Syngnathids are vulnerable ovoviviparous fishes in which males undergo repeated brooding within a reproductive season. The isotopic effects of diet on both breeders (pooled sexes) and parent-egg transmission have been demonstrated in a few fish species but never in syngnathids. Quantifying isotopic changes due to diet is necessary to assess parent-newborn conversions and to estimate accurate trophic enrichment factors (TEF). We assessed the isotopic (δ13C and δ15N) effects of 3 experimental diets on TEFs in seahorse Hippocampus guttulatus breeders and isotopic inheritance. Our results suggest that H. guttulatus follows an income-capital continuum pattern for parent-egg transmission. The isotopic variability in diets for breeders and the resulting experimentally derived TEFs were compared with fixed TEFs from reviews to estimate their impact on the relative contribution of potential prey sources in syngnathids from the Cíes archipelago (Atlantic Islands National Park, NW Spain). We estimated source contributions using stable isotope mixing models (SIMMs) by combining prey sources into ecologically informative groups and incorporating informative priors. We demonstrate that (1) most frequently used TEFs from reviews might not be suitable for all fish species, particularly syngnathids, and (2) dietary source variability has a great effect on source contribution estimates. This study is also the first to provide specific TEFs for syngnathids.

Isotopic stable analysis (SIA) is a powerful tool in the assessment of different types of ecological and physiological studies. For that, different preservation methods for sampled materials are commonly used prior to isotopic analysis. The effects of various preservation methods (freezing, ethanol and formaldehyde) were analyzed for C:N, and δ13C and δ15N signals on a variety of tissues including dorsal fins (three seahorse and two pipefish species), seahorse newborns (three seahorses species), and prey (copepods and different stages of Artemia) commonly used to feed the fishes under rearing conditions. The aims of the study were: (i) to evaluate isotopic effects of chemical preservation methods across different types of organisms and tissues, using frozen samples as controls, and (ii) to construct the first conversion models available in syngnathid fishes. The chemical preservation in ethanol and, to a lesser extent, in formaldehyde significantly affected δ13C values, whereas the effects on δ15N signatures were negligible. Due to their low lipid content, the isotopic signals in fish fins was almost unaffected, supporting the suitability of dorsal fins as the most convenient material in isotopic studies on vulnerable species such as syngnathids. The regression equations provided resulted convenient for the successful conversion of δ13C between preservation treatments. Our results indicate that the normalization of δ15N signatures in preserved samples is unnecessary. The conversion models should be applicable in isotopic field studies, laboratory experiments, and specimens of historical collections.

Isotopic stable analysis (SIA) is a powerful tool in the assessment of different types of ecological and physiological studies. For that, different preservative methods for the samples are commonly used prior to isotopic analysis. The effects of various preservative methods (drying, freezing, ethanol and formaldehyde) have been analyzed for C:N ratio, δ13C and δ15N on a variety of tissues including dorsal fins (three seahorse and two pipefish species), seahorse newborns (three seahorses species), and prey (copepods and different stages of Artemia) commonly used to feed the fishes in rearing conditions. The aims of the study were to: (i) evaluate isotopic effects of preservation methods across tissues; and (ii) construct the first conversion models available in syngnathid fishes. The preservation in ethanol and to a lesser extend in formaldehyde significantly affected δ13C values, whereas δ15N signatures were not affected significantly. Due to their low lipid content, the isotopic signals in fish fins were almost unaffected, supporting the suitability of dorsal fins as a convenient tool in isotopic studies on vulnerable species such as syngnathids. The regression equations provided permit the successful conversion of δ13C and δ15N values between preservative treatments. The conversion models can be applied to isotopic studies in the field and in the laboratory.

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