Scion-specific values within columns followed by the same letter are not significantly different according to Tukey-Kramer HSD Test (P < 0.05). * Significant effect at the 0.05 level. ** Significant effect at the 0.01 level. *** Significant effect at the 0.001 level.

plant viruses which have been previously recorded in Cyprus, including the aphid borne viruses Zucchini yellow mosaic virus (ZYMV), Papaya ringspot virus (PRSV-W), Watermelon mosaic virus (WMV), Cucurbit aphid-borne yellows virus (CABYV) and Cucumber mosaic virus (CMV), as well as the seed transmitted Cucurbit green mottle mosaic virus (CGMMV) (Papayiannis et al., 2005). Plants were also tested for the presence of two whitefly transmitted Crinivirus species that have been reported as associated with yellowing symptoms and vein clearing in cucurbit crops, Cucurbit yellow stunting disorder virus (CYSDV) and Cucumber vein yellowing virus (CVYV) using reverse transcription (RT) and polymerase chain reaction (PCR) and species specific primers and probes (Papayiannis et al., 2005,2010). Virus positive and negative controls were included in both serological and molecular tests. 2.5. Statistical analysis Contingency analysis and Pearson Chi-square tests were run to compare plant survival percentage on each rootstock with respect to non-grafted plants within each cultivar. Yield and quality data for each cultivar were subjected to one way analysis of variance (ANOVA) and means were separated by the Tukey-Kramer honestly significant difference (HSD) test. Principal component analysis (PCA) using the correlation matrix was performed on plant survival, morphological, yield, physical, chemical and phytochemical quality components to explore relationships among variables and treatments and also to determine which traits were the most effective for discriminating rootstock and cultivar combinations. PCA outputs included treatment component scores and variable loadings against each selected component. The first three principal components (PC1, PC2 and PC3) were selected for the ordination analysis, and the correlation between the original variables and the respective PC was calculated. The PCs with eigenvalues greater than 1 were selected (Dunteman, 1989), and loadings greater than |0.6| indicated significant correlations between the original variables and the extracted components (Matus et al., 1996). Analyses and calculations were carried out using the appropriate functions within SAS (SAS Institute, Inc., Cary, NC). 3. Results Shortly before harvest 20.0% of cv. Polynica plants grafted on ‘N101’ and 16.7–36.7% of cv. Raymond plants, depending on rootstock, were lost to wilting whereas no losses were incurred on non-grafted plants (Table 1). In cv. Elario plant decline was

observed on rootstocks ‘TZ148’, ‘Carnivore’ and ‘30900’ but it was limited and statistically non-significant. Plant wilting for all cultivars (Raymond, Elario, Polynica) was determined as of nonpathogenic origin. In the current work, unless otherwise specified, all subsequent reference to plant and fruit characteristics evaluated will relate strictly to the non-declined plants, i.e. the plants that survived their complete fruiting cycle. Fresh plant (scion) weight of cv. Raymond was significantly depressed only when grafted on rootstock ‘N101’ (Table 2). Irrespective of rootstock, grafted plants of cv. Raymond did not display visual symptoms of incompatibility and exhibited moreover the same vegetative growth as the non-grafted plants. By contrast, all cv. Polynica plants that survived grafting, exhibited explicit symptoms of incompatibility manifested as morphological asymmetry at the graft union and as reduced scion fresh weight. All rootstocks grafted with scions Raymond and Polynica had a negative effect on plant dry weight. In the case of cv. Polynica, plant dry weight reduction in relation to non-grafted plants was more severe (42.5%) than in cv. Raymond (7.9%–22.0%, depending on rootstock). Contrary to the above, plant fresh and dry weights of cv. Elario were not affected by any of the rootstocks used. Hypocotyl diameter of cv. Raymond was increased by grafting on all rootstocks but remained unaffected in cvs Elario and Polynica. Inversely, rootstock ‘N101’ reduced the 1st internode diameter of cvs Elario and Polynica but not of cv. Raymond. In all cultivars, the stem diameter quotient (1st internode/hypocotyl diameter) of the non-grafted plants was significantly higher compared to the grafted plants. Lower plant yield by 17.5% and fruit weight by 11.5% was observed for cv. Polynica grafted on ‘N101’ (Table 3). In contrast, yield, number of fruits per plant and fruit weight of cvs Elario and Raymond was not differentiated between grafted and nongrafted plants. No differences in fruit shape between grafted and non-grafted plants were observed in all cultivars. Harvest maturity for grafted plants of ‘Polynica’ and ‘Raymond’ was reached at 94 and 104–105 days after transplanting, respectively, whereas for non-grafted plants of both cultivars harvest maturity commenced 7–8 days later. On the contrary, harvest maturity for ‘Elario’ had similar timing on grafted (97–103 days) and non-grafted plants (99 days). Rind thickness was increased significantly (40.2%) only when cv. Elario was grafted on rootstock ‘TZ148’; flesh thickness in cvs Elario and Raymond was unaffected by grafting. By contrast, rind and flesh thickness of cv. Polynica grafted on ‘N101’ were decreased by 34.0% and 11.2%, respectively (Table 4). Loss of flesh firmness

