| Clubroot disease,caused by an obligate biotrophic pathogen Plasmodiophora brassicae,is an important soil-borne disease which distributes widely in the world.The understanding on mechanism of the host and P.brassicae interaction is of important theoretical and practical significance to effectively control clubroot disease.Previous studies on the interaction between P.brassicae and its host mainly focused on transcriptomics and proteomics analysis,but the involved metabonomics were limited.In the present study,metabolic changes of Arabidopsis thaliana and Brassica napus duing infection with P.brassicae were analyzed by liquid chromatography-mass spectrometry(LC-MS).The main results obtained are as following:(1)An effective method for in vitro inoculation of P.brassicae on A.thaliana without contamination was developped based on previously reported techniques.P.brassicae spores were extracted from callus cultures of clubroot gall sections,and then P.brassicae spores and A.thaliana seeds were co-cultured on 1/2MS medium.Microscopic observation showed that P.brassicae can reproduce in callus cells and complete the infection cycle in the roots of A.thaliana.This established method of P.brassicae inoculation on A.thaliana could effectively avoid contamination by other microbe.(2)The metabolic change of A.thaliana roots infected with P.brassicae at the late stage was assayed on 1/2MS medium via LC-MS technique.The results showed that the metabolites of flavonoids,lignans,terpenoids,steroidal glycosides,brassinosteroids and phospholipids were significantly increased during P.brassicae,and could play potential important roles in interaction between A.thaliana and P.brassicae at the late stage of infection.Pathway analysis revealed that fatty acid,aminosugars,and tryptophan metabolism of A.thaliana were effected by P.brassicae.(3)To clarify the early response of host to the infection of P.brassicae,metabonomics analysis of B.napus were conducted via LC-MS.Multivariate statistical methods of PLS-DA and OPLS-DA,and univariate statistical methods of T-test and Two-way ANOVA were used for pattern recognition and candidate biomarker screening.The results showed a significant difference of metabolites in B.napus at 1 day post inoculation(dpi),2 dpi and 5 dpi.Compared to other pathogen infection stages,metabolites of B.napus were most significantly changed at 1 dpi,which indicated that B.napus showed strong response to P.brassicae at the early infection.Some important metabolites were identified during P.brassicae infection.Those identified metabolites included phospholipids,flavonoids,lignans,terpenoids and alkaloids that were related to resistance to biotic and abiotic stresses.Indole-3-acetaldoxime,which was invoved in IAA synthesis,and 2-oxoarginine,4-guanidino-butyric acid and indole,which were related to arginine and aromatic amino acid synthesis and metabolism,were also identified as candidate biomarkers.These results revealed that host could exhibit a timely resistant response at both primary and secondry metabolism to promote the synthesis of resistance-related substances,once host recognized the pathogen infection,and that phospholipids and auxin played a key role in host-P.brassicae interaction at the early stage of infection.The results elso showed that the resistance-related secondry metabolites were changed significantly at both type and abundance,which indicated a more complicated function of them during pathogenic process of P.brassicae.Pathway analysis revealed that pathways related to the metabolism of amino acid,carbohydrate and cytochrome P450 and the biosynthesis of alkaloid were significantly changed at the early infection stage. |
PDF Full Text Request