Michael Lay-Yee

An ethylene-related cDNA from ripening apples

We report the isolation of a ripening-related apple cDNA which is complementary to a mRNA which may be involved in ethylene production. Poly(A)+ RNA was extracted from cortical tissue of ripe apple fruit (Malus domestica Borkh cv. Golden Delicious) and a cDNA library constructed in the plasmid vector pSPORT. The library was screened with pTOM13, a tomato cDNA clone thought to code for ACC oxidase in that fruit. An apple cDNA clone (pAP4) was isolated and sequenced. The 1182 bp cDNA insert includes an open reading frame of 942 bp, and shows strong homology with reported tomato and avocado sequences, both at the nucleic acid and amino acid levels. The polypeptide has a calculated molecular mass of 35.4 kDa and a calculated pI of 5.15. In apple cortical tissue, expression of pAP4-complementary RNA increased with ethylene production by the fruit during ripening. Expression was also enhanced in both ethylene-treated and wounded fruit.

Reduction of chilling injury in the sweet persimmon `Fuyu' during storage by dry air heat treatments

Abstract Sweet persimmon fruit ( Diospyros kaki cv. Fuyu L.) were heat-treated with dry air at 34–50°C for durations of 0.5–10 h, cooled immediately, and stored for 6.5 weeks at 0°C in air. After storage, fruit were held for 3 days at 20°C, and evaluated for quality. Extreme chilling injury (flesh gelling) was observed in non-heated fruit. Associated with development of chilling injury was increased ethylene production 1 day after removal from storage, and decreased fruit firmness, juiciness, titratable acidity, and colour ( L value, hue and chroma) 3 days after removal from storage. Increasing the temperature of the hot air heat treatment (HT) progressively reduced flesh gelling and alleviated chilling injury. However, external and internal browning increased with greater HT temperatures and duration. Least chilling injury and external browning were observed at 47°C for 0.5–3 h. However, the levels of browning observed in these treatments are likely to be commercially unacceptable. Thus, although hot air HTs confer significant protection against low temperature damage to persimmon fruit, heat damage needs to be reduced before such benefits can be realised commercially.

Antisense apple ACC-oxidase RNA reduces ethylene production in transgenic tomato fruit

Transgenic tomato (Lycopersicon esculentum) plants were produced which expressed antisense copies of an apple fruit ACC-oxidase RNA. In the fruit of the primary transformants, ethylene production was reduced by over 95% in one of the lines assessed, and to a lesser extent in the other lines. The line showing the greatest reduction in ethylene production showed a delay in the development of colour in the transgenic fruit. Northern analysis of steady-state RNA levels using strand-specific probes indicated that fruit of the low-ethylene line had very low levels of ACC-oxidase sense-RNA, and high levels of antisense RNA. In other lines, high levels of antisense RNA were not always associated with a reduction in the levels of either sense-transcript or ethylene production. The successful reduction in ethylene production confirms the usefulness of using tomato as a model system for testing specific ripening-related genes from heterologous fruit species such as apple.

Hot-water treatment for insect disinfestation and reduction of chilling injury of ‘Fuyu’ persimmon

Abstract ‘Fuyu’ persimmon ( Diospyros kaki L.) fruit were hot-water treated at temperatures ranging from 47 to 54 °C, for durations from 2.5 to 120 min (depending on temperature), with air- and water-treated fruit (20 °C for 120 or 60 min, respectively) as controls. Following treatment, fruit were stored at 0 °C in air for 6.5 weeks then held at 20 °C for 5 days and assessed for quality. Whereas a number of hot-water treatments caused damage in the form of skin or flesh browning, no damage was observed with certain treatments which showed potential as disinfestation treatments (47 °C for 90 and 120 min, 50 °C for 30 and 45 min, 52 °C for 20 and 30 min, and 54 °C for 20 min). These treatments also reduced the incidence and severity of chilling injury observed in fruit following cold storage, relative to that found in controls. Results suggest that hot-water treatment shows potential for both disinfestation and maintaining quality of persimmon fruit during cold storage.

Response of ‘Royal Gala’ apples to hot water treatment for insect control

Abstract The response of ‘Royal Gala’ apples ( Malus domestica Borkh.) to hot water treatments (HWTs) for the control of quarantine leafroller species was investigated. ‘Royal Gala’ apples from two orchards from each of two regions of New Zealand, and up to three harvest dates, were hot water treated at 44, 45 or 46°C for 35, 40 or 45 min. Following treatment, fruit were placed in cold storage at 0.5°C for 0, 4, 7 or 10 weeks, then held at 20°C for 7 days prior to assessment for quality. A number of HWTs were associated with damage in the form of skin browning and internal breakdown. Incidence of damage increased with increasing temperature and treatment duration, and with increasing length of time in cold storage. Incidence of HWT-associated damage varied between regions, harvest dates and orchards. Early harvest fruit had lower levels of damage than mid and late harvest fruit. A HWT of 44°C for 35 min, followed by 7 or 10 weeks of cold storage at 0.5°C, was found to be tolerated by all fruit tested. This treatment is effective in controlling the quarantine leafroller pests, Epiphyas postvittana (Walker), Planotortrix octo Dugdale and Ctenopseustis obliquana (Walker).

