Physiological and Ultrastructural Alterations Linked to Intrinsic Mastication Inferiority of Segment Membranes in Satsuma Mandarin (Citrus unshiu Marc.) Fruits

Chewing texture is important for fresh citrus fruits, and the mastication trait of a segment directly determines chewing texture. Roughing disorder impairs the quality of Satsuma mandarin fruits, and it is typically correlated with intrinsic mastication inferiority (IMI). This study explored the role of segment membranes (SMs) in IMI. Similar to IMI in roughing-disordered fruits, segment shear force significantly enhanced relative to controls (CK); cell layers and cell wall thickness increased also in inferior masticating SMs. The ‘Miyamoto Wase’ cultivar exhibited larger segment shear force and more SM cell layers than ‘Juxiangzao’. In SMs, vessel cells could be divided into outside layers where segments adjoin and inside layers where juice sacs grow from. The inside vessel cell layers in the inferior masticating SMs were denser. Vessels with a length of 200 to 300 μm and a diameter of 5 to 15 μm predominated in SMs. The average vessel diameter enlarged by 13% to 16.5% in inferior masticating SMs, depending on cultivars. Furthermore, there was a decrease in vessels with a diameter <5 μm and an increase in vessels >10 μm in the inferior masticating SMs. Between phenotypes, protopectin increased significantly throughout development of inferior masticating SMs, while water-soluble pectin increased during the later stages of development. In one inferior masticating SM sample, protopectin and water-soluble pectin levels were higher in the inner-ring area than those in the outer-ring area. Correspondingly, expression of CuPME21 which is involved in pectin hydrolysis was consistently upregulated in the inferior masticating SMs throughout fruit development. The findings in this work provide novel insights into citrus SM structure and its IMI.

Mitochondria: Key Organelles Accelerating Cell Wall Material Accumulation in Juice Sacs of Pummelo (Citrus grandis L. Osbeck) Fruits during Postharvest Storage

Granulation is a physiological disorder of juice sacs in citrus fruits, which develops through secondary cell wall formation. However, the synergistic changes in the cytoplasm of juice sac cells remain largely unknown. This study investigated the dynamic ultrastructure of juice sacs of “Guanxi” pummelo fruits by transmission electron microscopy and determined their cell wall material, soluble sugar, and organic acid contents. The results showed that lignin and hemicellulose are accumulated in juice sacs isolated from dorsal vascular bundles, while lignin and cellulose contribute to the granulation of juice sacs isolated from septal vascular bundles. The significant differences in lignin, cellulose, and hemicellulose contents between the two types of juice sacs began to be observed at 30 days of storage. Fructose levels were elevated in juice sacs isolated from the dorsal vascular bundles from 10 to 60 days. Sucrose contents significantly decreased in juice sacs isolated from the septal vascular bundles from 30 to 60 days. Meanwhile glucose, citric acid, and malic acid contents exhibited no apparent changes in both types of juice sacs. Based on the comprehensive analysis of the ultrastructure of both types of juice sacs, it was clearly found that plasma membrane ruptures induce cell wall material synthesis in intracellular spaces; however, cell wall substance contents did not significantly increase until the number of mitochondria sharply increased. In particular, sucrose contents began to decrease significantly just after the mitochondria amount largely increased in juice sacs isolated from the septal vascular bundles, indicating that mitochondria play a key role in regulating carbon source sugar partitioning for cell wall component synthesis.

