Harvested broccoli (Brassica oleracea) responds to high carbon dioxide and low oxygen atmosphere by inducing stress-response genes

Broccoli (Brassica oleracea L.) tissue held in a controlled atmosphere (CA; 10% carbon dioxide and 5% oxygen) senesces more slowly than tissue held in air. CA-treated broccoli tissues lose less water and soluble sugars, have lower protease activity, and have no significant loss of color (hue angle, chlorophyll content) for 96 h after harvest (20 °C, dark) compared to tissue held in air that starts to senesce and yellow after 48 h. The current study examined differential gene expression in broccoli tissues in response to postharvest CA treatment. This genetic analysis was undertaken to identify CA-responsive genes that may act as signaling elements and repress postharvest senescence processes. CA-responsive genes with up- and down-regulated expression (compared to air controls) were isolated after a 6 h CA treatment by differential display-polymerase chain reaction. The candidate CA-responsive genes included a number of novel genes without previously assigned functions, and genes of known function previously found to be regulated by stress (e.g. dehydration, salt stress, low temperature, and sugar starvation).     

UV-C treatment delays postharvest senescence in broccoli florets

Central broccoli heads (cv. de Cicco) were harvested and treated with UV-C light (4, 7, 10, or 14 kJ m⁻²). All treatments delayed yellowing and chlorophyll degradation at 20 °C but the irradiation dose of 10 kJ m⁻² allowed retaining the highest chlorophyll content yet had lower amounts of pheophytins than every treatment other than 7 kJ m⁻². This dose was selected to analyze the effect of UV-C on postharvest broccoli senescence at 20 °C. The UV-C treatment delayed yellowing, chlorophyll a and b degradation, and also the increase in pheophytins during storage. The activity of chlorophyll peroxidase and chlorophyllase was lower in UV-C treated broccoli. Instead, Mg-dechelatase activity increased immediately after the treatment, but after 4 and 6 d this activity was lower in UV-C treated florets than in controls. Treated broccoli also displayed lower respiration rate, total phenols and flavonoids, along with higher antioxidant capacity. The results suggest that UV-C treatments could be a useful non-chemical method to delay chlorophyll degradation, reduce tissue damage and disruption, and maintain antioxidant capacity in broccoli.     

Influence of hot water treatment on brown rot of peach and rapid fruit response to heat stress

Recent results on hot water as an alternative treatment open a new perspective in disease incidence reduction. In the present work peach fruit were wounded, inoculated with conidia of Monilinia laxa and 15 min, 3, 6, 12, 24 and 48 h after inoculation treated by dipping in hot water (HT) at 60 °C for 20 s. The effect of heat treatment on some cell wall genes involved in ripening such as β-galactosidase (β-GAL), pectin lyase (PL), polygalacturonase (PG) and pectin methyl esterase (PME), was analyzed by qRT-PCR. The expression levels of defense related genes, phenylalanine ammonia lyase (PAL) and chitinase (CHI), heat stress-related genes such as heat shock proteins 70 and 90 (HSP70, HSP90), and reactive oxygen species (ROS) scavenging genes were also evaluated by qRT-PCR. A 100% disease incidence reduction, as compared to untreated fruit, was obtained by treating 6 and 12 h after inoculation. Moreover, brown rot was inhibited by 85.7% when fruit were heat-treated 48 h after inoculation. The expression levels of cell wall genes (β-GAL, PL, PG and PME) showed a general decrease in HT fruit as compared to the control, whereas PAL, CHI, HSP70 and ROS-scavenging genes increased their expression level in HT samples with respect to the untreated ones. Our results show a curative activity of heat on peach inoculated with M. laxa 48 h before treatment. Each analyzed gene proved to be differentially expressed following heat treatment.     

Effects of heat treatment on chlorophyll degrading enzymes in stored broccoli (Brassica oleracea L.)

