Cold pre-treatment is effective for 1-MCP efficacy in ‘Tsugaru’ apple fruit

1-Methylcyclopropene (1-MCP) treatment maintains apple fruit quality during storage, but its efficacy is dependent on a number of conditions. ‘Tsugaru’ apples are a major early season cultivar in Japan, but because ‘Tsugaru’ fruit produce abundant ethylene, they have a short shelf-life, and efficacy of 1-MCP is not as high with ‘Tsugaru’ as with other cultivars. To improve 1-MCP efficacy, ‘Tsugaru’ fruit were pre-cooled at −1 °C or −3 °C for 24 h before 1-MCP treatment. Ethylene production decreased with the cold treatment, resulting in better storage after 1-MCP treatment. Although ethylene production was low at the end of 24 h of the cold pre-treatment, expression of ACS1, the ethylene receptor genes ERS1, ETR1(a), ETR1b, ETR2 and ETR5, and the cell wall degradation-related gene PG1 all increased with a 24 h cold treatment. It is assumed that these elevated gene expression levels were not caused by ethylene, but more directly by cold stimulus. Thus, a short period of cold stimulus suppresses ethylene production, but induces expression of some genes. 1-MCP treatment was more effective with some initial fruit chilling.     

Non-destructive monitoring of flesh softening in the black-skinned Japanese plums ‘Angeleno’ and ‘Autumn beaut’ on-tree and postharvest

There is a need to develop alternative harvest indexes for black skinned plums. The aim of this research was to analyze and compare the most commonly used indexes for deciding the harvest date for Japanese plum, and evaluate the effectiveness of new approaches for studying maturation. The ripening process was monitored on-tree and during postharvest in a non-destructive way, through the absorbance of chlorophyll (I_AD), the compression strength of the intact fruit, and the traditional parameters associated with ripening. Fruit were harvested at commercial ripeness and “tree-ripe”, and were stored for 10 d at 22 °C and 75-80% RH. The I_AD decreased during the last phase of development of the fruit on-tree, and it was related to the common indexes used for plums. ‘Angeleno’ showed a decrease of the I_AD 24% lower than that observed for ‘Autumn beaut’. The I_AD versus time showed the highest coefficients of determination when compared with the soluble solids concentration (SSC), flesh firmness, hue (H°) and chroma (C*) of the skin. The compression strength of the intact fruit was associated with flesh firmness, and to a lesser extent with the SSC for ‘Angeleno’, whereas for ‘Autumn beaut’ higher correlations for both the SSC and flesh firmness were observed. The C* of the skin on ‘Autumn beaut’ showed an erratic change during ripening; by contrast, for ‘Angeleno’, this index showed a clear trend. During postharvest it was observed that for ‘Angeleno’ fruit picked at commercial ripeness, the rate of change of the I_AD was practically the same as observed on-tree, while for ‘Autumn beaut’ the rates of change on-tree and at postharvest were 0.075 I_AD d⁻¹ and 0.024 I_AD d⁻¹, respectively. For the “tree-ripe” fruit, the rate of change was practically the same for both cultivars. Similar trends were observed for the compression strength.     

Effect of postharvest ethylene treatment on carotenoid accumulation and the expression of carotenoid biosynthetic genes in the flavedo of orange (Citrus sinensis L. Osbeck) fruit

Degreening with ethylene is a common postharvest practice in citrus fruit. In this work we have examined the effect of ethylene treatment on carotenoid content and composition, and on the expression of carotenoid biosynthetic genes in the flavedo of Navelate orange (Citrus sinensis L.) harvested at two ripening stages. The ethylene-induced fruit coloration and carotenoid content in the flavedo increased with the ripening stage of the fruit. Analysis of the changes in individual carotenoids revealed that ethylene stimulated an increase in phytoene, phytofluene, (9Z)-violaxanthin which is the main carotenoid in fully ripened orange peel, and the apocarotenoid β-citraurin, and decreased the concentration of chloroplastic carotenoids. These changes are consistent with the effect of ethylene on the expression of carotenoid biosynthetic genes, since it up-regulated the expression of phytoene synthase, ζ-carotene desaturase and β-carotene hydroxylase genes, sustained or transiently increased accumulation of phytoene desaturase, plastid terminal oxidase, β-lycopene cyclase and zeaxanthin epoxidase mRNAs, and decreased the expression of the ɛ-lycopene cyclase gene. These data indicate that exogenous ethylene reproduces and accelerates the physiological and molecular changes in the carotenoid biosynthesis naturally occurring during maturation of citrus fruit. On the other hand, gibberellic acid, which delays fruit degreening, reduced the ethylene-induced expression of early carotenoid biosynthetic genes and the accumulation of phytoene, phytofluene and β-citraurin.     

