Study On Programmed Cell Death And Related Genes Function Of Chestnut Blight Fungus

Chestnut blight fungus (Cryphonectria parasitica) is very destructive disease of chestnut tree; it causes the chestnut tree stem canker and one of the diseases of chestnut production. Because of the difficulty of control this disease; the chemical control of this disease is difficult. Study on the chestnut blight fungal cell death and related genes fuction, it is possible to find the new target of chestnut blight control and design the new disease control strategy to give the new theory of direction.Vegetative incompatibility induced programmed cell death between strains of chestnut blight fungal fusion cell The study of vegetative incompatibility induded cell death of chestnut blight fungus was done. In order to test the hypothesis whether vegetative incompatibility results in nuclear degradation, a characteristic of apoptosis, the agar electrophoresis was mainly used to detect the degeneration of fusion cell genome DNA of chestnut blight fungus, the cytological observation on the change of the nuclear in hyphae between incompatible strains, the production of the reactive oxygen species, the accumulation of lipid droplet and the identification of the live and dead cell. The results showed that the electrophoresis detected the DNA ladder, which provided the new evidence of vegetative incompatible cell death. Hyphal fusion cells between vegetative incompatible strains were compartmentalized by septal plugging and contained heavily degraded nuclear DNA. Reactive oxygen species accumulation decreased in hyphal fusion cells, the accumulation of lipid droplets increased and finally the hyphal fusion cell died. Morphological features of hyphal compartmentation and death during vegetative incompatibility can be described as a form of programmed cell death.Effect of exogenous hydrogen dioxide on four antioxidases of Chestnut Blight hypovirulent strain EP713 When plant pathology infected host plant, host plant produced oxide burst and protected itself from the fungi attacking. The exogenous oxide stress was used to mimic the influence of the host oxide burst, detected the change of antioxidase activity. The 10 mM H2O2 was used to treat the strain of EP155 and EP713 and the samples were taken at different treatment time. The proteins were extracted and detected the antioxide enzymes activity including the activity of catalase (CAT), superoxide dismutase (SOD) Glutathione peroxidase (GPx) and Glutathione reductase (GR). The results showed that the maximum activity of four kinds of antioxidases is different between EP155 and EP713, however, the time of the GR and GPx reaching maximum activity EP155 is later than EP713, and the activity of the four kinds of enzyme of EP155 is higher than EP713. These results showed that the activity and the time of four antioxdase to reach maximum activity are different.Hydrogen dioxide induced the cell death of chestnut blight conidia Chestnut blight fungus (C. parasitica) is one of the major diseases of chestnut tree worldwide. It is mainly spread by the conidia of this fungus. We were interested in the conidia response to the exogenous oxide stress. Sublethal and lethal dose of hydrogen dioxide was 10 mM and 150 mM; the germination rate of the conidia was analyzed and the cytology research was done along with the different treatment time. The cytology was mainly detected:the change of nuclear, reactive oxygen species and nitric oxide generation. The agar electrophoresis was detected the degeneration of genome DNA. The results have been shown that the change of the nuclear, the ROS generation increasing along with the treatment time, nitric oxide only appeared at the treated time of 30 min in 150 mM. The results of this experiment have been shown exogenous oxide stress induced the conidia of chestnut blight cell death, which is associated with the degeneration of nuclear and the generation of ROS and NO.Functional effects of ADP-ribosylation factor 1 from C. parasitica on its phenotypes ADP-ribosylation factor 1 gene CpARFl in C. parasitica was cloned and analyzed. The amino acid residues of this gene have conserved Golgi binding domain MXXE, the phylogenetic evolution is near to other ascomycytes. Using the gene silencing technique,99 transformants were attained,3 of these transformants were silenced mutants. The phenotypes of silenced mutants were detected, which were included in the sensitivity to CUSO4 and NaF, the accumulation of laccase, the reduction of conidia production and the decrease of pathogenicity. The results showed that CpARFl has influence on the sensitivity to CUSO4 and NaF, the production of laccase, the asexual reproduce and the pathogenicity.Functional effects of ubiquitin and ribosome L40 fusion protein from C. parasitica on its phenotypes Ubiquitin fusion protein (UFP) gene CpUFP in C. parasitica was cloned and analyzed. The amino acid residues of this gene have conserved ubiquitin conjugation enzyme binding domain, the phylogenetic evolution is near to other ascomycytes. Using the gene silencing technique,63 transformants were attained,2 of these transformants were silenced mutants. The phenotypes of silenced mutants were detected, which were included in the sensitivity to H2O2 and NaCl, the accumulation of laccase, the reduction of conidia production and the decrease of pathogenicity. The results showed that CpUFP has influence on the sensitivity to H2O2 and NaCl, the production of laccase, the asexual reproduce and the pathogenicity.