Effects of heat treatment on chlorophyll (Chl) degrading enzyme activities in stored broccoli (Brassica oleracea L.) were determined. Chl contents of broccoli treated at 50 °C for 1 h or without heat treatment (control) decreased after 4 days of storage at 15 °C, whereas the contents of broccoli treated at 50 °C for 2 h showed almost no change after 4 days of storage at 15 °C. Chlorophyllase activity of broccoli heat-treated for 1 or 2 h decreased during storage before the occurrence of yellowing, while the activity of the control showed a slight decrease on day 4. Chl oxidase activity of broccoli heat-treated for 2 h remained unchanged. Chl degrading peroxidase activity in the control markedly increased after 6 days of storage at 15 °C, but an increase in the activities in broccoli treated at 50 °C, especially for 2 h, was suppressed. Six anionic and two cationic isoperoxidases were detected in broccoli on day 0. One of the isoperoxidases, C₂ (Rf 0.3) cationic isoperoxidase, was involved in Chl degradation and the increase in C₂ level was greatly reduced by treatment at 50 °C for 2 h. These results indicate that heat treatment could reduce Chl degradation due to the suppression of Chl degrading enzyme activities.     

Yield response and N-fertiliser recovery of tomato grown under deficit irrigation

In search of new innovations for saving irrigation water, fruit yield response and N-fertiliser recovery of greenhouse grown spring-planted tomato (Lycopersicon esculentum Mill., cv. F1 Fantastic) were assessed as influenced by deficit irrigation, imposed using either conventional deficit irrigation (DI) or partial root drying (PRD). Three irrigation treatments were tested: (1) FULL, control treatment where the full amount of irrigation water, which was measured using Class-A pan evaporation data, was applied uniformly on the two halves of plant-root zone; (2) PRD, 50% deficit irrigation in which wetted and partially dry halves of the root-zone were interchanged every irrigation; (3) DI, conventional deficit irrigation maintained at 50% deficit, compared to FULL irrigation, with water applied on the both halves of the root-zone. During a growth period of 153 days, the highest fruit yield of 145.4 t ha⁻¹ was measured under FULL irrigation treatment, which was followed by PRD and DI treatments with statistically lower (P ≤ 0.01) yields of 114.6 and 103.4 t ha⁻¹, respectively. Irrigation water use efficiencies (IWUE) of both deficit treatments were significantly (P ≤ 0.01) higher (52.7% for PRD and 38.3% for DI) compared to FULL irrigation. Nitrogen-fertiliser recovery was over 70%, with no significant difference among the irrigation treatments. Both deficit treatments (DI and PRD) showed lower values of leaf water potential, photosynthetic rate and stomatal conductance compared to FULL irrigation. Before irrigation, xylem-sap abscisic acid (ABA) concentrations were 28% and 38% higher under water-stressed deficit treatments DI0 and PRD, respectively, compared to FULL irrigation, and the high ABA concentrations was maintained only under PRD effect, following irrigation. The results of this work suggest that PRD practices can be viable and advantageous compared to conventional techniques to minimise crop-yield reductions during deficit irrigation.     

Effects of postharvest treatments on gene expression in Prunus persica fruit: Normal and altered ripening

Peach (Prunus persica) fruit have a short shelf-life, and the most common method employed to delay ripening and increase their postharvest life is cold storage. However, after extended storage at low temperature some cultivars have alterated ripening processes, resulting in a lack of juice and a woolly texture. To improve our understanding of the molecular mechanisms involved in the responses of peach fruit to cold storage we determined gene expression changes of fruit (cv. O’Henry) under different postharvest conditions: ripening (5 days at 21 °C), cold storage (21 days at 4 °C) and induction of woolliness (21 days at 4 °C followed by 5 days at 21 °C).Cluster analyses of genes differentially expressed between treatments revealed unique patterns associated with biological processes that operate during postharvest treatments. Genes up-regulated during postharvest ripening and woolliness include components of ethylene, and aroma biosynthesis as well as oxidative stress response. During cold storage treatment and woolliness, several genes linked to the oxidative stress response increased in abundance, suggesting changes in redox status. Quantitative RT-PCR analysis showed a sequential increase levels of mRNAs encoding key components of cellular stress response. Moreover, after 21 days of cold storage, expression of genes encoding oxidoreductase, catalase, superoxide dismutase and gluthatione reductase was still significantly higher than before cold treatment, suggesting that fruit cells were able to respond to the increased production of ROS that was induced by extended cold storage. In the woolly fruit, up-regulation of stress response genes was accompanied by down-regulation of key components of metabolic pathways that are active during peach ripening. The altered expression pattern of these genes might account for the abnormal ripening of woolly fruit.     

