| Pine wood nematode disease is currently a major forest disease in the world.The pathogen pine wood nematode is currently a forestry pest that has a great threat in the world and is a devastating hazard to pine trees.The main hazards are Pinus massoniana,Loblolly pine,black pine,red pine and so on.In order to prevent the spread of pine wood nematode disease,we can start from the three aspects of pine wood nematode,pine tree and Monochamus alternatus.The prevention and control measures for pine wood nematode disease are currently based on chemical and physical control,supplemented by biological control.The main focus is on pine tree and the Monochamus alternatus which is medium of pine wood nematode disease.But the direct control of pine wood nematode is less.Esteya vermicola is the first reported endoparasitic fungus of the pinewood nematode(PWN),Bursaphelenchus xylophilus.It has high in vitro infectivity.As a potential application of biological control pine wood nematode,it has obtained a patent.The laboratory introduced this endoparasitic fungus of pine wood nematode with biocontrol potential from Korea.Based on the results of previous studies,this paper will test the fungus from four aspects: the difference of growth temperature,the difference of infestation temperature,the injection time in the field and symbiosis between the fungus and Monochamus alternatus.The purpose is to provide a theoretical basis for future,that is used for controlling pine wood nematode disease by fungus.By analyzing the colony diameter,germination rate,sporulation,the differences in different temperature culture conditions were clarified.According to the comprehensive analysis of the test results,E.vermicola can germinate and produce lunar spores with adherent nematode function under both 15°C and 25°C culture conditions.At 35°C,although the germination of spores eventually reached 100% on the 8th day,they did not produce lunar spores over time,resulting in loss of their ability to infect the pine wood nematode.Using the size of the colony as an indicator,it was determined that the treatment with the culture temperature of 25°C grew fastest,followed by the treatment with the temperature of 15°C,and the growth with the treatment of 35°C was the slowest.From the test results,the growth rates at 15°C and 35°C in the first three days are not significantly different,and from the 9th day,the strains grown at 35°C stopped growing.In this paper,different temperatures were used to demonstrate that E.vermicola had invasive activity against B.xylophilus at different temperatures,but it had different invading activities against B.xylophilus at different temperatures.The results show that the infectivity at low temperatures is significantly stronger than at high temperatures.The spores are more likely to adhere to the surface of the nematode so that infect the nematode better.The most effective at 15°C infection temperature,followed by 25°C and worst at 35°C.After 1 day,the attached nematode began to grow hyphae on its body surface.After 2 or 3 days,the mycelium began to grow,branching and spreading,and the new hyphae continued to produce spores for secondary growth.Spraying of spore suspensions at different times proved that endophytic fungi E.vermicola had invasive activity against B.xylophilus.And there are differences in the invasive activity of nematodes in three different treatments.Most of the pine trees in the untreated control group suffered serious damage,showing yellowing of pine needles,most withering and death.In the first month of the control group,pine trees began to show symptoms,and 2 pines died.The survival rate was 91.67%.In the following 2 months,the survival rate dropped from 91.67% to 52.5%.Most of the pines were seriously affected.After 3 months,the survival rate of treatment group was 76.67%,the inoculation of nematodes while spraying spores survived 84.17%.The treatment of spraying the spore suspension one week earlier started to show symptoms of withering after the second month of the inoculation of the nematode,and the survival rate was highest in the all groups after 3 months,which was 90.83%.The symptoms were slower and less severe than other group.The preliminary experiments on the two aspects that spore suspension spraying from the branches of Monochamus alternatus and soaking in the spore suspension provided a certain theoretical reference for the relationship between fungi and Monochamus alternatus.Although there was little difference between the treatments at the initial stage,as the time went on,the treatment group that directly sprayed the spores on the branches had a relatively higher survival rate.After 6 months,the survival rate of the treatment group which spores sprayed on the branches was the highest,between 67.5% and 80%.The survival rate of Monochamus alternatus soaked in the spore suspension treatment group was 65%-70%.The survival rate of the control group was only 40%-50%.The experiment of controlling pine wood nematode disease from different concentrations of spore suspensions proves that endophytic fungi E.vermicola can effectively inhibit pine wood nematode disease to a certain extent,and the control effect is superior to that using avermectin.The control effect increases with the concentration of spore suspension. |
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