By investigating the diet of the major prey of puffins we consider the potential effect of secondary consumption

It is also possible for molecular methods to shed light on questions that have historically been challenging or impossible to answer. For instance, as adult puffins forage at sea and do not leave identifiable components of prey in feces or in the form of a pellet, chick diet has been used as a best estimate for adult diet for birds in the Gulf of Maine, an assumption supported by similar levels of nitrogen isotopes in chick and adult blood. However, theory on optimal foraging predicts that as central-place foragers, adult puffins should feed their chicks a less diverse diet of high quality food while they feed on a more varied assortment of potentially lower quality prey. DNA-based dietary analysis of fecal samples offers the opportunity to document Gulf of Maine adult puffin diet and to test the similarity between adult and chick diet. Additionally, the diet of the main prey of puffin chicks is not well known and based on five >30 year old stomach content analyses. Molecularderived herring diet can simultaneously evaluate conventional stomach content analyses of diet while extending our knowledge of an important ecosystem. In this paper we apply next-generation sequencing of DNA barcodes from two genetic markers on puffin adult and chick fecal samples and herring stomach contents to study diet and describe the food chain in which these species exist. We compare the diet of puffin chicks using conventional field observations and molecular methods, test the assumption of common diet in puffin adults and chicks, and test for the effect of secondary consumption by considering the diets of chicks and herring. Further, we use the prey species detected by each genetic marker in diet samples to assess the efficacy of a multilocus technique as well as to make inferences about foraging ecology of puffins and herring. Since food webs provide the framework from which we draw conclusions about how an ecosystem functions, it is imperative that the studies used to construct food webs provide the most R428 accurate, unbiased, and repeatable estimated of diet possible. This paper provides the first analysis of multiple genetic markers across multiple taxa within a marine food web and demonstrates the broad utility of the technique as a tool for diet reconstruction in fish and birds. Our use of multi-locus pyrosequencing to reconstruct the diets of puffins and one of their primary food sources allowed for a more complete puffin food chain than known to date. The number and diversity of taxa identified from our molecular evaluation of diet is far greater than, though still consistent with, conventional diet studies in both herring and puffins. Many invertebrate taxa with planktonic life stages detected in herring diet were also found in puffin diet. We also found chick and adult puffins to have similar diets, although adult samples tended to have a higher proportion of invertebrate taxa.. This research demonstrates the general utility of next generation sequencing with multiple markers and highlights the use of this powerful tool for food web reconstruction. Failure to detect or misidentification of prey in predator diet can be a substantial hindrance to our understanding of how components of an ecosystem interact. For example, identifying commercially fished species consumed by puffins is important for the effective management of these stocks, as the impact of non-human predators on fish has historically been Reversine severely underestimated. The natural mortality rate of herring used in stock assessment models, for example, was less than 25% of the estimated consumption by mammals, piscivorous fish, and seabirds. Explicit consideration of the links between exploited species and the rest of the ecosystem, termed ecosystem-based management, has superseded historical stock-based resource management.

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