title: Rapid and accurate identification by real-time PCR of biotoxin-producing dinoflagellates from the family gymnodiniaceae.
abstract: The identification of toxin-producing dinoflagellates for monitoring programmes and bio-compound discovery requires considerable taxonomic expertise. It can also be difficult to morphologically differentiate toxic and non-toxic species or strains. Various molecular methods have been used for dinoflagellate identification and detection, and this study describes the development of eight real-time polymerase chain reaction (PCR) assays targeting the large subunit ribosomal RNA (LSU rRNA) gene of species from the genera Gymnodinium, Karenia, Karlodinium, and Takayama. Assays proved to be highly specific and sensitive, and the assay for G. catenatum was further developed for quantification in response to a bloom in Manukau Harbour, New Zealand. The assay estimated cell densities from environmental samples as low as 0.07 cells per PCR reaction, which equated to three cells per litre. This assay not only enabled conclusive species identification but also detected the presence of cells below the limit of detection for light microscopy. This study demonstrates the usefulness of real-time PCR as a sensitive and rapid molecular technique for the detection and quantification of micro-algae from environmental samples.
authors.affiliation: Cawthron Institute, 98 Halifax Street East, Private Bag 2, Nelson 7042, New Zealand. kirsty.smith@cawthron.org.nz.
authors.forename: Kirsty F
authors.initials: KF
authors.lastname: Smith
authors.affiliation: Tasmanian Herbarium, Tasmanian Museum and Art Gallery, Private Bag 4, Hobart, Tasmania 7001, Australia. dr.microbe@gmail.com.
authors.forename: Miguel
authors.initials: M
authors.lastname: de Salas
authors.affiliation: Cawthron Institute, 98 Halifax Street East, Private Bag 2, Nelson 7042, New Zealand. janet.adamson@cawthron.org.nz.
authors.forename: Janet
authors.initials: J
authors.lastname: Adamson
authors.affiliation: Cawthron Institute, 98 Halifax Street East, Private Bag 2, Nelson 7042, New Zealand. lesley.rhodes@cawthron.org.nz.
authors.forename: Lesley L
authors.initials: LL
authors.lastname: Rhodes
mesh_terms: D000818:Animals; D004247:DNA / Q000302:isolation & purification; D017931:DNA Primers; D004261:DNA Replication / Q000187:drug effects; D004141:Dinoflagellida / Q000235:genetics / Q000378:metabolism*; D004777:Environment; D004784:Environmental Monitoring; D018628:Gene Dosage; D057230:Limit of Detection; D008387:Marine Toxins / Q000096:biosynthesis*; D009520:New Zealand; D012335:RNA, Ribosomal / Q000096:biosynthesis / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D012623:Seawater; D013045:Species Specificity
chemical_list: D017931:DNA Primers; D008387:Marine Toxins; D012335:RNA, Ribosomal; D004247:DNA