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Development of a multiplex nested PCR method for detection of Xanthomonas axonopodis pv. manihotis in Cassava

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Abstract

Cassava (Manihot esculenta), an important multipurpose crop, represents a food source for millions of people and a source of industrial raw material in tropical regions. A limitation in its production is cassava bacterial blight (CBB), a disease caused by Xanthomonas axonopodis pv. manihotis (Xam). CBB is spread mainly by propagation of infected cuttings, therefore early pathogen detection and disease diagnosis are crucial to prevent its dispersal and the consequent negative impact on the crop. A previously reported detection method was inefficient at detecting isolates of Xam more recently collected in cassava fields in Colombia. Consequently, a new method for pathogen detection was developed using multiplex nested PCR. A set of primers was selected amongst a group designed for five housekeeping gene sequences (rpoB, gltA, ftsZ, gyrB and groEL), and for the region of the gene encoding for the C-terminal portion of TALE1Xam, a protein involved in virulence of Xam. In this work, a multiplex nested PCR based on the set of primers RpoB and Cterm was optimized to achieve an effective detection of Xam. The developed method successfully detects the presence of Xam in cassava plants collected in infected fields.

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Acknowledgements

We are thankful to Luis Miguel Rodriguez for support with databases, to Dr. Valerie Verdier (IRD, Montpellier, France) for providing reference strains and to Dr. Brian Staskawicz (University of California, Berkeley, USA) for providing the strains from Nigeria. This work was supported by the Ministerio de Agricultura de Colombia.

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Authors and Affiliations

  1. ManihotBiotec, Universidad Nacional de Colombia, Cr. 45 No, 26-85, Bogotá, Colombia

    Vivian Bernal-Galeano, Juan C. Ochoa & Camilo A. López

  2. Laboratorio de Micología y Fitopatología, Universidad de los Andes, Cr.1 No. 18-12, 111711, Bogotá, Colombia

    Vivian Bernal-Galeano, César Trujillo, Leidy Rache & Adriana Bernal

  3. Laboratorio de Interacciones Moleculares de Microorganismos Agrícolas, Universidad de los Andes, Cr.1 No.18-12, 111711, Bogotá, Colombia

    Leidy Rache & Adriana Bernal

Authors
  1. Vivian Bernal-Galeano
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  2. Juan C. Ochoa
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  3. César Trujillo
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  4. Leidy Rache
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  5. Adriana Bernal
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  6. Camilo A. López
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Correspondence to Camilo A. López.

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Section Editor: Alessandra A. de Souza

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Bernal-Galeano, V., Ochoa, J.C., Trujillo, C. et al. Development of a multiplex nested PCR method for detection of Xanthomonas axonopodis pv. manihotis in Cassava. Trop. plant pathol. 43, 341–350 (2018). https://doi.org/10.1007/s40858-018-0214-4

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  • DOI: https://doi.org/10.1007/s40858-018-0214-4

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