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Improved PCR for identification of members of the genus Xanthomonas

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Abstract

A PCR-based system was developed to reliably and robustly identify group I and II members of the genus Xanthomonas. Primer sets developed from three gene targets namely fyuA, ITS and gumD were evaluated in the study. Primer sets were evaluated using DNA extracted from 45 Xanthomonas strains representing 25 species broadly covering the genus. Fifteen non-Xanthomonas strains of plant-associated bacteria including phylogenetically closely related species Stenotrophomonas maltophilia and Xylella fastidiosa were also tested. The primers targeting fyuA amplified DNA from all xanthomonads except X. theicola, while the ITS primers amplified a DNA fragment of 254 bp in all 45 Xanthomonas strains; whereas no amplification was observed for non-xanthomonads. The gumD primers allowed efficient amplification of DNA in 38 out of 39 isolates from Group II, whereas no or very weak amplification occurred with DNA from Group I members. Internal controls of primers targeting bacterial 16S rDNA or plant 26S mitochondrial rDNA were successfully applied in multiplex PCRs for testing bacterial cultures or plant tissue, respectively. The findings give us a PCR based approach that can reliably and effectively differentiate xanthomonads from non-xanthomonads as well as separating the strains belonging to the two described groups of the genus Xanthomonas. The study thus offers valuable tools for disease surveillance and management. It can effectively be applied in rapid assessment of new disease occurrences, for which no specific detection tools could be in place.

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Acknowledgments

This work was made possible through funding from DANIDA under the ENRECA project LIFE—731. The contribution of Hanne, B. Nielsen of Danish Seed Health Centre is acknowledged and highly appreciated.

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Authors and Affiliations

  1. Danish Seed Health Centre for Developing Countries, Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen, Hoejbakkegaard Allé 3, 2630, Taastrup, Denmark

    John Adriko, Ernest Rashid Mbega, Carmen Nieves Mortensen, Ednar Gadelha Wulff & Ole Søgaard Lund

  2. National Agricultural Biotechnology Centre, National Agricultural Research Laboratories, P. O. Box 7065, Kampala, Uganda

    John Adriko

  3. National Banana Research Programme, National Agricultural Research Laboratories, P. O. Box 7065, Kampala, Uganda

    John Adriko, Wilberforce Kateera Tushemereirwe & Jerome Kubiriba

  4. African Seed Health Centre (AfSHC), Department of Crop Science, Sokoine University of Agriculture, P.O. Box 3005, Morogoro, Tanzania

    Ernest Rashid Mbega

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  1. John Adriko
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  2. Ernest Rashid Mbega
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Correspondence to John Adriko.

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Adriko, J., Mbega, E.R., Mortensen, C.N. et al. Improved PCR for identification of members of the genus Xanthomonas . Eur J Plant Pathol 138, 293–306 (2014). https://doi.org/10.1007/s10658-013-0329-x

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