Rapid Detection Of Ralstonia Solanacearum Using Recombinase Polymerase Amplification And Biocontrol Effect Of XCS1 Against R.solanacearum

Tobacco bacterial wilt is a bacterial soil-borne disease caused by Ralstonia solanacearum,which causes devastating damage to tobacco and huge economic losses in tobacco-growing areas.Therefore,it is particularly important to accurately detect pathogen and formulate comprehensive prevention and control measures in the early stage of tobacco bacterial wilt.This article aims is to establish a rapid,sensitive and specific RPA detection method for tobacco R.solanacearum,screen out antagonistic strains of tobacco R.solanacearum,and evaluate its antibacterial activity,providing theoretical basis for early field detection and sustainable green prevention of tobacco bacterial wilt.Based on recombinase polymerase amplification technology,specific primers and probes were designed with the Rip TALI-9 gene of R.solanacearum as the target gene.The RPA assay of R.solanacearum was established,which can amplify the target gene at a constant temperature of 38? within 20 minutes,and the detection result can be displayed in 10 minutes combined with lateral flow strip analysis.The RPA assay can detect the genomic DNA of tobacco bacterial wilt at a concentration of 1 pg/L or greater,susceptible plants with minimum bacterial concentration of 103 CFU/g,and soil containing minimum bacteria concentration of 104 CFU/g.These detection results are consistent with conventional PCR.After testing the infested plants and soil collected from the tobacco field,the RPA assay can detect the bacterial wilt of tobacco quickly and accurately in field samples.The antagonistic bacteria were isolated from the rhizosphere soil of tobacco in the plot of tobacco bacterial wilt by flat-pane confrontation method,and a strain of tobacco R.solanacearum biocontrol bacteria XCS1 was screened,which was identified as Pseudomonas fluorescens by physiological,biochemical and molecular biology.The inhibition zone diameter of the bacterial suspension with XCS1 was 19.2 mm,the optimal culture temperature was 30?,the initial pH of culture medium was 7,and the salt content was 2.5%.The biocontrol bacteria XCS1 can produce proteolytic enzymes and a particularly high level of siderophore.Besides,it can also produce VOCs with significant inhibitory effect on R.solanaceae.In addition,the control effect of biocontrol bacteria XCS1 on tobacco bacterial wilt was 56.1%by greenhouse pot experiments.