Evaluation of sxtA and rDNA qPCR assays through monitoring of an inshore bloom of Alexandrium catenella Group 1

Abstract

Alexandrium catenella (formerly A. tamarense Group 1, or A. fundyense) is the leading cause of Paralytic Shellfish Poisoning in North and South America, Europe, Africa, Australia and Asia. The quantification of A.catenella via sxtA, a gene involved in Paralytic Shellfish Toxin synthesis, may be a promising approach, but has not been evaluated in situ on blooms of A. catenella, in which cell abundances may vary from not detectable to in the order of 106 cells L−1. In this study, we compared sxtA assay performance to a qPCR assay targeted to a species-specific region of ribosomal DNA (rDNA) and an established fluorescent in situ hybridization (FISH) microscopy method. Passing-Bablok regression analyses revealed the sxtA assay to overestimate abundances when <5 cell equivalents A. catenella DNA were analysed, but otherwise was closer to microscopy estimates than the rDNA assay, which overestimated abundance across the full range of concentrations analysed, indicative of a copy number difference between the bloom population and a culture used for assay calibration a priori. In contrast, the sxtA assay performed more consistently, indicating less copy number variation. The sxtA assay was generally reliable, fast and effective in quantifying A. catenella and was predictive of PST contamination of shellfish.

From the sxtA4 Gene to Saxitoxin Production: What Controls the Variability Among Alexandrium minutum and Alexandrium pacificum Strains?

“…In field surveys, sxtA4 qPCR assays are found to be effective for identifying PST-producing dinoflagellates from mixed samples ( Murray et al, 2011 , 2019 ; Gao et al, 2015 ; Penna et al, 2015 ; Ruvindy et al, 2018 ). Indeed, the presence of sxtA4 seems to be a putative proxy to reveal the presence of toxic cells in blooms since there is a strong correlation between the detection of sxtA4 and the presence of toxins in A. ostenfeldii and A. pacificum ( Murray et al, 2011 ; Savela et al, 2016 ).…”

Dynamics of the Toxic Dinoflagellate Alexandrium pacificum in the Taiwan Strait and Its Linkages to Surrounding Populations

“…Paralytic shellfish toxins (PST) are among the most devastating biotoxins; consumption of PST-contaminated bivalves may cause tingling and numbness, respiratory difficulty, and even death [1]. The dinoflagellate genus Alexandrium includes several PST-producing species, of which Alexandrium catenella Kofoid (previously A. tamarense species complex Group I) and A. pacificum Litaker (previously A. tamarense species complex group IV) are widely distributed.…”

Assessing the Use of Molecular Barcoding and qPCR for Investigating the Ecology of Prorocentrum minimum (Dinophyceae), a Harmful Algal Species

“…The light microscopy counting method utilised in this study had a larger-thanaverage error rate, a high detection threshold of P. minimum (500 cells L −1 compared with 13 cells L −1 with qPCR), and comparatively fewer data points when compared with the molecular methods. Adoption of other light microscope counting methods, like the Utermöhl counting chamber [94], and the use of a DNA-based stain (i.e., a fluorescence in situ hybridisation (FISH) probe [95]) may have led to more accurate assessments of the abundance of P. minimum and the detection of cryptic species. For research into HAB ecology, a combination of the use of amplicon sequencing, to first determine the diversity of phytoplankton, particularly cryptic and small species, and then qPCR, to quantify the exact cell abundance, would appear to give optimal information for ecological inference and understanding of co-occurrence patterns.…”