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Table 4 Mean comparison for fruit rind and flesh thickness, flesh firmness, fruit hardness, skin netting and flesh CIELAB color components of melon cultivars Elario and Raymond grown non-grafted or grafted on rootstocks TZ148, N101, Carnivore and 30900, and cultivar Polynica grown non-grafted or grafted on rootstock N101. Source of variance

Elario TZ148 N101 Carnivore 30900 Nongrafted Raymond TZ148 N101 Carnivore 30900 Nongrafted Polynica N101 Nongrafted

Rind (mm)

Flesh thickness (mm)

Flesh firmness (kg)

Fruit hardness (kg-force)

Fruit skin netting

Lightness L*

Chroma C*

Hue angle h◦

Component a*

Component b*

(1–5)

(0–100)

(a*2 + b*2 )1/2

(0–360◦ )

(0–60)

(0–60)

11.57 a

37.94

5.68 b

14.67

2.78

71.61 a

29.13b

109.59

−9.80

27.60 bc

8.93 ab 9.35 ab 9.55 ab 8.26 b

36.53 34.53 34.13 35.16

5.55 b 6.34 b 5.74 b 8.32 a

14.65 14.52 14.66 14.82

2.97 3.29 3.38 3.04

69.42 ab 71.03 a 69.81 ab 68.68 b

30.80ab 31.67a 31.38a 29.07b

110.26 110.27 110.01 110.81

−10.71 −11.00 −10.76 −10.36

28.86 abc 29.68 a 29.47 ab 27.15 c

n.s. 41.19

***

n.s. 3.24 ab

***

72.23

16.11ab

n.s. 105.47

n.s. −4.34

***

3.42 ab

n.s. 15.49 bc

**

12.66 11.65 11.48 11.34 10.44

43.19 42.17 45.24 42.16

2.88 b 2.95 b 3.96 ab 4.41 a

15.97 abc 14.92 c 17.93 ab 18.61 a

2.96 b 3.23 ab 3.19 b 3.86 a

73.13 71.35 72.79 72.08

16.74ab 17.41a 15.72b 16.49ab

104.59 106.24 105.36 106.17

−4.22 −4.89 −4.19 −4.62

16.18 ab 16.70 a 15.12 b 15.81 ab

n.s. 7.23 b

n.s. 32.80b

***

**

**

3.71a

26.69 b

n.s. 111.82 a

n.s. −9.95

*

10.08 b

n.s. 70.48

*

3.54 b

10.97 a

36.92a

6.70 a

15.30 a

3.00b

69.43

29.61 a

110.61 b

−10.45

27.70 a

***

***

***

***

***

n.s.

***

***

n.s.

