European Journal of Plant Pathology

, Volume 143, Issue 4, pp 663–675 | Cite as

Rapid, specific and sensitive molecular detection assay for Alternaria helianthi that causes leaf blight disease in sunflower

  • R. L. Chavhan
  • V. R. Hinge
  • M. B. Chinchole
  • P. K. Chakrabarty
  • V. Y. Patade
  • H. B. Patil
Article

Abstract

The leaf blight disease of sunflower (Helianthus annuus) caused by Alternaria helianthi is a serious threat to its cultivation worldwide. Early and accurate detection of the pathogen is critical to efficient disease management and in avoiding further losses due to epidemics in sunflower. Conventional methods of detection and identification are time consuming, labour intensive, and lack specificity and sensitivity. A real-time PCR based TaqMan® probe assay was developed to target 156 bp Internal Transcribed Spacer (ITS) region of A. helianthi for its detection using fungal genomic DNA and infected plant tissues and seeds of sunflower. The specificity of the probe and primers was confirmed by testing their cross reactivity using genomic DNA of closely related Alternaria species isolated from 17 crop plants and 15 fungal species of other genera. No cross-reactivity could be detected with any of the other non-target fungal strains used in this study. The assay successfully detected as low as 1.0 pg fungal genomic DNA and up to 1 % infection in sunflower seed lots. To the best of our knowledge, this is the only probe based real-time PCR assay that enable high specificity and sensitivity for rapid detection of A. helianthi in infected seeds and plant tissues. The assay may also hold promise for application in effective bio-threat and risk mitigation program by early and accurate detection of the pathogen for effective management.

Keywords

TaqMan® Real-time PCR ITS Alternaria helianthi rDNA Sunflower 

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Copyright information

© Koninklijke Nederlandse Planteziektenkundige Vereniging 2015

Authors and Affiliations

  • R. L. Chavhan
    • 1
  • V. R. Hinge
    • 1
  • M. B. Chinchole
    • 1
  • P. K. Chakrabarty
    • 2
  • V. Y. Patade
    • 3
  • H. B. Patil
    • 1
  1. 1.Vilasrao Deshmukh College of Agricultural BiotechnologyLaturIndia
  2. 2.Indian Council for Agricultural ResearchNew DelhiIndia
  3. 3.Defence Institute of Bio-Energy Research, Defence Research & Development OrganisationNainitalIndia

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