Repeated non-invasive sampling of zebrafish Danio rerio sperm was conducted, sperm counts were obtained and a method for measurement of DNA damage in sperm was developed and validated (single-cell gel electrophoresis, comet, assay). DNA damage in sperm increased with concentration of hydrogen peroxide (H2O2, 0–200 µM), and in vitro exposure of sperm to 200 µM H2O2 produced 88·7 ± 3·9% tail DNA compared to unexposed controls [12 ± 0·7% tail DNA (mean ± s.e., n = 3)]. Frequency of sperm sampling (sampled every 2, 4 or 7 days) did not affect DNA damage in sperm, but sperm counts decreased 57 and 22% for fish sampled every 2 or 4 days, respectively.
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