PCR amplification of a middle repetitive element detects larval stone crabs (Crustacea: Decapoda: Menippidae) in estuarine plankton samples

Abstract

Planktonic larval dispersal and recruitment can be major determinants of the structure and dynamics of marine communities. However, these processes have been difficult to study because of their natural variability and the limitations of methods used to collect and analyze plankton samples. In particular, the use of microscopy to determine the composition of plankton samples is time-consuming and often limited by a lack of reliable morphological characters for species identification. The need for methods of greater accuracy and efficiency has led to the development of molecular approaches to plankton analysis, including detection by DNA hybridization, amplification of DNA from plankton samples by the polymerase chain reaction (PCR) and taxonomic characterization by ribosomal DNA sequence analysis. Here we describe a PCR-based method that detects larval crabs in estuarine plankton samples. This technique is unusually expedient and relatively cost-effective. It is based on the detection of a middle repetitive sequence characteristic of the stone crab Menippe mercenaria, as well as the closely related species M. adina. Amplification by PCR of a 585 base pair region of this sequence from plankton samples accurately indicates the presence of either species. Because of the high abundance of this sequence in the genome of Menippe, single larvae can be detected in typical plankton samples. Unlike methods based on 'universal' sequences (rRNA or regions of the mitochondrial genome), the amplification of a PCR product of the expected size is a reliable indication of the presence of the target species, and no further characterization is necessary. This technique is intended to facilitate the large-scale processing of plankton samples that is necessary for accurate determination of the temporal and spatial distributions of individual species in plankton communities.