***

15.41ab

24.76 b

n.s. non-significant effect. Scion-specific values within columns followed by the same letter are not significantly different according to Tukey-Kramer HSD Test (P < 0.05). * Significant effect at the 0.05 level. ** Significant effect at the 0.01 level. *** Significant effect at the 0.001 level.

was evident in the fruits of cv. Elario grafted on all rootstocks, in the fruits of cv. Raymond grafted on ‘N101’ and ‘Carnivor’, and in the fruits of cv. Polynica grafted on ‘N101’ (Table 4). Cultivar Raymond produced fruits with flesh as firm as the non-grafted plants when grafted on ‘TZ148’ and ‘30900’. The results of the present study revealed a significant correlation between whole fruit hardness and flesh firmness in the case of cv. Raymond (r = 0.51, p < 0.001) and cv. Polynica (r = 0.551, p < 0.001) but not for cv. Elario. Further analysis of data revealed that only rootstock ‘Carnivor’ combined with cv. Raymond and ‘N101’ combined with ‘Polynica’ reduced both whole fruit hardness and flesh firmness, in relation to non-grafted plants. No differentiation was observed among grafted and non-grafted plants of cv. Elario as regards fruit hardness. Fruit skin netting of cv. Raymond was decreased only when grafted on rootstocks ‘N101’ and ‘30900’. In contrast grafting cv. Polynica on ‘N101’ increased the netting on fruits. Cultivar Elario fruit skin netting was not affected by grafting. With respect to fruit coloration, cv. Elario grafted on ‘TZ148’ and ‘Carnivore’ yielded fruits of lighter flesh color (higher lightless values) than the non-grafted control (Table 4). Rootstocks ‘Carnivore’ and ‘30900’ affected positively the intensity of yellow color (b*) as well as the overall intensity of flesh color (C*) in cv. Elario. By contrast, in cv. Polynica flesh color component b* and overall chroma (C*) were decreased while hue angle (h◦ ) increased when grafted on ‘N101’ depicting a shift toward greenish-yellow. Rootstocks differentiated the content of the pulp in fructose, glucose, sucrose and total sugars of the three cultivars with respect to the non-grafted plants (Table 5). Pulp fructose content was lowered (20.8%) only when cv. Elario was grafted on rootstock ‘N101’ and increased only when cv. Raymond was grafted on ‘TZ148’ (25.7%), ‘Carnivore’ (29.1%) and ‘30900’ (43.9%). Glucose content was suppressed by 22.6% only when cv. Polynica was grafted on rootstock ‘N101’ and increased by 29.1% and 43.6% when cv. Raymond was grafted on rootstocks ‘Carnivore’ and ‘30900’, respectively. Sucrose content was lowered after grafting cv. Raymond on rootstocks ‘TZ148’ and ‘30900’ and increased only when cv. Elario was grafted on ‘N101’. Total sugar content was decreased only by grafting

cv. Polynica on ‘N101’ and increased only in cv. ‘Elario’ when grafted on rootstock ‘Carnivore’. Moreover, sweetness index (SI) was improved only for cv. Elario grafted on ‘Carnivore’ and cv. ‘Raymond’ grafted on ‘Carnivore’ or ‘30900’. The rest of the rootstocks did not have an effect on the SI of three scion cultivars. Consistent improvement of flesh SSC (Soluble Solids Content) with grafting was noted only for cv. Elario on all rootstocks. Suppression of the SSC was observed only for cv. Raymond when grafted on ‘30900’, whereas on all other rootstocks cv. Raymond remained unaffected (Table 5). The SSC of cv. Polynica were not altered by grafting. Dry matter content (DM) was improved only when cv. Elario was grafted on ‘N101’. Depression of DM was observed when cvs Raymond and Polynica were grafted on rootstocks ‘30900’ and ‘N101’, respectively. Regarding the sensory value of the fruits, grafting irrespective of rootstock yielded fruits of equal or better scores than the non-grafted plants. Specifically, in cv. Elario all rootstocks except ‘30900’ improved the sensory score; in cv. Raymond the sensory score was improved only on rootstock ‘N101’ but remained unaltered on all other rootstocks, and in cv. Polynica the sensory score was also improved by grafting on ‘N101’. The first three Principal Components (PC) derived from the collective analysis of means were associated with eigenvalues greater than 1; they explained 85.8% of the total variance, with PC1 accounting for 57.8% and PC2 for 18.4% (Table 6). PC1 was positively and significantly correlated with rind and flesh thickness, fruit hardness, flesh color lightness (L*), SSC, sucrose and total sugars content, percentage of plant decline, and fruit weight. It was negatively correlated with flesh firmness, flesh chroma C*, hue angle (h◦ ), component b*, the two monosaccharides, and plant fresh and dry weights. PC2 was positively correlated with fruit hardness, plant fresh and dry weights and the stem diameter quotient. Moreover, PC3, which explained 9.6% of total variance, was positively correlated with fruit dry matter content and negatively correlated with the stem diameter quotient and plant yield. In the loading plot (Fig. 1A) presenting the relationships among variables, it is demonstrated that the percentage plant decline is positively correlated

