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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. [ABSTRACT FROM AUTHOR]

Many omnivorous arthropods act as important pest control agents in agroecosystems. Assessing their prey choices helps to better understand their biocontrol potential in different environmental settings. To this end, multi‐target approaches of molecular gut content analysis allow studying their prey choices. Yet, DNA‐based analysis of trophic interactions is challenged by the question of whether the detected food DNA was ingested primarily as direct prey, or secondarily via the gut content of a consumed animal. Experimental data are needed to understand the potential bias derived through secondary predation. Here we address this issue for a phloem‐feeding aphid consumed by omnivorous carabids. In feeding experiments, Pseudoophonus rufipes beetles were offered grain aphids Sitobion avenae as primary prey, which had fed on barley seedlings. Gut content samples of carabids collected after 0, 3, 9, 12, 24, 32 or 64 h of digestion time and whole‐body aphids were screened for the presence of aphid and plant DNA. Samples positive for plant DNA were subsequently tested with Poaceae‐specific primers. None of the 7.8% gut content samples that tested positive for general plant DNA amplified with the Poaceae primers. Moreover, none of the whole‐body extracted aphids tested positive for plant DNA, neither with general nor with family‐specific primers. We, therefore, conclude that the plant DNA detections in our experiment can be ascribed to remnants of plant material in the guts of the field‐collected carabids. Our findings demonstrate that these phloem‐feeding aphids are an unlikely cause of false trophic link assignment because no secondarily consumed plant DNA, derived either from aphid gut content or host‐plant environmental DNA on aphid bodies, could be detected in carabid gut content samples. [ABSTRACT FROM AUTHOR]

Generalist predators such as carabid and cantharid beetles form an important component within natural enemy communities in arable land. Usually, predator-prey interactions are examined during the vegetation period, largely ignoring food web interactions occurring in the cold season. This is, however, when the larval stages of many polyphagous beetles develop whose survival critically depends on the availability of suitable prey. In this study, we examined intra- and extraguild feeding links in larval Cantharis spp. (Coleoptera: Cantharidae) and Nebria brevicollis (Coleoptera: Carabidae), both abundant cold-adapted invertebrate predators in European arable land. As these immature beetles are fluid feeders, which impedes a microscopic analysis of their gut content, multiplex polymerase chain reaction assays were developed to examine trophic linkages in the field. Collembolan DNA was detected in 49% and 54% of cantharid and carabid larvae, respectively. Earthworms were consumed by 34% of cantharid and 24% of carabid larvae. Significant differences in lumbricid and collembolan prey detection rates occurred between sampling dates in Cantharis spp. and N. brevicollis larvae, respectively, suggesting that these predators utilized different feeding strategies. In both predator taxa, however, only few individuals (0·2-1%) tested positive for DNA of intraguild prey, indicating that predator-predator trophic interactions were scarce in this community. Synthesis and applications. We present conclusive evidence that cold-adapted predatory beetle larvae are strongly linked to the detrital food chain by feeding on collembolans and earthworms. By improving the habitat conditions for detritivores in arable land by mulching, compost applications, or provision of plant cover during winter, their densities can be increased easily. Applying these measures year round will retain and sustain predatory beetles during their whole life cycle in arable land. Ultimately, we expect that these measures will enhance the ability of polyphagous beetle predators to provide their fundamental ecosystem service as regulators of agricultural pests. [ABSTRACT FROM AUTHOR]

White grubs (larvae of Coleoptera: Scarabaeidae) are abundant in below-ground systems and can cause considerable damage to a wide variety of crops by feeding on roots. White grub populations may be controlled by natural enemies, but the predator guild of the European species is barely known. Trophic interactions within soil food webs are difficult to study with conventional methods. Therefore, a polymerase chain reaction (PCR)-based approach was developed to investigate, for the first time, a soil insect predator-prey system. Can, however, highly sensitive detection methods identify carrion prey in predators, as has been shown for fresh prey? FreshMelolontha melolontha(L.) larvae and 1- to 9-day-old carcasses were presented toPoecilus versicolorSturm larvae. Mitochondrial cytochrome oxidase subunit I fragments of the prey, 175, 327 and 387 bp long, were detectable in 50% of the predators 32 h after feeding. Detectability decreased to 18% when a 585 bp sequence was amplified. Meal size and digestion capacity of individual predators had no influence on prey detection. Although prey consumption was negatively correlated with cadaver age, carrion prey could be detected by PCR as efficiently as fresh prey irrespective of carrion age. This is the first proof that PCR-based techniques are highly efficient and sensitive, both in fresh and carrion prey detection. Thus, if active predation has to be distinguished from scavenging, then additional approaches are needed to interpret the picture of prey choice derived by highly sensitive detection methods. [ABSTRACT FROM AUTHOR]

