Effects of water hardness on skeletal development and growth in juvenile fathead minnows

Skeletal growth in vertebrates is partially controlled by the availability of minerals. The correlation between aqueous calcium content and fish skeletal growth (“ossification”) is especially important as calcium absorption from water is an essential route to acquire this mineral. In this study, we test the hypothesis that a calcium-abundant aquatic environment will expedite larval skeletal development in fathead minnows beyond the skeletal development in a low-calcium environment. Fathead minnows were chosen for this study as they are a widely used toxicological model organism with well established rearing protocols. We divided three batches each of fathead minnow eggs from a colony supplied with calcium-abundant water (Ca = 65 ± 1.5 mg/L as CaCO3; hardness > 175 mg/L) and one with a low, yet adequate calcium content (Ca = 15 ± 0.11 mg/L as CaCO3; hardness < 50 mg/L) to rear fish. Half of each egg batch was exchanged between laboratories and fish were then reared to 60 days post-hatch under similar conditions. At 60 days, juveniles were sacrificed, cleared and stained. Ossification patterns for cranial (jaw) and caudal (fin) structures were quantified using NIH Image (http://rsb.info.nih.gov/nih-image/). Fish reared in calcium-abundant water had significantly increased survival but lower whole body mass when compared to their conspecifics in low-calcium water. Cranial ossification patterns did not vary. Caudal fin ossification was statistically significant, but slightly and surprisingly enhanced in juveniles reared in low-calcium water. All observed differences were independent of the origin of fish and imply differences in fish growth among cultures maintained under similar rearing conditions for the purpose of this study. Results of this investigation have implications for the rearing of model organisms for toxicological studies and need to be amended by physiological explorations.