Response of ‘Hayward’ kiwifruit to high-temperature controlled atmosphere treatments for control of two-spotted spider mite (Tetranychus urticae)

Abstract The response of ‘Hayward’ kiwifruit [ Actinidia deliciosa (A. Chev.) C.-F. Liang et A.R. Ferguson var. deliciosa ] to high-temperature controlled atmosphere (CA) treatments for control of two-spotted spider mite ( Tetranychus urticae Koch) was investigated. Mean LT 99 values for two-spotted spider mite inoculated onto kiwifruit and exposed to 0.4% O 2 , 20% CO 2 (CA) at 40 °C, were 5.4 ± 0.1 h and 8.1 ± 0.3 h for the non-diapausing and diapausing forms, respectively. ‘Hayward’ kiwifruit were subjected to 40 °C for 7 or 10 h in CA (treatments identified as giving 100% mortality for non-diapausing and diapausing two-spotted spider mites, respectively) or in air. Following treatment, fruit were cooled in ambient water or ambient air, stored at 0 °C in air for eight weeks, then held at 20 °C overnight and assessed for quality. Relative to non-treated controls, no significant damage was observed with fruit subjected to 40 °C air treatments. With CA treatments, no significant damage was observed with fruit treated for 7 h followed by hydrocooling. The 7 h CA treatment without hydrocooling and 10 h CA treatments with hydrocooling showed only slight damage, while the 10 h CA without hydrocooling had moderate fruit damage. The main disorders found were vascular browning and rots. Following storage, flesh firmness of fruit treated at 40 °C in air for 10 h or in CA for 7 and 10 h, with and without hydrocooling, was lower than that of non-treated controls.

The heat shock response is involved in thermotolerance in suspension-cultured apple fruit cells

Summary Cultured apple cells (Malus domestica) were heat-treated at temperatures from 24 to 42 °C and measurements carried out of cell viability and gene expression immediately after heat treatment. The effects on gene expression of temperature, heat treatment duration, and recovery at 25 °C after heat treatment were also studied. An increase in expression of heat shock protein (hsp) mRNA transcripts was detected within 5 °C of the culture growth temperature. Maximum levels of expression, which were maintained for at least 3 h on return to 25 °C after heat treatment, occurred for heat treatments at 38 °C. Cell viability was positively correlated with the expression of hsps. An increase in the expression of hsp transcripts was also associated with a 1 h 38 °C heat pre-treatment that made the apple cells tolerant to a subsequent 1 h 42 °C lethal heat treatment. This acquisition of thermotolerance occurred in the first 24 h after heat treatment and was demonstrated by greater viability of cells that received a pre-treatment prior to a lethal treatment when compared to cells that had received only the lethal heat treatment. The responses to heat stress in cultured apple cells are similar to those in other plant tissues grown at similar ambient temperatures. They suggest that a shift in temperature above growth temperature and its magnitude, rather than the reaching of a threshold temperature, is the stimulus that induces the heat shock response.

Combination high-temperature controlled atmosphere and cold storage as a quarantine treatment against Ctenopseustis obliquana and Epiphyas postvittana on 'Royal Gala' apples

Abstract Third and fifth instar brownheaded leafroller, Ctenopseustis obliquana (Walker), and lightbrown apple moth, Epiphyas postvittana (Walker), infested on apples were exposed to a high-temperature controlled atmosphere (HTCA), 2% O2, 5% CO2 at 40°C, and hot air (HA) at 40°C, and mortality responses determined. Mortality responses of third and fifth instar C. obliquana were similar regardless of treatment type (HTCA or HA). Mean lethal time (LT) estimates for C. obliquana larvae exposed to HTCA were less than those for larvae exposed to HA. No C. obliquana larvae survived 6 h exposure to either HTCA or HA. E. postvittana was more tolerant of both HTCA and HA treatments than was C. obliquana, and fifth instars were the more tolerant life stage. HTCA was significantly more effective against this species than was HA. Third and fifth instar E. postvittana were treated with two HTCA exposures (6 and 8 h) lethal to C. obliquana, and then exposed to cold storage (CS) at 0°C for 7 weeks. The combination of HTCA with CS enhanced treatment efficacy against both life stages. Although a few individuals of both life stages survived exposure to 6 h HTCA+CS, exposure to 8 h HTCA+CS resulted in complete kill. ‘Royal Gala’ apples (Malus domestica Bork.) subjected to HTCA or HA for 6 or 8 h, were either cooled by immersion in water at ambient temperature, or not cooled in water, and then exposed to CS for 7 weeks, before being transferred to 20°C for 7 days and assessed for quality. No significant damage was observed in control, HTCA- or HA-treated apples not cooled in water, though a very low incidence of cavitation and breakdown was observed in water-cooled apples.