• Intraguild predation within beetle communities was analysed in cereal fields. • Organic fertilization increased non-pest prey and lowered intraguild predation. • Lower intraguild predation correlated with lower pest spread, but not density. • No effect of organic fertilization was found on pest density or consumption. • Fertilization effects may offset pest predation from lower predator interference. Intraguild predation is an extreme form of competition, that depends on resource availability, niche overlap, behaviour and diversity. However, despite modelling and mesocosm experiments, field studies are scarce, and we know nothing of its temporal dynamics within real-world food webs, particularly in the context of biological control. To address this, we added manure to cereal fields to increase decomposer prey, and analysed the diet of over 6000 predators over the entire cropping season during two consecutive years. Our results show that manure increased available prey temporarily and reduced intraguild predation. With this organic fertilization, a reduction in pest spread between tillers within fields correlated with lower intraguild predation, while pest density remained unchanged across treatments, making the net effect on biological control unclear. We hypothesize that the mechanism responsible for why lowering intraguild predation reduces pest spread are changes in predator behaviour, that allow predators to exert more pressure on pests. Conversely, the absence of an effect on pest density may be linked to other effects of fertilization, such as increased pest and plant growth counteracting any increase in predation pressure. The high temporal resolution of our data showed how dynamic IGP is throughout the season, which stresses the importance of studying food webs over time, to generate a mechanistic understanding of biological control. [ABSTRACT FROM AUTHOR]

Intensified agriculture, a driver of biodiversity loss, can diminish ecosystem functions and their stability. Biodiversity can increase functional redundancy and is expected to stabilize ecosystem functions. Few studies, however, have explored how agricultural intensity affects functional redundancy and its link with ecosystem function stability. Here, within a continental‐wide study, we assess how functional redundancy of seed predation is affected by agricultural intensity and landscape simplification. By combining carabid abundances with molecular gut content data, functional redundancy of seed predation was quantified for 65 weed genera across 60 fields in four European countries. Across weed genera, functional redundancy was reduced with high field management intensity and simplified crop rotations. Moreover, functional redundancy increased the spatial stability of weed seed predation at the field scale. We found that ecosystem functions are vulnerable to disturbances in intensively managed agroecosystems, providing empirical evidence of the importance of biodiversity for stable ecosystem functions across space. [ABSTRACT FROM AUTHOR]

Plant diversity affects species richness and abundance of taxa at higher trophic levels. However, plant diversity effects on omnivores (feeding on multiple trophic levels) and their trophic and non-trophic interactions are not yet studied because appropriate methods were lacking. A promising approach is the DNA-based analysis of gut contents using next generation sequencing (NGS) technologies. Here, we integrate NGS-based analysis into the framework of a biodiversity experiment where plant taxonomic and functional diversity were manipulated to directly assess environmental interactions involving the omnivorous ground beetle Pterostichus melanarius. Beetle regurgitates were used for NGS-based analysis with universal 18S rDNA primers for eukaryotes. We detected a wide range of taxa with the NGS approach in regurgitates, including organisms representing trophic, phoretic, parasitic, and neutral interactions with P. melanarius. Our findings suggest that the frequency of (i) trophic interactions increased with plant diversity and vegetation cover; (ii) intraguild predation increased with vegetation cover, and (iii) neutral interactions with organisms such as fungi and protists increased with vegetation cover. Experimentally manipulated plant diversity likely affects multitrophic interactions involving omnivorous consumers. Our study therefore shows that trophic and non-trophic interactions can be assessed via NGS to address fundamental questions in biodiversity research. [ABSTRACT FROM AUTHOR]

