The Research Of Phytophthora Infestans On Population Genetic Structure And Mechanism Of Effector Proteins Promoting Late Blight Disease

Potato is the third most important food crop after rice and wheat,and China is the most potato production in the world which account for 22%and 26%of the world acreage and yield respectively.Late blight caused by Phytophthora infestans is the most devastating disease of potato,leading to around 1 billion US dollars of direct annual economic losses in China only.Resistant cultivars and fungicides are two main approaches used to control potato late blight.However,due to its fast evolution,resistant cultivars and fungicides are usually conquered by the pathogen in several years.Therefore,insights to the genetics of host pathogen interaction in molecular level and the evolution of the pathogen at population level are both needed for sustainable management of late blight.Pathogen population genetics aims to infer evolutionary history and decrypt effect of mutation,recombination,gene flow,genetic drift and selection on pathogen evolution by randomly analyzing a set of samples(usually more than 30 isolates)from natural or field population.Populations with large effective population size?high genetic variation and sexual reproduction tend to have high adaptability to the changes of environment,and quickly overcome resistant cultivars and fungicides.Gene flow can be affected by pathogen transmission mode and human activity(for example trade of agricultural products),it can bring high virulent or new genotype pathogen isolates to new places and leads to severe disease epidemic.Population analysis of mutation rate?selection?migration?genetic diversity?mating system?transmission mode?effective population size etc can provide information on pathogen genetic structure?evolutionary speed and capacity.In the first part of my dissertation,SSR markers were used to analyze genetic variation,population differentiation and gene flow in the 15 populations of P.infestance from different provinces.Results show that generic variation in allelic richness and gene and genotype diversity is relatively low in all of the populations assayed suggesting that the P.infestans populations in China is mainly reproduced by asexual reproduction even though large numbers of self-fertile isolates were found.On average,genetic diversity of pathogens from seed-tuber import places(for example southern winter crop region and central double-crop region)is higher than those from seed-tuber export places(for example northern single-crop region and southwestern mixed-crop region).The genetic differrentiation among populations is low(Fst=0.201).AMOVA analysis reveals that only 14.7%and 8.42%of total SSR variation are distibutted among populations within cropping region and among cropping regions,respectively.Indicate that both natural and human-mediated dispersals contribute to spatial poulation genetic structure of P.infestans.Isolation by distance analysis shows that gene flow between pathogen populations is negatively correlated with geographical distance and principal component analysis(PCA)based on Nei's genetic distance groups divided the 15 populations into 3 clusters.Two of the clusters contain populations from more than one cropping region.These results indicate that both nature and human mediated gene flow plays an important role on the population genetic structure of P.infestans.Therefore,in addition to control potato late blight with resistant cultivars and fungicides,attentions should be taken to reduce human-mediated gene flow by strick inspection and quarantine process to seed tubers.In molecular level,pathogen virulence and aggressiveness are two important factors determining the levels of disease epidemics.Over host pathogen coevolutionary history,plants have developd at least two immunity systems,i.e.PAMP/MAMP triggered immunity and effector triggered immunity to defense against pathogen infection and pathogens continue to evolve an array of apoplastic and cytoplasmic effectors to modulate host,suppress host immunity and increase pathogen pathogenicity.It is predicted that P.infestans can secrete at least 500 RXLR cytoplasmic effectors,but only less than 10%of them were functional analysised.Previous studies indicate that RXLR effector PiAVR2 interacts with putative phosphatase StBSL1,but mechanisms of the interaction are still unknown.The result in the second part of my thesis shows that StNRL 1 is a susceptibility factor of potato.It can promote P.infestans infection and suppresses INF1cell death after over-expression.PiAVR2 enhances StBSLl activity.The suppression of INF1cell death of BSL1 after gene silencing is similar to BRI1,indicates StBSL1 is a positive regulator of brassinosteroid(BR)pathway.PiAVR2 transgenic potato has the phenotype of BR pathway overactive,and promotes P.infestans infection.The PTI immunity antagonizes BR pathway.The over-expression of BHLH transcription factor StHBI1-like suppresses PTI immunity in Nicotiana.benthamiana,playing an important node of crosstalk between BR signalling and PTI in solanaceous plants.PiAVR2 interact with StBSL1,and makes BR pathway open,promotes the expression of BHLH transcription factor StHBI1-like,suppresses host immunity and promotes P.infestans infection.Susceptibility factors are a range of proteins that suppress plant immunity directly or support pathogen infection through altering metabolize indirectly.When pathogen effector interacts with and increase its activity,pathogen infection can be boosted.The third part of my dissertation shows that Pi02860 is one of the RXLR effectors localizing in plant cytoplasm.Either transient expression in N.benthamiana,or stable transgenic expression in potato,Pi02860 enhances leaf colonization of P.infestans.Pi02860 suppresses plant PTI immunity,but not CF4/AVR4 cell death.Pi02860 interacts with StNRL 1 on plant plasma membrane.NRL1 is the negative regulator of plant immunity.Silencing of NRL1 in N.benthamiana leads to reduced leaf colonization of P.infestans and promoted INF1 cell death.The suppression of INF1-mediated cell death by GFP-Pi02860 was retained on NRL1 silenced plants,indicates NRL1 plays very important role in the suppression of INF1 cell death of Pi02860.The over-expression of StNRL 1 promotes P.infestans infection and suppresses INF1 cell death,indicates StNRL 1 is a susceptibility factor of potato.P.infestans RXLR effector Pi02860 interacts with StNRL1,suppresses host immunity and promotes plant infection.This dissertation combines population and molecular genetic approachs to inverstigate the evolutionary mechanisms of P.infestans and pathogenicity of effecot proteins.Results from this dissertation could be used to guide the breeding and deployment of resistance gene,effective application of fungicides and design sustainable late blight management.