Conventional and real-time PCR-based assay for detecting pathogenic Alternaria brassicae in cruciferous seed. Thomas Guillemette, UMR 77 Pathologie Végétale, Faculté des Sciences, Angers, France; Béatrice Iacomi-Vasilescu, UMR 77 Pathologie Végétale, France, and USAMV, Department of Plant Protection, Bucharest, Romania; and Philippe Simoneau, UMR 77 Pathologie Végétale, France. Plant Dis. D-2004-0301-01R, 2004 (online). Accepted for publication 19 December 2003.

Alternaria brassicae is an important and widely distributed pathogen of crucifers. This fungus is responsible for the black spot disease that results in serious reductions in crop yields. Genetic control of the pathogen is not possible because most commercial cultivars are susceptible. Consequently, the use of pathogen-free seed is essential to limit the spread and incidence of the disease and also to reduce fungicide applications. Therefore, sanitary certification pro­grams for commercial cruciferous seed include A. brassicae detection. Diagnosis currently is obtained after plating seed on nutritive media, using incubation and morphological characterization of the fungus. This procedure is time-consuming—requiring at least 1 week to obtain a diagnostic result—and often not very accurate due to potential confusion with nonpatho­genic Alternaria spp. Therefore, there is an urgent need to develop alternative diagnostic tools such as molecular techniques based on polymerase chain reac­tion (PCR). In this article, we describe two molecular assays for detecting A. brassicae in cruciferous seed using conventional or real-time PCR. The two meth­ods perform equally well in terms of specificity, sensi­tivity, and speed. However, the real-time PCR assay is better suited for routine detection because no post-amplification manipulations are required. Further­more, this real-time PCR diagnosis method may be readily amenable to automation.

The current issue of APSNet, Volume 88, Number 5, May 2004, is at http://www.apsnet.org/pd/current/top.asp