Effects of Salicylic Acid and Methyl Jasmonate Treatments on Flavonoid and Carotenoid Accumulation in the Juice Sacs of Satsuma Mandarin In Vitro

Salicylic acid and jasmonic acid are two important plant hormones that trigger the plant defense responses and regulate the accumulation of bioactive compounds in plants. In the present study, the effects of salicylic acid (SA) and methyl jasmonate (MeJA) on flavonoid and carotenoid accumulation were investigated in the juice sacs of Satsuma mandarin in vitro. The results showed that SA treatment was effective to enhance the contents of eriocitrin, narirutin, poncirin, and β-cryptoxanthin in the juice sacs (p < 0.05). In contrast, the MeJA treatment inhibited flavonoid and carotenoid accumulation in the juice sacs (p < 0.05). Gene expression results showed that the changes of flavonoid and carotenoid contents in the SA and MeJA treatments were highly regulated at the transcriptional level. In addition, a transcriptional factor CitWRKY70 was identified in the microarray analysis, which was induced by the SA treatment, while suppressed by the MeJA treatment. In the SA and MeJA treatments, the change in the expression of CitWRKY70 was consistent with that of flavonoid and carotenoid biosynthetic key genes. These results indicated that CitWRKY70 might be involved in the regulation of flavonoid and carotenoid accumulation in response to SA and MeJA treatments in the juice sacs of citrus fruit.

The Involvement of Energy Metabolism and Lipid Peroxidation in Lignin Accumulation of Postharvest Pumelos

Lignification is especially prominent in postharvest pumelo fruit, which greatly impairs their attractiveness and commercial value. This study investigated the energy metabolism and lipid peroxidation and their relationship with accumulated lignin content in juice sacs of “Hongroumiyou” (HR) during 90 d of storage at 25 °C. The results indicated that, the alterations of energy metabolism in juice of sacs of postharvest pumelos was featured by a continuous decline in energy charge and ATP/ADP; an increase in succinic dehydrogenase (SDH) activity before 30 d and increases in activities of cytochrome c oxidase (CCO) and F0F1-ATPase before 60 d; but declines in activities of Ca2+-ATPase and H+-ATPase. Additionally, enhanced contents of H2O2, O2−, and –OH scavenging rate; increased malondialdehyde (MDA) content; and transformation of unsaturated fatty acids (USFA) to saturated fatty acids (USFA) and reduced USFA/SFA (U/S) could result in lipid peroxidation and membrane integrity loss. Moreover, correlation analysis showed that lignin accumulation was in close relation to energy metabolism and lipid peroxidation in juice sacs of postharvest pumelos. These results gave evident credence for the involvement of energy metabolism and lipid peroxidation in the lignin accumulation of HR pumelo fruit during postharvest storage.

Non-Destructive Identification and Estimation of Granulation in Honey Pomelo Using Visible and Near-Infrared Transmittance Spectroscopy Combined with Machine Vision Technology

Granulation is a physiological disorder of juice sacs in citrus fruit, causing juice sacs to become hard and dry and resulting in decreased internal quality of citrus fruit. Honey pomelo is a thick-skinned citrus fruit, and it is difficult to identify the extent of granulation by observation of the outer peel and fruit shape. In this study, a rapid and non-destructive testing method using visible and near-infrared transmittance spectroscopy combined with machine vision technology was applied to identify and estimate granulation inside fruit. A total of 600 samples in different growth periods was harvested, and fruit were divided into five classes according to five granulation levels. Spectral data were obtained for two ranges of 400–1100 nm and 900–1700 nm by visible and near-infrared transmittance spectroscopy. In addition, chemometrics were used to measure the chemical changes of soluble solid content (SSC), titratable acidity (TA), and moisture content (MC) caused by different granulation levels. Machine vision technology can rapidly estimate the external characteristics of samples and measure the physical changes in mass and volume caused by different granulation levels. Compared with using a single or traditional methods, the predictive performances of multi-category classification models (PCA-SVM and PCA-GRNN) were significantly enhanced. In particular, the model accuracy rate (ARM) was 99% for PCA-GRNN, with classification accuracy (CA), classification sensitivity (CS), and classification specificity (CSP) of 0.9950, 0.9750, and 0.9934, respectively. The results showed that this method has great potential for the identification and estimation of granulation. Multi-source data fusion and application of a multi-category classification model with the smallest number of input layers and acceptable high predictive performances are proposed for on-line applications. This method can be effectively used on-line for the non-destructive detection of fruits with granulation.

