Metabolomics Study During The Interaction Between Maize And Rhizoctonia Solani AG-1-IA

The infection of Rhizoctonia solani(R.solani)causes significant losses of maize production every year.Till now there is no effective control strategy.To this end,a thorough understanding of the interaction between maize and R.solani is needed.Numerous studies have shown that metabolomics is a powerful tool for revealing the mechanisms of interactions between crops and pathogens.In this study,we used metabolomics based on mass spectrometry and multivariate and univariate analysis to explore the mechanism of interaction between R.solani and maize and the values of R.solani for drug development.The sclerotia of R.solani AG-1-IA can not be formed in sealed PDA plates,but it can be naturally formed in non-sealed plates.The concentration of N2-acetyl-L-ornithine in the vegetative mycelia was the highest,which was less in the mycelia that did not form sclerotia,and was the least in the mycelia which formed sclerotia.The result suggested that a high level of N2-acetyl-L-ornithine was required for the vegetative growth of R.solani mycelia,while its concentration needed to be inhibited when R.solani mycelia became mature.When the sclerotia was formed,the up-regulation of PE(20:1(11Z)/14:1(9Z-)),PE(P-16:0/20:4(5Z,8Z,11 Z,13E)(15OH[S]))and PS(12:0/18:1(9Z))indicated that phospholipids played important roles in the process of sclerotia formation.The production of NO was enhanced by the formation of sclerotia while its metabolism in the mature mycelia without sclerotia was inhibited since the metabolic level of N(gamma)-nitro-L-arginine,an inhibitor of nitric oxide synthase,was increased when the sclerotia was not formed,and its concentration was decreased during sclerotia formation.The fact that the concentration of 9S,10 S,11R-trihydroxy-12 Z and 15Z-octadecadienoic acid was significantly higher in the undifferentiated group indicated that linoleic acids played important roles in the mature mycelia without sclerotia.Our results suggest that different levels of these metabolites may act as biomarkers for the developmental stages of R.solani AG-1-IA.The PAA had similar effects as R.solani on maize leaves on the aspect of the metabolism of traumatin which was up-regulated,while the syntheses of OPC-8:0 and 12-OPDA that are precursors for the synthesis of jasmonate acid,a plant defense signaling molecule,were down-regulated under both treatments.The enhanced synthesis of phytosphingosine in the two treated leaves indicated that reinforcing the cell wall is preferred by maize leaves to resist further infection.The up-regulation of vitexin 2'' O-beta-D-glucoside,rutin,gibberellin A14 aldehyde and DIBOA-glucoside in the two treated leaves suggested that maize leaves can utilize different types of resistant small molecules to resist R.solani Infection.However,discrepancy also existed in the influences exhibited by R.solani and PAA since in the R.solani infected leaf group the metabolic concentrations of canthaxanthin and zeaxanthin diglucoside were decreased while the expression of HOOA-PC was up-regulated.Conversely in the PAA treated leaf group,the synthesis of HOOA-PC was inhibited while the syntheses of canthaxanthin and zeaxanthin diglucoside was enhanced.Moreover,although syntheses of 15 metabolites were suppressed in both R.solani and PAA treated leaf tissues,the inhibitory effect exhibited by R.solani was stronger than that of PAA.The strengthened expressions of quercitrin and quercetin 3-O-glucoside in maize sheaths treated by the R.solani,suggested that flavonoids were the main resistant small molecules in sheaths.Furthermore,a significant decrease in the concentration of L-glutamate,which plays important roles in plant resistances to necrotrophic pathogens,only happened in the R.solani treated sheath tissues.The differentiated metabolite levels may have explained why maize sheaths were more susceptible to R.solani than leaves and the underlying mechanisms of R.solani pathogenesis.Metabolomics successfully revealed the metabolic patterns in R.solani when induced by different maize tissues.The induction of maize leaves significantly promoted the concentration of 2,3-Dinor-8-iso prostaglandin F2 alpha and 4-Ketoifosfamide.Meanwhile the expressions of CYP2 and EPHX2 related to the synthesis of 2,3-Dinor-8-iso prostaglandin F2 alpha and the expression levels of alcohol dehydrogenase(EC 1.1.1.1)related to the synthesis of 4-Ketoifosfamide were increased.The results showed that the infection on leaves was mediated by arachidonic acid.In addition,the cytochrome P450 pathway was used for detoxification of exogenous toxins.The induction of maize sheaths significantly enhanced the synthesis of L-Arginine and L-kynurenine,while is the intermediate metabolite in tryptophan metabolism.Meanwhile the expression levels of the upstream synthease indoleamine 2,3-dioxygenase [EC: 1.13.11.52] and the downstream consumption gene kynurenine aminotransferase [EC: 2.6.1.7] related to L-kynurenine were up-regulated.In the induction of maize roots,the concentrations of 3alpha,7alpha,12alpha-Trihydroxy-5beta-cholestanoate and related enzyeme alpha-methylacyl-Co A racemase(EC 5.1.99.4),stearidonic acid and related Acetyl Coenzyme C acyltransferase(EC 2.3.1.16)were significantly increased.The metabolites,the concentrations of which were significantly downregulated during the induction of the three maize tissues,were(1R)-Glutathionyl-(2R)-hydroxy-1,2-dihydronaphthalene,hexadecanoic acid,deoxycyti-dine and gibberellin A15.The expression levels of glutathione transferase(EC 2.5.1.18)associated with the synthesis of(1R)-Glutathionyl-(2R)-hydroxy-1,2-dihydronaphthalene and palmitoyl [protein] hydrolase which used for synthesis of hexadecanoic acid were decreased.The results showed that the infection on different maize tissues was closely related to the Cytochrome P450 metabolic pathway and was selective in different signal transduction pathways.Drug molecules were proved to exsit in the extract of R.solani mycelia.Twenty eight drug molecules with various physiological activities were found in the KEGG database.Among them four drug molecules,Anileridine,Proscillaridin,Bisoctrizole and Baricitinib,were validated by two drug mining algorithms.These pharmacological effects in the four drug molecules are used for pain inhibition,treatments of arrhythmia,absorption of ultraviolet radiation and treatments of rheumatoid arthritis.In the ISDB database four metabolites,(2S)-2beta-Pentyl-5alpha-[(R)-1-hydroxy-9-decenyl] oxolane-3beta-ol,2,4-Di-tert-butylphenol,Heptane-1,3,4,5,7-pentol and Lanyuamide II,were discovered.They are potential drug molecules with drug functions in nematode,anti-oxidation,inhibition of glucosidase and diastolic blood vessels.This study provided a new perspective for recognition of R.solani.