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To date, evaluating the diets of natural enemies like carabids has largely been limited to spatially explicit and short-term sampling. This leaves a knowledge gap for the intra-annual dynamics of carabid diets, and the provision and timing of delivery of natural pest control services. Season-long pitfall trapping of adult carabids was conducted in conventional winter wheat fields, from November 2018 to June 2019, in five French departments. Diagnostic Multiplex PCR of carabid gut contents was used to determine the dynamics of carabid diets. The overall detection rate of target prey DNA was high across carabid individuals (80%) but varied with the prey group. The rate of detection was low for pests, at 8.1% for slugs and 9.6% for aphids. Detection of intraguild predation and predation on decomposers was higher, at 23.8% for spiders, 37.9% for earthworms and 64.6% for springtails. Prey switching was high at the carabid community level, with pest consumption and intraguild predation increasing through the cropping season as the availability of these prey increased in the environment, while the detection of decomposer DNA decreased. Variation in diet through the cropping season was characterized by: (i) complementary predation on slug and aphid pests; and (ii) temporal complementarity in the predominant carabid taxa feeding on each pest. We hypothesize that natural pest control services delivered by carabids are determined by complementary contributions to predation by the different carabid taxa over the season.

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BACKGROUND: The use of DNA metabarcoding has become an increasingly popular technique to infer feeding relationships in polyphagous herbivores and predators. Understanding host plant preference of native and invasive herbivore insects can be helpful in establishing effective integrated pest management (IPM) strategies. The invasive Halyomorpha halys and native Pentatoma rufipes are piercing-sucking stink bug pests that are known to cause economic damage in commercial fruit orchards. RESULTS: In this study, we performed molecular gut content analysis (MGCA) on field-collected specimens of these two herbivorous pentatomids using next-generation amplicon sequencing (NGAS) of the internal transcribed spacer 2 (ITS2) barcode region. Additionally, a laboratory experiment was set up where H. halys was switched from a mixed diet to a monotypic diet, allowing us to determine the detectability of the initial diet in a time series of ≤3 days after the diet switch. We detected 68 unique plant species from 54 genera in the diet of two stink bug species, with fewer genera found per sample and a smaller diet breadth for P. rufipes than for H. halys. Both stink bug species generally prefer deciduous trees over gymnosperms and herbaceous plants. Landscape type significantly impacted the observed genera in the diet of both stink bug species, whereas season only had a significant effect on the diet of H. halys. CONCLUSION: This study provides further insights into the dietary composition of two polyphagous pentatomid pests and illustrates that metabarcoding can deliver a relevant species-level resolution of host plant preference. © 2024 Society of Chemical Industry.

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Molecular gut content analysis via diagnostic PCR or high-throughput sequencing (metabarcoding) of consumers allows unravelling of feeding interactions in a wide range of animals. This is of particular advantage for analyzing the diet of small invertebrates living in opaque habitats such as the soil. Due to their small body size, which complicates dissection, microarthropods are subjected to whole-body DNA extraction-step before their gut content is screened for DNA of their food. This poses the problem that body surface contaminants, such as fungal spores may be incorrectly identified as ingested food particles for fungivorous species. We investigated the effectiveness of ten methods for body surface decontamination in litter-dwelling oribatid mites using Steganacarus magnus as model species. Furthermore, we tested for potential adverse effects of the decontamination techniques on the molecular detection of ingested prey organisms. Prior to decontamination, oribatid mites were fed with an oversupply of nematodes (Plectus sp.) and postmortem contaminated with fungal spores (Chaetomium globosum). We used diagnostic PCR with primers specific for C. globosum and Plectus sp. to detect contaminants and prey, respectively. The results suggest that chlorine bleach (sodium hypochloride, NaClO, 5%) is most efficient in removing fungal surface contamination without significantly affecting the detection of prey DNA in the gut. Based on these results, we provide a standard protocol for efficient body surface decontamination allowing to trace the prey spectrum of microarthropods using molecular gut content analysis.

