The ecology and biology of oysters (Ostreidae) across the tropics is poorly understood. Morphological plasticity and shared characteristics among tropical oysters have resulted in the misidentification of species, creating challenges for understanding basic species-specific biological information that is required for restoration and aquaculture. Genetic barcoding has proven essential for accurate species identification and understanding species geographic ranges. To reduce the costs of molecular species identification we developed multiplex PCR assays using the cytochrome c oxidase subunit I (COI or cox1) barcoding gene for the rapid identification of five species of oysters within the genus Saccostrea commonly found in Australia and/or the Indo-Pacific: Saccostrea glomerata , Saccostrea lineage B, Saccostrea lineage F, Saccostrea lineage G, and Saccostrea spathulata (lineage J).
Multiplex assays were successful in species-specific amplification of targeted species. Non-destructive DNA sampling by extracting oyster pallial fluid was also tested on adult oysters collected from the Noosa estuary in Queensland to assess whether oysters were able to be identified non-destructively. DNA concentrations as low as 1 ng/ μL still amplified in most cases, allowing for identification. The development of these primers has enabled the first successful spawning runs of Saccostrea lineage G, a species with high aquaculture potential.
These multiplex assays will be essential tools for species identification in future studies , and we expect that they will be widely applicable after being tested for non-specific amplification of other locally occurring species in other locations. The multiplex assays developed in this study outline easily replicable methods for the development of additional species-specific primer sets for other Saccostrea species, which will be instrumental in unravelling the taxonomic ambiguities within this genus in tropical regions.