title: Coupling Spore Traps and Quantitative PCR Assays for Detection of the Downy Mildew Pathogens of Spinach (Peronospora effusa) and Beet (P. schachtii).
abstract: Downy mildew of spinach (Spinacia oleracea), caused by Peronospora effusa, is a production constraint on production worldwide, including in California, where the majority of U.S. spinach is grown. The aim of this study was to develop a real-time quantitative polymerase chain reaction (qPCR) assay for detection of airborne inoculum of P. effusa in California. Among oomycete ribosomal DNA (rDNA) sequences examined for assay development, the highest nucleotide sequence identity was observed between rDNA sequences of P. effusa and P. schachtii, the cause of downy mildew on sugar beet and Swiss chard in the leaf beet group (Beta vulgaris subsp. vulgaris). Single-nucleotide polymorphisms were detected between P. effusa and P. schachtii in the 18S rDNA regions for design of P. effusa- and P. schachtii-specific TaqMan probes and reverse primers. An allele-specific probe and primer amplification method was applied to determine the frequency of both P. effusa and P. schachtii rDNA target sequences in pooled DNA samples, enabling quantification of rDNA of P. effusa from impaction spore trap samples collected from spinach production fields. The rDNA copy numbers of P. effusa were, on average, ≈3,300-fold higher from trap samples collected near an infected field compared with those levels recorded at a site without a nearby spinach field. In combination with disease-conducive weather forecasting, application of the assays may be helpful to time fungicide applications for disease management.
authors: Steven J
authors: SJ
authors: Klosterman
authors: Amy
authors: A
authors: Anchieta
authors: Neil
authors: N
authors: McRoberts
authors: Steven T
authors: ST
authors: Koike
authors: Krishna V
authors: KV
authors: Subbarao
authors: Hermann
authors: H
authors: Voglmayr
authors: Young-Joon
authors: YJ
authors: Choi
authors: Marco
authors: M
authors: Thines
authors: Frank N
authors: FN
authors: Martin
mesh_terms: D001483:Base Sequence; D027461:Beta vulgaris / Q000382:microbiology*; D017931:DNA Primers / Q000235:genetics; D004275:DNA, Ribosomal / Q000235:genetics; D057230:Limit of Detection; D008969:Molecular Sequence Data; D044742:Peronospora / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D010935:Plant Diseases / Q000382:microbiology*; D060888:Real-Time Polymerase Chain Reaction; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013045:Species Specificity; D018724:Spinacia oleracea / Q000382:microbiology*; D013170:Spores / Q000302:isolation & purification*
chemical_list: D017931:DNA Primers; D004275:DNA, Ribosomal