Abundance and diversity of natural enemies increases with higher landscape complexity. However, more species can also increase negative interactions such as intraguild predation (IGP), which could be detrimental to pest control. Direct observations of these trophic interactions are still lacking. In the present study, we employed DNA-based gut content analyses of coccinellid beetles to assess IGP (direct and coincidental) and aphid consumption. Species-specific and group-specific primer pairs were used to unveil the interactions of the most abundant coccinellid species, three cereal aphids and their associated parasitoids. In order to determine whether the landscape complexity promotes these interactions, we used replicated wheat fields in two contrasting landscape contexts over the season in central Chile. With higher abundance of coccinellids, we observed greater aphid consumption in complex contexts. The consumption of parasitized aphids was mostly detected early in complex contexts, suggesting a detrimental effect on the parasitoid population. Following molecular analysis, we can state that the aphids are important feeding resources for the coccinellid assemblage and are actively foraging on this important pest. Finally, this study demonstrates that direct IGP is a common interaction in this system, independently of the landscape context during the complete season. Nevertheless, the native Eriopis chilensis was found to be the most frequently consumed intraguild prey. In contrast with expected predictions, we found little evidence that the landscape context increased negative interactions, such as IGP and IGP-C and in turn that these decrease the control of Sitobion avenae. [ABSTRACT FROM AUTHOR]

Most ecosystem functions and related services involve species interactions across trophic levels, for example, pollination and biological pest control. Despite this, our understanding of ecosystem function in multitrophic communities is poor, and research has been limited to either manipulation in small communities or statistical descriptions in larger ones.Recent advances in food web ecology may allow us to overcome the trade‐off between mechanistic insight and ecological realism. Molecular tools now simplify the detection of feeding interactions, and trait‐based approaches allow the application of dynamic food web models to real ecosystems. We performed the first test of an allometric food web model's ability to replicate temporally nonaggregated abundance data from the field and to provide mechanistic insight into the function of predation.We aimed to reproduce and explore the drivers of the population dynamics of the aphid herbivore Rhopalosiphum padi observed in ten Swedish barley fields. We used a dynamic food web model, taking observed interactions and abundances of predators and alternative prey as input data, allowing us to examine the role of predation in aphid population control. The inverse problem methods were used for simultaneous model fit optimization and model parameterization.The model captured >70% of the variation in aphid abundance in five of ten fields, supporting the model‐embodied hypothesis that body size can be an important determinant of predation in the arthropod community. We further demonstrate how in‐depth model analysis can disentangle the likely drivers of function, such as the community's abundance and trait composition. Analysing the variability in model performance revealed knowledge gaps, such as the source of episodic aphid mortality, and general method development needs that, if addressed, would further increase model success and enable stronger inference about ecosystem function.The results demonstrate that confronting dynamic food web models with abundance data from the field is a viable approach to evaluate ecological theory and to aid our understanding of function in real ecosystems. However, to realize the full potential of food web models, in ecosystem function research and beyond, trait‐based parameterization must be refined and extended to include more traits than body size. Dynamic food web models could aid our understanding of ecosystem functions driven by trophic interactions, but are rarely tested against data. The authors confront a trait‐based food web model with field data, and show that it can explain observed abundance dynamics of an arthropod herbivore and provide insight into ecosystem functioning. [ABSTRACT FROM AUTHOR]

Primary succession on bare ground surrounded by intact ecosystems is, during its first stages, characterized by predator‐dominated arthropod communities. However, little is known on what prey sustains these predators at the start of succession and which factors drive the structure of these food webs. As prey availability can be extremely patchy and episodic in pioneer stages, trophic networks might be highly variable. Moreover, the importance of allochthonous versus autochthonous food sources for these pioneer predators is mostly unknown. To answer these questions, the gut content of 1,832 arthropod predators, including four species of carabid beetles, two lycosid and several linyphiid spider species caught in early and late pioneer stages of three glacier forelands, was screened molecularly to track intraguild and extraguild trophic interactions among all major prey groups occurring in these systems. Two‐thirds of the 2,310 identified food detections were collembolans and intraguild prey, while one‐third were allochthonous flying insects. Predator identity and not successional stage or valley had by far the strongest impact on the trophic interaction patterns. Still, the variability of prey spectra increased significantly from early to late pioneer stage, as did the niche width of the predators. As such the structure of pioneer arthropod food webs in recently deglaciated Alpine habitats seems to be driven foremost by predator identity while site and early successional effects contribute to a lesser extent to food web variability. Our findings also suggest that in these pioneer sites, predatory arthropods depend less on allochthonous aeolian prey but are mainly sustained by prey of local production. [ABSTRACT FROM AUTHOR]

