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Strengths and weaknesses of microarray approaches to detect Pseudo-nitzschia species in the field

  • Molecular tools for monitoring Harmful Algal Blooms
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Abstract

The planktonic diatom genus Pseudo-nitzschia contains several genetically closely related species. Some of these can produce domoic acid, a potent neurotoxin. Thus, monitoring programs are needed to screen for the presence of these toxic species. Unfortunately, many are impossible to distinguish using light microscopy. Therefore, we assessed the applicability of microarray technology for detection of toxic and non-toxic Pseudo-nitzschia species in the Gulf of Naples (Mediterranean Sea). Here, 11 species have been detected, of which at least 5 are potentially toxic. A total of 49 genus- and species-specific DNA probes were designed in silico against the nuclear LSU and SSU rRNA of 19 species, and spotted on the microarray. The microarray was tested against total RNA of monoclonal cultures of eight species. Only three of the probes designed to be species-specific were indeed so within the limits of our experimental design. To assess the effectiveness of the microarray in detecting Pseudo-nitzschia species in environmental samples, we hybridized total RNA extracted from 11 seasonal plankton samples against microarray slides and compared the observed pattern with plankton counts in light microscopy and with expected hybridization patterns obtained with monoclonal cultures of the observed species. Presence of species in field samples generally resulted in signal patterns on the microarray as observed with RNA extracted from cultures of these species, but many a-specific signals appeared as well. Possible reasons for the numerous cross reactions are discussed. Calibration curves for Pseudo-nitzschia multistriata showed linear relationship between signal strength and cell number.

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Acknowledgments

The authors thank Francisco Rodriguez Hernandez, Jixin Chen and Sylvie Tesson who provided some of Pseudo-nitzschia strains used in this study and the EU-FP7 project MIDTAL for funding. We also thank Pasquale de Luca for technical advice and Linda K. Medlin (coordinator of the MIDTAL project) and Adriana Zingone for scientific discussion.

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  1. Laboratory of Ecology and Evolution of Plankton, Stazione Zoologica Anton Dohrn, Villa Comunale I, 80121, Naples, Italy

    Lucia Barra, Maria Valeria Ruggiero, Diana Sarno, Marina Montresor & Wiebe H. C. F. Kooistra

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  1. Lucia Barra
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  2. Maria Valeria Ruggiero
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  3. Diana Sarno
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  4. Marina Montresor
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  5. Wiebe H. C. F. Kooistra
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Correspondence to Maria Valeria Ruggiero.

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Responsible editor: Robert Duran

Authors Barra Lucia and Ruggiero Maria Valeria contributed equally to the paper.

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Barra, L., Ruggiero, M.V., Sarno, D. et al. Strengths and weaknesses of microarray approaches to detect Pseudo-nitzschia species in the field. Environ Sci Pollut Res 20, 6705–6718 (2013). https://doi.org/10.1007/s11356-012-1330-1

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