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Table 5 Mean comparison for fruit pulp fructose, glucose, sucrose and total sugar content, sweetness index, soluble solids and dry matter content and taste score of melon cultivars Elario and Raymond grown non-grafted or grafted on rootstocks TZ148, N101, Carnivore and 30900, and cultivar Polynica grown non-grafted or grafted on rootstock N101. Source of variance

Fructose (␮g/g)

Glucose (␮g/g)

Sucrose (␮g/g)

Total sugars (␮g/g)

Sweetness Index

Soluble solids (%)

Dry matter (%)

Taste score (1–5)

Elario TZ148 N101 Carnivore 30900 Non-grafted

12.3 ab 11.8 b 15.0 a 14.9 a 14.9 a

12.3 ab 11.8 b 14.9 a 14.4 ab 14.1 ab

30.7 ab 32.6 a 27.9 ab 26.3 b 25.3 b

55.3 ab 56.2 ab 57.8 a 55.5 ab 54.3 b

5832 b 5915 ab 6164 a 5938 ab 5829 b

11.54 a 11.89 a 11.69 a 11.55 a 10.44 b

11.01 ab 11.24 a 10.52 ab 10.81 ab 10.02 b

3.37 a 3.73 a 3.58 a 3.17 ab 2.57 b

Raymond TZ148 N101 Carnivore 30900 Non-grafted Polynica N101 Non-grafted

**

**

**

*

*

***

*

***

10.8 ab 10.2 bc 11.1 ab 12.3 a 8.6 c

9.5 bc 9.3 bc 10.2 ab 11.2 a 8.3 c

35.9 b 36.9 ab 37.8 ab 35.7 b 40.0 a

56.2 c 56.4 c 59.0 ab 59.3 ab 56.9 bc

5933 b 5927 b 6209 a 6271 a 5910 b

12.20 ab 12.21 12.53 ab 11.81 b 12.93 a

10.93 ab 10.59 ab 10.99 ab 10.34 b 11.35 a

3.72 ab 4.43 a 3.98 ab 3.81 ab 3.67 b

***

***

*

***

***

*

*

*

14.3 14.0 n.s.

9.7 b 12.4a

30.3 30.2 n.s.

54.3 b 56.6 a

5938 6062 n.s.

10.98 10.75 n.s.

9.00 b 10.20 a

4.1 a 2.8 b

***

***

**

**

n.s. non-significant effect. Scion-specific values within columns followed by the same letter are not significantly different according to Tukey-Kramer HSD Test (P < 0.05). * Significant effect at the 0.05 level. ** Significant effect at the 0.01 level. *** Significant effect at the 0.001 level. Table 6 Eigenvalues, relative and cumulative proportions of total variance, and correlation coefficients for each trait with respect to three principal components (PCs). Item

PC1

PC2

PC3

Eigenvalue Percentage of variance Cumulative variance Eigenvectors Rind thickness Flesh thickness Flesh firmness Fruit hardness La Ca hb ba Fructose Glucose Sucrose Total sugars SSC Fruit dry matter Decline incidence (%) Plant fresh weight Plant dry weight Diameter quotient Yield per plant Fruit weight

11.56 57.79 57.79

3.68 18.42 76.22

1.92 9.59 85.80

0.70 0.92 −0.84 0.55 0.87 −0.96 −0.96 −0.95 −0.86 −0.80 0.94 0.56 0.82 0.40 0.70 −0.62 −0.60 −0.31 0.25 0.97

0.39 0.28 0.46 0.75 −0.10 −0.01 −0.16 0.00 −0.19 0.21 0.10 0.33 0.12 0.55 −0.50 0.76 0.78 0.66 0.48 0.15

0.13 −0.01 0.01 0.12 0.15 0.22 −0.04 0.24 −0.01 0.36 −0.06 0.29 0.38 0.59 −0.15 0.10 −0.03 −0.62 −0.79 −0.08

