| Tobacco black shank is a fungal soil-borne disease caused by Phytophthora parasitica var.nicotianae,which has highly prevalent in China.Tobacco black shank disease often mixed with tobacco root black rot,tobacco nematode disease,bacterial wilt and other diseases,resulting serious reduction in tobacco yield.For a while,due to the restrictions of tobacco growing areas,the control methods to tobacco farmers are relatively simple.In addition,unreasonable application of medicine and fertilizer not only increases the pressure of control tobacco black shank disease,but also leads to ecological hazards such as the decrease of soil fertility.In this context,the isolation of Plant growth-promoting rhizobacteria(PGPR)and the application of biological control have brought a new idea for the control of tobacco black shank disease.Based on the selection of biocontrol strains that antagonize P.parasitica var.nicotianae,this paper conducts a preliminary study on its biological control characteristics,and provides basis for the application of PGPR strains in field control of tobacco black shank disease.The main results are as follows:1.61 rhizosphere soil samples of healthy tobacco plants in different habitats were collected in Shaanxi and Sichuan province.Through high temperature treatment,253strains of rhizosphere bacteria with exuberant growth vitality were obtained.4 strains of biocontrol bacteria with antibacterial rate higher than 85%were obtained by confrontation test against the target of P.parasitica var.nicotianae.The resistance growth test showed that strain DY-6-6 had strong growth activity,under p H 6.0～8.0 and temperature range of26～38℃.Moreover,it showed growth activity after ultraviolet irradiation and low temperature treatment.Finally,through physiological and molecular biological identification,strain DY6-6 was identified as Bacillus velezensis.2.The malondialdehyde(MDA)content of pathogen treated with B.velezensis DY-6-6 metabolites was 73.01%higher than that of normal mycelia,and the protein content of the bacteria decreased by 60.56%,indicating that the metabolites caused damage to the mycelium cell membrane;Compared with the control group,the activities of catalase(CAT)and superoxide dismutase(SOD)of tobacco black shank fungus decreased by 63.76%and70.54%respectively,which showed that the metabolism of disease-resistant enzyme activity was inhibited.In addition,after treatment,the hyphae and sporangium morphology showed obvious abnormal expansion,distortion and fracture,and the number of sporangia was also inhibited.Except for the significant antagonistic effect on tobacco black shank,the inhibition rate of B.velezensis DY-6-6 to P.capsici、P.infestans and other 10 kinds of pathogenic fungi were more than 50%.At the same time,B.velezensis DY-6-6 has ability to produce cellulase,protease and nitrogen fixation,and the biofilm production is highly active.After 48 hours of culture,it produces a dense wrinkled biofilm with central depression.High dose of the broad spectrum fungicides will inhibite the growth of B.vaselezensis DY-6-6,but biofilm was produced in all the plates and bacteriostatic rate was lower than 10%in the compound test with 100%dilution of 58%Mn-Zn methacrylate wettable powder,50%carbendazim and 1%avermectin granules.3.The growth-promoting effect of B.velezensis DY-6-6 on tobacco is as follows:The germination rate of tobacco seed could reach 100%if 10～30%metabolites were immersed to promote germination,but the high concentration of metabolites above 30%could inhibit seed germination;In the seedling stage,the fresh weight,leaf number and height of tobacco seedlings were significantly higher than those of the control group,and the rates of diseased seedlings and weak seedlings were all less than 4%.In pot experiment,the fresh weight and dry weight of tobacco leaves treated by root irrigation with fermentation broth increased by 67.73%,50.00%,and the fresh weight of roots increased by 62.40%respectively.In addition,the maximum colonization of B.velezensis DY-6-6 in the rhizosphere soil of tobacco plants treated with 58%Mn-Zn methacrylate within 40 days was 6.26×10~6 CFU/g,which was not significantly different from that of the control group.Finally,through the indoor potted tobacco control effect test,it is clear that the relative preventive effect of B.velezensis DY-6-6 on tobacco black shank can reach 70.3%,which is equivalent to the preventive effect of 58%Mn-Zn methacrylate;the relative therapeutic effect is 59.2%,slightly lower than 58%Mn-Zn methacrylate.In field application,B.velezensis DY-6-6 can promote growth and prevent diseases before transplanting,supplemented by supplementary spraying treatment during the onset period,finally effectively control tobacco black shank and reduce the amount of fungicides,achieve the balanced development of tobacco industry between sustainable green agriculture. |
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