The ALEX CHIP—Development of a DNA chip for identification and monitoring of Alexandrium

Harmful Algae Blooms (HABs) threaten humans, ecosystems, fishery, tourism, and aquaculture, and the occurrence of single cells in mixed phytoplankton assemblages is often difficult to detect. The genus Alexandrium has undergone steady taxonomic revision since its first description, and identification of its species has been confused because of overlapping morphological features and minute differences. The design of molecular probes from the 18S to 28S rDNA has shown great potential for distinguishing of species or even clades, but using these probes in a whole-cell hybridization format is tedious and time-consuming. Solid-phase methods, such as DNA microarrays, offer the potential to analyze multiple targets in a single experiment. This study describes the development of a DNA microarray for detection of several species belonging to the genus Alexandrium. Nine probes from other hybridization methods (fluorescence- in-situ-hybridization [FISH] and sandwich hybridization assay [SHA]) were tested on the microarray, and one new probe was developed for Alexandrium minutum. The specificity of the probes was tested by hybridization with 18S and 28S PCR-fragments from pure cultures and by analysis of filtered and spiked seawater samples from the Weser estuary (German Bight). Some published SHA and FISH probes did not work in a microarray format. A hybridization protocol was established, and the subset of the best performing probes for each species or clade was determined and recommended for classification and monitoring of field samples in the high throughput microarray format.