Boldface factor loadings are considered highly weighed. a L: lightness. C: Chroma. b: yellowness; SSC: soluble solids content. b h: hue angle.

with rind and flesh thickness, fruit hardness, flesh color lightness (L*), sucrose, total sugars, SSC and fruit weight. Furthermore, percentage plant decline was negatively correlated with flesh firmness, flesh color parameters C*, h◦ , and b*, pulp fructose and glucose content, plant fresh and dry weights. The score plot against PC1 and PC2 of all cultivar × rootstock combinations has associated treatments with two main groups: (a) a group clustered in the positive side of PC1 (quadrants 2 and 4) that included only grafted plants of cv. Raymond, and (b) a second group clustered in the negative side of PC1 (quadrants 1 and 3) with the grafted and non grafted plants of cv. Elario (Fig. 1B). Also, non-grafted cv. Polynica (quadrant 1), non-grafted cv. Raymond (quadrant 2) and cv. Polynica grafted on ‘N101’ (quadrant 3) appeared discrete and independent of the two main groups. Quadrant 1 of the score plot included treatments that yielded high plant fresh and dry weights and firmer fruits of high glucose content.

Treatments in quadrant 2 were characterized by high fruit weight, rind thickness, fruit hardness and sucrose content. The lower negative side of PC1 (quadrant 3) included treatment combinations which produced fruits characterized by high C*, b*, h* and fructose content. Treatments in the lower right quadrant (quadrant 4) had the highest percentage plant decline. 4. Discussion Grafting scion cultivars Raymond and Polynica on interspecific Cucurbita rootstocks was the cause of non-pathological plant decline in the period prior to harvest. The absence of plant decline among grafted plants of cv. Elario suggests that this cultivar is likely more resilient to physiological stress, hence its collapse is triggered by more extreme conditions (e.g. higher temperature). Moreover, the absence of decline in grafted cv. Elario precludes the hypothetical co-effect of the grafting procedure (i.e. self-grafting) on decline, in agreement to previous reports on self-grafted vs. non-grafted melon plants which have not identified significant differences between the two (Guan et al., 2014, 2015; Schultheis et al., 2015; Zhao et al., 2011). Minuto et al. (2010) suggested that plant collapse in the absence of soil-pathogens could be triggered by the transportation of auxins from the scion to the susceptible rootstock, eliciting de novo ethylene production and reactive oxygen species (ROS) formation in the root; this may result in the inhibition of root function and development, and may eventually lead to plant decline as the rootstock fails to support scion demand for water and minerals. Aloni et al. (2008b) hypothesized a discrete two-stage scenario during re-establishment of vascular connections wherein incompatibility could lead to plant decline: a) the initial stage, during which the vascular tissues of rootstock and scion may fail to connect successfully, and b) a later stage, at which the grafted plant is unable to compensate for the oxidative stress at the graft interface. Compatibility issues implicated in the phenomenon of plant decline in the absence of a phytopathogenic agent, have been further associated with the suppression of the vegetative fresh weight of the melon scion grafted on various interspecific rootstocks (Edelstein et al., 2004). In the current study, plants of cv. Polynica that survived grafting, suffered severe limitation of vigour and fresh weight (56.7%) compared to the non-grafted plants. By contrast, despite the significant plant decline incurred in all cv. Raymond rootstock combinations, fresh weight of the surviving plants was not depressed, except on rootstock ‘N101’; accordingly,

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Fig. 1. (A) Principal component loading plot and (B) scores of principal component analysis for plant survival, plant yield and morphometric traits, and melon fruit physicochemical quality characteristics as a function of rootstock (R1, TZ148; R2, N101; R3, Carnivore; R4, 30900; NG, non-grafted) and cultivar (Pol, Polynica; Ray, Raymond; El, Elario). Declined, plant decline incidence (%); PFW, plant fresh weight; PDW, plant dry weight; Quotient, plant1st internode/hypocotyl diameter; PY, plant yield; FW, fruit weight; Thickness, fruit flesh thickness; Firmness, fruit flesh firmness; Hardness, fruit hardness; DM, fruit dry matter; C* , flesh chroma; b* , flesh yellowness; h◦ , flesh hue angle; L* , flesh lightness; SSC, soluble solids content; FRU, fructose content; GLC, glucose content; SUC, sucrose content; TS, total sugar content.

