| There are various soil borne fungal diseases that endanger the growth of crops.Wheat stem rot is a cereal disease caused by Fusarium pseudograminearum,which occurs in many arid and semi-arid Growing region in the world.In recent years,this disease has become increasingly common,causing serious losses to wheat yields in multiple regions.Peanut southern blight is a kind of soil borne fungal disease caused by Sclerotium rolfsii Sacc.It can infect many important crops besides peanuts.The disease expands with the expansion of peanut planting area,and it is serious in Shandong,Liaoning,Shanxi,Henan and other places.This study aims to screen antagonistic strains with high antibacterial activity and stable effects.The main content and research results are as follows:1.Isolation and screening of biocontrol bacteria.265 strains of bacteria were isolated from the wheat plant and the rhizosphere soil,13 strains of bacteria that had inhibitory effect on Sclerotium rolfsii Sacc were screened through plate confrontation,and then F pseudograminearum and Sclerotium rolfsii Sacc were selected for repeated screening,and 4strains of bacteria with significant inhibitory effect were obtained: Soil1-2,Soil1-3,Soil1-59,Soil2-93.The inhibition rates of Sclerotium rolfsii Sacc were 83.15%,78.09%,72.73%,and68.78%,respectively,The antibacterial rates against F pseudograminearum were 88.54%,81.77%,69.70% and 53.51% respectively.According to comprehensive identification,strain Soil1-2 is Bacillus velezensis,strain Soil1-3 is Bacillus subtilis,strain Soil1-59 is Bacillus atrophaeus,and strain Soil2-93 is Bacillus pumilus.2.Determination of bacteriostasis of biocontrol bacteria fermentation filtrate.The bacteriostatic spectrum of the 4 strains of bacteria selected was determined,and the results showed that they also had different degrees of inhibitory effects on the other 8 pathogenic fungi.According to the growth curve of four strains of bacteria,fermentation filtrates with incubation time of 24,48,and 72 hours were prepared to determine the bacteriostasis of 10 pathogenic bacteria.The aseptic fermentation filtrate had a poor inhibitory effect on Colletotrichum gloeosporioides,Fusarium oxysporum and Fusarium stratified,but had a significant inhibitory effect on all seven other pathogenic bacteria.The inhibitory rates of the fermentation filtrates of the four strains of bacteria against Sclerotium rolfsii Sacc were100%,and the inhibitory rates against F pseudograminearum were: Soil1-2,Soil1-3>Soil1-59,Soil2-93,and 72 h>48 h.Based on this result,the inhibitory effects of fermentation filtrate culture media with a mixture ratio of 10,20,30,and 40% on Sclerotium rolfsii Sacc and F pseudograminearum were measured.The results showed that the inhibitory rate increased with the increase of the mixture concentration.The fermentation filtrates of strains Soil1-2 and Soil1-3 reached their highest level of inhibition on the growth of the two disease hyphae at a lower mixture concentration.The inhibitory rates of strains Soil1-59 and Soil2-93 against F pseudograminearum were slightly lower,but they also had significant inhibitory effects.3.Study on the antibacterial mechanism of four biocontrol bacteria.The extracellular enzyme production test showed that both strains Soil1-2 and Soil1-3 could produce protease,amylase,and cellulase;The strain Soil1-59 can produce protease and amylase,but cannot produce cellulase;The strain Soil2-93 can produce protease and cellulase,but cannot produce amylase.Bacterial volatile gas inhibition test showed that the four biocontrol strains could significantly inhibit the growth of Sclerotium rolfsii Sacc hyphae,but could not inhibit the growth of F pseudograminearum hyphae.The crude protein extracted from the fermentation supernatant of four strains of bacteria by ammonium sulfate precipitation method had a inhibition rate of 19.51 to 34.23% on the growth of Sclerotium rolfsii Sacc mycelium;The inhibition rates against F pseudograminearum varied greatly,with the inhibition rates of strains Soil1-2 and Soil1-3 being 79.59% and 83.52%,and the inhibition rates of strains Soil1-2 and Soil1-3 being 17.24% and 27.79%.4.Pot culture control effect measurement.Under greenhouse conditions,the control effects of 72 h fermentation broth of strains Soil1-2 and Soil1-3 on wheat crown rot were tested,as well as the control effects of 48 h fermentation broth of four biocontrol strains on southern blight of peanut.The results of wheat pot cultivation showed that there was no significant difference in the disease index between the treatment group and the control group.Compared with the fermentation broth treatment,the disease index was slightly lower with a control effect of 10%.The control effects of strains Soil1-2,Soil1-3,Soil1-59,and Soil2-93 on southern blight of peanut were the highest on the fifth day after treatment,with protective and control effects of 51.06%,40.42%,63.83%,and 41.58%,respectively;The treatment and prevention effects were 50.35%,29.06%,47.96%,and 36.11%,respectively.The strains Soil1-2 and Soil1-59 have better control effects on southern blight of peanut,and their protective effects are superior to those of treatment.5.The growth promoting effect of biocontrol bacteria on peanut seedlings.After continuously treating peanut seedlings with biocontrol bacteria fermentation broth for two weeks,various indicators of peanut plants were measured.The results showed that the fermentation broth of four biocontrol bacteria had no significant growth promoting effect on peanut seedlings.6.Effects of biocontrol bacteria on defense enzyme activities in peanut seedlings.The activities of polyphenol oxidase(PPO),catalase(CAT),and phenylalanine ammonia lyase(PAL)in peanut leaves were measured on the 1st,4th,and 7th days after peanut seedlings were irrigated with bacterial fermentation broth.The enzyme activity in peanut leaves in the treatment group was significantly higher than that in the control group,and the overall variation pattern of PPO enzyme activity showed a first increase and then a decrease;The change trend of enzyme activity of CAT and PAL is similar,that is,gradually decreases with time.In summary,the four strains of biocontrol bacteria isolated from the rhizosphere soil of wheat have strong inhibitory ability to the growth of the hyphae of Sclerotium rolfsii Sacc and F pseudograminearum.The results of the pot experiment on wheat stem rot were poor.The pot experiment on peanut white spot had significant control effects,but no growth promoting ability.The enzyme activity measurement showed that the activity of defense enzymes in peanut leaves was improved.It has been proved that the four strains of biocontrol bacteria have great biocontrol potential in the control of southern blight of peanut. |
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