Abstract: Piscivorous birds frequently display sex‐specific differences in their hunting and feeding behavior, which lead to diverging impacts on prey populations. Cormorants (Phalacrocoracidae), for example, were previously studied to examine dietary differences between the sexes and males were found to consume larger fish in coastal areas during autumn and winter. However, information on prey partitioning during breeding and generally on sex‐specific foraging in inland waters is missing. Here, we assess sex‐specific prey choice of Great Cormorants (Phalacrocorax carbo) during two subsequent breeding seasons in the Central European Alpine foreland, an area characterized by numerous stagnant and flowing waters in close proximity to each other. We developed a unique, noninvasive approach and applied it to regurgitated pellets: molecular cormorant sexing combined with molecular fish identification and fish‐length regression analysis performed on prey hard parts. Altogether, 364 pellets delivered information on both, bird sex, and consumed prey. The sexes differed significantly in their overall prey composition, even though Perca fluviatilis, Rutilus rutilus, and Coregonus spp. represented the main food source for both. Albeit prey composition did not indicate the use of different water bodies by the sexes, male diet was characterized by higher prey diversity within a pellet and the consumption of larger fish. The current findings show that female and male cormorants to some extent target the available prey spectrum at different levels. Finally, the comprehensive and noninvasive approach has great potential for application in studies of other piscivorous bird species. [ABSTRACT FROM AUTHOR]

Abstract: The “habitat heterogeneity hypothesis” predicts positive effects of structural complexity on species coexistence. Increasing habitat heterogeneity can change the diversity (number of species, abundances) and the functional roles of communities. The latter, however, is not well understood as species and individuals may respond very differently and dynamically to a changing environment. Here, we experimentally test how habitat heterogeneity affects generalist arthropod predators, including epigaeic spiders, carabid and staphylinid beetles, under natural conditions by assessing their diversity and directly measuring their trophic interactions (which provide a proxy for their functional roles). The experiment was conducted in spring barley fields in Southern Sweden where habitat heterogeneity was manipulated by increasing within‐field plant diversity. Increased habitat heterogeneity triggered rapid changes in the feeding behaviour of generalist predators characterized by lower trophic specialization at both network (H2’, degree of interaction specialization in the entire network) and species level (d’, degree of interaction specialization at the species level). We presume that this is because spatial separation resulted in relaxed competition and allowed an increased overlap in resources used among predator species. Predators collected from heterogenous habitats also showed greater individual‐level dietary variability which might be ascribed to relaxed intraspecific competition. Our results provide conclusive evidence that habitat heterogeneity can induce rapid behavioural responses independent of changes in diversity, potentially promoting the stability of ecosystem functions. A plain language summary is available for this article. [ABSTRACT FROM AUTHOR]

Molecular methods allow noninvasive assessment of vertebrate predator-prey systems at high taxonomic resolution by examining dietary samples such as faeces and pellets. To facilitate the interpretation of field-derived data, feeding trials, investigating the impacts of biological, methodological and environmental factors on prey DNA detection, have been conducted. The effect of meal size, however, has not yet been explicitly considered for vertebrate consumers. Moreover, different noninvasively obtained sample types remain to be compared in such experiments. Here, we present a feeding trial on abundant piscivorous birds, Great Cormorants ( Phalacrocorax carbo), to assess meal size effects on postfeeding prey DNA detection success. Faeces and pellets were sampled twice a day after the feed of large (350-540 g), medium (190-345 g) and small (15-170 g) fish meals contributing either a large (>79%) or small (<38%) share to the daily consumption. Samples were examined for prey DNA and fish hard parts. Molecular analysis of faeces revealed that both large meal size and share had a significantly positive effect on prey DNA detection rate postfeeding. Furthermore, large meals were detectable for a significantly longer time span with a detection limit at ~76 hr and a 50% detection probability at ~32 hr postfeeding. In pellets, molecular methods reliably identified the meal consumed the previous day, which was not possible via morphological analysis or when examining individual faeces. The less reliable prey DNA detection of small meals or meal shares in faeces signifies the importance of large numbers of dietary samples to obtain reliable trophic data. [ABSTRACT FROM AUTHOR]

