A DNA methylation age predictor for zebrafish.

Changes in DNA methylation at specific CpG sites have been used to build predictive models to estimate animal age, predominantly in mammals. Little testing for this effect has been conducted in other vertebrate groups, such as bony fish, the largest vertebrate class. The development of most age-predictive models has relied on a genome-wide sequencing method to obtain a DNA methylation level, which makes it costly to deploy as an assay to estimate age in many samples. Here, we have generated a reduced representation bisulfite sequencing data set of caudal fin tissue from a model fish species, zebrafish (Danio rerio), aged from 11.9-60.1 weeks. We identified changes in methylation at specific CpG sites that correlated strongly with increasing age. Using an optimised unique set of 26 CpG sites we developed a multiplex PCR assay that predicts age with an average median absolute error rate of 3.2 weeks in zebrafish between 10.9-78.1 weeks of age. We also demonstrate the use of multiplex PCR as an efficient quantitative approach to measure DNA methylation for the use of age estimation. This study highlights the potential further use of DNA methylation as an age estimation method in non-mammalian vertebrate species.

A Comparison of Buffered Tricaine Methanesulfonate (MS-222) and Isoeugenol Anesthesia for Caudal Fin Clipping in Zebrafish (Danio rerio).

The purpose of this study was to compare the safety and efficacy of buffered tricaine methanesulfonate (MS-222) and isoeugenol for the anesthesia of zebrafish undergoing caudal fin clipping. Eighty 9 mo Danio rerio (AB strain) zebrafish were allocated to one of 2 equal groups: buffered MS-222 (168 mg/L, n = 40) or isoeugenol (20 mg/L, n = 40). The time to induction of anesthesia was significantly shorter in the isoeugenol group (141 ± 70 s) than in the MS-222 group (207 ± 103 s). The time to recovery from anesthesia was also shorter in the MS-222 group (373 ± 125 s) than in the isoeugenol group (491 ± 176 s). No obvious displays of distress or aversion to anesthesia were observed in either group. No difference was detected in the proportion of zebrafish that became anesthetized with either drug. One male zebrafish in the buffered MS-222 group was found dead at the 1-h post-procedural monitoring time point, but there was no difference between groups in the proportion of fish that survived anesthesia to the end of experiment. In conclusion, the safety and efficacy of buffered MS-222 (168 mg/L) and isoeugenol (20 mg/L) was similar for zebrafish undergoing anesthesia for caudal fin clipping.