| Green mold, caused by Penicillium digitatum, is the most serious postharvest disease of citrus. Citral and octanal are natural antibacterial substance with great potential, widely exists in plant essential oil, reportedly exerts strong anti-fungal activity against postharvest pathogenic fungi. In our recent papers, citral and octanal were observed to have good anti-fungal effects against the mycelium of P. digitatum. However, the mechanism of citral and octanal against the spores of P. digitatum was rather limited. Referring to P. digitatum spores as research object, this research investigate the effect of citral and octanal to the germination of P. digitatum spores. The targets were investigated by determining the following:(i) the morphology of spores using optical microscope,(ii) membrane permeability and integrity: the cell constituents’ release, the extracellular conductivity and the integrity of membrane,(iii) detecting the hydrogen peroxide content and reactive oxygen species(ROS),the activity of catalase(CAT), peroxidase(POD) and superoxide dismutase(SOD) relating membrane lipid peroxidation. The main results are as follows:(1). Finally, through screening, the citrus fruit inoculated back methods was used to obtain P. digitatum spores. The spore yield per fruit was 3.62±0.05 g. This method can obtain large amounts of spores with high viability, purity and uniform morphology, which can be used for subsequent analysis.(2). Integrated water agar culture and liquid water culture(PDB) to determine the affection of citral and octanal treatment on P. digitatum spores germination rates, the highest treatment concentrations were 4.00 μL/m L(citral) and 2.00 μL/mL(octanal), and the spore germination rates were 11.07 % and 15.29 %, which were significantly lower than the control group(75.82 %, 72.62 %; p <0.05).(3). The affection of citral and octanal with different concentrations on the morphology of P. digitatum spores. Optical microscope observations showed that the spores of P. digitatum in control were normal and homogenous, which represented uniform shape, complete structure, and rich materials. The treatments group showed shriveled, depressed, fold, and rupture of spores, and the extent of the damage and the number of spores were increased with the increasing of treatment time. These results revealed that citral and octanal could damage the structure of spores.(4). Effect of spores’ cell membrane permeability and integrity show that, after the treatment of citral and octanal, the extracellular conductivity and OD260 nm, the integrity of membrane were changed. The rate were raised 130.37 μs/cm, 0.507,-33.6 % by citral treated; 62.1 μs/cm, 0.19,-32.8 % by octanal treated.(5) The determination of H2O2, ROS, MDA, and the enzyme activity of catalase(CAT), peroxidase(POD), superoxide(SOD) that were from P. digitatum spores. The results showed that after citral and octanal treatment for 120 min, the citral treatment content of H2O2 increased 2.5 times, ROS levels increased 2.9 times and total MDA content increased by 9 times. The octanal treatment content of H2O2 increased 4.1 times and ROS levels increased 2.3 times, total MDA content increased by 5 times. The 4.00 μL/mL citral treatment of 20 ~ 30 min, CAT activity more than 9.5 times, POD activity increased nearly 11 times, the activity of SOD increased 14 times; The 2.00 μL/mL octanal treatment group three enzyme activity are increased in multiples were 10, 5.1 and 2.7 times(p<0.05). However, the enzyme activities were at a low level when the treatments after 60 min, eventually enzyme activity were lost. After the treatment of citral or octanal, the content of H2O2 and ROS were overproduction, MDA excessive accumulation, what indicated that citral, octanal have lead spore eventuated oxidative stress, and caused lipid peroxidation in P. digitatum spores, then make the spores died, loss of their germination possibly.This paper proved that citral and octanal can inhibit the germination of P. digitatum spores, the damaged cell membrane, influenced the exchange of materials through the P. digitatum spores membrane, caused a large number of ROS be produced, MDA excessive accumulation, resulting in oxidative stress leads to spore membrane lipid peroxidation and accelerated apoptosis spores, inhibiting spore germination ultimately. |