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In many agroecosystems, brown marmorated stink bugs (Halyomorpha halys) (Hemiptera: Pentatomidae) are polyphagous pests that cause significant economic losses to numerous crops every year. Insectivorous birds may provide a means of sustainable predation of invasive pests, such as H. halys. In forest margins surrounding peach, pecan, and interplanted peach-pecan orchards, we monitored H. halys populations with pheromone-baited traps, mist-netted birds, and collected avian fecal samples for molecular gut content analysis. We screened 257 fecal samples from 19 bird species for the presence of H. halys DNA to determine whether birds provide the biological control of this pest. Overall, we found evidence that four birds from three species consumed H. halys, including Northern cardinal (Cardinalis cardinalisis), Tufted titmouse (Baeolophus bicolor), and Carolina wren (Thryothorus ludovicianus). Halyomorpha halys captured in traps increased over time but did not vary by orchard type. Although incidence of predation was low, this may be an underestimate as a result of our current avian fecal sampling methodology. Because birds are members of the broader food web, future studies are needed to understand avian ecosystem services, especially in terms of pest control, including H. halys and other pest species.

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The Hibiscus mealybug, Nipaecoccus viridis (Newstead), has recently established in Florida citrus and become a pest of concern given secondary pest outbreaks associated with management of citrus greening disease. Chemical controls used to manage other citrus arthropod pests are not as effective against N. viridis due to its waxy secretions, clumping behavior, and induced cellular changes to host plant tissue which increase microhabitats. Populations of this mealybug pest are regulated by natural enemies in its native region, but it remains unclear if resident natural enemies in Florida citrus could similarly suppress N. viridis populations. This investigation: 1) established species-specific primers for N. viridis based on the mitochondrial gene Cytochrome-oxidase 1 (COI), 2) determined duration of N. viridis DNA detectability in a known predator, the mealybug destroyer (Cryptolaemus montrouzieri Mulsant), by using identified primers in molecular gut content analysis, and 3) screened field-collected predators for the presence of N. viridis DNA. The detection rate of N. viridis DNA was >50% at 36 h after adult C. montrouzieri feeding but DNA was no longer detectable by 72 h after feeding. Field-collected predators were largely comprised of spiders, lacewings, and C. montrouzieri. Spiders, beetles (primarily C. montrouzieri), and juvenile lacewings were the most abundant predators of N. viridis, with 17.8, 43.5, and 58.3 of field-collected samples testing positive for N. viridis DNA, respectively. Our results indicate that Florida citrus groves are hosts to abundant predators of N. viridis and encourage the incorporation of conservation or augmentative biological control for management of this pest.

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Wolf spiders are typically the most common group of arthropod predators on both lake and marine shorelines because of the high prey availability in these habitats. However, shores are also harsh environments due to flooding and, in proximity to marine waters, to toxic salinity levels. Here, we describe the spider community, prey availabilities, and spider diets between shoreline sites with different salinities, albeit with comparatively small differences (5‰ vs. 7‰). Despite the small environmental differences, spider communities between lower and higher saline sites showed an almost complete species turnover. At the same time, differences in prey availability or spider gut contents did not match changes in spider species composition but rather changed with habitat characteristics within a region, where spiders collected at sites with thick wrack beds had a different diet than sites with little wrack. These data suggest that shifts in spider communities are due to habitat characteristics other than prey availabilities, and the most likely candidate restricting species in high salinity would be saline sensitivity. At the same time, species absence from low-saline habitats remains unresolved.

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As one of the most abundant predators of insects in terrestrial ecosystems, spiders have long received much attention from agricultural scientists and ecologists. Do spiders have a certain controlling effect on the main insect pests of concern in farmland ecosystems? Answering this question requires us to fully understand the prey spectrum of spiders. Next-generation sequencing (NGS) has been successfully employed to analyze spider prey spectra. However, the high sequencing costs make it difficult to analyze the prey spectrum of various spider species with large samples in a given farmland ecosystem. We performed a comparative analysis of the prey spectra of Ovia alboannulata (Araneae, Lycosidae) using NGS with individual and mixed DNA samples to demonstrate which treatment was better for determining the spider prey spectra in the field. We collected spider individuals from tea plantations, and two treatments were then carried out: (1) The DNA was extracted from the spiders individually and then sequenced separately (DESISS) and (2) the DNA was extracted from the spiders individually and then mixed and sequenced (DESIMS). The results showed that the number of prey families obtained by the DESISS treatment was approximately twice that obtained by the DESIMS treatment. Therefore, the DESIMS treatment greatly underestimated the prey composition of the spiders, although its sequencing costs were obviously lower. However, the relative abundance of prey sequences detected in the two treatments was slightly different only at the family level. Therefore, we concluded that if our purpose were to obtain the most accurate prey spectrum of the spiders, the DESISS treatment would be the best choice. However, if our purpose were to obtain only the relative abundance of prey sequences of the spiders, the DESIMS treatment would also be an option. The present study provides an important reference for choosing applicable methods to analyze the prey spectra and food web compositions of animal in ecosystems.

