High-Resolution Melting of 18S rDNA sequences (18S-HRM) for discrimination of bivalve's species at early juvenile stage: application to a spat survey

摘要

It is particularly difficult to identify bivalve species at early development stages (larvae and spat) by morphological examination. Targeting genetic marker commonly used for species identification, we developed a cheap and user friendly tool for bivalve identification through High-Resolution Melting (HRM) analysis. Based on the polymorphism of the nuclear 18S ribosomal RNA gene among 11 orders of bivalve, the 18S-HRM method allowed discriminating species of the Ostreida, Mytilida and Pectinida orders. In this study, this method proved to be a very sensitive tool able to authenticate one species that differed by only one mutation from another one. We applied the method by identifying bivalve species in spat collections from different benthic substrates moored at subtidal and intertidal areas in euhaline conditions. This field survey showed that the settlement of the invasive species Crassostrea gigas was limited vertically to intertidal substrates. In conclusion, the 18S-HRM analysis is a fast, closed-tube method with high resolution and cost effectiveness, and is thus a good alternative method to identify bivalve species. This HRM assay can by applied for species identification of bivalves in the context of ecological management as well as food traceability for bivalves with economical interests.