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Title: PCR-based assays for the detection of Puccinia horiana on chrysanthemums

item Pedley, Kerry

Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/19/2009
Publication Date: 11/10/2009
Citation: Pedley, K.F. 2009. PCR-based assays for the detection of Puccinia horiana on chrysanthemums. Plant Disease. 93:1252-1258.

Interpretive Summary: Chrysanthemum white rust, caused by Puccinia horiana, is an economically important disease that infects most chrysanthemum cultivars grown worldwide for cut flower and potted plant production. The disease is particularly problematic to commercial chrysanthemum propagation in greenhouses and nurseries where environmental conditions foster its spread. The disease is not established in the U.S. and is listed by USDA, APHIS as a quarantine significant pest. Current identification protocols for P. horiana rely upon both macroscopic symptoms and morphological features of the pustules as well as microscopic examination of the teliospores. For this reason, during the initial stages of infection the pathogen can often elude detection. In greenhouses and nurseries, early and accurate identification of white rust is essential to limit the spread of the pathogen and to ensure clean stock for propagation. The identification of plant pathogenic fungi based on molecular markers is advantageous because it does not require the presentation of fully developed disease symptoms or morphological features of a pathogen, and therefore can be used to confirm the presence of a pathogen during the early stages of an infection. This work describes the development of conventional and real-time PCR assays for the detection of the pathogen. Both assays were specific and sensitive, enabling detection of P. horiana in infected plant tissue prior to the development of symptoms.

Technical Abstract: Puccinia horiana, the causal agent of chrysanthemum white rust, is a pathogen of quarantine status in many countries where Chrysanthemum × morifolium cultivars are grown. Current identification protocols for white rust rely upon macroscopic symptom development and microscopic examination of infected leaves for teliospores. Symptoms become visible 7 – 10 days after initial infection under favorable conditions followed by the production of telia. Infected plants can therefore evade detection before symptoms and fruiting bodies are evident. Here I describe the development of conventional and real-time polymerase chain reaction assays using primers designed to amplify portions of the ribosomal internal transcribed spacer (ITS) regions. Both assays were capable of detecting the presence of P. horiana in asymptomatic tissue. The primers were also tested using DNA isolated from leaf tissue infected with chrysanthemum brown rust, caused by P. tanaceti, but did not show any cross reactivity.