A metabarcoding approach for the feeding habits of European hake in the Adriatic Sea

Abstract

European hake (Merluccius merluccius) is one of the most economically important fish for the Mediterranean Sea. It is an important predator of deep upper shelf slope communities currently characterized by growth overexploitation: the understanding of hake's diet might support next generation management tools. However, all current European hake diet studies depend on the morphological identification of prey remains in stomach content, with consequent limitations. In this study, we set up a metabarcoding approach based on cytochrome oxidase I PCR amplification and Miseq Illumina paired‐end sequencing of M. merluccius stomach content remains and compared the results to classic morphological analyses. A total of 95 stomach contents of M. merluccius sampled in the North‐Central Adriatic Sea were analyzed with both the metabarcoding and morphological approaches. Metabarcoding clearly outperformed the morphological method in the taxonomic identification of prey describing more complex trophic relationships even when considering the morphological identification of 200 stomach contents. Statistical analysis of diet composition revealed a weak differentiation among the hake's size classes, confirming an opportunistic feeding behavior. All the analyses performed showed the presence of a core of shared prey among the size classes and a cloud of size‐specific prey. Our study highlights the exceptional potential of metabarcoding as an approach to provide unprecedented taxonomic resolution in the diet of M. merluccius and potentially of other marine predators, due to the broad‐spectrum of detection of the primers used. A thorough description of these complex trophic relationships is fundamental for the implementation of an ecosystem approach to fisheries.

Feeding studies take guts – critical review and recommendations of methods for stomach contents analysis in fish

“…Dietary studies using DNA metabarcoding is rapidly proliferating, opening new techniques for the exploration of trophic interactions and population and community dynamics (Jakubavičiūtė et al ., ; Kress et al ., ; Valentini et al ., ). Hence, DNA‐based analysis of stomach contents is a promising approach, especially from the possibility of providing a high taxonomic resolution of prey (Jakubavičiūtė et al ., ; Riccioni et al ., ). However, there are still some shortcomings of this approach.…”

“…DNA degradation during digestion is another limiting factor for DNA barcoding of stomach contents (Kress et al ., ; Sakaguchi et al ., ). Other constraints relate to the situation that the results mainly are limited to species richness and frequency of occurrence of prey due to the semi‐quantitative nature of the metabarcoding analysis of stomach contents (Harms‐Tuohy et al ., ; Riccioni et al ., ). Secondary prey ( i.e ., prey of the primary prey) may also be detected from the barcoding, resulting in an overestimation of prey richness (Harms‐Tuohy et al ., ; Sakaguchi et al ., ).…”

“…Secondary prey ( i.e ., prey of the primary prey) may also be detected from the barcoding, resulting in an overestimation of prey richness (Harms‐Tuohy et al ., ; Sakaguchi et al ., ). Nevertheless, DNA metabarcoding provides a new and promising complementary approach to traditional morphological methods and the combination of visual and metabarcoding analyses of stomach contents will likely prove to be a powerful tool (Jakubavičiūtė et al ., ; Riccioni et al ., ). For fluid‐feeding parasitic fishes like such as catfishes in the family Trichomycteridae with mucophagic, lepidophagic or hematophagic diets that cannot be visually identified, the molecular approach is a particularly useful alternative (Bonato et al ., ).…”

Molecular Ecological Network Analyses: An Effective Conservation Tool for the Assessment of Biodiversity, Trophic Interactions, and Community Structure

“…Only recently has molecular network analysis been applied to the assessment of ecological systems. Metabarcoding studies in this context have focused on varying scales: the diet of European hake (Riccioni et al, 2018); coral reef fish (Casey et al, 2019); freshwater ecosystems (Compson et al, 2019); aquatic macroinvertebrates (Bush et al, 2019); and bat (Clare et al, 2019), forest (Evans et al, 2016), and herbivore (Kartzinel et al, 2015) communities. However, to the best of our knowledge (see Sousa et al, 2019 for a review), to date a metabarcoding study has yet to use fecal eDNA to reconstruct ecological networks and unravel trophic interactions among carnivores, omnivores, and herbivores within a terrestrial mammal community to inform biodiversity assessments.…”

A powerful long metabarcoding method for the determination of complex diets from faecal analysis of the European pond turtle (Emys orbicularis, L. 1758)

“…HTS has been used to assess the diet composition of a wide taxonomic range of animals. Animals whose diets have been successfully investigated have included mammals (Buglione et al., 2018; De Barba et al., 2014; Esnaola et al., 2018; Robeson et al., 2017), birds (Crisol‐Martinìez et al., 2016; Han & Oh, 2018), reptiles (Caut et al., 2019; Kartzinel & Pringle, 2015; Koizumi et al., 2017), fish (Barbato et al., 2019; Harms‐Tuohy et al., 2016; Riccioni et al., 2018) and arthropods (Kamenova et al., 2018; Kennedy et al., 2020; Krehenwinkel et al., 2016). For most species, faecal samples, in contrast to stomach contents, can easily be obtained, with a minimal level of interaction and harm inflicted on the studied animal.…”