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<p><i>Bipolaris oryzae</i> is a major rice seed-borne pathogen causing significant yield losses globally. Ensuring disease free seeds is the most practical and economic way of managing this pathogen. Conventional methods involve lengthy incubation periods, while PCR-based molecular diagnosis requires specialized equipment and hence, not amenable for on-site testing. To address this, a field-level diagnostic approach utilizing Loop-mediated isothermal amplification (LAMP) was developed. The LAMP primers were designed targeting exon 2 of the Glycosyl Transferase Family 90 protein of <i>B. oryzae</i>. The proposed LAMP assay demonstrated exceptional sensitivity, capable of detecting <i>B. oryzae</i> template at concentrations as low as 10&#xa0;fg within 30&#xa0;min at 60&#xa0;°C. To assess specificity, genomic DNA from 22 different phytopathogens was tested and the assay specifically detected <i>B. oryzae</i> alone. The assay was further validated by using ten weed species collected from <i>B. oryzae</i>-infected rice fields. The presence of <i>B. oryzae</i> in eight weed species was confirmed using the developed LAMP protocol except <i>Brachypodium distachyon</i> and <i>Phalaris minor</i>. Also, the assay was successfully employed to detect <i>B. oryzae</i> in the seeds of thirteen rice varieties. The novel LAMP assay enables rapid detection of <i>B. oryzae</i> in rice seeds, plants and soil, thereby, making it ideal diagnostic assay for seed health laboratories, plant quarantine stations and on-site detection of the pathogen in seeds and plants.</p>

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Colorimetric on-site detection of Bipolaris oryzae in rice seeds and its ecological monitoring through Loop-mediated Isothermal Amplification assay

  • 期刊论文

  • 发表日期：

    2025年3月19日

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  • Logeshwari Rajendran,
  • Gopalakrishnan Chellappan,
  • Kamalakannan Ayyanar,
  • Ramalingam Jegadeesan,
  • Saraswathi Ramasamy,
  • Naveena Sirivella

摘要

Bipolaris oryzae is a major rice seed-borne pathogen causing significant yield losses globally. Ensuring disease free seeds is the most practical and economic way of managing this pathogen. Conventional methods involve lengthy incubation periods, while PCR-based molecular diagnosis requires specialized equipment and hence, not amenable for on-site testing. To address this, a field-level diagnostic approach utilizing Loop-mediated isothermal amplification (LAMP) was developed. The LAMP primers were designed targeting exon 2 of the Glycosyl Transferase Family 90 protein of B. oryzae. The proposed LAMP assay demonstrated exceptional sensitivity, capable of detecting B. oryzae template at concentrations as low as 10 fg within 30 min at 60 °C. To assess specificity, genomic DNA from 22 different phytopathogens was tested and the assay specifically detected B. oryzae alone. The assay was further validated by using ten weed species collected from B. oryzae-infected rice fields. The presence of B. oryzae in eight weed species was confirmed using the developed LAMP protocol except Brachypodium distachyon and Phalaris minor. Also, the assay was successfully employed to detect B. oryzae in the seeds of thirteen rice varieties. The novel LAMP assay enables rapid detection of B. oryzae in rice seeds, plants and soil, thereby, making it ideal diagnostic assay for seed health laboratories, plant quarantine stations and on-site detection of the pathogen in seeds and plants.