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Quantitative and specific detection of the biocontrol agent, Serratia plymuthica, in plant extracts using a real-time TaqMan® assay

  • Microbial Genetics • Original Paper
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Abstract

A Serratia plymuthica-specific TaqMan® assay was designed based on the consensus nucleotide sequence from the 3′- end of the luxS gene present in all S. plymuthica strains tested. The specificity of the assay was demonstrated by testing 21 Serratia spp. strains and 30 isolates belonging to various species that can potentially coexist with S. plymuthica in the same environment. Positive reactions in the TaqMan® assay were observed only for S. plymuthica isolates and not for other bacteria. The TaqMan® assay could detect down to 1.95 ng of S. plymuthica DNA, down to 5 bacterial cells per reaction (100 cfu ml−1) in vitro, down to 50 bacterial cells per reaction (1,000 cfu ml−1) in spiked potato root extracts and down to 5 bacterial cells per reaction (100 cfu ml−1) in spiked potato haulm extracts. We used this assay to quantify S. plymuthica A30 cells in potato and tomato haulms and roots grown from S. plymuthica A30-inoculated potato seed tubers and tomato seeds. The results were comparable with the spread-plating of plant extracts on a newly developed S. plymuthica A30 selective medium (CVTR2Arif). The TaqMan® assay can be used to quantify S. plymuthica isolates in different ecosystems and in complex substrates.

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Acknowledgements

The project was financed by STW Foundation (Technologiestichting STW, The Netherlands) via grant no. 10306 “Curing seed potatoes from blackleg causing bacteria”. We would like to thank Joeke Postma (Plant Research International, Wageningen University and Research Centre, The Netherlands), Paolina Garbeva (Netherlands Institute of Ecology, NIOO-KNAW, The Netherlands) and Sylwia Jafra (Intercollegiate Faculty of Biotechnology, University of Gdansk and Medical University of Gdansk, Poland) for providing us with the Serratia spp. strains and helpful discussions. Thanks are indebted to M. C. M. Perombelon (ex. SCRI, UK) for his valuable comments on this manuscript and his editorial work.

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  1. Plant Research International, P.O. Box 69, 6700 AB, Wageningen, The Netherlands

    Robert Czajkowski & Jan M. van der Wolf

  2. Netherlands Institute of Ecology (NIOO-KNAW), Droevendaalsesteeg 10, 6708 PB, Wageningen, The Netherlands

    Robert Czajkowski

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  1. Robert Czajkowski
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Correspondence to Jan M. van der Wolf.

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Czajkowski, R., van der Wolf, J.M. Quantitative and specific detection of the biocontrol agent, Serratia plymuthica, in plant extracts using a real-time TaqMan® assay. J Appl Genetics 53, 457–467 (2012). https://doi.org/10.1007/s13353-012-0106-0

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