Natural essential oil compounds with heat treatment to control stem-end rot on grapefruit during postharvest handling and marketing

2015 Annual Report for GS15-149

Project Type: Graduate Student
Funds awarded in 2015: $10,948.00
Projected End Date: 12/31/2016
Grant Recipient: University of Florida
Region: Southern
State: Florida
Graduate Student:
Major Professor:

Natural essential oil compounds with heat treatment to control stem-end rot on grapefruit during postharvest handling and marketing

Summary

Screening of essential oils against mycelial growth of L. theobromae showed carvacrol and thymol had effective inhibition on mycelial growth of L. theobromae. In amended PDA medium, carvacrol and thymol inhibited mycelial growth in a concentration-dependent manner, with higher concentration having a higher efficiency. Both carvacrol and thymol showed effectively curative and preventative activities against Diplodia stem end rot on ‘Ruby Red’ grapefruit in a concentration-dependent manner, with 1.0% of essential oil in wax showing the highest efficiency. No injury was detected when coating fruit with shellac wax containing carvacrol or thymol, even at high concentration (1.0%).

 

 

Objectives/Performance Targets

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Objective 1. Screen essential oil compounds for their efficacy against L. theobromae in vitro. Study the effects of promising compounds to inhibit mycelium growth and conidial germination of L. theobromae.

  1. Screening of essential oils against mycelial growth of L. theobromae

Benzaldehyde, carvacrol, citral, cinnamaldehyde, eugenol, eucalyptol, menthol, and trans-cinnamaldehyde were evaluated against mycelial growth of L. theobromae isolated from infected citrus fruit in vitro. Exposure to benzaldehyde, citral, cinnamaldehyde, eugenol, eucalyptol, and trans-cinnamaldehyde volatiles (50 μL/L) did not inhibit mycelia growth. However, carvacrol volatile at 50 μL/L effectively inhibited mycelial growth, with the percent inhibition 98% (Figure 1-A). In amended PDA medium, carvacrol and thymol at 0.1 mg/mL inhibited mycelial growth by 93% and 97%, respectively, while menthol had no antifungal activity against L. theobromae (Figure 1-B).

  1. Studying the effects of carvacrol and thymol to inhibit mycelial growth of L. theobromae

Carvacrol and thymol were amended in PDA medium at difference concentrations (0.01, 0.02, 0.05, 0.1, 0.2, and 0.5 mg/mL) to test their antifungal activities against mycelial growth of L. theobromae. In amended PDA medium, carvacrol and thymol inhibited mycelial growth in a concentration-dependent manner, with higher concentration having a higher efficiency. Half-inhibition concentration of carvacrol and thymol against mycelium growth was 0.036 and 0.028 mg/mL, respectively (Figure 2).

Objective 2. Using the most effective essential oil compounds from objective 1, evaluate their ability in both ambient and heated solutions to control SER on inoculated grapefruit and on naturally infected fruit.

  1. Optimal Diplodia inoculation method

In this study, ‘Ruby Red’ grapefruit were inoculated in three sites to test the infection efficiency: stem, equator, or blossom. Fruit were wounded with a sterilized nail (2 mm deep by 2 mm wide) on one of these three sites. Two types of inoculums were tested for their infection ability: mycelial suspension (10 μL) or mycelial plug (3 mm in diameter). Mycelial suspension was injected into each wound site, while mycelial plug was placed on each wound site. And then, fruit were incubated in two conditions: room condition (75°F and 90% RH) or high temperature condition (85°F and 90%). Result showed that fruit inoculated at blossom with mycelial plug and incubated in high temperature condition had 100% infection with uniform lesion area after 48h. This method was chosen for the next experiment. When mycelial plugs were removed 12h after inoculation, fruit also developed 100% infection 48h after inoculation. That indicated fruit was already infected 12h after inoculation, although the lesions showed up 48h after inoculation.