the surviving grafted plants were asymptomatic and appeared as vigorous as the non-grafted control. Similarly, cv. Elario displayed vegetative growth and scion fresh weight unaffected by grafting on all rootstocks examined. Our results demonstrate that rootstock effect on scion fresh weight does not always expose physiological incompatibility or reflect the sensitivity of a melon scion to non-pathological decline. However, the negative rootstock effect on plant dry weight of cvs Raymond and Polynica but not of cv. Elario could be inferred as plausible index of incompatibility. The results of the present study consequently suggest that both plant

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fresh and dry weights must be accounted for a more reliable discernment of rootstock-scion physiological incompatibility. Traka-Mavrona et al. (2000) speculated that the dissonance among Cucurbita and Cucumis stem diameters impairs the connection between their vascular bundles and may be considered accountable for the reported low survival rate of grafted plants. Our results indicate that the stem diameter quotient (1st internode diameter/hypocotyl diameter) is not highly reliable as an index of physiological incompatibility and for prognostigating plant decline. In all of our rootstock-scion combinations the stem diameter quotient was significantly higher in non-grafted compared to the grafted plants, notwithstanding the wide and discrete scion-sensitive variation in decline rate: ‘Elario’ = no decline; ‘Raymond’ = 16.7–36.7%; ‘Polynica’ = 20%. This finding is partly consistent with Edelstein et al. (2004) who demonstrated that differences between the scion and rootstock diameters, determined at an early vegetative stage, are not related with plant survival. However, in the current study, the attenuation effect of grafting on the 1st internode’s diameter, relative to the hypocotyl’s, determined at about ten days before the end of the fruiting cycle, has demonstrated an intriguing trend: the relative mean difference in stem diameter quotient between non-grafted and grafted plants ranged from 29.1% in cv. Elario, to 41.5% in cv. Raymond and 44.0% in cv. Polynica. This differentiation is in accord to the discrete sensitivity of the three scions to physiological decline, manifested in their respective decline rates. Rootstock-mediated melon yield increase, has been reported inadvertently in studies focusing on the potential utility of grafting as an effecting tool for controlling soil borne pathogens (Crinò et al., 2007). Other studies, wherein parallel experiments were conducted in both infested and pathogen-free soil environments, revealed that differences in area yield (tons/ ha) between grafted and non-grafted treatments were largely shaped by disease incidence, as yield differentiation was recorded only in the presence of soil pathogens (Cohen et al., 2002). In the study of Traka-Mavrona et al. (2000), performed on a pathogen-free substrate, grafted plant survival rates ranged 8–91% depending on rootstock × scion combination, but differences in yield per plant were not found between the surviving grafted and the non-grafted plants. In our current study, also executed in a disease-free soil environment, rootstock × scion incompatibility was reflected in the yield per plant and the mean fruit weight of cv. Polynica but not of cvs Raymond and Elario. Hence, contrary to Traka-Mavrona et al. (2000), our results demonstrate that non-pathological rootstockscion incompatibility, manifested as high plant death rate, in certain rootstock × scion combinations may also compromise the yield attributes of the grafted plants that survive their fruiting cycle. The distinctive yield behavior of cv. Polynica in response to grafting reflects its inability to support its crop load, resulting in the production of fruits of inferior weight, rind and flesh thickness. Increase in the thickness of fruit rind or flesh in response to grafting onto intespecific rootstocks has not been previously reported in any melon studies. Traka-Mavrona et al. (2000) showed that hybrid rootstocks ‘TZ148’ and ‘Mammoth’ did not affect the fruit flesh thickness of scions ‘Thraki’, ‘Peplo’, ‘Lefko Amynteou’ and ‘Kokkini Banana’. In the current study, thickening of the rind but not of the flesh was recorded only in ‘Elario’ when grafted onto ‘TZ148’. Such thickening of the rind, provided the absence of incompatibility, may prove desirable for the postharvest performance of the melon fruit. Given the imperative use of disease-resistant rootstocks for managing soil pathogenicity, the implications of rootstock-scion interaction on fruit quality characteristics are of prime commercial significance. In understanding the physiological basis of non-pathological plant decline it is furthermore critical to investigate possible expression of latent incompatibility as impairment of fruit quality of the scion. Flesh firmness constitutes one of the

