Study On The Mechanism Of Enhancing Rice Drought Resistance By Endophytic Fungus P. Sylvestris B3

Plant fungal endophytes can establish a symbiotic relationship with plants.The symbiotic advantage of endophytic fungi is mainly reflected in promoting plant growth and enhancing host stress tolerance.Endophytic fungus B3,isolated from the inner bark of Bischofia polycarpa,can establish a symbiotic relationship with rice and peanuts.Endophytic fungus B3 can promote the nitrogen absorption of rice.Well-developed aerenchyma induced by B3 helps the rice absorb mineral elements under flooding conditions.These growth-promoting effects of B3 on rice laid a substance basis for the rice to survive under the abiotic stress.However,it is still unclear whether B3 can enhance rice drought tolerance and the underlying mechanisms.Therefore,we conducted pot experiments to explore the effects of B3 on rice's physiological characteristics under natural drought conditions,and PEG-6000was used to simulating drought stress at the seedling stage to further explore its underlying physiological mechanism.To explore the effect of endophytic fungus B3 on the physiological phenotype of rice under drought conditions.First,we applied the fungal suspension of B3 to the rice root surface to observe the colonization of B3 in the root of rice.Secondly,we carried out outdoor pot experiments,set up natural drought stress at the seedling stage,the tillering stage,and the reproductive stage.We measured the physiological and biochemical indexes related to drought in different treatment groups at these three growth periods.Such as relative water content of rice leaves,growth parameters(plant height,root length,shoot dry weight,and root dry weight),photosynthetic parameters(chlorophyll content,net photosynthetic rate,stoma conductance,intercellular CO2 concentration,and transpiration rate),antioxidant enzyme(superoxide dismutase,catalase,and peroxidase)activities,osmotic substances(proline,soluble protein,and soluble sugar)content,and yield.The results showed that B3 could successfully colonize rice roots.The leaves of the non-inoculation treatment were curled obviously under drought stress.No obvious curl was found in rice leaves treated with endophytic fungus B3.Under drought stress,the relative water content,photosynthesis(chlorophyll content and net photosynthetic rate),osmotic substances(proline,soluble protein,and soluble sugar)content,and antioxidant enzyme(superoxide dismutase,catalase,and peroxidase)activities of rice leaves treated with endophytic fungus B3 were significantly higher than those of untreated treatment.The stomatal conductance and transpiration rate of rice treated with endophytic fungi B3 were substantially lower than those of the uninoculated treatment.In addition,under drought stress at the seedling stage and reproductive stage,the yield per plant of rice treated with endophytic fungi B3 increased by 43.43%and 17.85%,respectively,compared with the untreated group,which may be related to the enhanced photosynthesis of rice under drought stress by endophytic fungi B3.The above results indicate that endophytic fungus B3 may reduce the oxidative damage caused by drought stress on rice by increasing the antioxidant enzyme activity of rice.Endophytic fungi B3 may increase the water content of rice leaves under drought stress by increasing osmotic substance content in rice and reducing water loss.Endophytic fungus B3 may promote rice growth under drought stress by improving rice photosynthesis.Endophytic fungus B3 eventually increased rice yield by improving rice's physiological state under drought conditions.To further explore the effect of endophytic fungus B3 on photosynthetic physiological characteristics of rice under drought stress.PEG-6000 was used to conduct drought stress treatment on rice at the four-leaf stage,and photosynthetic pigments(chlorophyll and carotenoid)content and chlorophyll fluorescence parameters were measured.At the same time,the physiological indexes related to drought resistance were also measured,such as leaf relative water content,antioxidant enzyme(superoxide dismutase,catalase,and peroxidase)activity,and osmotic substance(proline,soluble protein,and soluble sugar)content.The results showed that under drought stress,compared with the control,endophytic fungus B3 treatment increased the chlorophyll and carotenoid contents of rice by 31.19%and 13.01%,respectively.Under drought stress,compared with the endophytic fungus B3treatment group,the OJIP curve of the non-inoculated treatment showed K point,and the light ABS/RC absorbed by the unit reaction center was higher,indicating that the oxygen release complex of rice in the non-inoculation treatment group was damaged under drought stress.Under drought stress,compared with non-inoculation treatment,endophytic fungus B3 reduced ABS/RC by 18.76%.The decrease of ET0/CS0,ET0/RC,and?E0 indicated that the electron transfer of PS? was strongly inhibited under drought stress.Under drought stress,ET0/CS0,ET0/RC,and?E0 in the endophytic fungus B3 treatment group increased by 96.10%,64.99%,and 41.11%,respectively,compared with those in the non-inoculation treatment.These results indicated that B3 could alleviate the inhibition of drought on the rice electron transport chain.The above results showed that endophytic fungus B3 under drought stress enhanced rice photosynthesis by increasing photosynthetic pigments(chlorophyll and carotenoid)content,the activity of the PS? reaction center(oxygen-releasing complex),and the electron transfer ability of PS?.In addition,the enhanced antioxidant enzyme activity and osmotic substance content of endophytic fungus B3 could further protect the photosynthetic apparatus of rice leaves.When we carried out drought stress on rice at the seedling stage,we found that the rice treated with B3 had a higher relative water content.Endophytic fungus B3can secrete abscisic acid in vitro culture conditions.Abscisic acid is a stress hormone,which plays an essential role in plant defense against the external environment.It has been reported that abscisic acid can induce stomatal closure.We speculate that higher leave relative water content might be related to the stomatal closure caused by B3.So we used a microscope to observe the open and closure of stomata in leaves and detected the contents of ABA and H₂O₂and their gene expression to explore whether B3 reduces water loss by inducing stomatal closure in leaves and whether stomatal changes are related to ABA and H₂O₂.And by adding exogenous ABA and H₂O₂ and their inhibitors,respectively,we tried to explore whether ABA and H₂O₂ were involved in the process of stomatal closure induced by B3 and the upstream,downstream relationship between the two signaling substances.The results showed that B3 induced rice to close stomata through ABA and H₂O₂ pathways,and the ABA was located upstream of the H₂O₂.All in all,the experimental results provide direct evidence for enhancing rice drought tolerance by B3.We believe that is related to the increased ABA level indued by B3 and the induction of H₂O₂ synthesis.Stomatal closure induced by B3 may make an essential contribution to the improvement of physiological level and yield under drought stress.