| Green mold, caused by Penicillium digitatum Sacc, is the most damaging postharvest disease of citrus fruit and can lead to a loss of 90% during postharvest handling. Limonene plays important role in the process of green mold, but the mechanism of it is less clear. Referring to P.digitatum spores as research object, this research investigate the effect of limonene to P.digitatum in vivo and in vitro and the flow direction of limonene adding in culture. The main results are as follows:(1). A linear relationship between different levels(104, 105, 106, 107 spores/mL) of spore concentration of P.digitatum and optical density(OD) was determined. The line showed that they had a good relationship and it can be used to accurate quantify the spore concentration for later experiment.(2). The effect of spore germination by limonene in water agar was tested. Results showed that low concentration(0-0.50μL/mL) of limonene promoted spore germination and high concentration(over 1.0μL/mL) inhibit it at a spore concentration of 106/mL or 107/mL. The maximum promoting concentration is 0.25μL/mL and 0.50μL/mL, respectively.(3). The effect of spore germination by limonene in PDB was tested. Results showed that low concentration(0-0.50μL/mL) of limonene promoted spore germination and high concentration(over 1.0μL/mL) inhibit it at a spore concentration of 106/mL and 107/mL. The maximum promoting concentration is 0.25μL/m 0.50μL/mL, respectively. Spoers were germinated completely after culturing spoers for 11 h.(4). Effect of exogenous limonene on the process of green mold on citrus was tested. As a result, it showed that citrus without sterilization and wound would not decay and citrus with wound began to decay after 4 days and had a decay incidence of 98.9% after 6 days. Wounding the oil gland or between oil gland of citrus and incubated for 48 h, 60 h and 72 h, the lesion diameter was 2.12 cm, 3.34 cm, 6.64 cm and 1.16 cm, 1.58 cm, 4.71 cm respectively. Supplementing limonene in oil gland and inoculating spores for 60 h, we found 0.25-2.0μL/mL of limonene can promote the process of green mold and 4.0μL/mL of limonene will slow down the process. And supplementing limonene between oil gland, we found that 0.25-4.0μL/mL of limonene can promote the process of green mold but 4.0μL/mL of limonene will slow down the promotion. With adding more limonene on wound in oil gland and wound between oil gland, difference of lesion diameter on both group was decrease, and there was no significant difference when limonene concention was up to 2.0μL/mL(5). Changes of limonene after culturing spores for hours in PDB was qualified by GC-MS. Results showed that limonene did not transform to other terpene at a low concentration(no more than 0.50μL/ml) and transform to α-terpineol at a high concentration(more than 1.0μL/mL).(6). The concentration of intracellular and extracellular limonene was quantified by GC after culturing spores for 13 hours in PDB with adding different concentration of limonene. Results showed that intracellular limonene would not increase after adding limonene more than 0.50μL/ml and 1.0μL/mL for spore concentration of 106 spores/mL and 107 spores/mL. The sum of intracellular and extracellular limonene had no difference with the quantity of limonene initial.(7). The concentration of intracellular and extracellular limonene was quantified by GC after culturing spores for different hours in PDB with adding 0.25μL/ml of limonene. Results showed that intracellular limonene would not increase after culturing 11 hours and 7 hours for spore concentration of 106 spores/mL and 107 spores/mL. The sum of intracellular and extracellular limonene had no difference with the quantity of initial limonene concentration.This study proved that certain concentration of limonene can stimulate the germination of P.digitatum spore and accelerate the process of green mold on citrus. Limonene was more likely to execute the function as a signal molecule or stimulator rather than nutrient... |
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