A Rapid Identification Method for Germplasm Materials Resistant to Citrus Huanglongbing Disease

[Objectives]This study was conducted to develop a rapid identification method for citrus germline materials resistant to Huanglongbing disease and lay a basis for accelerating citrus breeding for resistance to Huanglongbing and increasing the breeding efficiency.[Methods]Thirty-six citrus germplasms suspected to be resistant to citrus Huanglongbing disease were collected.The method of direct high grafting to citrus trees infected with Huanglongbing pathogen was adopted.The resistance of the test materials was identified and evaluated by field symptoms combined with quantitative PCR.It was defined as the top grafting identification method.[Results]The test materials that were grafted in spring started to germinate after one month,and three months late(June 5,2018)typical mottled yellowing on leaves was observed on KH-14 for the first time.After four months(July 5,2018)of top grafting,typical mottled yellowing occurred on 23 materials,and 11 materials showed no such symptom.After six months(September 4,2018)of top grafting,although the growth of KH-18,KH-12,KHY-4,KHY-5 and KHY-6 were normal,yellowing was observed on their leaves.Only KH-21 grew well,and showed no yellow shoots and yellowing leaves.It was identified as the material with resistance to Huanglongbing disease.Quantitative PCR tests on the above six materials showed that KH-21 was negative and other five were positive.Real-time fluorescence quantitative PCR test indicated that the average Huanglongbing bacteria amount in KH-21 was 1870.0 cell/μg DNA,and the average Huanglongbing bacteria amount in the control material was 372285.5 cell/μg DNA,indicating KH-21 was resistant to Huanglongbing bacteria.[Conclusions]The method for infecting bacteria by top grafting takes six months,can detect large amount of seedlings,and is time-saving,efficient,cost-saving and accurate.This method can quickly identify the resistance of citrus varieties to citrus Huanglongbing disease,and can be popularized and used in the identification of citrus Huanglongbing disease resistance.