  1. Evaluating carvacrol and thymol in both ambient and heated solution to control SER on inoculated grapefruit

Preliminary experiment showed that both carvacrol and thymol at low concentration (0.2%) in water resulted in phytotoxic peel injury, while no injury was detected when coating fruit with shellac wax containing carvacrol or thymol, even at high concentration (1.0%). Considering the commercial application practicality, we changed essential oil dipping treatments to essential oil coating treatments.

  1. Evaluating carvacrol and thymol in shellac wax to control SER on inoculated grapefruit

In this study, carvacrol and thymol were mixed in a commercial shellac wax (590HS) to determine their curative and preventive activities against Diplodia SER on ‘Ruby Red’ grapefruit. For curative activity, fruit were inoculated and incubated at 85°F and 90% for 12h and the mycelial plugs were removed. And then, the fruit were coated with shellac wax containing carvacrol or thymol at different concentrations (0.1%, 0.2%, 0.5%, or 1.0%) on packingline and incubated at 85°F and 90% again. For preventative activity, fruit were coated as above, inoculated, and incubated at 85°F and 90%. Both carvacrol and thymol showed effectively curative and preventative activities against Diplodia SER on ‘Ruby Red’ grapefruit in a concentration-dependent manner, with 1.0% essential oil in wax showing the highest efficiency. For curative activity, coating fruit with shellac wax containing 1.0% of carvacrol or thymol reduced lesion area by 59% or 37%, respectively, compared to the control after 48h (Figure 3). For preventative activity, coating fruit with shellac wax containing 1.0% of carvacrol or thymol reduced lesion area by 63% or 52%, respectively, compared to the control after 48h (Figure 4). Although thymol showed higher efficiency than carvacrol to inhibit mycelial growth in vitro, it was less effective to control Diplodia SER on fruit. That may be related to high penetrability of carvacrol or fruit disease resistance induced by carvacrol.

Objective 3. Test the effects of essential oil compounds in ambient or heated solutions on fruit internal and external quality during postharvest handling and simulated marketing conditions.

  1. Coating fruit with shellac wax containing carvacrol or thymol fruit internal and external quality during storage

Fruit were coated with shellac wax containing 1.0% of carvacrol or thymol on packingline and stored at 50°F and 90%. Fruit internal and external quality will be evaluated after one month and two months storage.

Accomplishments/Milestones

In the current study, we found out the two most effective essential oil compounds, carvacrol and thymol, against mycelial growth of L. theobromae in vitro. When carvacrol and thymol were mixed in commercial shellac wax and applied on ‘Ruby Red’ grapefruit inoculated with Diplodia, the showed both curative and preventative activities to control stem end rot, with higher concentration showing higher efficiency. No peel injury was detected in the treatment. In the next study, we will evaluate the effects of essential oil coating on fruit internal and external quality during storage, including weight loss, color, firmness, titratable acid, soluble solid content, chilling injury, and natural decay.

Impacts and Contributions/Outcomes

Stem-end rot (SER), caused by L. theobromae, is often the most important postharvest disease of citrus fruit in warm and humid regions such as Florida. This disease is exacerbated by commercial degreening practices used to improve peel color of early season fruit. Currently, use of synthetic fungicides is the main method to control this disease. However, consumer concerns with pesticide usage and the potential for pathogen resistance to these fungicides limits their availability and use in many citrus-producing regions and is a concern for the southeast region.

The current study identified an effective method to control postharvest Diplodia SER on ‘Ruby Red’ grapefruit instead of fungicide treatment. Coating fruit with shellac containing carvacrol or thymol showed both curative and preventative activity against SER on grapefruit inoculated with Diplodia. Essential oils are aromatic oily liquids extracted from plant organs that are recognized as safe for human health and the environment. The US Food and Drug Administration has listed some essential oils as food for human consumption. Our finding from this this study could provide a safe and effective strategy to control Diplodia SER in Florida degreening process.