Abstract:

Objective: Injecting agents into pine trees is an effective measure to prevent pine wilt disease. Avermectin and emamectin benzoate are the two most widely used trunk injection drugs at present. However, if these two drugs are used for a long time, it is bound to lead to drug resistance of pine wood nematode. At the same time, there is also a need for more efficient, convenient and lasting new agents in production. This paper aims to explore the potency of a new compound medicament of 2% avermectin and 6%·fluopyram in prevention and treatment of pine wilt disease, in order to find an alternative trunk injection in place of avermectin and emamectin benzoate. Method: The toxicity of a new compound agent of 2% avermectin ·6% fluopyram was compared with 5% avermectin and 5% fluopyram. An indoor nematicidal experiment was conducted through the nematode immersion method to analyze the lethal effect of the three agents on pine wood nematode. The three agents were evenly sprayed with three agents in the LC₂₀ concentration on a Petri dish containing Botrytis cinerea, the feeding conditions of pine wood nematodes on B. cinerea were observed, the number of pine wood nematodes was counted, and the reproductive rate (Pf/Pi) under LC₂₀ concentration was calculated. The eggs at the bottom of the Petri dish were collected, and different concentrations of agents were added to the Petri dish. After 24 h, the eggs hatching was observed, the egg hatching rate was calculated, and the morphology of eggs was observed with Zeiss fluorescence microscope. Pine wood nematode was mixed with the agent and placed in a temperature gradient incubator, and the nematicidal activity of 2% avermectin and 6% fluopyram on pine wood nematode under different temperature conditions was analyzed. Result: The results showed that after the pine wood nematode was treated with three agents for 24 hours, the LC₅₀ values of 2% avermectin and 6%·fluopyram, avermectin 5%, and fluopyram 5% were 2.020 8, 51.153 6, and 21.607 1 mg·L^-1, respectively, and the LC₂₀ values were 0.447 9, 12.031 8, and 8.558 2 mg·L^-1, respectively. The nematicidal effect was 2% avermectin and 6% fluopyram > 5% fluopyram > 5% avermectin. The compound agent had a significant virulence synergism. After six days of spray treatment with LC₂₀ concentration of the three agents, the number of pine wood nematode in the 2%·avermectin and 6% fluopyram treatment and 5% fluopyram treatment group was only 1 425 ± 250, and 1 675 ± 377, far lower than that in 5% avermectin group (10 925 ± 504) and the control group (> 20 000). The experiment of hatching rate showed that the three agents could significantly inhibit the hatching of eggs, and the hatching rate of eggs decreased with the increase in the concentration of the agents. After 24 h, at 10 mg·L^-1, the inhibitory effect on egg hatching was 2% avermectin and 6% fluopyram > 5% avermectin > 5% fluopyram, and the hatching rate was 20.32%, 21.36%, and 22.89%, respectively. After being treated with 2% avermectin and 6% fluopyram at a concentration of 10 mg·L^-1 for 24 h vacuoles were produced in pine nematode eggs and the eggs died. In the temperature gradient experiment, within 15 ℃-35 ℃, the nematicidal activity of 2% avermectin and 6% fluopyram increased with the temperature increase in each 5 ℃. Conclusion: In summary, the new compound agent of 2% avermectin and 6% fluopyram has significant nematicidal activity against B. xylophilus and virulence synergism, and the toxicity is significantly higher than that of the two agents alone. The compound agent has the potential to become a new trunk injection for the prevention of pine wilt disease.

Key words: pine wilt disease, fluopyram, avermectin, sublethal concentration, median lethal concentration