Christopher B. Watkins

Influence of Salicylic Acid on H2O2 Production, Oxidative Stress, and H2O2-Metabolizing Enzymes (Salicylic Acid-Mediated Oxidative Damage Requires H2O2)

We investigated how salicylic acid (SA) enhances H2O2 and the relative significance of SA-enhanced H2O2 in Arabidopsis thaliana. SA treatments enhanced H2O2 production, lipid peroxidation, and oxidative damage to proteins, and resulted in the formation of chlorophyll and carotene isomers. SA-enhanced H2O2 levels were related to increased activities of Cu,Zn-superoxide dismutase and were independent of changes in catalase and ascorbate peroxidase activities. Prolonging SA treatments inactivated catalase and ascorbate peroxidase and resulted in phytotoxic symptoms, suggesting that inactivation of H2O2-degrading enzymes serves as an indicator of hypersensitive cell death. Treatment of leaves with H2O2 alone failed to invoke SA-mediated events. Although leaves treated with H2O2 accumulated in vivo H2O2 by 2-fold compared with leaves treated with SA, the damage to membranes and proteins was significantly less, indicating that SA can cause greater damage than H2O2. However, pretreatment of leaves with dimethylthiourea, a trap for H2O2, reduced SA-induced lipid peroxidation, indicating that SA requires H2O2 to initiate oxidative damage. The relative significance of the interaction among SA, H2O2, and H2O2-metabolizing enzymes with oxidative damage and cell death is discussed.

Responses of early, mid and late season apple cultivars to postharvest application of 1-methylcyclopropene (1-MCP) under air and controlled atmosphere storage conditions

The potential for commercial application of 1-methylcyclopropene (1-MCP) to maintain quality of ‘McIntosh’, ‘Empire’, ‘Delicious’ and ‘Law Rome’ apples under air and controlled atmosphere (CA) storage conditions was investigated. These cultivars represent early, mid and late season apples with ripening rates ranging from fast to slow. 1-MCP gas concentrations used were 0.5, 1 and 2 m ll 1 , generated from measured amounts of Ethylbloc™ powder. Fruit of each cultivar were removed from storage at 6 week intervals during 30 weeks in air, or at 8 week intervals during 32 weeks in CA, and evaluated after 1 and 7 days at 20°C. Effects of 1-MCP were greater in CA than air storage. A dose response of internal ethylene concentrations and flesh firmness to 1-MCP was found in ‘McIntosh’ and ‘Law Rome’, but ‘Delicious’ and ‘Empire’ ripening was generally prevented by all 1-MCP concentrations. 1-MCP reduced superficial scald incidence, and accumulations of a-farnesene and conjugated trienols during air storage. The results indicate that the efficacy of 1-MCP is affected by cultivar and storage conditions, and that successful commercial utilization of the chemical will require understanding of these relationships.

A Reevaluation of the Key Factors That Influence Tomato Fruit Softening and Integrity

The softening of fleshy fruits, such as tomato (Solanum lycopersicum), during ripening is generally reported to result principally from disassembly of the primary cell wall and middle lamella. However, unsuccessful attempts to prolong fruit firmness by suppressing the expression of a range of wall-modifying proteins in transgenic tomato fruits do not support such a simple model. ‘Delayed Fruit Deterioration’ (DFD) is a previously unreported tomato cultivar that provides a unique opportunity to assess the contribution of wall metabolism to fruit firmness, since DFD fruits exhibit minimal softening but undergo otherwise normal ripening, unlike all known nonsoftening tomato mutants reported to date. Wall disassembly, reduced intercellular adhesion, and the expression of genes associated with wall degradation were similar in DFD fruit and those of the normally softening ‘Ailsa Craig’. However, ripening DFD fruit showed minimal transpirational water loss and substantially elevated cellular turgor. This allowed an evaluation of the relative contribution and timing of wall disassembly and water loss to fruit softening, which suggested that both processes have a critical influence. Biochemical and biomechanical analyses identified several unusual features of DFD cuticles and the data indicate that, as with wall metabolism, changes in cuticle composition and architecture are an integral and regulated part of the ripening program. A model is proposed in which the cuticle affects the softening of intact tomato fruit both directly, by providing a physical support, and indirectly, by regulating water status.