Chlorophyll fluorescence for non-destructive measurement of flavonoids in broccoli

Chlorophyll fluorescence (ChlF) by excitation with two radiation beams of different wavelengths can be used for non-destructive quantitation of epidermal flavonoids in leaves. The method was tested on marketable broccoli heads, but including red light (685 nm) and green light (530 nm) in addition to ultraviolet (UV) radiation (382 nm) and blue light (450 nm) for excitation of ChlF. As a reference, the content of flavonoids was measured with high performance liquid chromatography. To induce flavonoids, postharvest treatment during 12 days with various combinations of visible and UV radiation was tried, but the achieved changes in the flavonoid levels were not statistically significant. Instead a large natural variation of flavonoid content among the broccoli heads was used to correlate it with the fluorescence data. It was possible to estimate the content of flavonoids in flower buds by means of ChlF: relative epidermal absorbance of blue light was well correlated with flavonoid content (r = 0.69, p < 0.001). For quercetin alone the correlation was higher (r = 0.77, p < 0.001). On the other hand, relative epidermal absorbance of UV radiation or green light had a much weaker correlation with flavonoid content (r ≤ 0.40, p < 0.05). Within the broccoli heads, flower buds had 15 times higher levels of flavonoids than the floret stalks, which in turn had five times higher contents than the main stem. Repeated ChlF measurements on individual broccoli heads during cold storage could monitor small but significant changes in flower buds, possibly indicating a breakdown of constituents absorbing UV and green light.     

Vitamin C in broccoli (Brassica oleracea L. var. italica) flower buds as affected by postharvest light,UV-B irradiation and temperature

In this study, the changes in vitamin C, l-ascorbic acid (AA) and l-dehydroascorbic acid (DHA) levels in broccoli flower buds were examined during pre-storage and storage periods, simulating refrigerated transport with wholesale distribution and retail, respectively. Broccoli heads were pre-stored for 4 or 7 days at 0 °C or 4 °C in the dark and then stored for 3 days at 10 °C or 18 °C. During storage the broccoli heads were exposed for 12 h per day to three different levels of visible light (13, 19 or 25 μmol m⁻² s⁻¹) or a combination of visible light (19 μmol m⁻² s⁻¹) and UV-B irradiation (20 kJ m⁻² d⁻¹), or they were stored in the dark. The vitamin C content in broccoli flower buds during storage was significantly affected by pre-storage period and temperature. Higher vitamin C levels in flower buds after storage were observed for broccoli heads pre-stored for 4 days or at 0 °C as compared to those pre-stored for 7 days or at 4 °C. Storage temperature also affected vitamin C in broccoli flower buds, with higher levels observed for broccoli stored at 10 °C than at 18 °C. Hence, vitamin C in broccoli flower buds was demonstrated to decrease together with increasing pre-storage period, pre-storage temperature and storage temperature. AA in broccoli flower buds was influenced mainly by storage temperature and to a minor extent by pre-storage temperature. The DHA level and DHA/AA ratio were stable in flower buds of broccoli pre-stored for 7 days, whereas increasing tendencies for both DHA level and ratio were observed after pre-storage for 4 days. These results indicate a shift in the ascorbate metabolism in broccoli flower buds during storage at low temperatures, with its higher rate observed for broccoli pre-stored for shorter time. There were no effects of the light and UV-B irradiation treatments on vitamin C, AA and DHA levels in broccoli flower buds.     

Sucrose feeding of Cut Dendrobium inflorescences promotes bud opening,inhibits abscission of open flowers,and delays tepal senescence

Inflorescences of Dendrobium cv. Khao Sanan were held in distilled water (controls) or in solutions containing two antimicrobial compounds (silver nitrate and 8-hydroxyquinoline sulfate) with or without 4 g L⁻¹ sucrose or 4 g L⁻¹ glucose. Sugar + antimicrobial compounds promoted bud opening and largely prevented abscission of open flowers. It also resulted in a delay of tepal senescence. No clear effect was found of sucrose feeding on the concentrations of sucrose, glucose and fructose in the tepals of open flowers, but an increase in sucrose, glucose, and fructose concentrations was found in the column + labellum. It is concluded that the effect of sugar feeding on abscission and tepal senescence might relate to a reduction of ethylene sensitivity. It was previously concluded that tepal senescence in Dendrobium flowers was not regulated in the tepals but in the column. It is not clear how the increased in sugar levels in the column + labellum relates to the delay of tepal senescence after sugar feeding.     

