Effects of euthanasia methods on stable carbon (δ C value) and nitrogen (δ N value) isotopic compositions of fry and juvenile rainbow trout Oncorhynchus mykiss.

Rationale: Carbon and nitrogen stable isotope analyses of fish tissues are now commonly used in ecological studies but mostly require the sacrifice of the animal. Ethical considerations recommend the use of anesthetics for tissue sampling. This study examines how anesthetics affect stable isotope ratios of fish compared with other euthanasia methods.

Methods: Rainbow trout fry and juveniles were sacrificed using ice-freezing (as this common method used to kill fish does not affect natural isotopic ratios), electronarcosis or an overdose of chemical anesthetics (2-phenoxyethanol, benzocaine and clove oil). For fry, we sampled the whole animal whereas, for juveniles, white dorsal muscle, liver, red blood cells, plasma, external tegument and pectoral fin were sampled. Isotopic ratios and the elemental compositions of carbon and nitrogen were then measured.

Results: The δ N values, and the C and N contents of all considered tissues as well as δ C values of muscle, liver, red blood cells and plasma, were not affected by the use of chemical anesthetics. Clove oil and to a lesser extent 2-phenoxyethanol and benzocaine decreased δ C values of whole fry and juvenile external tegument and pectoral fin. The use of electronarcosis drastically affects the δ C and δ N values of all fish tissues.

Conclusions: Anesthetics should be avoided for δ C analysis when tissues are in contact with the water containing the anesthetic. Ice-immersion has to be preferred when approved by guidelines. If not, benzocaine and 2-phenoxyethanol should be preferred over clove oil. Electronarcosis should not be used to kill fish until further investigations are performed.