Ethanol fermentation- and ethylene physiology-related gene expression profiles in Red Delicious apples stored under variable hypoxic conditions and protocols

From Firenze University Press Journal: Advances in Horticultural Science

E. Salamé, Crop Science Research Center, Scuola Superiore Sant’Anna

S. Brizzolara, Crop Science Research Center, Scuola Superiore Sant’Anna

M. Rodriguez, Department of Agronomy, Food, Natural Resources, Animals and Environment

M. Iob, Marvil Engeneering

P. Tonutti, Crop Science Research Center, Scuola Superiore Sant’Anna

B. Ruperti, Department of Agronomy, Food, Natural Resources, Animals and Environment

Dynamic Controlled Atmosphere (DCA) is beneficial in maintaining specific quality parameters but, due to the extreme oxygen levels applied, can cause adverse effects on the fruit by inducing excessive anaerobic metabolism and the production of off-flavors. The metabolic adaptation and responses of apples (Malus domestica Borkh.) cv. Red Delicious to static or dynamic oxygen concentrations (0.3 and 0.8%, with sequential shifts) during cold storage for 7 months were studied by monitoring quality parameters and the expression of genes involved in sugar, fermentative metabolism, and ethylene physiology. Ethanol content reached the highest levels (around 400 mg/kg FW) under 0.3% oxygen concentration and fruit firmness appeared to be reduced in samples accumulating the highest levels of ethanol. Oxygen switch was effective in reducing the ethanol concentrations with timing-dependent variable effects. The expression of fermentative (alcohol dehydrogenase, lactate dehydrogenase, pyruvate decarboxylase) and sugar metabolism (β-amylase; phosphofructokinase; sucrose synthase) genes resulted to be differently affected by the hypoxic conditions imposed, in particular during the early stages of storage. Sucrose synthase expression appeared to be highly sensitive to changes in low oxygen concentration. Ethylene biosynthesis (ACC synthase and oxidase) genes showed marked differences in their expression in relation to the static and dynamic protocols and the hypoxic conditions, as well as six Ethylene Responsive Factors (ERF) genes, some of them possibly involved in the oxygen sensing mechanism operating in fruit tissues.

Dynamic Controlled Atmosphere (DCA) represents one of the latest technical innovations for the long storage of apples (and few other fruit crops) (Tonutti, 2015). With this technology, fruits are kept under extremely low oxygen concentrations (0.4 kPa or lower) that are beneficial for maintaining specific quality parameters (e.g., flesh firmness, acidity). However, by activating anaerobic metabolism, an accumulation of ethanol takes place. Low concentra­tions of ethanol are desirable in terms of improving organoleptic traits, reducing the incidence of chilling injuries (e.g., superficial scald) and limiting ethylene biosynthesis (Dixon and Hewett, 2000; Scott et al., 2000; Weber et al., 2020). Yet, the accumulation of excessive ethanol results in the appearance of off­fla­vors and physiological disorders (Pedreschi et al., 2009). Thus, based on different stress indicators (chlorophyll fluorescence ­CF­, respiratory quotient ­RQ­, and ethanol concentration), oxygen must be promptly adjusted (increased) to reach “safe” con­centrations. The imposed extreme hypoxic conditions induce selective responses of apple tissues starting from the modulation of gene expression involved in particular in primary metabolism and hormone (mainly ethylene) physiology (Cukrov et al., 2019). In Granny Smith, one of the apple cultivar most fre­quently stored in DCA, differential expression of sucrose synthase (SuSy), alcohol dehydrogenase (ADH) and pyruvate­related metabolism (lactate dehydrogenase, LDH, pyruvate decarboxylase, PDC, and alanine aminotransferase, AlaAT) genes was detected when comparing 0.4 with 0.8 kPa oxygen concentration (Cukrov et al., 2016). When, according to the DCA protocol, oxygen level is increased from the lowest applied concentrations, molecular and metabolic rearrangements rapidly occur, with changes in both primary and secondary metabolism (Brizzolara et al., 2019), indicating that highly reac­tive mechanisms and oxygen sensors are present in apple cortex. The expression of genes involved in fer­mentative metabolisms (e.g., ADH), in secondary metabolism (e.g., phenylpropanoid pathway), hor­monal responses and regulatory mechanisms (ethyl­ene biosynthesis, ERFs) resulted to be affected by the oxygen switch. The duration of the storage and the oxygen con­centrations applied obviously play a key role in deter­mining the fruit metabolic responses and the dynam­ics of fermentative metabolite accumulation. In addi­tion, apple varieties react differently to extremely low oxygen conditions during storage, in particular in terms of fermentation, ethanol production and accu­mulation (Thewes et al., 2019; Brizzolara et al., 2020; Thewes et al., 2021 a; Park et al., 2022). Zanella and Stürz (2015) showed that, differently from eight other varieties, ‘Red Delicious’ apples react signifi­cantly and accumulate higher ethanol levels under hypoxia, and in a specific comparison between Granny Smith and Red delicious (Brizzolara et al., 2017), it was reported that the latter considerably accumulated ethanol under both ULO (0.9 kPa oxy­gen) and DCA (0.2­0.55 kPa oxygen) conditions, as also observed by Lumpkin et al. (2014). Among important commercial apple varieties, the responses of Red Delicious to controlled atmosphere (CA) and DCA still need to be compared and clarified, which makes this cultivar a genotype of interest in terms of both applied aspects and physiological stud­ies related to DCA conditions and different oxygen regimes and concentration adjustment protocols.

DOI: https://doi.org/10.36253/ahsc-14180

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