Differential effect of regulated deficit irrigation on growth and photosynthesis in young peach trees intercropped with grass

The effects of a moderate soil water deficit on several shoot growth variables (1st and 2nd order shoot growth and final leaves number, final height and final number of 2nd order shoots) and on net photosynthesis were studied in young peach trees during the two years following plantation (January 2014). Trees were either fully irrigated (C), subjected to moderate water deficit (RDI) or subjected to moderate water deficit and associated with a grass-legume mixture on the entire orchard floor (RDI +G). Irrigation was scheduled according to soil water potential target ranges in order to keep C trees above −0.02 MPa, i.e. at field capacity, and RDI and RDI + G trees between −0.04 MPa and −0.06 MPa. The level of water deficit obtained was moderate but yet significantly reduced by 50% overall tree growth in 2014 in RDI. This reduction was enhanced when water deficit lasted longer and when it was associated with grass in RDI + G. No reduction in growth variables occurred in RDI in 2015 due to the shorter duration of water deficit. Overall reduction was observed in 2015 in RDI + G mostly due to a carry-over effect of the previous year. Net photosynthesis was reduced by the longer and more intense water deficit in 2014, but was not reduced during the soil water deficit in 2015. An indicator of plant process sensitivity to water deficit, taking into account the variable reduction with regards to the control, the water deficit intensity and its duration was used to classify shoot growth variables and net photosynthesis according to their sensitivity to water deficit. Variables could be classified according to the following order of ascending sensitivity: net photosynthesis <1st order final leaf number < final tree height <1st order final shoot length <2nd order final leaf number <2nd order final shoot number <2nd order final shoot length. Applying a moderate water deficit combined with full grass cover drastically reduces overall tree size due to grass competition.     

Pre-harvest calcium application increases biomass and delays senescence of broccoli microgreens

Microgreen consumption has been steadily increasing in recent years due to consumer awareness of their unique color, rich flavor, and concentrated bioactive compounds. However, industrial production and marketing is limited by their short shelf-life associated with rapid deterioration in product quality. This study investigated the effect of pre-harvest calcium application on the post-harvest quality and shelf-life of broccoli microgreens. Broccoli microgreen seedlings were sprayed daily with calcium chloride at concentrations of 1, 10 and 20 mM, or water (control) for 10 days. The fresh-cut microgreens were packaged in sealed polyethylene film bags. Package headspace atmospheric conditions, overall visual quality and tissue membrane integrity were evaluated on days 0, 7, 14, and 21, during 5 °C storage. Results indicated that the 10 mM calcium chloride treatment increased the biomass by more than 50%, and tripled the calcium content as compared to the water-treated controls. Microgreens treated with 10 mM calcium chloride spray exhibited higher superoxide dismutase and peroxidase activities, lower tissue electrolyte leakage, improved overall visual quality, and reduced microbial growth during storage. Furthermore, calcium treatment significantly affected expression of the senescence-associated genes BoSAG12, BoGPX6, BoCAT3 and BoSAG12. These results provide important information for commercial growers to enhance productivity and improve postharvest quality and shelf-life, potentially enabling a broadening of the retail marketing of broccoli microgreens.     

