Rhodosporidium paludigenum induces resistance and defense-related responses against Penicillium digitatum in citrus fruit

Induced disease resistance against plant pathogens is a promising non-fungicidal decay control strategy. In this study, a potential biocontrol yeast, Rhodosporidium paludigenum, was investigated for its induction of disease resistance against Penicillium digitatum in citrus fruit. The results showed that R. paludigenum is the most effective yeast among three selected yeasts in stimulating the resistance of citrus fruit to green mold. When R. paludigenum was applied 48–72 h before inoculation with P. digitatum, disease incidence and disease severity in citrus fruit significantly decreased. Application of R. paludigenum at concentrations of 1 × 10⁸ and 1 × 10⁹ cells mL⁻¹ respectively resulted in 49.6% and 52.5% reductions in the percentage of infections. Induction of resistance to P. digitatum by R. paludigenum treatment significantly enhanced the activities of defense-related enzymes, including β-1,3-glucanase, phenylalanine ammonia-lyase, peroxidase, and polyphenoloxidase, which may be an important mechanism by which the biocontrol yeast reduces the fungal disease of citrus fruit caused by P. digitatum.     

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.     

1-MCP regulates ethylene biosynthesis and fruit softening during ripening of ‘Tegan Blue’ plum

To investigate the effects of postharvest application of 1-MCP on ethylene production and fruit softening, activities of ethylene biosynthesis and fruit softening enzymes were measured during postharvest ripening of plum (Prunus salicina Lindl. cv. Tegan Blue) fruit after being exposed to 1-MCP (0, 0.5, 1.0 or 2.0 μL L⁻¹) at 20 ± 1 °C for 24 h. Following the treatments, fruit were allowed to ripen at ambient temperature (20 ± 1 °C), and ethylene production in fruit, activities of ACS and ACO, ACC content and fruit softening enzymes (PE, EGase, exo-PG and endo-PG) in fruit skin and pulp were recorded at different intervals. Postharvest application of 1-MCP significantly delayed and suppressed the climacteric ethylene production with reduction in the activities of ethylene biosynthesis enzymes (ACS, ACO) and ACC content, and fruit softening enzymes (PE, EGase, exo-PG and endo-PG) in the skin as well as in pulp tissues. The reduction was more pronounced with increased concentrations of 1-MCP. 1-MCP treated fruit showed different rates of fruit softening and activities of ethylene biosynthesis enzymes in the skin and pulp tissues which warrant further investigation on regulation of gene expression related to these enzymes with the inhibitory effect of 1-MCP.     

Degreening behavior in ‘Fallglo’ and ‘Lee × Orlando’ is correlated with differential expression of ethylene signaling and biosynthesis genes

Two citrus types (‘Fallglo’ and ‘Lee × Orlando’) exhibiting differential fruit degreening response when treated with ethylene were selected. Fruit were harvested at commercial maturity but at different developmental periods (Harvest I, II and III). Rate of color change was greater in ‘Fallglo’ than in ‘Lee × Orlando’ when fruit were treated with 5 μL L⁻¹ of ethylene for 24 h. After 24 h of transfer of fruit to ethylene-free storage, rate of change decreased in ‘Fallgo’ and exhibited varied response in ‘Lee × Orlando’ depending on harvest date. ‘Fallglo’ fruit from Harvests I and II were completely degreened at the end of storage for 7 d; however ‘Lee × Orlando’ were not and were green in color. No difference in seedling triple response was observed between ‘Fallglo’ and ‘Lee × Orlando’ and sequences of the four ethylene receptors were identical between them. Expression of genes involved in ethylene biosynthesis and signaling pathways were studied in flavedo to test if differences in these pathways were correlated with differential ethylene sensitivity of the citrus types. Basal levels of ACS2 and ACO expressions declined as maturity progressed, and ethylene-induced expression of ACS1 and ACO were influenced by fruit maturity. At Harvests I and II, ethylene-induced increase in ACS1 and ACO expressions and ACC levels were greater in ‘Fallglo’ than in ‘Lee × Orlando’. Ethylene treatment influenced MACC content only during Harvest I in ‘Lee × Orlando’. MACC levels were generally higher in ‘Lee × Orlando’ than in ‘Fallglo’. Expressions of ETR1 and ETR2 were ethylene responsive in ‘Fallglo’ and only ETR1 expression was ethylene responsive in ‘Lee × Orlando’. Ethylene had more impact on ETR1 expression in ‘Fallglo’ than in ‘Lee × Orlando’. Ethylene had a negative effect on ETR3 expression which was more pronounced in ‘Lee × Orlando’ than in ‘Fallglo’. Expressions of ERS1, CTR1, EIN2, EIL1 and EIL2 were not affected by ethylene in both citrus types. Expression of chlorophyllase gene and rate of total chlorophyll degradation were higher in ‘Fallglo’ than in ‘Lee × Orlando’ during ethylene treatment. Differential degreening behavior of ‘Fallglo’ and ‘Lee × Orlando’ correlated with peel maturity, and factor(s) downstream of ethylene signaling but upstream of ethylene biosynthesis play a role in the differential sensitivity.     