Ethylene synthesis,ripening capacity,and superficial scald inhibition in 1-MCP treated ‘d’Anjou’ pears are affected by storage temperature

A continuing challenge for commercializing 1-methylcyclopropene (1-MCP) to extend the storage life and control superficial scald of ‘d’Anjou’ pear (Pyrus communis L.) is how to initiate ripening in 1-MCP treated fruit. ‘D’Anjou’ pears harvested at commercial and late maturity were treated with 1-MCP at 0.15 μL L⁻¹ and stored either at the commercial storage temperature −1.1 °C (1-MCP@−1.1 °C), or at 1.1 °C (1-MCP@1.1 °C) or 2.2 °C (1-MCP@2.2 °C) for 8 months. Control fruit stored at −1.1 °C ripened and developed significant scald within 7 d at 20 °C following 3–5 months of storage. While 1-MCP@−1.1 °C fruit did not develop ripening capacity due to extremely low internal ethylene concentration (IEC) and ethylene production rate for 8 months, 1-MCP@1.1 °C fruit produced significant amounts of IEC during storage and developed ripening capacity with relatively low levels of scald within 7 d at 20 °C following 6–8 months of storage. 1-MCP@2.2 °C fruit lost quality quickly during storage. Compared to the control, the expression of ethylene synthesis (PcACS1, PcACO1) and signal (PcETR1, PcETR2) genes was stable at extremely low levels in 1-MCP@−1.1 °C fruit. In contrast, they increased expression after 4 or 5 months of storage in 1-MCP@1.1 °C fruit. Other genes (PcCTR1, PcACS2, PcACS4 and PcACS5) remained at very low expression regardless of fruit capacity to ripen. A storage temperature of 1.1 °C can facilitate initiation of ripening capacity in 1-MCP treated ‘d’Anjou’ pears with relatively low scald incidence following 6–8 months storage through recovering the expression of certain ethylene synthesis and signal genes.     

Ethylene biosynthesis in apricot: Identification of a ripening-related 1-aminocyclopropane-1-carboxylic acid synthase (ACS) gene

Apricots are climacteric fruits with a high susceptibility to flesh softening and loss of flavor during postharvest storage, and most of the ripening processes are regulated by ethylene, which also has an effect on its own biosynthesis. To understand this process in apricot, inhibition of ethylene biosynthesis and perception was performed for studying key genes involved in the ethylene biosynthetic pathway. Apricots, cv. “Patterson”, were harvested with yellow-green ground color and immediately treated with either the ethylene perception inhibitor 1-methyl cyclopropene (1-MCP) at 10 μL L⁻¹ or the ethylene biosynthesis inhibitor aminoethoxyvinylglycine (AVG) at 1 g L⁻¹. After treatment, quality and physiological attributes such as firmness, color, total soluble solids, acidity, fruit weight, ethylene production and respiration rates were evaluated every 2 d until they ripened at 20 °C. Gene expression analysis was performed by quantitative polymerase chain reaction (qPCR). Both ethylene inhibitors were effective in reducing ethylene production, respiration rate and fruit softening. Three 1-aminocyclopropane-1-carboxylic-acid synthase (ACS) genes were characterized, but only the expression of ACS2 was highly reduced by ethylene inhibition, suggesting a key role in ethylene synthesis at ripening. Contrarily, ACS1 and ACS3 showed a higher expression under ethylene inhibition suggesting that the corresponding genes are individually regulated in a specific mode as observed in other climacteric fruits. Finally, changes in 1-aminocyclopropane-1-carboxylic-acid oxidase genes did not show a consistent pattern of ethylene modulation.     

Effects of chlorine dioxide treatment on respiration rate and ethylene synthesis of postharvest tomato fruit

Tomato fruit at the mature green stage were treated with ClO₂ gas in a sealed container for 12 h, and then stored at 23 °C with 85% relative humidity (RH) for 23 d. Respiration rate, respiration-related enzymes including phosphohexose isomerase (PHI), succinate dehydrogenase (SDH), and glucose-6-phosphate dehydrogenase (G-6-PDH) and 6-phosphogluconate dehydrogenase (6-PGDH), ethylene production, and the expression of LeAOX1a, LeCOX1, LeACS2, LeACS4 and LeACO1 genes were measured. The results showed that application of ClO₂ gas was effective in reducing total respiration, cytochrome pathway respiration and the expression of LeCOX1, but no significant reduction in the activities of respiration-related enzymes was observed during storage. Fruit treated with ClO₂ resulted in lower ethylene production. Furthermore, the expression of ethylene biosynthesis related genes, including LeACS2, LeACS4 and LeACO1 was reduced by the ClO₂ treatment. These results indicate that ClO₂ treatment might delay the ripening of tomato fruit, possibly by a mechanism involving suppression of respiration rate and ethylene biosynthesis.     

