Geographic and seasonal comparison of LSU rDNA copy numbers in Alexandrium catenella life cycle stages

Alexandrium catenella is one of many cyst-forming Alexandrium harmful algal bloom species (HABs) that cause paralytic shellfish poisoning. Identifying and quantifying A. catenella cysts from environmental samples using microscopy is time-consuming and challenging due to their shared morphological similarities with other cysts. In this study, we used molecular methods to increase efficiency of quantifying cysts from environmental samples by employing a qPCR assay that targeted the LSU ribosomal gene. To optimize cyst quantification, we made a regional, seasonal, and life cycle stage investigation of A. catenella rDNA copy number content. To our knowledge, this is the first study to make such a comparison. Our experimental design included A. catenella vegetative strains and cysts collected from the Gulf of Maine (GOM), Kodiak Island, Alaska, and Puget Sound, Washington. The results revealed that there were no significant differences regionally in rDNA copy numbers between cysts and vegetative cells, however, sampling period was a significant predictor of cyst rDNA copy number. The validity of using the qPCR assay for making cyst abundance estimates was confirmed by comparing qPCR estimates to microscopic counts in sediment samples collected during NOAA A. catenella cyst surveys in the GOM. The application of qPCR methods instead of manual counting to enumerate cysts is recommended as an efficient, cost effective, and time saving approach for monitoring cyst abundances, particularly for large surveys. The results of this study have implications for molecular quantification of not only A. catenella, but also other HAB species with similar life cycle strategies.