Use of citral incorporated in postharvest wax of citrus fruit as a botanical fungicide against Penicillium digitatum

The antifungal activity of citral against Penicillium digitatum, the causal agent of citrus green mold, was tested by in vitro and in vivo experiments. In vitro assays showed that the minimum inhibitory concentration and the minimum fungicidal concentration (MFC) were both 4000 μL L⁻¹. Results of in vivo tests demonstrated that wax + citral (1× MFC) treatment did not effectively inhibit the growth of P. digitatum in Ponkan mandarin fruit, whereas wax + citral (10× MFC) treatment significantly decreased the incidence of green mold after 6 days of storage at 25 ± 2 °C. Wax + citral (10× MFC) treatment remarkably increased the content of vitamin C and antioxidant enzyme activities such as catalase, superoxidase dismutase, and peroxidase but decreased the activities of phenylalanine ammonia lyase, polyphenol oxidase, and malonaldehyde. The treatment had minor effects on the pH, coloration index, and total soluble solids. This study provided theoretical data for the practical application of citral on citrus fruit quality during postharvest storage.     

Influence of time from harvest to 1-MCP treatment on apple fruit quality and expression of genes for ethylene biosynthesis enzymes and ethylene receptors

A strong potent inhibitor of ethylene action, 1-methylcyclopropene (1-MCP) maintains apple fruit quality during storage. To understand the influence of time after harvest until 1-MCP treatment, we studied expression patterns of genes for ethylene biosynthesis enzymes and ethylene receptors in two apple cultivars, ‘Orin’ and ‘Fuji’, which differ in ethylene production. Ethylene production and expression of MdACS1, MdERS1, and MdERS2 were suppressed in all 1-MCP-treated ‘Fuji’ fruit, but in ‘Orin’, the later 1-MCP was applied after harvest, the less was the suppression of ethylene production and expression of these genes. In fruit in which 1-MCP had low efficacy (e.g., ‘Orin’ treated at 7 DAH), ethylene production and the level of MdERS1 were briefly reduced by 1-MCP treatment at 2 days after treatment, then began to increase. Since ethylene receptors negatively regulate the ethylene signalling pathway, the increased levels of ethylene production and ethylene receptors after 1-MCP treatment might reduce 1-MCP efficacy.     

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.     

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.     

Genotypic differences in post-storage photosynthesis and leaf chloroplasts in response to ethylene and 1-methylcyclopropene in Aglaonema

This study investigated the effects of ethylene in storage and 1-methylcyclopropene (1-MCP) pretreatment on post-storage leaf senescence as measured by changes in photosynthesis and chloroplast degradation of two Aglaonema cultivars. Potted plants of ‘Chalit's Fantasy’ and ‘White Tip’ with or without 1-MCP treatment (600 nL L⁻¹ 1-MCP for 6 h) were exposed to 3.0 μL L⁻¹ ethylene, while being stored for 5 d at 16 °C in the dark, and then placed under an indoor environment for further observation. Plants that did not receive 1-MCP and ethylene served as controls. Ethylene did not affect the stomatal conductance in either cultivar. Ethylene reduced the net CO₂ assimilation rate and F_v/F_m (potential photochemical efficiency of photosystem II) in ‘White Tip’, but not in ‘Chalit's Fantasy’. Chloroplast number in a palisade or spongy mesophyll cell did not differ among treatments in ‘Chalit's Fantasy’. However, ethylene-treated ‘White Tip’ had fewer chloroplasts in the mesophyll cells, had more and larger plastoglobules in the chloroplasts, and had looser granal stacking with enlarged thylakoid lumens. ‘Chalit's Fantasy’ plants that were treated with 1-MCP before exposure to ethylene had higher net CO₂ assimilation rates and stomatal conductance than the control or plants that were exposed to ethylene without 1-MCP pretreatment. 1-MCP pretreatment mitigated the injurious effect of ethylene on ‘White Tip’ by increasing net CO₂ assimilation rate and F_v/F_m, and maintaining the quantity and structural integrity of chloroplasts.     