In trophic studies on piscivorous birds, it is vital to know which kind of dietary sample provides the information of interest and how the prey can be identified reliably and efficiently. Often, noninvasively obtained dietary samples such as regurgitated pellets, feces, and regurgitated fish samples are the preferred source of information. Fish prey has usually been identified via morphological analysis of undigested hard parts, but molecular approaches are being increasingly used for this purpose. What remains unknown, however, is which dietary sample type is best suited for molecular diet analysis and how the molecular results compare to those obtained by morphological analysis. Pellets, feces, and regurgitated fish samples of Great Cormorants ( Phalacrocorax carbo sinensis) were examined for prey using both morphological hard part analysis and molecular prey identification. The sample types and methods were compared regarding number of species detected (overall and per sample) as well as the prey species composition and its variability among individual samples. Via molecular analysis, significantly higher numbers of prey species were detected in pellets, feces, and fish samples. Of the three sample types, pellets contained the most comprehensive trophic information and could be obtained with the lowest sampling effort. Contrastingly, dietary information obtained from feces was least informative and most variable. For all sample types, the molecular approach outperformed morphological hard part identification regarding the detectable prey spectrum and prey species composition. We recommend the use of pellets in combination with molecular prey identification to study the diet of piscivorous birds. [ABSTRACT FROM AUTHOR]

1. Studies of the impact of predator diversity on biological pest control have shown idiosyncratic results. This is often assumed to be as a result of differences among systems in the importance of predator-predator interactions such as facilitation and intraguild predation. The frequency of such interactions may be altered by prey availability and structural complexity. A direct assessment of interactions among predators is needed for a better understanding of the mechanisms affecting prey abundance by complex predator communities. 2. In a field cage experiment, the effect of increased predator diversity (single species vs. three-species assemblage) and the presence of weeds (providing structural complexity) on the biological control of cereal aphids were tested and the mechanisms involved were investigated using molecular gut content analysis. 3. The impact of the three-predator species assemblages of aphid populations was found to be similar to those of the single-predator species treatments, and the presence or absence of weeds did not alter the patterns observed. This suggests that both predator facilitation and intraguild predation were absent or weak in this system, or that these interactions had counteracting effects on prey suppression. Molecular gut content analysis of predators provided little evidence for the latter hypothesis: predator facilitation was not detected and intraguild predation occurred at a low frequency. 4. The present study suggests additive effects of predators and, therefore, that predator diversity per se neither strengthens nor weakens the biological control of aphids in this system. [ABSTRACT FROM AUTHOR]

The applicability of species-specific primers to study feeding interactions is restricted to those ecosystems where the targeted prey species occur. Therefore, group-specific primer pairs, targeting higher taxonomic levels, are often desired to investigate interactions in a range of habitats that do not share the same species but the same groups of prey. Such primers are also valuable to study the diet of generalist predators when next generation sequencing approaches cannot be applied beneficially. Moreover, due to the large range of prey consumed by generalists, it is impossible to investigate the breadth of their diet with species-specific primers, even if multiplexing them. However, only few group-specific primers are available to date and important groups of prey such as flying insects have rarely been targeted. Our aim was to fill this gap and develop group-specific primers suitable to detect and identify the DNA of common taxa of flying insects. The primers were combined in two multiplex PCR systems, which allow a time- and cost-effective screening of samples for DNA of the dipteran subsection Calyptratae (including Anthomyiidae, Calliphoridae, Muscidae), other common dipteran families (Phoridae, Syrphidae, Bibionidae, Chironomidae, Sciaridae, Tipulidae), three orders of flying insects (Hymenoptera, Lepidoptera, Plecoptera) and coniferous aphids within the genus Cinara. The two PCR assays were highly specific and sensitive and their suitability to detect prey was confirmed by testing field-collected dietary samples from arthropods and vertebrates. The PCR assays presented here allow targeting prey at higher taxonomic levels such as family or order and therefore improve our ability to assess (trophic) interactions with flying insects in terrestrial and aquatic habitats. [ABSTRACT FROM AUTHOR]

Pioneer communities establishing themselves in the barren terrain in front of glacier forelands consist principally of predator species such as carabid beetles and lycosid spiders. The fact that so many different predators can co-inhabit an area with no apparent primary production was initially explained by allochthonous material deposited in these forelands. However, whether these populations can be sustained on allochthonous material alone is questionable and recent studies point towards this assumption to be flawed. Intraguild predation ( IGP) might play an important role in these pioneer predator assemblages, especially in the very early successional stages where other prey is scarce. Here, we investigated IGP between the main predator species and their consumption of Collembola, an important autochthonous alternative prey, within a glacier foreland in the Ötztal ( Austrian Alps). Multiplex PCR and stable isotope analysis were used to characterize the trophic niches in an early and late pioneer stage over 2 years. Results showed that intraguild prey was consumed by all invertebrate predators, particularly the larger carabid species. Contrary to our initial hypothesis, the DNA detection frequency of IGP prey was not significantly higher in early than in late pioneer stage, which was corroborated by the stable isotope analysis. Collembola were the most frequently detected prey in all of the predators, and the overall prey DNA detection patterns were consistent between years. Our findings show that IGP appears as a constant in these pioneer predator communities and that it remains unaffected by successional changes. [ABSTRACT FROM AUTHOR]

