HOFFMAN, G.G.*; WILBUR, A.E.: A single-step multiplex PCR identification assay to distinguish megalopae of Callinectes sapidus from Callinectes similis in plankton samples.

Studying recruitment variation in economically important species such as the blue crab, Callinectes sapidus, requires the collection and processing of large numbers of plankton samples and settlement traps. Long time-course collections can meet the sampling needs of such investigations, but accurate and specific species information requires efficient identification of the target species. Traditionally this need has been satisfied by microscopic examination of less than perfectly preserved larval individuals. Larval decapods are particularly difficult to identify to the species level based on morphological criteria alone and this limitation is amplified when congeners such as C. sapidus and C. similis are present in the collections. Here a single-step multiplexed PCR technique is described to efficiently and accurately discriminate the target decapod crustaceans based on the size of mitochrondrial cytochrome oxidase I PCR products. The assay was developed using easily identified adults of the both species from a wide geographic area, tested for specificity, and the PCR products were sequenced to verify the results. Single megalopae are used as a template in PCR reactions to discriminate commercially important blue crabs from the very similar larvae of C.similis. This assay demonstrates the capability of discriminating target megalopae from non-target animals using a single step reaction that can be adapted to process large numbers of collected larvae.