| Tripterygium wilfordii Hook.f,a precious medicinal plant,are classified into Corey kindo genus Euonymus,which spread across Fujian,Zhejiang,Anhui,Jiangxi and Taiwan province.The existence of endophytic bacteria has been comprehensively found in hygeian plants,which mainly distribute in tissues and organs,interacting and coexisting with the parasitifer.To date,the symbiosis of plant and endophytic bacteria has been proved common in nature.In the micro-ecosystems of plants,endophytic bacteria have essential roles in physiological as well as biochemical activities,some of which have been demonstrated beneficial to the host by phosphate-solubilizing,nitrogen-fixing,siderophore-producing and Indole acetic acid(IAA)-secreting properties.Thus the growth and the disease-resistant capabilities of the host species can be promoted.In recent years,endophytic bacteria which have significant affects in secondary metabolism have been isolated from variant medicinal plants.These were indicated to have the ability of producing active-substances,synthesizing active-materials of the host and accelerating the conversion in secondary metabolisms.In this research,any two of endophytic bacterial strains isolated from T.wilfordii were co-cultured.Capability of growth-promoting and pathogenic-resistant properties of mixtures of endophyitc strains were examined.Further,influences of mixtures of endophytic bacteria on growth of T.wilfordii issue culture seedlings and suspension cells were assessed by co-inoculating plants and adding endophytic bacteria elicitors to the cell suspensionsystem.The results are indicated as follows.Ⅰ.By comparing mixture of strains with corresponding single strains,effects of interactions between endophytic bacteria on growth-promoting traits were evaluated.The results indicated that Interactions between co-cultured bacteria obviously changed the growth-promoting index.Compared with the single strains,abilities of phosphate solubilization and IAA secretion of most mixtures of the strains had great improvement.The strains that were co-cultured with LY4 were found to have larger increment of phosphate solubilization.The strain mixtures including LG3,LJ3 and LJ12 showed higher secretion of IAA.Higher ACC deaminase activity were tested from the strain mixtures including LJ12.The strains LJ12 and LY3 had important contribution to siderophore production of the responding strain mixtures.The growth-promoting index of the mixtures of most of the strains showed no significant correlation to those of the single strains.The growth-promoting properties of the mixtures of the endophytic bacteria did not depend on the abilities of the single strains.Ⅱ.Strain LG6 were indentified as Pseudomonas chlororaphis with 100%similaritythrough 16S rDNA Sequence Identification and homology analysis,while starin LY3 showed 99%homology to Pantoea ananatis.Strain LJ3 and LJ12 shared 100%similarity with Bacillus anthracis.Ⅲ.LG6,LY3 along with their mixed strains displayed optimal in disease-resistant,while LJ 3 and LJ 12 shown no bio-activity in this assessment.It was remarkable that the mixture of all strains did not performed the best in disease-resistant,which indicated that this capability was not defined by the numbers of the active strains.The activity of zymotic fluid was more significant than Sterile fermentation broth.Ⅳ.Endophytic bacterial strains LG6,LY3,LJ3 and LJ12 and their mixtures were made into living strains,crude extracts of dead thallus and bacterium solution,and then were added into the culture systems of plant suspension cells.Growth status of suspension cells after addition of endophytic strains were analyzed by examining fresh weight and activity of suspension cells,pH value,electrical conductivity,soluble sugar and NO3-content of culture media.The results indicated that living endophytic strains had a great effect on the growth of suspension cells.Living strains of LY3,LJ3 and LJ12 and LY3+LJ12 and LG6+LY3+LJ3+LJ12 promoted cell fresh weight compared to control.Living strains of LY3 and majority of mixtures lowed pH values of culture media.Compared to those of control,electrical conductivity of culture media treated by adding most of living strains was increased.Living endophytic strains were also found to change soluble sugar content and increase NO3-content of culture media compared to the control.Different from living strains,crude extracts of dead thallus and bacterium solution disturbed cell growth and decreased cell fresh weight compared to the control.Ⅴ.Four strains of endophytic bacteria and their mixture exerted effects on the growth of T.wilfordii seedlings.When inoculated with LJ12,the fresh weight,dry weight,height and number of leaves of tissue culture seedlings increased evidently.The inoculation of h2 and all strains in mix-treatments promoted the growth of the seedlings significantly.Under the mix-treatments,h4,h5 and h6 shown a negative tendency in all physiological data comparing to the blank.All host plants treated by single strain shown better growth-promoting activity than those treated by mixed strains.Ⅵ.The chlorophyll contents of the seedlings decreased after inoculated strains,except LG6,LY3,h3 and h4 have augments in the content of chlorophyl A,while all of which shown negative effects in the content of chlorophyl B.It was also noticeable that only LG6 performed an increasing in the contents of all chlorophyll comparing to the blank.Generally speaking,inoculated with single strains displayed better than inoculated with mixed strains.The Pn after inoculated with hl,h4,h6 exhibited higher comparing to the blank,while the Gs shown a lower grade after inoculated with LG6 single strain.It is pivotal to say that the Gs of the seedlings performed positive while vaccinated with the other three single strains as well as the samples treated with the mixed strains.An decreasing in Tr value was observed while the seedlings were treated with single strains comparing to the blank,however those treated with mixed strains revealed a higher level except h1 in the mean time.The augments of Tr value after treated with mixed strains indicated an increasing of the loss of transpiration water per unit,which also fitted the fact that the Cr value decreased simultaneously.A tendency of declining in qp were witnessed after inoculation,which demonstrated both treated with single strain and multiple(mix)strains performed negatively in this factor.Besides,Those inoculated with LG6 and LJ12 exhibited a rise in NPQ. |
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