Histological findings, cadmium bioaccumulation, and isolation of expressed sequence tags (ESTs) in cadmium-exposed, specific pathogen-free shrimp, Litopenaeus vannamei postlarvae

摘要

Cadmium (Cd) is an ubiquitous environmental pollutant of increasing worldwide concern. It has become one of the most hazardous heavy metals in aquatic environments and could threaten aquatic organisms, including marine shrimp. Shrimp are sensitive to Cd and have been found to accumulate it in their bodies in proportion to environmental concentrations. The effects of Cd on the biology and gene expression of the commercially important Litopenaeus vannamei are unknown. The overall hypothesis is that Cd exerts effects on shrimp at both biological and molecular levels. These changes may provide a way to identify genes responsible for toxicity, detoxification and/or tolerance to Cd exposure both acute and chronic. To test the hypothesis, a small-scale pilot project was initiated to obtain baseline information on histological changes associated with Cd treatment and to develop the genomics tools needed to identify genes associated with Cd exposure. The specific objectives of this study were to (1) observe histopathologic changes in control- and Cd-treated postlarvae (PL) during a 48-h period, (2) measure Cd concentrations in Cd-exposed and untreated PLs, and (3) isolate expressed sequence tags (ESTs) from control and Cd-treated PLs for future genomics studies. Specific pathogen-free L. vannamei PLs at stage 42 (PL42, from the Kona Line of the United Slates Marine Shrimp Farming Program) were used in a waterborne bioassay to determine histological changes in PLs exposed to a range of concentrations of CdCl2 (0, 0.01, 0.1, 1.0, and 10.0 ppm) for 48 h. Results for objective (1) indicated variable response of individual shrimp to Cd exposure at different concentrations. All but 2 animals from 10-ppm group died by 24 h. Histological lesions were limited to the integument, musculature, gills, hepatopancreas, and midgut-hindgut. For objective (2), results showed that the Cd levels in control PLs at 0 h were low and remained relatively low throughout the study. There was a dose- and time-dependent relationship of waterborne Cd exposure and accumulation. Results from objective (3) suggested differential gene expression in control- and Cd-treated PLs as reflected by the number of ESTs homologous to genes with different molecular function isolated from approximately 1,100 clones each from the control and Cd-exposed (1 ppm) cDNA libraries. These ESTs contributed to the establishment of an EST database for L. vannamei (ShrimpESTBase). Homology searches of the nucleotide and translated amino acid sequences of ESTs isolated from the control and Cd-treated (I ppm) cDNA libraries identified a significant number of clones similar to (a) known housekeeping genes and genes involved in immune recognition, signal transduction and effector function, (b) other shrimp ESTs of unknown function, (c) ESTs from other species, (d) predicted, unknown or unnamed proteins from other species, and (e) no homology to any sequence in the GenBank database. Some ESTs (similar to 30%) from the Cd-treated library showed homology to unique sequences representing potential transposable elements. The results provide baseline information on the potential effects of Cd on shrimp health and growth and suggest a complex interaction between environmental conditions, water, feed, stress, and genetic background of PLs and should be further investigated under both laboratory and commercial conditions. Moreover, data suggest L. vannamei may be useful as bioindicators for the condition of their natural environment.