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The invasive Drosophila suzukii feeds and reproduces on various cultivated and wild fruits and moves between agricultural and semi-natural habitats. Hedges in agricultural landscapes play a vital role in the population development of D. suzukii, but also harbor a diverse community of natural enemies. We investigated predation by repeatedly exposing cohorts of D. suzukii pupae between June and October in dry and humid hedges at five different locations in Switzerland. We sampled predator communities and analyzed their gut content for the presence of D. suzukii DNA based on the COI marker. On average, 44% of the exposed pupae were predated. Predation was higher in dry than humid hedges, but did not differ significantly between pupae exposed on the ground or on branches and among sampling periods. Earwigs, spiders, and ants were the dominant predators. Predator communities did not vary significantly between hedge types or sampling periods. DNA of D. suzukii was detected in 3.4% of the earwigs, 1.8% of the spiders, and in one predatory bug (1.6%). While the molecular gut content analysis detected only a small proportion of predators that had fed on D. suzukii, overall predation seemed sufficient to reduce D. suzukii populations, in particular in hedges that provide few host fruit resources.

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BACKGROUND: Modern pest control management systems are based on the support of naturally occurring arthropod predators, as it has been shown that such predators offer an important ecosystem service. However, most naturally occurring arthropod predators are generalists (euryphagous). Their role in the biological control of specific pests has been recognized but remains poorly studied. Here, we focused on the naturally occurring arthropod predators of psyllids - the main insect pest of pear trees. We investigated the abundance of psyllids and all of their potential enemies in an abandoned pear orchard on a weekly basis from early spring to early summer. In addition, employing polymerase chain reaction diagnostics and specific primers, we investigated the predation rate on psyllids in all predators collected. RESULTS: We found four predatory groups: spiders were the most abundant (60%, N = 756), followed by coccinellid beetles, anthocorid bugs and cantharid beetles. Anthocorids and spiders had the highest predation rates among the predatory groups. Among spiders, >50% of foliage-dwelling spiders (belonging to the genera Philodromus and Clubiona; N = 206) were positive for psyllids and showed a numerical response to the abundance of psyllids. CONCLUSION: We conclude that foliage-dwelling spiders are, of the four groups, the most important natural enemies of psyllids on pear trees during spring in Central Europe, as they outnumber specialized Anthocoris bugs. © 2021 Society of Chemical Industry.

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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.

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BACKGROUND: Generalist predators play a key role in the biocontrol of insect pests in agricultural systems. However, predators are subject to frequent mortality events due to periodic disturbance regimes such as crop planting and harvest, which inevitably affect the population development of predators. Conservation of predators in this critical period is important for double-cropping systems such as winter wheat and summer maize, the most widely used cropping system in North China. RESULTS: Planting Cnidium monnieri flower strips at field borders could not only serve as a bridge habitat to conserve the dominant predator Propylaea japonica in wheat fields during harvest but also help the predator immigrate to adjacent maize fields. The predator abundance was 7-fold higher on flower strips than that on natural vegetation strips during the wheat postharvest period and before the maize plant emergence for about a month, and its abundance in maize fields planted with flower strips was nearly 2-fold higher than that in maize fields planted with natural vegetation strips. Moreover, 77.56% of predators that entered maize fields were proven to originate from flower strips. CONCLUSION: Our findings provided evidence that manipulating flower strips as a bridge habitat in wheat-maize rotation fields could conserve P. japonica during crop phenophase changes, and we quantitatively testified that the proportion of this predator in maize fields derived from flower strips. In practice, such a strategy may also be applied in other double-cropping and triple-cropping systems. © 2020 Society of Chemical Industry.

