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Loop-mediated isothermal amplification assay for pre-symptomatic stage detection of Xanthomonas axonopodis pv. punicae infection in pomegranate

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A Correction to this article was published on 06 June 2020

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Abstract

Bacterial blight in pomegranate caused by Xanthomonas axonopodis pv. punicae (Xap) is an increasing threat to pomegranate cultivation in India. To prevent economic losses, it is pivotal to detect the infection in latent stages rather than in symptomatic stages. We have developed an enhanced loop-mediated isothermal amplification (LAMP) technique for the detection of latent Xap infection in pomegranate using six sets of pathovar-specific primers. The DNA-intercalating dye ethidium bromide was standardized for visualizing positive amplification in the LAMP assay. The optimum reaction time was 30 min or more at 65 °C, and assay sensitivity was enhanced down to femtogram (fg) amounts of template DNA (0.153 fg) in the LAMP assay. In artificially inoculated plants, the enhanced LAMP assay detected Xap on the 7th day post inoculation (dpi), while PCR detected Xap only after the 11th dpi. Finally, the specificity of the LAMP assay for Xap was validated against 31 bacterial strains belonging to 22 species. The LAMP assay developed in this study is highly specific, sensitive and robust and can be used as an improved alternative for PCR-based methods for the early detection of Xap infection in pomegranate. Early detection of Xap in pomegranate ensures quality fruits with increased yield by adopting precautionary measures that ensure improved monetary benefit to farmers.

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Acknowledgements

Our team would like to thank A. Satish (Dept. of Soil Science, UASB) for assisting in GIS mapping construction. We are grateful to Basavaraj (Dept. of Plant Pathology, UASB), Shankarappa (Dept. of Horticulture, Bagalkot), and Vinay Kumar (Dept. of Soil Science, UASB) for their technical assistance. The authors are highly thankful to Dr. Rashmi Aggarwal, Head and Principal Scientist, Dr. K. K. Mondal Principal Scientist, Division of Plant Pathology, ICAR-IARI New Delhi, India, for providing a bacterial culture (Xanthomonas campestris pv. campestris) and Edwin Raj (Plant Physiology and Biotechnology, UPASI Tea Research Institute) for his support in statistical analysis.

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Authors and Affiliations

  1. Department of Plant Pathology, University of Agricultural Sciences, Bangalore, India

    M. K. Prasannakumar, P. Buela Parivallal, K. Priyanka, M. E. Puneeth & K. T Rangaswamy

  2. ICAR-Indian Agricultural Research Institute, Regional Station, Wellington, The Nilgiris, India

    C. Manjunatha

  3. Department of Genetics and Plant Breeding, College of Agriculture, V. C. Farm, Mandya, India

    H. B. Mahesh

  4. Rice Pathology Laboratory, All India Coordinated Rice Improvement Programme, Gangavathi, India

    D. Pramesh

  5. Centre for Advanced Studies in Botany, University of Madras, Guindy Campus, Chennai, India

    Karthik S. Narayan & Venkatesh Babu Gopal

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  1. M. K. Prasannakumar
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  2. P. Buela Parivallal
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  3. C. Manjunatha
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  9. M. E. Puneeth
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  10. K. T Rangaswamy
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Correspondence to M. K. Prasannakumar.

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Prasannakumar, M.K., Buela Parivallal, P., Manjunatha, C. et al. Loop-mediated isothermal amplification assay for pre-symptomatic stage detection of Xanthomonas axonopodis pv. punicae infection in pomegranate. Australasian Plant Pathol. 49, 467–473 (2020). https://doi.org/10.1007/s13313-020-00720-w

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