Metabolite profile comparison of a graft chimera ‘Hongrou Huyou’ (Citrus changshan-huyou + Citrus unshiu) and its two donor plants

Background: Chimeras synthesized artificially by grafting are crucial to the breeding of perennial woody plants. ‘Hongrou Huyou’ (Citrus changshan-huyou + Citrus unshiu) is a new graft chimera originating from the junction where a Citrus changshan-huyou (“C”) scion was top-grafted onto a stock Satsuma mandarin ‘Owari’ (C. unshiu, “O”). The chimera was named OCC because the cell layer constitutions were O for Layer 1(L1) and C for L2 and L3. In this study, profiles of primary metabolites, volatiles and carotenoids derived from different tissues in OCC and the two donors were investigated, with the aim of determining the relationship between the layer donors and metabolites. Results: The comparison of the metabolite profiles showed that the amount and composition of metabolites were different between the peels and the juice sacs, as well as between OCC and each of the two donors. The absence or presence of specific metabolites (such as the carotenoids violaxanthin and β-cryptoxanthin, the volatile hydrocarbon germacrene D, and the primary metabolites citric acid and sorbose) in each tissue was identified in the three phenotypes. According to principal component analysis (PCA), overall, the metabolites in the peel of the chimera were derived from donor C, whereas those in the juice sac of the chimera came from donor O. Conclusion: The profiles of primary metabolites, volatiles and carotenoids derived from the peels and juice sacs of OCC and the two donors were systematically compared. The content and composition of metabolites were different between the tissues and between OCC and the each of the two donors. A clear donor dominant pattern of metabolite inheritance was observed in the different tissues of OCC and was basically consistent with the layer origin; the peel of the chimera was derived from C, and the juice sacs of the chimera came from O. These profiles provide potential chemical markers for genotype differentiation, citrus breeding assessment, and donor selection during artificial chimera synthesis.

Metabolite profile comparison of a graft chimera ‘Hongrou Huyou’ (Citrus changshan-huyou + Citrus unshiu) and its two donor plants

Background: Chimeras synthesized artificially by grafting are crucial to the breeding of perennial woody plants. ‘Hongrou Huyou’ (Citrus changshan-huyou + Citrus unshiu) is a new graft chimera originating from the junction where a Citrus changshan-huyou (“C”) scion was top-grafted onto a stock Satsuma mandarin ‘Owari’ (C. unshiu, “O”). The chimera was named OCC because the cell layer constitutions were O for Layer 1(L1) and C for L2 and L3. In this study, profiles of primary metabolites, volatiles and carotenoids derived from different tissues in OCC and the two donors were investigated, with the aim of determining the relationship between the layer donors and metabolites. Results: The comparison of the metabolite profiles showed that the amount and composition of metabolites were different between the peels and the juice sacs, as well as between OCC and each of the two donors. The absence or presence of specific metabolites (such as the carotenoids violaxanthin and β-cryptoxanthin, the volatile hydrocarbon germacrene D, and the primary metabolites citric acid and sorbose) in each tissue was identified in the three phenotypes. According to principal component analysis (PCA), overall, the metabolites in the peel of the chimera were derived from donor C, whereas those in the juice sac of the chimera came from donor O. Conclusion: The profiles of primary metabolites, volatiles and carotenoids derived from the peels and juice sacs of OCC and the two donors were systematically compared. The content and composition of metabolites were different between the tissues and between OCC and the each of the two donors. A clear donor dominant pattern of metabolite inheritance was observed in the different tissues of OCC and was basically consistent with the layer origin; the peel of the chimera was derived from C, and the juice sacs of the chimera came from O. These profiles provide potential chemical markers for genotype differentiation, citrus breeding assessment, and donor selection during artificial chimera synthesis.