Assessment of soybean yield with altered water-related genetic improvement traits under climate change in Southern Brazil

Water deficit is a major factor responsible for soybean yield gap in Southern Brazil and tends to increase under climate change. An alternative to reduce such gap is to identify soybean cultivars with traits associated to drought tolerance. Thus, the aim of this study was to assess soybean adaptive traits to water deficit that can improve yield under current and future climates, providing guidelines for soybean cultivar breeding in Southern Brazil. The following soybean traits were manipulated in the CSM-CROPGRO-Soybean crop model: deeper root depth in the soil profile; maximum fraction of shoot dry matter diverted to root growth under water stress; early reduction of transpiration under mild stress; transpiration limited as a function of vapor pressure deficit; N₂ fixation drought tolerance; and sensitivity of grain filling period to water deficit. The yields were predicted for standard and altered traits using climate data for the current (1961–2014) and future (middle-century) scenarios. The traits with greater improvement in soybean yield were deeper rooting profile, with yield gains of ≈300 kg ha⁻¹, followed by transpiration limited as a function of vapor pressure deficit and less drought-induced shortening of the grain filling period. The maximum fraction of shoot dry matter diverted to root and N₂ fixation drought tolerance increased yield by less than 75 kg ha⁻¹, while early reduction of transpiration resulted in a small area of country showing gains. When these traits were combined, the simulations resulted in higher yield gains than using any single trait. These results show that traits associated with deeper and greater root profile in the soil, reducing transpiration under water deficit more than photosynthesis, creating tolerance of nitrogen fixation to drought, and reducing sensitivity of grain filling period to water deficit should be included in new soybean cultivars to improve soybean drought tolerance in Southern Brazil.     

Nitrogen uptake and N-use efficiency of Mediterranean cotton under varied deficit irrigation and N fertilization

Efficient N management is essential to optimize yields and reduce degradation of the environment, but requires knowledge of deficit irrigation effects on crop yields and crop N outputs. This study assessed the N content and N-use efficiency of cotton over the 2008 and 2009 growing seasons in a single field site of the Thessaly Plain (central Greece). The experiment consisted of nine treatments with three fertilizer rates (60, 110 and 160 kg N ha⁻¹) split into three irrigation levels (approx. 1.0, 0.7 and 0.4 of the amount applied by the producer). Reduced water supply induced a shift in the distribution of N within the plant with seeds becoming an N sink under conditions of water stress. Total crop N increased linearly with irrigation level and reached a maximum average of 261 and 192 kg N ha⁻¹ in 2008 and 2009, respectively. Fertilizer application did not trigger a crop N or yield response and indicated that N inputs were in excess of crop needs. Variation in weather patterns appeared to explain annual differences of nitrate-N in the top soil and N uptake by the crop.The index of lint production efficiency (iNUE) detected crop responses caused by irrigation and annual effects, but failed to account for excessive N inputs due to mineral fertilizer applications. A maximum average iNUE of 9.6 was obtained under deficit irrigation, whereas an iNUE of 8.1 was obtained under 40 cm irrigation when crop N uptake was not excessive (192 kg ha⁻¹ in 2009). In contrast, NUE, as an estimator of N recovery efficiency, identified excessive fertilizer inputs as N losses to the environment and indicated that 60 kg N ha⁻¹ was a rate of high N removal efficiency and long-term N balance. However, NUE failed to account for crop N responses to irrigation and weather/management patterns. In this case study, neither index was able to detect all the factors influencing the N mass balance and both were required in order to provide a comprehensive evaluation of the environmental performance of our cropping system.     

Impact of edible coatings and mild heat shocks on quality of minimally processed broccoli (Brassica oleracea L.) during refrigerated storage

The effects of edible coatings and mild heat shocks on quality aspects of refrigerated broccoli were studied. Minimally processed broccoli was coated with either chitosan or carboxymethyl-cellulose with or without a previous application of a mild heat shock of 1.5 min at 50 °C. Product was packaged in multilayered polyolefin bags and stored at 5 °C for 18 d. Quality parameters such as weight loss, texture, colour, ascorbic acid content, total chlorophyll content, oxygen concentration inside the bags, browning potential, mesophilic aerobic counts, and sensory quality, were evaluated during storage. Edible coatings exhibited a beneficial impact on broccoli quality. The weight loss in uncoated broccoli was found to be between 2 and 5 times higher compared to coated samples. During storage, coated florets from both thermally and non-thermally treated samples, presented higher retention of the (?a*/b*) ratio indicating better green colour retention and a reduced rate of floret yellowing. Chitosan coating always presented the lower ascorbic acid degradation rates (twofold lower compared with control samples). Broccoli texture for uncoated samples increased significantly during storage. However, for carboxymethyl-cellulose coated broccoli a slight increase in texture was observed while for chitosan coated broccoli no significant changes in texture were observed throughout the storage period. After the edible coating application the microbial broccoli load dropped by around 1.5 and 0.9 logarithmic units in chitosan and carboxymethyl-cellulose films, respectively. During storage, the application of chitosan coating significantly reduced total microbial counts in the thermally and non-thermally treated uncoated samples. Among the assayed edible coatings, chitosan effectively maintained quality attributes and extended shelf life of minimally processed broccoli. The single application of a mild heat shock had a measurable influence in reducing weight loss, enzymatic browning in broccoli stems, and in delaying yellowing of broccoli florets. Moreover, chitosan coating combined with a mild heat shock showed the best performance for long-term refrigerated storage of minimally processed broccoli.     

