title: Rapid and sensitive detection of Phytophthora colocasiae responsible for the taro leaf blight using conventional and real-time PCR assay.
abstract: Conventional and real-time PCR assays were developed for sensitive and specific detection of Phytophthora colocasiae, an oomycete pathogen that causes leaf blight and corm rot of taro. A set of three primer pairs was designed from regions of the RAS-related protein (Ypt1), G protein alpha-subunit (GPA1) and phospho-ribosylanthranilate isomerase (TRP1) genes. In conventional PCR, the lower limit of detection was 50 pg DNA, whereas in real-time PCR, the detection limit was 12.5 fg for the primer based on Ypt1 gene. The cycle threshold values were linearly correlated with the concentration of the target DNA (range of R(2) = 0.911-0.999). All the primer sets were successful in detecting P. colocasie from naturally infected leaves and tubers of taro. Phytophthora colocasiae was detected from artificially infested samples after 18 and 15 h of postinoculation in conventional and real-time PCR assay, respectively. The developed PCR assay proved to be a robust and reliable technique to detect P. colocasiae in taro planting material and for assessing the distribution of pathogen within fields, thus aid in mitigating taro leaf blight.
authors.affiliation: Division of Crop Protection, Central Tuber Crops Research Institute, Thiruvananthapuram, India.
authors.forename: Vishnu S
authors.initials: VS
authors.lastname: Nath
authors.forename: Vinayaka M
authors.initials: VM
authors.lastname: Hegde
authors.forename: Muthulekshmi L
authors.initials: ML
authors.lastname: Jeeva
authors.forename: Raj S
authors.initials: RS
authors.lastname: Misra
authors.forename: Syamala S
authors.initials: SS
authors.lastname: Veena
authors.forename: Mithun
authors.initials: M
authors.lastname: Raj
authors.forename: Suresh K
authors.initials: SK
authors.lastname: Unnikrishnan
authors.forename: Sree S
authors.initials: SS
authors.lastname: Darveekaran
mesh_terms: D031060:Colocasia / Q000382:microbiology*; D017931:DNA Primers / Q000235:genetics; D008828:Microbiological Techniques / Q000379:methods*; D008969:Molecular Sequence Data; D010838:Phytophthora / Q000235:genetics / Q000302:isolation & purification*; D010935:Plant Diseases / Q000382:microbiology*; D018515:Plant Leaves / Q000382:microbiology; D035281:Plant Tubers / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA
chemical_list: D017931:DNA Primers
keywords: Phytophthora colocasiae; SYBR green assay; disease management; qPCR; single-copy genes; specific detection