Sandwich Hybridization Assay (SHA) As a Novel Genetic Approach for Rapid Identification and Quantification of Red Drum (Sciaenops ocellatus) Eggs

There has been a recent shift from microscopy to molecular approaches for identifying and quantifying marine plankton, driven by a need to expedite sample processing for research and management purposes while increasing detection sensitivity and accuracy.  One such method is sandwich hybridization assay (SHA) which uses DNA probes to directly detect unpurified and unamplified large sub-unit (LSU) ribosomal RNA (rRNA) of a target organism.  The resulting colorimetric response can be used to infer organism abundance.  SHA applications have thusfar focused on phytoplankton and zooplankton.  Here, we present a novel application of SHA for detecting finfish eggs using the important recreational gamefish red drum, Sciaenops ocellatus, as our test species.  This novel molecular advancement has direct management implications for egg production studies that identify spawning locations, study spawning behavior, and calculate indices of abundance.  Initial sequencing of S. ocellatus LSU rRNA and closely-related Sciaenids indicated low genetic divergence, so probe design used the more variable internally transcribed spacer (ITS) region.  Probe validations studies, including standard curves, cross-reactivity tests, and field sampling over an annual cycle in S. ocellatus spawning grounds are planned.  To determine effects of developmental stage, eggs sampled over 24 hours (spawning to hatching) will be evaluated.