Combined effect of ultrasound,mild heat shock and citric acid to retain greenness,nutritional and microbiological quality of minimally processed broccoli (Brassica oleracea L.): An optimization study

Response surface methodology (RSM) and Box–Behnken design were used to study the combined hurdle effect of mild heat time (1–5 min) at 50 °C, ultrasonic processing time (0–10 min) and citric acid concentration (0–2%) on the quality of refrigerated broccoli after 10 d of storage at 5 °C. Treatment effects were evaluated on weight loss, superficial colour (hue angle (H°) and total colour difference (ΔE)), headspace gas composition (O₂ and CO₂), overall browning potential, chlorophyll content, ascorbic acid content, mesophilic counts and overall visual quality (OVQ) and optimize the process by means of the desirability function. Predicted models were found to be significant with high regression coefficients (91–97%). High regression coefficients indicated that second-order polynomial models could be used to predict and optimize the quality retention in minimally processed broccoli during storage. The mesophilic counts, ascorbic acid content and the overall visual quality were significantly influenced by the three independent variables either independently or interactively. Both thermal and ultrasonic treatments were found to be critical factors influencing changes in chlorophyll content, O₂ concentration inside the package, hue angle and ΔE. On the other hand, thermal treatment and citric acid concentration were found to be significant on overall browning potential. By using the desirability function approach and considering superficial colour parameters, O₂ concentration, mesophilic counts, browning potential, ascorbic acid and chlorophyll content, the optimum processing conditions were 7.5 min of ultrasonic treatment, 3 min of a heat shock treatment and a citric acid concentration of 1.5%. These results were in good agreement with the maximum found from the canonical analysis performed from the response surface when only considering sensorial analysis. Under these optimal processing conditions it is possible to employ citric acid treatment in combination with ultrasonic and thermal treatments as hurdles for retention of green colour, nutritional quality, microbial control and for extending shelf life of refrigerated minimally processed broccoli.     

Wound response in orange as a resistance mechanism against Penicillium digitatum (pathogen) and P. expansum (non-host pathogen)

Penicillium digitatum is the most devastating postharvest pathogen of citrus. In addition, Penicillium expansum is the main pathogen of pome fruit, although recent studies have demonstrated its ability to infect oranges under some conditions. In this study, we evaluated wound response in ‘Valencia’ oranges harvested at three different maturity stages and the effect of wound response on the establishment of both pathogens when fruit were stored at two different temperatures (20 and 4 °C). The effect of wounding and pathogen inoculation on lignin content, was also quantified. Lastly, the expression of several phenylpropanoid pathway-related genes was also analyzed by semi-quantitative RT-PCR. Results indicated that, in general, P. digitatum exhibited lower decay incidence and severity as time between wounding and inoculation increased. Decay incidence and severity were higher in fruit from the over-mature harvest than in fruit from immature and commercial harvests. P. expansum was able to infect fruit at 20 °C but lesions were small compared to lesion size of fruit stored at 4 °C. Lignin content in wounded fruit (control) and in samples wounded and inoculated with P. expansum was highest in fruit from the immature harvest at 7 d post-wounding and inoculation. Wounded fruit had higher expression of pal1, comt1 and pox1 genes at 48 h than at 24 h. However, samples inoculated with P. digitatum showed lower expression at 48 h than at 24 h. Our results indicated that maturity and storage temperature play an important role in orange wound response.