Identification Of The Pathogen Of Chinese Chestnut Yellow Crinkle Disease And Its Control

In recent years,a disease,named "Small leaf of chestnut",has occurred in northeast Hebei Province.It spread too rapidly to threatens the healthy development of chestnut industry.To identify of the pathogen of the disease and explore a set of effective comprehensive prevention and control techniques.16S r DNA sequence of the pathogen was amplified,cloned,and sequenced.Blast showed that the similarity of the pathogen with Candidatus phytoplasma castaneae CnYC(EU599362)and Ca.P.castaneae(AB054986)was 99.44% and99.60%,respectively.The round or nearly circular phytoplasmas were observed in phloem of branches and leaves by Transmission Electron Microscope that phytoplasma,with the size of 0.1-0.7 ?M.The results showed the pathogen of "Small leaf of chestnut" is Ca.P.castaneae,and the "Small leaf of chestnut" in eastern and northern Hebei is Chinese chestnut Yellow Crinkle(CnYC).The virtual RFLP analysis based on 16 S r DNA sequence showed that the pathogen Ca.P.castaneae belonged to 16 Sr XIX-A subgroup and the strain number was CnYC-Hebei.Using the specific sequence of phytoplasma 16 S r DNA gene to detect chestnut leaves and new shoots showing chestnut yellowing and shrinkage disease in different growth stages by nested PCR amplification,the results showed that the average detection rates of diseased leaves in the budding stage,leaf development stage,flowering stage,maturity stage and pre-deciduous stage were 0,0,20.04%,47.52%,and 66.27%,respectively.The average detection rates in the early stage were 0,33.18% and 52.28%.This indicates that the detection rate of chestnut phytoplasma increases with the extension of the growth period.The control effect of seven antibiotics on chestnut yellow shrinkage disease was detected by hole hanging bag infusion method.The control effect of Tetracycline hydrochloride were 85.34%,The control effects of tetracycline,polypeptide tetracycline and roxithromycin were more than 62.41%.Then,Comprehensive prevention and control test was complete with tetracycline hydrochloride andagricultural cultivation measures.The control effect of comprehensive prevention and control test significantly better than the control.This study provides technical support for the prevention and control of the CnYC disease.