Physical and mechanical changes in strawberry fruit after high carbon dioxide treatments

‘Pajaro’ strawberry (Frageria x ananassa Duch.) fruit were exposed to 5‐40% CO2 for 0‐3 days, followed by normal cold storage at 0°C for up to 3 weeks. Strawberry fruit were firmer in air storage at 0°C than at harvest. Firmness was further enhanced by CO2 treatments. Adhesion between cells was measured by the application of tensile tests to plugs of tissue, followed by the examination of fracture surfaces using low temperature scanning electron microscopy. These tests indicated that cell-to-cell adhesion increased by 60% as a result of CO2 treatments. However, there were no differences in the density, electrolyte leakage, propensity for cells to rupture in hypertonic solutions, water potential, osmotic potential or turgor of CO2-treated and control fruit. Electrical impedance spectroscopy was used to assess changes in the electrical resistance of the apoplast and symplast. Carbon dioxide treatments reduced the resistance of the apoplast (resistance at 50 Hz) below that of control fruit, but did not affect the resistance of the symplast (resistance at 1 MHz). This result suggests that concentrations of H and HCO3 increased in the apoplast, although no change was detected in the symplast. We speculate that the mechanism for CO2-induced firmness enhancement in strawberry is due to changes in the pH of the apoplast. Such changes in pH may promote the precipitation of soluble pectins and thus enhance cell-to-cell bonding in strawberry fruit.

Firmness and concentrations of acetaldehyde, ethyl acetate and ethanol in strawberries stored in controlled and modified atmospheres

Abstract ‘Pajaro’ strawberries ( Fragaria × ananassa Duch.) were stored at 0 °C in a range of controlled atmosphere (CA) conditions with CO 2 concentrations up to 24%, O 2 concentrations down to 1%, or a combination of 10% CO 2 and 2% O 2 . Elevated CO 2 concentrations resulted in firmer fruit, while low O 2 did not affect texture. Off-flavours developed after 3 days of storage at 20% CO 2 , but decreased when fruit was subsequently held for 24 h at 20 °C. However, off-flavours were persistent after CA storage for 7 days or more. Off-flavours were related to increases in ethyl acetate and ethanol concentrations but not to acetaldehyde. Beneficial atmospheres of close to 10% CO 2 and 2% O 2 resulted in a firmer texture and delayed ripening with no off-flavour development. However, fruit quality was poor when similar atmospheres were developed in modified atmosphere (MA)-producing polythene bags. Rapid imposition of CA resulted in better quality fruit than when MAs around the fruit were developed gradually.

A GH3-like gene, CcGH3, isolated from Capsicum chinense L. fruit is regulated by auxin and ethylene*

Auxin, which has been implicated in multiple biochemical and physiological processes, elicits three classes of genes (Aux/IAAs, SAURs and GH3s) that have been characterized by their early or primary responses to the hormone. A new GH3-like gene was identified from a suppressive subtraction hybridization (SSH) library of pungent pepper (Capsicum chinense L.) cDNAs. This gene, CcGH3, possessed several auxin- and ethylene-inducible elements in the putative promoter region. Upon further investigation, CcGH3 was shown to be auxin-inducible in shoots, flower buds, sepals, petals and most notably ripening and mature pericarp and placenta. Paradoxically, this gene was expressed in fruit when auxin levels were decreasing, consistent with ethylene-inducibility. Further experiments demonstrated that CcGH3 was induced by endogenous ethylene, and that transcript accumulation was inhibited by 1-methylcyclopropene, an inhibitor of ethylene perception. When over-expressed in tomato, CcGH3 hastened ripening of ethylene-treated fruit. These results implicate CcGH3 as a factor in auxin and ethylene regulation of fruit ripening and suggest that it may be a point of intersection in the signaling by these two hormones.

