COI Metabarcoding Provides Insights into the Highly Diverse Diet of a Generalist Salamander, Salamandra salamandra (Caudata: Salamandridae)

Abstract

DNA metabarcoding has proven to be an accessible, cost-effective, and non-invasive tool for dietary analysis of predators in situ. Although DNA metabarcoding provides numerous benefits in characterizing diet—such as detecting prey animals that are difficult to visually identify—this method has seen limited application in amphibian species. Here, we used DNA metabarcoding to characterize the diet of fire salamanders (Salamandra salamandra) (Linnaeus, 1758) in three distinct regions across the northwestern Iberian Peninsula. To test the efficiency of COI-based metabarcoding in determining salamanders’ diet diversity, we compared our COI-based results with results from traditional diet studies from neighboring and distant populations, as well as with recent findings obtained in a DNA metabarcoding study using 18S. Two COI primers were used in combination to investigate the potential impact of primer bias in prey detection. Our COI metabarcoding approach increased taxonomic resolution and supported a generalist diet in S. salamandra. Between primers, there were no significant differences in the diversity and richness of prey detected. We observed differences in the prevalence of prey identified between sampling regions both in our study and in other studies of S. salamandra diet. This COI metabarcoding study provides recommendations and resources for subsequent research using DNA metabarcoding to study amphibian diets.

The Trophic Niche of Two Sympatric Species of Salamanders (Plethodontidae and Salamandridae) from Italy

“…Salamandra salamandra behaved exactly the opposite from Speleomantes , preferring mostly slow-moving worm-like prey [ 40 , 76 ]. This type of prey consumed by S. salamandra accounted for 79% of its diet ( Table 1 ), corroborating similar results obtained from different populations throughout its distribution [ 13 , 39 , 53 , 77 , 78 ]. Among the stomach contents recognized from S. salamandra , we observed a very low frequency of prey with hard cuticles, and generally they were <1 cm in size ( Table 1 ).…”

“…Among the stomach contents recognized from S. salamandra , we observed a very low frequency of prey with hard cuticles, and generally they were <1 cm in size ( Table 1 ). In a recent study, it was shown that soft-bodied prey can be underrepresented when analyzing the species’ diets through stomach flushing [ 13 ]. In our study, we observed that most (>63%) of the prey consumed by S. salamandra exclusively consisted of soft-bodied prey ( Table 1 ), which means that these taxa are not necessarily underrepresented in this type of study.…”

“…As we discussed for S. italicus (see above), this inconsistency may have been due to the delay between the foraging event and the inspection of salamanders’ stomachs. In their study, Marques, Mata and Velo-Antón [ 13 ] investigated the trophic niche of S. salamandra , analyzing their feces. This means that the prey had undergone the entire digestive process and that the salamanders had assimilated most of the wet mass by defecating only the solid residues that they had not been able to digest.…”

“…The trophic niche of these two species has only been assessed in the absence of potential competitors e.g. [ 12 , 13 , 14 ], leaving unknown the mechanisms and the extent to which these species are able to modify their trophic niche to coexist with competitors. Speleomantes italicus is one of the eight species of Plethodontidae occurring in Europe, and its distribution encompasses the Apennine chain, from northern Tuscany to Abruzzo (Italy), where it occurs in forested areas and subterranean environments as well [ 15 , 16 ].…”

“…The larvae feed upon multiple aquatic invertebrate species, but they can also adopt cannibalism when prey are scarce [ 14 , 37 , 38 ]. Adults forage in terrestrial environments and they often prey upon “worm-like” prey such as annelids, diplopods (among arthropods), and “slugs”, defined as apparently shell-less terrestrial gastropods (among mollusks) [ 13 , 39 , 40 ].…”