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Development of a recombinant Arxula adeninivorans cell bioassay for the detection of molecules with progesterone activity in wastewater

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Abstract

This study describes the development of a bioassay to detect the presence of progesterone and progesterone-like molecules in wastewater samples. The basis of the bioassay is the integration of the human progesterone receptor gene into the yeast Arxula adeninivorans for the constitutive synthesis of the receptor. After incubation, binding of the analyte to the receptor induces the production of a reporter protein. Two reporter proteins were compared for detection parameters such as half-maximal activity (EC50), limit of detection (LoD) and limit of quantification (LoQ). When the extracellular phytase K was used, an EC50 value of 155 ng L−1 and a LoD of 27 ng L−1 progesterone were obtained after 4 h incubation, while use of the fluorescent dsRED as the reporter protein, resulted in an EC50 of 320 ng L−1 and a LoD of 65 ng L−1 after 20 h incubation. Use of phytase K as the reporter protein offers decreased incubation time and increased sensitivity; however the dsRED reporter system is less labor-intensive. Additionally, the affinity of known agonists and antagonists of the human progesterone receptor was determined. The utility of this bioassay was confirmed by measuring total progesterone equivalent concentration of samples from a wastewater treatment plant. The A. adeninivorans-based transactivation assay was able to measure concentrations of about 311 ng L−1 in the influent stream but could not detect progesterone activity in effluent. One key feature of the assay is the robustness of A. adeninivorans, which allows sample measurement without any sample preparation.

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Acknowledgments

We would like to thank Ruth Franz for her technical support. The research work was supported by grant from the BMWi (Grant No. KF2131610MD0 and KF2131626SB4).

Conflict of interest

The authors declare no conflict of interest.

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Authors and Affiliations

  1. Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Corrensstr. 3, 06466, Gatersleben, Germany

    Alexandre Chamas, Annabel Nieter, Ha Thi Minh Pham, Martin Giersberg & Gotthard Kunze

  2. Institute of Biotechnology, Vietnam Academy of Science and Technology, 18 Hoang Quoc Viet, Cau Giay, Ha Noi, Vietnam

    Ha Thi Minh Pham

  3. QuoData GmbH, Prellerstr. 14, 01309, Dresden, Germany

    Karina Hettwer & Steffen Uhlig

  4. New diagnostics GmbH, Moosstr. 92c, 85356, Freising, Germany

    Kirsten Simon

  5. School of Biological Sciences, University of Canterbury, Private Bag 4800, Christchurch, 8041, New Zealand

    Keith Baronian

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  1. Alexandre Chamas
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  2. Annabel Nieter
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  9. Gotthard Kunze
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Correspondence to Gotthard Kunze.

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Chamas, A., Nieter, A., Pham, H.T.M. et al. Development of a recombinant Arxula adeninivorans cell bioassay for the detection of molecules with progesterone activity in wastewater. Anal Bioanal Chem 407, 8109–8120 (2015). https://doi.org/10.1007/s00216-015-8985-7

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