Development of an Improved Isolation Approach and Simple Sequence Repeat Markers To Characterize Phytophthora capsici Populations in Irrigation Ponds in Southern Georgia

摘要

Phytophthora capsici, the causal agent of Phytophthora blight, is a major concern in vegetable production in Georgia and many other states in the United States. Contamination of irrigation water sources by P. capsici may be an important source of inoculum for the pathogen. A simple method was developed in this study to improve the efficiency of recovering P. capsici from fruits used as baits in irrigation ponds. In contrast to direct isolation on agar plates, infected fruit tissues were used to inoculate stems of pepper seedlings, and the infected pepper stems were used for isolation on agar plates. With isolation through inoculation of pepper stems, the frequency of recovering P. capsici from infected eggplant and pear fruits increased from 13.9% to 77.7% and 8.1% to 53.5%, respectively, compared with direct isolation on agar plates. P. capsici was isolated from seven out of nine irrigation ponds evaluated, with most of the ponds containing both A1 and A2 mating types and a 4:5 ratio of A1 to A2 when isolates from all ponds were calculated. All P. capsici isolates were pathogenic on squash plants, and only a small proportion (8.2%) of the isolates were resistant or intermediately sensitive to mefenoxam. Simple sequence repeats (SSRs) were identified through bioinformatics mining of 55,848 publicly available expressed sequence tags of P. capsici in dbEST GenBank. Thirty-one pairs of SSR primers were designed, and SSR analysis indicated that the 61 P. capsici isolates from irrigation ponds were genetically distinct. Cluster analysis separated the isolates into five genetic clusters with no more than two genetic groups in one pond, indicating relatively low P. capsici genetic diversity in each pond. The isolation method and SSR markers developed for P. capsici in this study could contribute to a more comprehensive understanding of the genetic diversity of this important pathogen.