Study On The Host Drought-resistance Mechanisms Of The Enhanced Symbiotic Relationship Between Arbuscular Mycorrhizal Fungi And Peanut By Phomopsis Liquidambaris

Legumes are the most important crop species worldwide and account for 27%of the world's production of major crops.Leguminous plants can form symbiotic relationships with arbuscular mycorrhizal fungi(AMF).As the endophytic partners of AMF,endophytic fungi and AMF have a close interaction relationship.As one of the most important leguminous cash crops in the world,peanut has been planted in the red soil area of low hills in southern China.The serious continuous monocropping problem reduces the yield of peanut,which leads to soil acidification and the deficiency of soil nutrients.However,seasonal drought in the late growth period of peanut in this region further severely restricted the pod development and yield of peanut.Our previous field experiments in Yingtan,Jiangxi Province showed that the application of plant endophytic fungus Phomopsis liquidambaris B3(B3)could significantly increase the yield of continuous cropping peanut.However,the mechanism of the symbiotic interaction between B3 and AMF in peanut and the drought resistance of mycorrhizal symbiont under drought conditions are still unclear.Therefore,this paper hypothesized that B3 could improve the drought resistance of peanut by promoting the co-occurrence of AMF and peanut In order to prove this hypothesis,we constructed a peanut-B3-AMF tripartite symbiosis system to study the plant-microbial interactions and their ecological functions during different growth periods of peanut under continuous cropping conditions.In order to study the effect of B3 on root-dwelling AMF community assembly and drought resistance of peanut under continuous cropping conditions,firstly,the soil water status of continuous cropping in different growth stages of peanut was adjusted under greenhouse potting conditions.Amplicon sequencing of AM fungal community structure showed that B3 promoted the symbiosis of AMF(Gigaspora)with host.The colonization of B3 was only detected in the early stage of peanut growth.With the significant differential colonization of Gigaspora,the physiological indexes of plant drought resistance(including stomatal conductance,photosynthesis,transfer of photosynthetic products to roots and root development),growth and yield of peanut plants under drought conditions were significantly enhanced.Based on the results of linear discriminant analysis,we selected two AMF to carry out the experiment of B3-AMF co-inoculation under the condition of sterilized continuous cropping soil.It was found that B3 and different AMFs had diametrically opposite interactions.Among them,the cooperative relationship between B3 and Gigaspora margarita(Gigaspora margarita,G.margarita)improves the physiological indicators of drought resistance and the potassium content of the leaves of the host plant in the later stage of drought treatment(60 days).At this point in time,in the co-inoculation group,the easily extractable-glomalin soil-associated protein(E-GRSP)produced by AMF was significantly accumulated in the plant rhizosphere environment.This implies that the improvement of peanut drought resistance may be related to the accumulation of E-GRSP.Furthermore,after sterilizing the peanut rhizosphere soil obtained in the previous experiment,peanuts were cultivated again,and it was found that the peanuts cultivated in the sterile soil produced by the co-inoculation treatment showed significantly improved physiological indicators of drought resistance and leaf potassium.The E-GRSP extracted and purified from the treated soil can chelate a variety of mineral elements including potassium.Correlation analysis showed that the protein content was significantly positively correlated with the physiological indicators of plant drought resistance and the formation of soil aggregates.In addition,peanut root exudates and B3 metabolites affected by B3 did not have significant differences in the development of different AMFs in the pre-symbiotic phase.Based on the experimental results of natural soil and sterilized soil,soil p H,available phosphorus,soluble sugar content and phosphorus level of host roots are significantly related to the colonization of Gigaspora.This part of the results showed that B3 can improve the drought resistance of peanuts by affecting the AMF community composition in the roots of peanuts and specifically promoting Gigaspora.Based on the foregoing research results,in order to further study the nutrient interaction mechanism and regulation factors of the triple symbiosis system“B3-G.margarita-peanuts”under continuous cropping conditions,this study firstly used the split-root inoculation system to explore the B3-G.margarita at different time points in the early stage of peanut growth.Compared with AMF,B3 is pre-colonized in the roots of peanuts and has a significant preference for directly using the hexose carbon source supplied by the host.The high expression of the host's fatty acid synthesis and transport genes(RAM2/STR1/STR2)indicates that the plant supplies a large amount of fatty acids to AMF to facilitate its mycelial development and energy storage.The presence of B3 allows AM fungi to maintain a vegetative growth state(the ratio of PLFAs C16:1?5/NLFAs C16:1?5).In return,AMF enhances the translocation of polyphosphate from the root to the root and the expression of the phosphorus transport gene(Gigm T)in the arbuscular region.In the split-root inoculation experiment with different levels of inorganic phosphorus(KH₂PO₄),as the concentration of KH₂PO₄increased,the expression of peanut symbiotic genes was significantly reduced,and the nutrient absorption of plants no longer depended on symbiotic fungi.Moreover,the interaction of the two on the growth promotion effect of the host under low-phosphorus conditions gradually becomes the growth inhibitory effect on the host as the level of KH₂PO₄increases.From the perspective of the host's photosynthetic product supply,the increased level of inorganic phosphorus leads to a decline in the host's supply of non-structural sugars or fatty acids to B3 and AM fungi.At the same time,especially AM fungi,under the conditions of medium and high phosphorus,it retains more polyphosphoric acid in the exophytic hyphae and reduces the expression of phosphorus transport genes in the arbuscular position.This part of the results shows that KH₂PO₄level regulates the triple interaction between AMF-B3-peanuts from the perspective of nutrient exchange.Based on the results of previous studies,in order to further study the ecological function of this protein,we studied the drought resistance mechanism of G.margarita-peanut symbiont.Our results showed that peanuts cultivated in the sterilized soil with AM fungi-inoculated history showed significantly enhanced leaves K accumulation(by 28.3%),drought resistance performance and pod yield(by 50.3%and 52.1%).Through collecting different types of mineral-sequestering glomalin from living AM fungal hyphae,the peanut drought resistance was improved only when the K-sequestering glomalin was added.Moreover,we found that peanut root exudates could prime the dissociation of glomalin-bound K and further satisfy the requirement of crops towards K under drought stress.This part of the results shows that the enhanced peanut drought resistance should be attributed to the K-sequestering glomalin.Based on the previous results,we further investigated the effects of peanut continuous cropping regime on the interaction between fungi and bacteria in soil aggregates and the potential mechanism of nutrient exchange.A pot experiment using two soil types showed that the monocropping regime specifically influenced the overall abunbance and community structure of AM fungi and actinomycetes within macroaggregates(by 54.54%,87.38%,57.51% and 44.32%).Structural equation modelling(SEM)analysis showed that soil p H is the most important determinant factor to restrict the causal relationship between AM fungi and actinomycetes(R²=0.62 and R²=0.64).Under non-monocropped regime,however,the synergistic AM fungi-actinomycetes interactions are mainly attributed to the priming effect of easily extracted-glomalin related soil protein(E-GRSP),which enhanced the biomass and phosphatase secretion of actinomycetes.In return,the activated actinomycetes improved the transfer of phosphorus from AM fungi to peanut.These results suggested that monocropping regime restricted the AM fungi-actinomycetes interactions and their potential functions.Overall,this research allows us to better understand the interaction of endophytic fungi-AM fungi-host triple symbionts in traditional agricultural systems from the ecological perspective of plant-microbe interactions.Meanwhile,our study emphasized the ecological importance of optimizing agricultural practices and enhancing arbuscular mycorrhizal symbiosis to maintain the sustainable development of agroecosystems in facing with the agricultural challenges.