Molecular characterization of two banana ethylene signaling component MaEBFs during fruit ripening

EIN3 Binding F-box protein (EBF) is an essential signaling component necessary for ethylene response. However, little information is available on EBF genes during banana fruit ripening. Two EBF genes designated MaEBF1 and MaEBF2 were isolated and characterized from banana fruit. Subcellular localization analysis showed that MaEBF1 and MaEBF2 were both nuclear proteins. Expression of MaEBF1 and MaEBF2 in fruit with four ripening characteristics revealed that MaEBF2 was enhanced by ethylene during fruit ripening, while MaEBF1 changed only slightly. Moreover, the MaEBF2 promoter was activated after ethylene treatment, further supporting its involvement in fruit ripening. More importantly, MaEBF2 was shown to physically interact with MaEIL5, using yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) assays. Together, these results suggest that MaEBF may be involved in banana fruit ripening, at least partly via interaction with MaEIL5. Our findings expand our understanding of the regulatory network of ethylene signaling cascade in banana fruit ripening.     

Postharvest ultraviolet-C irradiation suppressed Psy 1 and Lcy-β expression and altered color phenotype in tomato (Solanum lycopersicum) fruit

Mature green cherry tomato fruit were harvested and treated with ultraviolet-C (UV-C) irradiation at a predetermined dose of 4.2 kJ m⁻², and stored at 18 °C for 35 days. The effects of UV-C treatment on color change, pigment contents, and the expression of major genes involved in carotenoid metabolism, including Psy 1, Pds, Lcy-β, and Lcy-ɛ, encoding phytoene synthase, phytoene desaturase, lycopene β-cyclase and lycopene ɛ-cyclase, respectively, were examined. The UV-C treated fruit developed a pink red color in contrast to the normal orange red color of control fruit. Lycopene accumulation during ripening in UV-C treated fruit was significantly inhibited but its final content was not affected. However, both accumulation and final content of β-carotene were significantly suppressed in UV-C treated fruit. The lower content of β-carotene, leading to a higher lycopene to β-carotene ratio, is probably responsible for the altered color phenotype in UV-C treated fruit. Psy 1, a major gene involved in lycopene synthesis was inhibited by UV-C irradiation. Significantly suppressed expression of Lcy-β gene was also observed in UV-C treated fruit. Thus it is possible that the lower transformation from lycopene to carotenes contributed to the relatively stable content of lycopene.     

Characterization of ethylene biosynthesis and its regulation during fruit ripening in kiwifruit,Actinidia chinensis ‘Sanuki Gold’

Ethylene biosynthesis in kiwifruit, Actinidia chinensis ‘Sanuki Gold’ was characterized using propylene, an ethylene analog, and 1-methylcyclopropene (1-MCP), an inhibitor of ethylene perception. In fruit harvested between a young stage (66 days after pollination) (DAP) and an early commercial harvesting stage (143 DAP), 2 days of exposure to propylene were sufficient to initiate ethylene biosynthesis while in fruit harvested at commercial harvesting stage (154 DAP), 4 days of propylene treatment were required. This observation suggests that response of ethylene biosynthesis to propylene treatment in kiwifruit declined with fruit maturity. Propylene treatment resulted in up-regulated expression of AC-ACO1, AC-ACO2, AC-SAM1 and AC-SAM2, prior to the induction of AC-ACS1 and ethylene production, confirming that AC-ACS1 is the rate limiting step in ethylene biosynthesis in kiwifruit. Treatment of fruit with more than 5 μL L^?1 of 1-MCP after the induction of ethylene production subsequently suppressed ethylene production and expression of ethylene biosynthesis genes. Treatment of fruit with 1-MCP at harvest followed with propylene treatment delayed the induction of ethylene production and AC-ACS1 expression for 5 days. These observations suggest that in ripening kiwifruit, ethylene biosynthesis is regulated by positive feedback mechanism and that 1-MCP treatment at harvest effectively delays ethylene production by 5 days.     