Physiological response of ‘Larry Ann’ plums to cold storage and 1-MCP treatment

The aim of this work was to study the specific effects of low temperature and 1-MCP treatment on ethylene metabolism and oxidative behaviour in plums (Prunus × salicina cv. Larry Ann). Control fruit were stored at 20 °C or 0 °C and the 1-MCP (625 nL L^?1) treated fruit at 0 °C. Changes in the kinetics of ethylene production upon removal were related to changes in ACC metabolism (ACC and MACC levels), oxidative behaviour (H₂O₂ content) and enzymatic antioxidant potential (SOD, CAT and POX enzymes) during cold storage. Low temperature stress inhibited the synthesis of MACC, which appeared to be the basic process that regulated ACC and ethylene production at ambient temperature. Although 1-MCP treatment inhibited ethylene production and ACC accumulation in the cold, it did not inhibit the accumulation of MACC. Neither cold nor 1-MCP treatment induced oxidative stress. Nevertheless, the 1-MCP treatment significantly impaired the increase in POX activity observed during cold storage. Collectively these results showed the underlying role that ACC metabolism plays in the ripening behaviour of cold-stored plums, confirming previous results. The results also indicate that MACC and malonyl transferase activity are the key regulatory factors that control ripening and possibly some ethylene-related disorders such as chilling injury in cold-stored plums.     

1-MCP pretreatment prevents bud and flower abscission in Dendrobium orchids

Dendrobium orchid inflorescences were treated for 4 h at 25 °C with or without 100–500 nl/l 1-MCP and were then placed in water at 25 °C to follow abscission. In controls, depending on the experiment, 20–80% of the floral buds and 0–20% of the open flowers abscised within 1 week. The 1-MCP pretreatment largely prevented this abscission. If flowers were exposed to 1.0 μl/l ethylene for 3 days, all floral buds and all open flowers abscised within the 3 days of treatment. 1-MCP treatment just prior to ethylene treatment largely prevented the ethylene effect. Treatment with STS was as effective as treatment with 1-MCP. Dendrobium inflorescences are usually shipped by air in cardboard boxes lined with plastic film. The stem ends are placed in plastic tubes filled with water. After shipment and placement in water, a considerable percentage of the buds, and some flowers, abscise. This is probably due to elevated ethylene concentrations inside the boxes. Treatment of the inflorescences with 100–500 nl/l 1-MCP prior to simulated air transport largely prevented abscission during vase life. 1-MCP treatment inhibited ethylene production of the inflorescences by lowering both ACC synthase in open flowers and ACC oxidase activity in floral buds.     

Internal ethylene concentrations in apple fruit at harvest affect persistence of inhibition of ethylene production after 1-methylcyclopropene treatment

Factors that affect the efficacy of 1-methycyclopropene (1-MCP) treatment of apples [Malus sylvestris (L.) Mill var. domestica (Borkh.) Mansf.] include cultivar and maturity. In this study, ‘McIntosh’, ‘Cortland’ and ‘Empire’ apples were categorized by internal ethylene concentrations (IECs) at harvest, treated with 1 μL L⁻¹ 1-MCP, and the IECs of individual fruit followed at 30 d intervals during air storage at 0.5 °C for 90 d. IECs at harvest ranged from <0.5 μL L⁻¹ to ≥100 μL L⁻¹, 51 < 100 μL L⁻¹, and 10 < 50 μL L⁻¹ for ‘McIntosh’, ‘Cortland’ and ‘Empire’, respectively. 1-MCP treatment resulted in a decrease of IECs in fruit of all cultivars by day 30 after harvest. During subsequent storage IECs remained low in fruit with <1 μL L⁻¹ at harvest, but in ‘McIntosh’, ‘Cortland’ increased in proportion to IECs at harvest, but not in ‘Empire’. The importance of initial IECs in fruit on the persistence of 1-MCP inhibition of ethylene production was confirmed in a further experiment, in which IECs in untreated and 1-MCP treated ‘McIntosh’ and ‘Empire’ apples were measured for up to 194 d. 1-MCP also decreased 1-aminocyclopropene-1-carboxylic acid (ACC) concentrations in fruit. The results of our study are consistent with the hypothesis that IEC modulates the sensitivity of climacteric fruit to 1-MCP.     

