| Ginseng (Panax ginseng C. A. May) is a perennial herb medicinal plant of the Araliaceae Panax. As a traditional medicine, it has a very high medicinal and economic value. The government paid attention to the forest resources protection seriously and prohibited deforestation for plant ginseng strictly in recent years, the model of non-forest planted has been the development of direction of ginseng industry in China. Due to the changes of the growth environment of ginseng, the gray mold disease incidence and the severity of farmland rise radically, and restricted the healthy development of the ginseng industry severely. Supported by the national Agro-scientific Research in the PISSRlic Interest——The Study of Infestation Regularity and Security Precautionary Technology of the Disease and Pest on Ginseng and based on the original foundation of laboratory, three aspects have been studied: comparison of the biological characteristics, analysis of genetic diversity, pathogenic mechanism. The results were as follows:1. Differences of biological characteristics and pathogenicity of 16 Botrytis cinerea isolates from northeast China to Panax ginseng were compared. The results showed that the effects of different conditions on the mycelium growth of B. cinerea strains were significant. On the same condition, differences were significant in mycelium growth among tested strains’ except light. The optimum media for growth of all strains include PDA and PSA. PSA was the optimum medium for the mycelium growth of isolates PJAR1, PJAL3, PYCL1 and PJLL1. PDA was the optimum medium for the mycelium growth of isolates FSYL1, PJAL1, PJAL2, PYLL1, PBLL1, PQAL1, PHLL1 and LSYF1, while PSA was optimum for other strains. Temperatures for mycelium growth of each tested isolates ranges from 5℃ to 25℃. The optimum temperature of mycelium growth of the isolate FSYL, PJLL2, PRHL1, PHLL1 and LSYF2 was 25 ℃， while that for others was 20 ℃. The ranges of pH for mycelium growth of the tested isolates were the same, from 3 to 11. As for mycelium growth of PYCL1 and LSYF2, the favorable pH value was 6. The optimum pH of mycelium growth of the isolates (PJAR1, PJAL2, PJAL3 and PYLL1) was 4, while that for the other isolates was 5. The optimum carbon sources of mycelium growth were glucose and maltose. The favorable nitrogen sources of mycelial growth of tested isolates were yeast extract, beef extract and peptone. There was no significant effect of light on the mycelium growth of B. cinerea strains. The results also showed that all tested isolates could cause grey mould on Panax ginseng, strawberry, tomato, respectivily. However, there was difference in pathogenicity among the different isolates from the same host, and the pathogenicity difference was not obviously related to the localities of isolates. Most of strong virulent strains were sclerotium type.2. Using ISSR-PCR molecular markers, genetic diversity analysis of ginseng grey mold and tag system is established. The optimum conditions for ISSR-PCR were 1.0 U Taq DNA polymerase,0.2 mmol-L-1 dNTPs,2.5 mmol-L-1 Mg2+,0.4 μmol·L-1 primers,30 ng templates DNA in 25 μ1 reaction system and UBC 810 as premier. The diversity of 23 B. cinerea isolates with 13 primers was analyzed, and the results showed that 190 fragments were amplified.154 polymorphic loci were accounted for 81.1% in the total amplified fragments. The genetic diversity among 23 B. cinerea isolates was analyzed according to the amplified results and a molecular dendrogram was constructed. The similarity coefficient of 23 B. cinerea isolates was at 0.53-1.00 and they were divided into 6 groups at the level of 0.814. In this paper, the author has established the befitting ISSR-PCR reaction system for B. cinerea and found there was manifest interspecific genetic diversity of B. cinerea isolates. This technique can be used to analyze the interspecific genetic diversity of B. cinerea in the future study. Genetic group division and strain geographical source had certain relevance. The relationship between genetic lineages and pathogenicity of B. cinerea was complicated. It was found that there is no certain corresponding relationship between genetic lineages and pathogenicity.3. Conditions for in vitro cell wall degradation enzymes (CWDEs) and inoculation of B. cinerea CWDEs activity change has carried on the system research, the results showeing that: After the ginseng of grey mould of CWDEs treatment, the typical water symptom was formed. PG and PMG showed maximum activities in the second day after inoculation. Cx and β-glucosidase showed maximum activities in the fourth day after inoculation. But PGTE and PMTE were not detected. Based on the CWDEs produced in vitro conditions, the results revealed that 4 kinds of CWDEs had a specific optimal culture conditions. Stationary culture made more for the production of CWDEs compared to the shake culture. The activities Cx and β-glucosidase reached a peak when the isolate was cultured 9 d, while PG and PMG reached a peak when the isolate was cultured 12 d. It was found that the best culture temperature for PG and PMG was 15℃ and the best for Cx and P-glucosidase was 20 ℃. In regarding to pH value,4 kinds of CWDEs showed maximum activities at culture pH 5.4. Based on six different virulent strains and CWDEs correlation analysis, the results indicate that the highly virulent isolates had more mycelial growth than the moderately virulent isolates, with different media. Pathogenic factors of B. cinerea were investigated in regard to fungal growth and the production of the cell wall-degrading enzymes, PG, PMG, p-glucosidase and Cx. PG, PMG, Cx and p-glucosidase activities produced by the highly virulent isolates in in shake cultures and in decayed leaves were greater than those of the moderately virulent isolates. In pectin shake culture, PG and PMG activities produced by the most highly virulent isolate was respectively 2.2 and 1.7 times greater, when compared to the most weakly virulent isolate. In inoculated ginseng leaves, PG and PMG activities of the most highly virulent strain were respectively 2.1 U/mg and 3.2 U/mg, while those of the most weakly virulent isolates were respectively 0.3U/mg and 1.4U/mg. In CMC liquid medium, Cx and β-glucosidase activities produced by the most highly virulent isolates were respectively 1.8 and 2.0 times greater, when compared to the most weakly virulent isolates. In inoculated ginseng leaves, Cx and β-glucosidase activities produced by the most highly virulent isolates were respectively 5.5 and 2.1 times greater, when compared to the most weakly virulent isolates. The relationship between the pathogenieity and the CWDEs activities was analsed. The results showed that there was some correlation between the expansion rate of the lessions ceaused by B.einerea and the activities of CWDEs and PG plays an important role. |
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