Optical Imaging and Molecular Sequencing of a Preserved Collection of Bivalve Larvae

Abstract

From the 1980s through 1995, scientists at numerous marine, coastal, estuarine, and freshwater laboratories spawned bivalves to provide larvae for use in identifying species based on larval hinge structures and gross shell morphometry. These larvae were preserved in 95% ethanol and stored in sample vials, many of which dried out over the years. Advantage was taken of 50 of 56 species from this collection (and two additional species that were not in the collection for a total of 52 species) to explore the use of optical techniques (polarized light and a full-wave compensation plate) to highlight birefringence patterns of larval shells to discriminate individual species. Representative images of various developmental stages of 77% (40/52) of the larval bivalve species in the collection were successfully imaged. Similarities across birefringence patterns were observed at the taxonomic ordinal and familial level. Molecular polymerase chain reaction techniques were used in an effort to sequence many of the dried-out specimens and they successfully identified 19% (10/52) of the larval bivalve species with matches in GenBank. Here it has been demonstrated that optical techniques are efficient for imaging dried-out larval bivalve shells for classification purposes and we present successful sequences of 10 species of bivalve larvae from the preserved collection.