The Competition Of Antagonistic Strain HT-6 Against Phytophthora Infestans And Its Inducible Inhibition Mechanism

Potato is the fourth largest crop after corn,wheat,and rice.It is rich in protein and many other nutrients needed by the human.As the country with the largest potato planting area and the largest yield,China has launched the potato staple food strategy in 2015.The planting area has increased year by year.However,the late blight caused by Phytophthora infestans seriously restricts the development of the potato industry.The prevention and control measures of potato late blight in production mainly include the selection of disease-resistant varieties,rationalized agricultural cultivation,and the use of chemical pesticides.Biological control has attracted more and more attention due to its low toxicity,high efficiency,and non-pollution characteristics.The HT-6 strain isolated by our laboratory has strong inhibitory effect on P.infestans.Its inhibition rate against mycelium was 89.22%,but it works only when the P.infestans exists in the experiment,and the inhibition rate of the its sterile fermentation broth was 0.Our laboratory has done a fundamental research on the biochemical mechanism of bacteria HT-6against P.infestans.It was found that when the P.infestans was inhibited by the HT-6 strain,the increase in the content of soluble sugar and starch in the mycelium is significantly inhibited,while the decrease in the content of soluble protein is significantly promoted.We also made an initial analysis of anti-oomycetes substances of HT-6 strain after induced by P.infestans.It is proved that the antagonistic substance,a polar substance,is soluble in methanol,ethyl acetate and water.And its methanol extract can increase the defensive enzyme activity and soluble protein content of potato tubers.The taxonomic status of the bacterium HT-6 is not yet clear.The stability of the antagonistic substances produced by bacterium HT-6 induced by P.infestans is not clear.The effect of this strain on the activity of zoospores of P.infestans has not been reported,such as motility ability,biofilm formation ability and the ability to infect potato tuber slices.In addition,the HT-6 strain can exert its inhibitory effect on P.infestans through the production of cell wall degrading enzymes,but no other antagonistic mechanisms that have not been reported.This research focuses on the problems mentioned above,and the main achievements are as follows:1.The biological taxonomic status of the bacteria HT-6 was identified.The colony characteristics of HT-6 on different media were observed,and the physiological and biochemical characteristics of this strain were determined;the strain phylogenetic tree was constructed based on the 16S r DNA sequence.It was confirmed that the bacteria HT-6 was Bacillus subtilis HT-6;2.The stability of the HT-6 bacterial solution to the environment after being induced and stimulated by P.infestans was clarified.When the bacterial liquid of this strain was treated with a high temperature of 100℃for 60 min,pH value of 6-8 for 24 h,30 W ultraviolet irradiation for 8 h,continuous culture 10 generations,the inhibition rates were still more than 80%;3.The various activities of zoospores of P.infestans obtained from different culture fluids were evaluated.It was found that the release rate of zoospore prepared from tap water was the highest（46.77%）,and the zoospores prepared from 10%V8 culture fluids had the largest swimming rate（23.25%）,the strongest biofilm-forming ability(OD₅₄₀=1.107)and the highest infection ability（disease index 87.51）;4.The effect of bacterial HT-6 on the various activities of zoospores of P.infestans was explored.When the zoospores from 10%V8 culture broth were treated with different concentrations of HT-6 bacterial solution,it was found that the inhibitory effect of bacteria HT-6 on zoospore activity was directly proportional to the concentration of the bacterial solution.When the bacterial solution OD₆₀₀was 1.433,the release ability of zoospores was greatly hindered,and the diseases index on tuber slices and swimming rate were 23.55,2.97%,respectively.The biofilm-forming ability is completely inhibited.The bacteria HT-6 can strongly inhibit the activity of zoospores;5.It is revealed that the bacterium HT-6 has a variety of inhibition mechanisms against P.infestans.Bacterium HT-6 can not only inhibit P.infestans through space and nutrition competition,but also can produce cell wall degrading enzymes（cellulase 6.638 U/m L,β-1,3-glucan enzyme 2.913 U/m L）through the induction of Phytophthora infestans.And the space and nutrition competition dominates.