Fish are both consumers and prey, and as such part of a dynamic trophic network. Measuring how they are trophically linked, both directly and indirectly, to other species is vital to comprehend the mechanisms driving alterations in fish communities in space and time. Moreover, this knowledge also helps to understand how fish communities respond to environmental change and delivers important information for implementing management of fish stocks. DNA‐based methods have significantly widened our ability to assess trophic interactions in both marine and freshwater systems and they possess a range of advantages over other approaches in diet analysis. In this review we provide an overview of different DNA‐based methods that have been used to assess trophic interactions of fish as consumers and prey. We consider the practicalities and limitations, and emphasize critical aspects when analysing molecular derived trophic data. We exemplify how molecular techniques have been employed to unravel food web interactions involving fish as consumers and prey. In addition to the exciting opportunities DNA‐based approaches offer, we identify current challenges and future prospects for assessing fish food webs where DNA‐based approaches will play an important role.

Although a significant proportion of plant tissue is located in roots and other below-ground parts of plants, little is known on the dietary choices of root-feeding insects. This is caused by a lack of adequate methodology which would allow tracking below-ground trophic interactions between insects and plants. Here, we present a DNA-based approach to examine this relationship. Feeding experiments were established where either wheat (Triticum aestivum) or maize (Zea mays) was fed to Agriotes larvae (Coleoptera: Elateridae), allowing them to digest for up to 72 h. Due to the very small amount of plant tissue ingested (max = 6.76 mg), DNA extraction procedures and the sensitivity of polymerase chain reaction (PCR) had to be optimized. Whole-body DNA extracts of larvae were tested for the presence of both rbcL and trnL plastid DNA using universal primers. Moreover, based on cpDNA sequences encoding chloroplast tRNA for leucine (trnL), specific primers for maize and wheat were developed. With both, general and specific primers, plant DNA was detectable in the guts of Agriotes larvae for up to 72 h post-feeding, the maximum time of digestion in these experiments. No significant effect of time since feeding on plant DNA detection success was observed, except for the specific primers in maize-fed larvae. Here, plant DNA detection was negatively correlated with the duration of digestion. Both, meal size and initial mass of the individual larvae did not affect the rate of larvae testing positive for plant DNA. The outcomes of this study represent a first step towards a specific analysis of the dietary choices of soil-living herbivores to further increase our understanding of animal–plant feeding interactions in the soil.

Carabid beetles are abundant in temperate agroecosystems and can play a pivotal role as biocontrol agents. While there is good knowledge regarding their effects on invertebrate pests in some systems, comparably little is known on the rate of seed feeding under field conditions. Molecular approaches are ideally suited for investigating carabid feeding interactions; to date, however, they have only been applied to animal prey. We sampled adult carabid beetles in organic cereal fields in three regions along a Central European transect. Regurgitates from populations of the three most common species,Poecilus cupreus, Pseudoophonus rufipesandPterostichus melanarius, were screened for plant DNA, cereal aphids, collembolans and earthworms. The frequency of carabid individuals positive for plant DNA was high (> 70%) and independent of carabid species, sex, region and the time point of sampling. Detections for non-pest and pest prey were comparably lower, with 21.6% for collembolans, 18.1% for earthworms and 4.2% for aphids, respectively. Despite the prolonged detection period of plant DNA in carabid guts, as compared to animal prey, these first results suggest that weed seeds form an important part of the adult carabid diet. It would also lend support to the hypothesis that seed-feeding carabids are biocontrol agents of weeds, with effects of regulation on the weed seedbank that depend on behavioural and contextual factors including carabid species preferences for weed seed species, their life stage and tillage practices.

Plant roots represent an important food source for soil‐dwelling animals, but tracking herbivore food choices below‐ground is difficult. Here, we present an optimized PCR assay for the detection of plant DNA in the guts of invertebrates, using general plant primers targeting the trnT‐F chloroplast DNA region. Based on this assay, we assessed the influence of plant identity on the detectability of ingested plant DNA in Agriotes click beetle larvae. Six different plant species were fed to the insects, comprising a grass, a legume and four nonlegume forbs. Moreover, we examined whether it is possible to amplify DNA of decaying plants and if DNA of decayed plant food is detectable in the guts of the larvae. DNA of the ingested roots could be detected in the guts of the larvae for up to 72‐h post‐feeding, the maximum digestion time tested. When fed with living plants, DNA detection rates differed significantly between the plant species. This may be ascribed to differences in the amount of plant tissue consumed, root palatability, root morphology and/or secondary plant components. These findings indicate that plant identity can affect post‐feeding DNA detection success, which needs to be considered for the interpretation of molecularly derived feeding rates on plants. Amplification of plant DNA from decaying plants was possible as long as any tissue could be retrieved from the soil. The consumption of decaying plant tissue could also be verified by our assay, but the insects seemed to prefer fresh roots over decaying plant material.

Soil‐dwelling pests inflict considerable economic damage in agriculture but are hard to control. A promising strategy to reduce pest pressure on crops is to increase the plant diversity in agroecosystems. This approach, however, demands a sound understanding of species' interactions, which is widely lacking for subterranean herbivore–plant systems. Here, we examine the effects of plant diversification on wireworms, the soil‐dwelling larvae of click beetles that threaten crops worldwide. We conducted a field experiment employing plant diversification by adding either wheat or a mix of six associated plants (grasses, legumes, and forbs) between rows of maize to protect it from Agriotes wireworms. Wireworm feeding behavior, dispersal between crop and associated plants, as well as maize damage and yield were examined. The former was assessed combining molecular gut content and stable isotope analysis. The pests were strongly attracted by the associated plants in August, when the crop was most vulnerable, whereas in September, shortly before harvest, this effect occurred only in the plant mix. In maize monoculture, the larvae stayed in the principal crop throughout the season. Larval δ¹³C signatures revealed that maize feeding was reduced up to sevenfold in wireworms of the vegetationally diversified treatments compared to those of the maize monoculture. These findings were confirmed by molecular analysis, which additionally showed a dietary preference of wireworms for specific plants in the associated plant mix. Compared to the monoculture, maize damage was reduced by 38% and 55% in the wheat and plant mix treatment, which translated into a yield increase of 30% and 38%, respectively. The present findings demonstrate that increasing the plant diversity in agroecosystems provides an effective insurance against soil pests. The underlying mechanisms are the diversion of the pest from the principle crop and a changed feeding behavior. The deployment of diverse mixes of associated plants, tailored to the specific preferences of the soil herbivores, provides a promising strategy for managing subterranean pests while maintaining crop yield.