An immunological study of predation on hatchery-reared, juvenile red drum ( Sciaenops ocellatus, Linnaeus): description of an ELISA and predator-prey studies in nature

Abstract

This report is a continuation of an on-going study to develop immunological methods for eventual use in determining the predation mortality of newly released, hatchery-reared red drum ( Sciaenops ocellatus, Linnaeus). Using a specific goat antiserum produced to a purified 80 kDa red drum glycoprotein, we detected the glycoprotein routinely in soluble extracts of red drum by Western blots. To supplement the immunoblotting, we proceeded to develop a highly sensitive and specific ELISA (enzyme-linked immunosorbent assay). The major problem in developing the ELISA was defining conditions to eliminate a natural inhibitor in soluble extracts of red drum that prevented the 80 kDa protein from binding to microtiter plates. The technical difficulties for a successful ELISA were resolved by adjusting extracts to pH 4.7 and 0.3 M NaCl, based on conditions developed for purification of the 80 kDa protein on a cationic-exchange gel by fast protein liquid chromatography (FPLC). The defined parameters eliminated the inhibition and resulted in optimal binding of the glycoprotein to the polymer surface of plates for ELISA. Approximately 10 h after the release of tens of thousands of red drum fingerlings at two sites in Biscayne Bay, FL, USA, a center-bag haul seine was used to sample the fish and capture predators. Two species, Sphyraena barracuda (Walbaum), great barracuda, and Strongylura notata (Poey), redfin needlefish, were the major predators. ELISA and Western blots were used to identify visually difficult or unidentifiable Sciaenops ocellatus in gut contents of the predators. Based on nine samples from seven Strongylura notata, and 10 samples from eight Sphyraena barracuda, 100% of the samples were identified as red drum in the needlefish and 50% in the great barracuda. These studies confirm the feasibility of using immunological methods to identify otherwise unidentifiable prey in gut contents of predators in nature.