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Detection of host plant DNA from sap-feeding insects can be challenging due to potential low concentration of ingested plant DNA. Although a few previous studies have demonstrated the possibility of detecting various fragments of plant DNA from some sap-feeders, there are no protocols available for potato leafhopper, Empoasca fabae (Harris) (Hemiptera: Cicadellidae), a significant agricultural pest. In this study we focused on optimizing a DNA-based method for host plant identification of E. fabae and investigating the longevity of the ingested plant DNA as one of the potential applications of the protocol. We largely utilized and modified our previously developed PCR-based method for detecting host plant DNA from grasshopper and the spotted lanternfly gut contents. We have demonstrated that the trnL (UAA) gene can be successfully utilized for detecting ingested host plant DNA from E. fabae and determining plant DNA longevity. The developed protocol is a relatively quick and low-cost method for detecting plant DNA from E. fabae. It has a number of important applications-from determining host plants and dispersal of E. fabae to developing effective pest management strategies.

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While carabid beetles have been shown to feed on a variety of crop pests, little is known about their species assemblages in US annual ryegrass crops, where invertebrate pests, particularly slugs, lepidopteran larvae and craneflies, incur major financial costs. This study assesses the biological control potential of carabid beetles for autumn- and winter-active pests in annual ryegrass grown for seed by: (a) investigating the spatial and temporal overlap of carabids with key pests; and (b) molecular gut content analysis using qPCR. Introduced Nebria brevicollis was the only common carabid that was active during pest emergence in autumn, with 18.6% and 8.3% of N. brevicollis collected between September and October testing positive for lepidopteran and cranefly DNA, respectively, but only 1.7% testing positive for slug DNA. While pest DNA was also detected in the guts of the other common carabid species-Agonum muelleri, Calosoma cancellatum and Poecilus laetulus-these were active only during spring and summer, when crop damage by pests is less critical. None of the four carabid species was affected by disk tilling and only N. brevicollis was significantly associated with a vegetated field margin. However, as its impact on native ecosystems is unknown, we do not recommend managing for this species.

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Studies of predation can contribute greatly to understanding predator-prey relationships and can also provide integral knowledge concerning food webs and multi-trophic level interactions. Both conventional polymerase chain reaction (cPCR) and quantitative PCR (qPCR) have been employed to detect predation in the field because of their sensitivity and reproducibility. However, to date, few studies have been used to comprehensively demonstrate which method is more sensitive and reproducible in studies of predation. We used a Drosophila melanogaster-specific DNA fragment (99 bp) to construct a tenfold gradient dilution of standards. Additionally, we obtained DNA samples from Pardosa pseudoannulata individuals that fed on D. melanogaster at various time since feeding. Finally, we compared the sensitivity and reproducibility between cPCR and qPCR assays for detecting DNA samples from feeding trials and standards. The results showed that the cPCR and qPCR assays could detect as few as 1.62 × 103 and 1.62 × 101 copies of the target DNA fragment, respectively. The cPCR assay could detect as few as 48 hr post-feeding of the target DNA fragment. But the qPCR assay showed that all spiders were positive after consuming prey at various time intervals (0, 24, 48, 72, and 96 hr). A smaller proportion of the technical replicates were positive using cPCR, and some bands on the agarose gel were absent or gray, while some were white and bright for the same DNA samples after amplification by cPCR. By contrast, a larger proportion of the technical replicates were positive using qPCR and the coefficients of variation of the Ct value for the three technical replicates of each DNA sample were less than 5%. These data showed that qPCR was more sensitive and highly reproducible in detecting such degraded DNA from predator's gut. The present study provides an example of the use of cPCR and qPCR to detect the target DNA fragment of prey remains in predator's gut.

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The data presented here are related to the article entitled "Soil functions are affected by transition from conventional to organic mulch-based cropping system"[1]. Data were collected in 2016 in a processing tomato field located near Perugia, Italy. In details, data were collected in three differently managed processing tomato cropping systems: conventional integrated (INT); traditional organic with cover crops and conventional tillage (ORG); and organic coupled with conservation agriculture, with mulch-based cover crop and no-tillage (ORG+). We report data on the impact of each cropping system on crop biomass and yield, soil physicochemical properties, size and structure of soil microbial community, soil invertebrate biodiversity and habitat provision (predator-prey trophic interactions).

