Establishment Of A Monospore Isolation System And Identification Of Physiological Races Of Cruciferous Root Tuberculosis Bacteria

Clubroot disease caused by Plasmdiophora brassicae Woron.is a destructive worldwide disease that is difficult to control in cruciferous vegetable production.One of the main reasons is the genetic diversity of P.brassicae,which leads to the differentiation of physiological races.It is of great significance for monitoring the physiological differentiation of P.brassicae and disease resistance breeding to define the types and geographical distribution of the physiological microflora.This paper aims to establish a set of simple,stable and widely adaptable technical system for the isolation of P.brassicae from different regions in China.The single-spore strains obtained were identified by Williams identification system.The main results are as follows:1.A set of simple,stable,high survival rate,high morbidity and wide adaptability technical system of P.brassicae single-spore separation was established.By improving the agar block method and replacing the conventional agar block with the agarose coagulation layer,the problem that the agarose block method contains many impurities and is difficult to distinguish the single spore was solved,and the optimal concentration of agarose coagulation layer for the separation of the single spores of P.brassicae was determined to be 1.3%.It was determined that the best seedling age for single-spore inoculation was the Chinese cabbage susceptible variety "juxin" in 81-well box for 24 h,and the best culture condition was dark culture for 24 h after inoculation.Based on the comparison of three kinds of single-spore inoculation methods,including agarose gelation method,drop method and agar block method,the result proved that agarose method was the best single-spore inoculation method with the disease incidence up to 17.45%,which is significantly higher than that of drop method and agar block method.2.The established agarose method was used to separate single spore strains of P.brassicae from 20 different sampling areas in China.275 strains of P.brassicae single spore strain were obtained,and 205 strains of single spore strain were expanded and preserved.3.The differentiation types and geographical distribution of physiological subspecies of brassica brassica in 8 regions of China were defined.By Williams identification system,the physiological species of 40 single spore strains were identified.Ten different physiological subspecies were identified,including P2,P4,P5,P6,P7,P8,P9,P10,P12 and P14.Among them,P4 was the dominant species,and the frequency of occurrence in the sample strain was 42.5%.Secondly,a total of 9 single spore strains were P5,which appeared in 22.5% of the sample strains.The dominant species in four plateau areas,namely,yichang hubei,wulong chongqing,kunming yunnan and guangyuan sichuan,were P4.P4 and P5 were the dominant physiological race in three plains and hilly regions,including Qingdao Shandong,Xinye Henan and Shunyi Beijin,which showed relatively weak infection and propagation ability to identify hosts.Through the research of this paper,a set of simple and stable single spore separation system of P.brassicae was established.By identifying the pathogenic types and distribution of physiological races of P.brassicae in 8 different regions of China,we can provide resistant materials for disease resistant breeding.