Development of Genus-Specific PCR Primers for Molecular Monitoring of Invasive Nostocalean Cyanobacteria.

Abstract

The geographical range of invasive cyanobacteria with high toxigenic potential is widening because of eutrophication and global warming, thus, monitoring their appearance is necessary for safe water quality control. Most invasive cyanobacteria are nostocalean species, and their accurate identification by classical morphological methods may be problematic. In this study, we developed polymerase chain reaction (PCR) primers to selectively identify five invasive cyanobacterial genera, namely, Chrysosporum, Cuspidothrix, Cylindrospermopsis, Raphidiopsis, and Sphaerospermopsis, using genetic markers such as rbcLX, rpoB, rpoC1, and cpcBA, and determined the amplification conditions for each pair of primers. The primer performances were verified on single or mixed nostocalean cyanobacterial isolates. The five primers allowed selective identification of all the target genera. In field samples collected during summer, when cyanobacteria flourished in the Nakdong River, the respective PCR product was observed in all samples where the target genus was detected by microscopic analysis. Besides, weak bands corresponding to Sphaerospermopsis and Raphidiopsis were observed in some samples in which these genera were not detected by microscopy, suggesting that the cell densities were below the detection limit of the microscopic method used. Thus, the genus-specific primers developed in this study enable molecular monitoring to supplement the current microscopy-based monitoring.