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The aim of this study was to develop and validate a high resolution melting (HRM) method for the rapid, accurate identification of the various harmful diatom  Pseudo-nitzschia  species in marine environments.  Pseudo-nitzschia  has a worldwide distribution and some species are toxic, producing the potent domoic acid toxin, which poses a threat to both human and animal health. Hence, it is important to identify toxic  Pseudo-nitzschia  species. A pair of primers targeting the LSU rDNA of the genus  Pseudo-nitzschia  was designed for the development of the assay and its specificity was validated using 22 control DNAs of the  P. calliantha ,  P. delicatissima/P. arenysensis  complex and  P. pungens . The post-PCR HRM assay was applied to numerous unidentified  Pseudo-nitzschia  strains isolated from the northwestern Adriatic Sea (Mediterranean Sea), and it was able to detect and discriminate three distinct  Pseudo-nitzschia  taxa from unidentified samples. Moreover, the species-specific identification of  Pseudo-nitzschia  isolates by the HRM assay was consistent with phylogenetic analyses. The HRM assay was specific, robust and rapid when applied to high numbers of cultured samples in order to taxonomically identify  Pseudo-nitzschia  isolates recovered from environmental samples.

A high resolution melting method for the molecular identification of the potentially toxic diatom Pseudo-nitzschia spp. in the Mediterranean Sea