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most important sensory traits of melon fruit (Guan et al., 2014). It is by convention often construed as the hardness of the intact melon fruit. However, the current study demonstrates that the hardness of intact melon fruit does not always correlate with the state of actual flesh firmness, as in the current case of cv. Elario. In terms of actual flesh firmness, grafting melon cultivars on Cucurbita spp. rootstocks has been reported to confer positive (Guan et al., 2014), neutral (Colla et al., 2010a) or negative (Zhao et al., 2011) effects, subject to widespread rootstock × scion interaction. While unequivocal association between cultivar susceptibility to plant decline and loss of fruit hardness or flesh firmness could not be established in the context of the current study, a trend for loss of flesh firmness was evident among the rootstock × scion combinations examined. Given that the commercial scions employed in this study were selected after having demonstrated decline under aggravated field conditions in preliminary work, the observed trend for loss of flesh firmness in response to grafting could represent an index of latent rootstock-scion incompatibility; this is exemplified particularly in the case of cv. Elario wherein plant decline under the current experimental conditions was minimal yet its flesh firmness was compromised on all rootstocks examined. The possible association between loss of flesh firmness and physiological incompatibility in melon deserves further investigation as it might enlighten on the mechanisms of plant decline. The odorous melon scions used in the current study are characterized by a climacteric ripening behavior (Pech et al., 2008). The production of ROS and the triggering of endogenous ethylene production caused by the transportation of auxins from the scion to the rootstock, as proposed by Minuto et al. (2010), further to its effect on rootstock function, may elicit an autocatalytic acceleration of fruit ripening, entailing cell wall metabolic events that lead to tissue softening, progressing from the locular centre toward the periphery of the pericarp (Nishiyama et al., 2007). The shortening of the time period from planting to harvest maturity observed in the current study underpins this scenario. The comprehensive acceleration of ripening in tandem to the autocatalytic production of ethylene can stress rootstock-scion and overall source-sink relations to the point of collapse. Moreover, this effect was observed only on grafted plants of the ‘Polynica’ and ‘Raymond’, both having demonstrated plant decline, but not on ‘Elario’ which showed no decline. Fruit sweetness, defined practically as the soluble solids content of the juice, constitutes a major sensory feature of melon quality (Yamaguchi et al., 1977). In the current study, consistent enhancement of SSC with grafting was observed in the case of cv. Elario which, moreover, did not suffer plant decline; hence, neither an explicit physiological incompatibility nor a latent one, expressed as SSC suppression, could be supported for this scion across rootstocks. The SSC of scions ‘Raymond’ and ‘Polynica’, which demonstrated explicit incompatibility and plant decline, remained largely unaffected. It is worth stressing that even rootstock ‘N101’ did not affect the SSC of the latter two scions, notwithstanding its detrimental effect on their crop stand. Our results suggest that rootstock effect on fruit SSC is not associated with physiological incompatibility and its potential effect on plant survival. Melon SSC seems rather a subject to wide rootstock-scion interaction as demonstrated in melon grafting studies conducted in diseasefree environments (Guan et al., 2014; Traka-Mavrona et al., 2000). The current study additionally demonstrated that the total and individual soluble carbohydrate contents of ripe melon fruits, as well as their sweetness index derived from the relative contribution of individual sugars to sweetness (Brown and Summers, 1985; Elmstrom and Davis, 1981), are also subject to rootstock × scion interaction. Working with muskmelon cv. Zhongmi1 grafted on hybrid rootstocks ‘Ribenstrong’ and ‘Shengzhen’, Liu et al. (2009) also showed that carbohydrate accumulation was rootstock-mediated. The discrete case of cv. Polynica in the cur-

