| Potato(Solanum tuberosum L)belongs to Solanaceae of Solanum,and an annual dicotyledonous herbaceous plant.It's the fourth largest food crop after rice,corn and wheat.In January 2015,the Ministry of Agriculture of the PRC proposed the potato main grain strategy.Therefore,developing potato industry,preventing potato diseases,and increasing potato yield are an inevitable choice for achieving the goal.Late blight caused by Phytophthora infestans(Mont.)de Bary is a worldwide destructive disease in potato production,which seriously threatens the yield of potato.The use of antagonistic bacteria and their metabolites to control plant diseases,promote plant growth and improve soil microbial community structure has attracted researchers' attention.In recent years,it has been found that the compound microbial preparation could overcome the disadvantages of the single antagonistic bacteria,such as the unstable antibacterial activity and the poor effect of disease prevention and so on.In previous experiments,compound fermentation(CFBF-I)from four strains of antagonistic bacteria(Sy11,SR13-2,ST-1 and HT-6)were used to control potato late blight on detached leaves and tuber slices as well as potted plants in our laboratory and obtained good disease protection effect.But all the four strains of Sy11,SR13-2,ST-1 and HT-6 were not isolated from potato plant,and might be difficult to colonize in potato during practical disease prevention in future.Later,our lab acquired bacterium strain W-7 from the leaf infected by P.infestans in farm field of Keshan,Heilongjiang province,which showed very strong suppression to mycelia growth and sporangia germination of P.infestans.In this paper,the antagonistic strain W-7 was added in compound fermentation CFBF-1 and made it into compound fermentation CFBF-II in order to further enhance the disease control ability of CFBF-II on potato plant and promote its colonization ability on potato plants.The main research contents were as follows:(1)Plate streak method and dual culture method were applied to clear that whether five strains of Sy11,SR13-2,ST-1,and HT-6 could be cultured together,and the way of their complex fermentation for obtaining the compound ferment liquid(CFBF-II),along with influence of CFBF-II on the physiological and biochemical aspects of P.infestans.(2)The methods of soaking,irrigation and plate counting were used to explore the disease prevention effect of CFBF-II on potato in vitro and potato plants,and the effects on plant growth promotion and rhizosphere microbial community,as well as that of CFBF-II on chlorophyll content of potato leaves.(3)The effects of CFBF-II on soluble protein and defense-related enzymes in potato resistance to late blight were studied by Coomassie brilliant blue method and SDS-PAGE electrophoresis.The main research results are as follows:1.There was no obvious antagonism between 5 strains of antagonistic bacteria,which could be used for compound fermentation and acquired compound ferment bacterial fluid(CFBF-II).Compared with the CFBF-I and single HT-6 strain with the strongest inhibitory activity among 5 strains,CFBF-II showed stronger suppression effect on P.infestans mycelia growth,and the inhibition rate was 83.8%for culturing 11 th day after treatmet.There was significant diffence between CFBF-II and CFBF-I(P<0.05).The content of soluble protein in mycelia treated with CFBF-II was 72.8% which was greatly lower than that of the control group,and evident lower that of CFBF-I,but no significant difference between them.Microscopic examination showed that the mycelia deformation rate was 73.96% in CFBF-II,significantly higher than that in CFBF-I(39.76%)(P<0.05).The inhibition rates of direct sporangium germination and zoospore release after CFBF-II teatment were 33.4% and 32.5%,respectively,remarkable higher than that of CFBF-I,there were sigficant differences between them.2.The effect of CFBF-I and CFBF-II on seed potato germination was significantly better than that of Control and HT-6 bacterial fluid.Moreover,the bud hight after treatment with CFBF-II was up to 17.0mm,and the bud diameter reached 4.1mm,in maximum.The germination rate of seed potato was 41.18%,significantly better higher that of CFBF-I(29.41%).CFBF-II treatment had the best effect in control potato late blight on tuber slices and detached leave compared with CFBF-I and HT-6 treatments as well as water control,the protection effects were 63.8% and 80.0% on tuber slices and leaves,respectively.They were higher than that of CFBF-I,which were 25.0% and 10.9%,respectively.In comparison with LB liquid medium treatment,plant height,stem diameter,leaf number,leaf area,taproot length and fibrous root number after CFBF-II treatment were increased by 14.8%,26.5%,16.6%,32.9%,33.7% and 25.1%,respectively on the 55 th day.In addition,on the 50 th day after CFBF-II treatment,the number of bacteria increased in potato rhizosphere soil,but the number of fungi,Fusarium oxysporum and actinomycetes was decreased evidently.And CFBF-II ttreatment also significantly increased the total content of chlorophyll a+b in potato leaves.3.The content of soluble protein in potato tubers increased after CFBF-II treatment,which was significantly better than the promotion effect of CFBF-I.Especially the treatment combining induction of CFBF-II with challenge inoculation of P.infestans could enhance the content of soluble protein in potato tubers in greater extent than the treatment only induction of CFBF-II.SDS-PAGE electrophoresis results showed that soluble protein in potato tubers between CFBF-II,CFBF-I,and HT-6 as well as control was no significant difference in number and content(brightness)of protein band in late blight resistant potato varieties(Dabaihua).While in the potato varieties(Fevorita)which is susceptible to late blight,the number of protein band after induction with CFBF-II,CFBF-I,and HT-6 as well as control was fewer than that in Dabaihua)cultivar,and the band brightness became also weak.In addition,the band brightnees in areas from 14.4 KD to 24 KD and from 97.4 KD to 100 KD on SDS-PAGE electrophoresis after combining induction with CFBF-I or HT-6 and challenge inoculation pathogen was decreased greatly even disappeared in comparison with treatment only induction no inoculation.Only the CFBF-II induction with inoculation significantly increased the band brightnees in two areas compared with only induction with CFBF-II.It indicated that there exsited the relationship between CFBF-II induction and P.infestans infection on protein content of potato tuber,but no relation between CFBF-I or HT-6 induction and P.infestans infection.In addition,treatment with CFBF-II induction could enhance the activities of three resist-related enzymes POD,PPO and PAL in potato tuber slices,especially the CFBF-II induction in combination inoculation increased three enzyme activities greater than that only induction treatment.Among them,POD activity reached peak at 6th day after induction,but both PPO and PAL activities at 4th day.It suggested that PPO and PAL might play role in potato against P.infestans before POD.The main research conclusions :(1)There was no antagonistic effect between 5 strains of antagonistic bacteria,and CFBF-II showed better effect than single strain and CFBF-I in inhibiting Phytophthora infestans.It could lead high mycelia aberration rate and significantly decrease protein content of mycelia.(2)CFBF-II had no adverse effects on potato tubers and leaves in vitro,and could achieve good results in the prevention of late blight.(3)CFBF-II hadremarkable effects on plant height,stem diameter,leaf number,leaf area and root system,and could improve root microbial community.It could greatly enhance the chlorophyll content of potato leaf.(4)CFBF-II could raise soluble protein content and defense enzyme POD,PPO,and PAL activity,in which POD and PAL might play role before POD enzyme in induction potato against late blight. |
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