Detection of Alexandrium minutum dinoflagellate in environmental samples using electrochemical genosensor

Dinoflagellates are aquatic microorganisms bearing two dissimilar flagella that inhabit both salt and fresh waters. These microorganisms are mostly harmless, however, under certain conditions, some species rapidly reproduce forming water blooms that not only discolour the waters but also compromise the health of every organism in the vicinity, as some dinoflagellates produce potent toxins deemed unsafe for human health (e.g. Alexandrium minutum).

In this work, a disposable electrochemical genosensor for the detection of the toxic dinoflagellate Alexandrium minutum was developed. The analytical platform methodology consisted in a sandwich format heterogeneous hybridization of complementary DNA sequences assay. The 70 bp A. minutum-specific targeting probe, the 45 bp fluorescein isothiocyanate-labelled signalling DNA probe and the 25 bp thiolated-DNA-capture probe were designed, after analysing public databases. To maximize the complementary DNA hybridization and to avoid the formation of strong secondary structures, a mixed mercaptohexanol (MCH) and self-assembled monolayer (SAM) A. minutum-specific DNA-capture probe was immobilized onto disposable screen-printed gold electrodes (SPGE).

Using chronoamperometric measurements, the enzymatic amplification of the electrochemical signal was obtained with a concentration range from 0.12 to 1.0 nM, a LD of 24.78 pM with a RSD < 5.2 %. This electrochemical genosensor was successfully applied to the selective analysis of the targeted A. minutum specific region of denatured genomic DNA, extracted from toxic dinoflagellates present in the Atlantic Ocean, and human epithelium cells.