Mode of action of nitric oxide in inhibiting ethylene biosynthesis and fruit softening during ripening and cool storage of ‘Kensington Pride’ mango

The mode of action of nitric oxide (NO) in inhibiting ethylene biosynthesis and fruit softening during ripening and cool storage of mango fruit was investigated. Hard mature green mango (Mangifera indica L. cv. ‘Kensington Pride’) fruit were fumigated with 20 μL L⁻¹ NO for 2 h at 21 °C and allowed to ripen at 21 ± 1 °C for 10 d, or stored at 13 ± 1 °C for 21 d. During ripening and cool storage, ethylene production and respiration rate from whole fruit were determined daily. The 1-aminocyclopropane-1-carboxylic acid (ACC) content, activities of ACC synthase (ACS), ACC oxidase (ACO), and fruit softening enzymes such as pectin esterase (PE), endo-1,4-β-d-glucanase (EGase), exo- and endo-polygalacturonase (exo-PG, endo-PG) as well as firmness and rheological properties of pulp were determined at two- and seven-day intervals during ripening and cool storage, respectively. NO fumigation inhibited ethylene biosynthesis and respiration rate, and maintained higher pulp firmness, springiness, cohesiveness, chewiness, adhesiveness, and stiffness. NO-fumigated fruit during cool storage and ripening had lower ACC contents through inhibiting the activities of both ACS and ACO in the fruit pulp. NO-fumigated fruit showed decreased activities of exo-PG, endo-PG, EGase, but maintained higher PE activity in pulp tissues during ripening and cool storage. In conclusion, NO fumigation inhibited ethylene biosynthesis through inhibition of ACS and ACO activities leading to reduced ACC content in the fruit pulp which consequently, reduced the activities of fruit softening enzymes during ripening and cool storage.     

Low oxygen and 1-MCP pretreatments delay superficial scald development by reducing reactive oxygen species (ROS) accumulation in stored ‘Granny Smith’ apples

‘Granny Smith’ apples are highly susceptible to superficial scald, a symptom of chilling injury. For many crops, low temperature storage results in oxidative stress and chilling injury, caused by increased production of superoxide anions which in turn leads to the generation of other dangerous reactive oxygen species (ROS). Application, prior to cold storage, of low oxygen (LO2, <0.5%) atmospheres, ethanol (<2% vapour) or 1-methylcyclopropene (1-MCP, 0.5 μL L⁻¹) at 20 °C, was effective in reducing superficial scald in fruit following 24 weeks of cold storage. ROS levels were measured by confocal laser-scanning microscopy of apple peel treated with the fluorescent probe 2′,7′-dichlorodihydrofluorescein diacetate. In control fruit, ROS levels increased during cold storage and shelf-life and were very high after only 8 weeks, whereas in 1-MCP-, ethanol- and LO2-treated fruit, ROS levels remained low throughout storage. Gene-expression levels of ROS-scavenging enzymes were induced by the various pretreatments: catalase (MdCAT) was induced by LO2 treatment, whereas Mn superoxide dismutase (MdMnSOD) was induced by 1-MCP treatment. Polyphenol oxidase (MdPPO) gene expression levels were associated with scald symptom development and were highest in control fruit. Ethylene levels and expression of ethylene biosynthesis genes were correlated with α-farnesene levels and <alpha>-farnesene synthase (MdAFS) gene expression in the variously treated fruit. Accumulation of the α-farnesene oxidation product, 6-methyl-5-hepten-2-one (MHO), was highest in control fruit after 8 weeks, in accordance with ROS accumulation. The LO2 pretreatment mechanism might involve production of anaerobic metabolites, causing a delay in ethylene and α-farnesene biosynthesis and oxidation; this is different from the mechansism of action of 1-MCP, even though both consequently reduce ROS accumulation and scald symptoms.     

Chilling-injury of harvested tomato (Solanum lycopersicum L.) cv. Micro-Tom fruit is reduced by temperature pre-treatments

Heat-shocks were used to reduce the development of chilling injury symptoms during ripening of tomato fruit (Solanum lycopersicum L. cv. Micro-Tom). Mature green tomatoes were immersed in 30-50 °C water for 3-9 min before being chilled at 2.5 °C for 0, 0.5, 1, 2, 3, or 14 days, and then held at 20 °C for an additional 7-14 days. The affect of both heat-shock and chilling treatments were independent of fruit weight. Measured at 20 °C after 14 days of chilling, fruit exposed to 40 °C for 7 min exhibited reduced chilling injury symptoms, as measured by their advanced ripening score and decreased rate of ion leakage into an isotonic 0.2 M mannitol solution. Reduced rates of leakage from the symplastic compartment probably contributed to the 2-fold decrease in the amount of ions in the apoplastic space, when compared to the control. A subsequent paper will report the results of metabolic profiling of Micro-Tom tomato fruit subjected to treatments that significantly decreased their development of chilling injury symptoms.