In glacier forelands spiders constitute a large proportion of the invertebrate community. Therefore, it is important to be able to determine the species that can be found in these areas. Linyphiid and theridiid spider identification is currently not possible in juvenile specimens using traditional morphological based methods, however, a large proportion of the population in these areas are usually juveniles. Molecular methods permit identification of species at different life stages, making juvenile identification possible. In this study we tested a molecular tool to identify the 10 most common species of Linyphiidae and Theridiidae found in three glacier foreland communities of the Austrian Alps. Two multiplex PCR systems were developed and over 90% of the 753 field-collected spiders were identified successfully. The species targeted were found to be common in all three valleys during the summer of 2010. A comparison between the molecular and morphological data showed that although there was a slight difference in the results, the overall outcome was the same independently of the identification method used. We believe the quick and reliable identification of the spiders via the multiplex PCR assays developed here will aid the study of these families in Alpine habitats. [ABSTRACT FROM AUTHOR]

Abstract: Soils harbour an extraordinary diversity of animals which are functionally connected by their feeding interactions. Determining these feeding links, however, is difficult due to the opaque habitat the animals live in and because many prey remains found in predator gut contents cannot be identified morphologically. DNA-based gut content analysis overcomes these hurdles. This approach opens new possibilities to empirically examine below-ground feeding links by measuring prey DNA detection frequencies in various predators. Comparing prey DNA detection rates between different predator taxa, however, is not straightforward as digestion rates and thus post-feeding prey DNA detection intervals can be affected by predator identity. In the present study we examined the effect of predator identity for a range of soil-dwelling adult and larval beetles as well as lithobiid and geophilid centipedes. These predators were fed with white grubs (scarabaeid beetle larvae) and their post-feeding prey DNA detection intervals were established for small, medium and large prey DNA fragments. In geophilids the 50% detection probability for the medium prey DNA fragment was 98 h post-feeding whereas it ranged between 22 and 34 h in beetles. Among the different beetle species, the magnitude of the differences in detection intervals was strongly affected by the size of the prey DNA fragment targeted: the intervals showed the highest similarity in the medium-sized prey DNA fragment which was most frequently detected. Predator biomass and meal size rarely affected post-feeding prey DNA detection intervals. Our findings suggest that prey DNA detection success is similar within taxonomically related species of soil-dwelling macro-invertebrates such as beetles and centipedes. Between these groups prey detection can be adjusted according to taxon-specific digestion rates for better comparisons. Moreover, the magnitude of the predator identity effect is modulated by assay sensitivity: highly sensitive assays which target small- to medium-sized prey DNA fragments are recommended as these minimize the interspecific differences in post-feeding prey DNA detection intervals. [Copyright &y& Elsevier]

Molecular methods have become an important tool for studying feeding interactions under natural conditions. Despite their growing importance, many methodological aspects have not yet been evaluated but need to be considered to fully exploit the potential of this approach. Using feeding experiments with high alpine carabid beetles and lycosid spiders, we investigated how PCR annealing temperature affects prey DNA detection success and how post-PCR visualization methods differ in their sensitivity. Moreover, the replicability of prey DNA detection among individual PCR assays was tested using beetles and spiders that had digested their prey for extended times postfeeding. By screening all predators for three differently sized prey DNA fragments (range 116-612 bp), we found that only in the longest PCR product, a marked decrease in prey detection success occurred. Lowering maximum annealing temperatures by 4 °C resulted in significantly increased prey DNA detection rates in both predator taxa. Among the three post-PCR visualization methods, an eightfold difference in sensitivity was observed. Repeated screening of predators increased the total number of samples scoring positive, although the proportion of samples testing positive did not vary significantly between different PCRs. The present findings demonstrate that assay sensitivity, in combination with other methodological factors, plays a crucial role to obtain robust trophic interaction data. Future work employing molecular prey detection should thus consider and minimize the methodologically induced variation that would also allow for better cross-study comparisons. [ABSTRACT FROM AUTHOR]