Cultivar variation in response of strawberry fruit to high carbon dioxide treatments

The influence of cultivar on responses of strawberry fruit to 20 kPa CO2 or air storage at 2 °C was measured for 7 days. CO2 storage increased firmness of all cultivars over air storage, but the degree of increase was affected by cultivar and days in storage. CO2-treated fruit were lighter colored and less intensely red than air-treated fruit. Accumulations of acetaldehyde, ethanol and ethyl acetate in CO2-treated fruit were strongly affected by cultivar, with those in ‘Annapolis’ having the lowest and those in ‘Governor Simcoe’ having the highest concentrations. Genetic variation in this response provides useful material for investigating the metabolic basis of CO2 effects in strawberry fruit.

Metabolic changes in 1-methylcyclopropene (1-MCP)-treated ‘Empire’ apple fruit during storage

Abstract‘Empire’ apple fruit are more susceptible to flesh browning at 3.3°C if treated with 1-methylcyclopropene (1-MCP), an inhibitor of ethylene perception. To better understand the metabolic changes associated with this browning, untargeted metabolic profiling with partial least squares analysis has been used to visualize changes in metabolic profile during hypoxic controlled atmosphere (CA) storage, ethylene insensitivity, and disorder development. Overall, most carbohydrates and organic acids were not appreciably affected, but the levels of amino acids and volatile metabolites were significantly affected, by 1-MCP treatment. Sorbitol and levels of some amino acids were elevated towards the end of storage in 1-MCP treated fruit. CA storage reduced the levels of many volatile components and 1-MCP reduced these levels further. Additionally multiple metabolites were associated with the development of flesh browning symptoms. Unlike other volatile compounds, methanol levels gradually increased with storage duration, regardless of 1-MCP treatment, while 1-MCP decreased ethanol production. Results reveal metabolic changes during storage that may be associated with development of flesh browning symptoms.

Postharvest treatments of fresh produce

Postharvest technologies have allowed horticultural industries to meet the global demands of local and large-scale production and intercontinental distribution of fresh produce that have high nutritional and sensory quality. Harvested products are metabolically active, undergoing ripening and senescence processes that must be controlled to prolong postharvest quality. Inadequate management of these processes can result in major losses in nutritional and quality attributes, outbreaks of foodborne pathogens and financial loss for all players along the supply chain, from growers to consumers. Optimal postharvest treatments for fresh produce seek to slow down physiological processes of senescence and maturation, reduce/inhibit development of physiological disorders and minimize the risk of microbial growth and contamination. In addition to basic postharvest technologies of temperature management, an array of others have been developed including various physical (heat, irradiation and edible coatings), chemical (antimicrobials, antioxidants and anti-browning) and gaseous treatments. This article examines the current status on postharvest treatments of fresh produce and emerging technologies, such as plasma and ozone, that can be used to maintain quality, reduce losses and waste of fresh produce. It also highlights further research needed to increase our understanding of the dynamic response of fresh produce to various postharvest treatments.

Arabidopsis AtNAP regulates fruit senescence

Arabidopsis has been used as a model system to study many aspects of plant growth and development. However, fruit senescence in Arabidopsis has been less investigated and the underlying molecular and hormonal (especially ethylene) regulatory mechanisms are not well understood. It is reported here that the Arabidopsis silique has characteristics of a climacteric fruit, and that AtNAP, a NAC family transcription factor gene whose expression is increased with the progression of silique senescence, plays an important role in its senescence. Silique senescence was delayed for 4–5 d in the atnap knockout mutant plants. The ethylene climacteric was delayed for 2 d in the atnap silique and the associated respiratory climacteric was suppressed. Exogenous ethylene stimulated respiration in the wild type, but not in the atnap mutant. The decoupling of the ethylene and respiratory climacterics in the atnap mutant suggests that AtNAP is required for ethylene stimulation of respiration. qPCR analyses revealed that the expression patterns of genes involved in ethylene biosynthesis, perception, and signalling, ACS2, ETR1, CTR1, EIN2, EIN3, and ERF1, were also altered in the atnap mutant. The effects of exogenous ABA, SA, 6-BA, and NAA on ethylene production and respiration in siliques of the wild type and atnap mutant were also investigated. A model involving ABA-AtNAP-controlled stomatal opening in regulating ethylene-stimulated respiration in fruit senescence is presented.