rent experimental context, whose grafting resulted in the decrease of fruit total sugar content, could simply reflect its lower photosynthetic capacity stemming from retarded vegetative growth. Overall, no systematic pattern could be deduced to connect rootstock effect on sugar accumulation to physiological incompatibility and plant decline. Principal Component Analysis formulated a comprehensive framework encompassing all rootstock-scion combinations in respect of plant decline rate, morphological, yield and quality parameters. Differentiation of scion types was obtained along PC1, with Ananas type cv. Raymond concentrated on the positive side and Galia types ‘Elario’ and ‘Polynica’ concentrated on the negative side. Scion response to grafting differentiated further along PC2. All cv. Elario treatments clustered in a single group indicating no differentiation between grafted and non-grafted plants. Contrarily, in the case of cv. Raymond, and more emphatically so in the case of cv. Polynica, the PCA highlighted the substantial grafting effect as the non-grafted control was distanced from the grafted population. Based on the current findings, it is possible to identify at least three scion types with respect to plant decline caused by non-pathological rootstock-scion incompatibility: (a) scions relatively resilient to plant decline, whose surviving individuals are macroscopically asymptomatic, produce similar yield but moderately compromised fruit quality vs. the non-grafted plants (e.g. cv. Elario); (b) scions susceptible to plant decline, whose surviving individuals nonetheless remain macroscopically asymptomatic, produce similar yield but moderately compromised fruit quality vs. the non-grafted plants (e.g. cv. Raymond); (c) scions expressing explicit incompatibility culminating in plant decline, whose surviving individuals demonstrate suppressed yield and fruit quality compared to the non-grafted plants (e.g. cv. Polynica). 5. Conclusion Based on our results we propose that the expression of incompatibility as plant collapse shortly before harvest is a onetime event that does not necessarily reflect on the yield or fruit quality parameters of the asymptomatic surviving plants. However, the negative rootstock effect on plant dry weight, the consistent loss of mesocarp firmness and the attenuation of the 1st internode’s diameter relative to the hypocotyl’s in grafting combinations selected on the basis of reported sensitivity to plant collapse, suggest they may serve as plausible indices of physiological incompatibility in melon. References Aloni, B., Karni, L., Deventurero, G., Levin, Z., Cohen, R., Katzir, N., Lotan-Pompan, M., Edelstein, M., Aktas, H., Turhan, E., Joel, D.M., Horev, C., Kapulnik, Y., 2008a. Possible mechanisms for graft incompatibility between melon scions and pumpkin rootstocks. Acta Hortic. 782, 313–324. Aloni, B., Karni, L., Deveturero, G., Levin, Z., Cohen, R., Kazir, N., Lotan-Pompan, M., Edelstein, M., Aktas, H., Turhan, E., Joel, D.M., Horev, C., Kapulnic, Y., 2008b. Physiological and biochemical changes at the rootstock-scion interface in graft combinations between Cucurbita rootstocks and a melon scion. J. Hortic. Sci. Biotechnol. 83, 777–783. Aloni, B., Cohen, R., Karni, L., Aktas, H., Edelstein, M., 2010. Hormonal signaling in rootstock-scion interactions. Sci. Hortic. 127, 119–126. Andrews, P.K., Marquez, C.S., 1993. Graft incompatibility. Hortic. Rev. 15, 183–232. Brown, A.C., Summers, W.L., 1985. Carbohydrate accumulation and colour development in watermelon. J. Am. Soc. Hort. Sci. 110, 683–687. Cohen, R., Horev, C., Burger, Y., Shriber, S., Hershenhorn, J., Katan, J., Edelstein, M., 2002. Horticultural and pathological aspects of Fusarium wilt management using grafted melons. HortScience 37, 1069–1073. Colla, G., Rouphael, Y., Cardarelli, M., Massa, D., Salerno, A., Rea, E., 2006. Yield, fruit quality and mineral composition of grafted melon plants grown under saline conditions. J. Hortic. Sci. Biotechnol. 81, 146–152. Colla, G., Suãrez, C.M.C., Cardarelli, M., Rouphael, Y., 2010a. Improving nitrogen use efficiency in melon by grafting. HortScience 45, 559–565. Colla, G., Rouphael, Y., Cardarelli, M., Salerno, A., Rea, E., 2010b. The effectiveness of grafting to improve alkalinity tolerance in watermelon. Environ. Exp. Bot. 68, 283–291.

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