1-MCP partially alleviates dehydration-induced abscission in cut leaves of the fern Nephrolepis cordifolia

Fern leaves, also called fronds, are often used in bouquets. Leaves of the sword fern (Nephrolepis cordifolia) consist of a central vascular tissue, with numerous leaflets (pinnae) at each side. Leaves that have been cut and immediately placed in water show abscission of the pinnae, starting from about day 4 of vase life, with 50% pinnae abscission on day 13. The onset of pinnae abscission was hastened by a period of dehydration (3, 6, 9 or 12 h at 25 °C). The time to 50% pinnae abscission was between 7.0 and 4.7 days after 3 h and 12 h of dehydration, respectively. Dehydration treatments might induce air emboli in the xylem, but in these experiments did not inhibit water uptake. Dehydration did increase the rate of ethylene production of the cut leaves throughout vase life. A 3 h treatment with 1-MCP at concentrations of 200 or 300 nL L^?1 prior to the period of dehydration reduced the rate of ethylene production and reduced the rate of abscission. 1-MCP treatments were also effective if given after the period of dehydration. The data show that pinnae abscission limits the vase life of cut leaves of the sword fern, and that a short period of water stress drastically increases the rate of abscission. The increase in pinnae abscission was correlated with an increase in ethylene production. As 1-MCP alleviated the effect of dehydration on pinnae abscission, the dehydration effect involved ethylene perception. The data suggest that a small water stress induced an autocatalytic rise in ethylene production which was the direct cause of the increase in pinnae abscission.     

1-MCP controls ripening induced by impact injury on apricots by affecting SOD and POX activities

The influence of 1-MCP on the response of apricots to mechanical injury (impact) and the potential involvement of oxidative stress was investigated. Apricots (Prunus armeniaca L. cv. Marietta) picked at an early ripening (commercial harvest) stage (11–11.5 °Brix) were dropped from 30 cm onto a flat, hard surface to simulate an impact injury; fruit were treated with 500 nl 1⁻¹ 1-MCP for 20 h at 20 °C before or after the impact injury. Injured fruit showed a substantial rise in ethylene production after 4 days, while in fruit treated with 1-MCP, this increase started after 6 days, with a production rate lower than that of injured fruit. Increase in the respiration rate was delayed for 1-MCP-treated injured fruit in comparison with untreated injured ones. Tissue softening was reduced by 1-MCP treatment, showing less tissue deformability. Scanning EM analysis of injured tissue revealed healthier cells in 1-MCP treated apricots. 1-MCP-treated the increase of superoxide dismutase activity (SOD) due to mechanical injury in the first 4 days and this behaviour was related to ethylene production. Peroxidase activity (POX) increased in injured tissue immediately but then remained stable; 1-MCP, particularly when applied before the impact, increased POX activity. These results indicate that using 1-MCP can control ripening acceleration of apricots induced by mechanical injury. SOD, POX, and ethylene relationships are discussed.     

The effects of ethylene, 1-MCP and AVG on sprouting in sweetpotato roots

Exogenous ethylene is commonly used as a commercial sprouting inhibitor of potato tubers. The role of ethylene in the control of sprouting of sweetpotato roots, however, is not known. The aim of this study was to investigate the role of ethylene in control of sprouting in sweetpotato roots by observing the effect of an ethylene synthesis inhibitor, aminoethoxyvinylglycine (AVG), and the ethylene antagonist, 1-methylcyclopropene (1-MCP), in the presence and absence of exogenous ethylene on root sprouting and associated sugar accumulation. Continuous exposure to 10 μl L⁻¹ ethylene, 24 h exposure to 625 nl L⁻¹ 1-MCP or dipping in 100 μl L⁻¹ AVG all inhibited sprout growth in sweetpotato roots of two varieties over 4 weeks of storage at 25 °C. The observations that both ethylene on its own and 1-MCP, which inhibits ethylene action, inhibit sprout growth indicate that while continuous exposure to exogenous ethylene leads to sprout growth inhibition, ethylene is also required for sprouting. In potato tubers ethylene is required to break dormancy, while continuous exposure inhibits sprout growth.Monosaccharide concentrations in ethylene, 1-MCP or AVG treated roots were lower than in untreated roots, and for ethylene treated roots this was associated with higher respiration rates. This is consistent with the activation of some additional process by ethylene which uses energy through sugar metabolism. 1-MCP and AVG both inhibited this increase in respiration rate and counteracted the decrease in monosaccharide concentrations. 1-MCP presumably counteracts the ethylene stimulation of this process, while the effect of AVG is attributed to its possible inhibitory effects on protein synthesis.