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Diaphorina citri Kuwayama is the vector of the pathogenic bacteria Candidatus Liberibacter spp., the causative agent of Huanglongbing (HLB), the most serious disease of citrus worldwide. Because predatory insects have been historically neglected in biological control programs for D. citri, the impact of generalist predators on D. citri population densities is little understood. A useful tool to evaluate the dynamics of predator-prey interactions is molecular analysis of predators' gut content. We constructed a specific molecular marker to detect D. citri DNA in the gut content of predator insects, for use in estimating the predation rate of field-collected predators in citrus orchards on D. citri. Bioassays of the DNA half-life detection time were carried out with two predatory species, the ladybird beetle Hippodamia convergens Guérin-Méneville and the lacewing Chrysoperla externa (Hagen). The D. citri DNA half-life detection time (DT50) was 6.11 h for H. convergens and 5.46 h for C. externa. One hundred and seven field-collected predators were used for gut-content analysis (52 larvae/adults of ladybirds and 55 larvae of lacewings). The assays showed that 17.3% of ladybirds but no lacewings tested positive for D. citri DNA. These results show that generalist predators can contribute to biological control of D. citri and should be considered for use in pest management programs in citrus orchards.

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The contribution of generalist insect predators to the control of the two-spotted spider mite, Tetranychus urticae Koch (Acari: Tetranychidae), an herbivorous pest of many crops, is poorly understood. One of the common insect predators in strawberries is the generalist predatory bug Anthocoris nemorum L. (Hemiptera: Anthocoridae), which has the potential to contribute to the control of pest populations. The feeding of adult A. nemorum on T. urticae was assessed by sampling individuals from an organic strawberry field in Denmark, and using PCR gut content analysis to detect remains of T. urticae within their gut. In the lab, we assessed that the DNA half-life detectability was 21.5 h. Significant numbers of field-collected A. nemorum tested positive for T. urticae prey DNA, with very high numbers in June (62.8%) and August (38.8%). This study presents conclusive evidence that the generalist predator A. nemorum can contribute to the decrease of T. urticae densities in strawberry fields, although the actual contribution in the present study is probably limited because predator populations were relatively low compared to T. urticae. The abundance of T. urticae did not increase significantly during the period of sampling, suggesting that a complex of natural enemies can achieve biological control of T. urticae in protected strawberries.

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To establish if fishes' consumption of jellyfish changes through the year, we conducted a molecular gut-content assessment on opportunistically sampled species from the Celtic Sea in October and compared these with samples previously collected in February and March from the Irish Sea. Mackerel Scomber scombrus were found to feed on hydrozoan jellyfish relatively frequently in autumn, with rare consumption also detected in sardine Sardina pilchardus and sprat Sprattus sprattus. By October, moon jellyfish Aurelia aurita appeared to have escaped predation, potentially through somatic growth and the development of stinging tentacles. This is in contrast with sampling in February and March where A. aurita ephyrae were heavily preyed upon. No significant change in predation rate was observed in S. sprattus, but jellyfish predation by S. scombrus feeding in autumn was significantly higher than that seen during winter. This increase in consumption appears to be driven by the consumption of different, smaller jellyfish species than were targeted during the winter.

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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.

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Terrestrial predators on marine shores benefit from the inflow of organisms and matter from the marine ecosystem, often causing very high predator densities and indirectly affecting the abundance of other prey species on shores. This indirect effect may be particularly strong if predators shift diets between seasons. We therefore quantified the seasonal variation in diet of two wolf spider species that dominate the shoreline predator community, using molecular gut content analyses with general primers to detect the full prey range. Across the season, spider diets changed, with predominantly terrestrial prey from May until July and predominantly marine prey (mainly chironomids) from August until October. This pattern coincided with a change in the spider age and size structure, and prey abundance data and resource selection analyses suggest that the higher consumption of chironomids during autumn is due to an ontogenetic diet shift rather than to variation in prey abundance. The analyses suggested that small dipterans with a weak flight capacity, such as Chironomidae, Sphaeroceridae, Scatopsidae and Ephydridae, were overrepresented in the gut of small juvenile spiders during autumn, whereas larger, more robust prey, such as Lepidoptera, Anthomyidae and Dolichopodidae, were overrepresented in the diet of adult spiders during spring. The effect of the inflow may be that the survival and growth of juvenile spiders is higher in areas with high chironomid abundances, leading to higher densities of adult spiders and higher predation rates on the terrestrial prey next spring.

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