| Camellia oleifera anthracnose is one of the major diseases in China's Camellia producing areas. It occurs broadly and causes serious drop in buds and leaves, sometimes resulting in the death of the tree. The Camellia anthrax prevention and treatment by planting resistant varieties and chemical control to control the growing problem of pest resistance, and chemical pesticide residues pose a serious hazard. Characteristics of biological control for its safety, efficiency and no pollution has become an important way of plant diseases, widespread concem.Endophytic bacteria Y13is isolated from the healthy leaves of Camellia which has good control effect to Camellia anthrax, the strain was identified as Bacillus subtilis.The study found that endophytic bacteria Y13has better control effect to Camellia anthracnose. But the control effect is not stable in field experiment.so In order to effectively take advantage of this strain, take endophytic bacteria Y13as research object,the paper studied Y13'colonization in Camellia oleifera and mechanism to Camellia anthrax. The main results are as follows:(1) Colonization of Endophytic bacteria Y13in the Camellia. Endophytic bacteria Y13was induced by rifampicin and obtained Y13R by antibiotics labeled method.By pot experiment,cameilia seedlings were inoculated with Y13by sprayings pouring root and dipping root.and determinded Y13's quantity on different time.Different inoculated methods made Y13'colonization in Camellia.the Y13strain can only be detected in the oil tea leaves and stems,not detected in roots by spray leaf inoculation.and Y13's quantity incresed at first time and decreased at end.on the10d the quantity of Y13in the leaves reached its peak,5.88×103cfu/g. On the25d Y13can not be detected in the stems,but in the leaves Y13'quantity can reach1-8×103cfu/g.By dip root treatment,Y13'quantity decreased gradually in roots and stems but increased in leaves.and decreased on the20d.the quantity of Y13in leaves reached1-28X103cfu/g or so on the25d. Irrigation is similar to dipping root. According to the colonization site of Y13in the Camellia oleifera,the quantity of Y13in the leaves is larger than roots and stems. According to the treatment,spray is better than pouring root and dipping root. (2) Characteristics and separation of antibacterial substance inY13.By plate confrontation culture, determinded the inhibitory effect of Y13sterile filtrate to the Camellia anthrax up to61.2%, which showed that Y13secreted active material to extracellular, indicating effective inhibition of Y13secretion of active substances to the extracellular, the use of organic solvents to extract the active substances results show that:the active substances is not soluble in organic solvents, can be precipitated by methanol and ethanol, indicating that the polarity of the substance is large, using PH precipitation method extract active meterial showed that:Active substances can be precipitated under acidic conditions PH2-3precipitated the strongest activity, using different concentrations of ammonium sulfate precipitation method to extract the active substance, the results show that the active substances can be precipitated by ammonium sulfate. and the activity of substance is highest in70%ammonium sulfate, The activity of salt analyte reached39.8%after dialysis.so we can conjecture the active substance is protein.on activity in crude extracts of the physicochemical properties, the results showed that the antagonistic activity of substance stable to heat, was not sensitive to the ultraviolet radiation stable in neutral pH conditions, unstable in acid and alkali conditions, easy to produce precipitation under acidic conditionspartially sensitive to proteinase K is not sensitive to trypsin.(3) Study on the induced resistant mechanism of Y13to Camellia anthrax. By Y13'induce and Camellia anthrax treatment, changes of Camellia seedlings'resistance enzymes PAL,POD,SOD,PPO and content of MDAand AOX production rate were determinded on different inoculation time.The results show that, After induction treatment of Y13, the activities of PAL, POD, PPO and SOD in the plant leaves inereased obviously. the PAL activity of vaccination Y13and anthrax reached their maximum3days after the treatment, and showed2.9fold higher more than the water control treatment. When only anthrax vaccine or Y13, the PAL activity reached a peak in5days,1.7and2.3fold higher more than the water control treatment respectively; the activity of POD and SOD after induction come to the highest in5days, increased of15.5-29.5and8.5-20.5units of one activity respectively, compared with the control treatment; PPO activity reached its peak3days after the induction, but decreased rapidly7days latter.02-production rate reached its peak1day after the induction,increased of80.6%, compared with the control treatment,and then tended to normal level10days latter. MDA content Vaccination Y13Individually and simultaneously inoculation Y13and anthrax decreased rapidly after induction treatment lday. decreased of36%, compared with the control treatment3days latter.Y13induced resistance gene expression in Camellia, thus increased the Camellia defense enzymes SOD, POD, PPO activity, reduced the Camellia reactive oxygen species, reduced membrane lipid peroxide level, thereby enhanced the Camellia comprehensive resistance ability. |
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