Molecular analysis of predation on parasitized hosts

Predation on parasitized hosts can significantly affect natural enemy communities, and such intraguild predation may indirectly affect control of herbivore populations. However, the methodological challenges for studying these often complex trophic interactions are formidable. Here, we evaluate a DNA-based approach to track parasitism and predation on parasitized hosts in model herbivore-parasitoid-predator systems. Using singleplex polymerase chain reaction (SP-PCR) to target mtDNA of the parasitoid only, and multiplex PCR (MP-PCR) to additionally target host DNA as an internal amplification control, we found that detection of DNA from the parasitoid, Lysiphlebus testaceipes, in its aphid host, Aphis fabae, was possible as early as 5 min. post parasitism. Up to 24 h post parasitism SP-PCR proved to be more sensitive than MP-PCR in amplifying parasitoid DNA. In the carabid beetles Demetrias atricapillus and Erigone sp. spiders, fed with aphids containing five-day-old parasitoids, parasitoid and aphid DNA were equally detectable in both predator groups. However, when hosts containing two-day-old parasitoids were fed to the predators, detection of parasitoid prey was possible only at 0 h (immediately after consumption) and up to 8 h post consumption in carabids and spiders, respectively. Over longer periods of time, post-feeding prey detection success was significantly higher in spiders than in carabid beetles. MP-PCR, in which parasitoid and aphid DNA were simultaneously amplified, proved to be less sensitive at amplifying prey DNA than SP-PCR. In conclusion, our study demonstrates that PCR-based parasitoid and prey detection offers an exciting approach to further our understanding of host-parasitoid-predator interactions.     

Secondary predation: quantification of food chain errors in an aphid-spider-carabid system using monoclonal antibodies

"Secondary predation" occurs when one predator feeds on a second predator, which has in turn eaten a target prey. Detection of prey remains within predators using monoclonal antibodies cannot distinguish between primary and secondary predation, potentially leading to quantitative and qualitative food chain errors. We report the first fully replicated experiments to measure secondary predation effects, using an aphid-spider-carabid system. Aphids, Sitobion avenae, were fed to spiders, Lepthyphantes tenuis, which were allowed to digest their prey for a range of time intervals. The spiders were then fed to carabids, Poecilus (=Pterostichus) cupreus, which were again allowed to digest their prey for set periods. The anti-aphid monoclonal antibody used to identify S. avenae remains in P. cupreus was one that detected an epitope that increased in availability over the first few hours of digestion, amplifying the signal, extending detection periods and thus increasing the chances of detecting secondary predation. Despite this, and the fact that spiders are known to digest their prey more slowly than many other predators, detection of secondary predation was only possible if the carabids were killed immediately after consuming at least two spiders which were, in turn, eaten immediately after consuming aphids. As this scenario is unlikely to occur frequently in the field it was concluded that secondary predation is unlikely to be a serious source of error during field studies.     

The significance of facultative scavenging in generalist predator nutrition: detecting decayed prey in the guts of predators using PCR

Gut-content analyses using molecular techniques are an effective approach to quantifying predator-prey interactions. Predation is often assumed but scavenging is an equally likely route by which animal DNA enters the gut of a predator/scavenger. We used PCR (polymerase chain reaction) to detect scavenged material in predator gut homogenates. The rates at which DNA in decaying slugs (Mollusca: Pulmonata) and aphids (Homoptera: Aphididae) became undetectable were estimated. The detectability of DNA from both carrion types in the guts of the generalist predator Pterostichus melanarius (Coleoptera: Carabidae) was then determined. The effects of carrion age and weight, as well as beetle sex, on detection periods, were quantified. Laboratory trials measured prey preference of beetles between live and decaying prey. Further experiments measured, for the first time, feeding by P. melanarius on dead slugs and aphids directly in the field. In both field and laboratory, P. melanarius preferentially fed on dead prey if available, but preference changed as the prey became increasingly decayed. Disappearance rates for slug carrion in wheat fields and grasslands were estimated and P. melanarius was identified as the main scavenger. Comparison of the retention time for dead slugs in the field, with the detection period for decaying slug material in the guts of the predators, showed that PCR-based techniques are not able to distinguish between predated and scavenged food items. This could potentially lead to overestimation of the impact of predation on slugs (and other prey) by carabids. Possible implications of facultative scavenging by invertebrate predators for biocontrol and food-web research are discussed.     

Aphid cards – Useful model for assessing predation rates or bias prone nonsense?

Predation on pest organisms is an essential ecosystem function supporting yields in modern agriculture. However, assessing predation rates is intricate, and they can rarely be linked directly to predator densities or functions. We tested whether sentinel prey aphid cards are useful tools to assess predation rates in the field. Therefore, we looked at aphid cards of different sizes on the ground level as well as within the vegetation. Additionally, by trapping ground-dwelling predators, we examined whether obtained predation rates could be linked to predator densities and traits. Predation rates recorded with aphid cards were independent of aphid card size. However, predation rates on the ground level were three times higher than within the vegetation. We found both predatory carabid activity densities as well as community weighted mean body size to be good predictors for predation rates. Predation rates obtained from aphid cards are stable over card type and related to predator assemblages. Aphid cards, therefore, are a useful, efficient method for rapidly assessing the ecosystem function predation. Their use might especially be recommended for assessments on the ground level and when time and resource limitations rule out more elaborate sentinel prey methods using exclosures with living prey animals.     

The influence of time and temperature on molecular gut content analysis： Adalia bipunctata fed with Rhopalosiphum padi

Gut content analysis is a useful tool when studying arthropod predator-prey interactions. We used polymerase chain reaction （PCR） technique to examine how detection of prey DNA in the gut content of predators was influenced by digestion time and temperature. Such knowledge is critical before applying PCR-based gut content analysis to field collected predators. Larvae of the two-spotted ladybeetle （Adalia bipunctata L.） were fed with the bird cherry-oat aphid （Rhopalosiphum padi L.） at either 21℃ or 14℃. After consuming one aphid, the predators were allowed to digest the prey for a range of time periods up to 24 hours. The influence of temperature on A. bipunctata feeding behavior was also recorded. From the fed larvae, total DNA was extracted and PCR reactions with R. padi specific primers were run. The number ofA. bipunctata that tested positive for R. padi DNA was negatively related to the length of digestion time. Temperature influenced larval feeding behavior but did not have a significant effect on R. padi DNA detection. After pooling the data from both temperature treatments we estimated the time point when R. padi DNA could be amplified from 50% of the fed A. bipunctata by PCR to be 4.87 hours. With such a rapid decrease in prey DNA detection success, positive PCR reactions will most likely be the result of predation events occurring shortly before capture. If a defined digestion temperature range has proven not to influence prey detection, PCR data obtained from predators collected within that particular range can be interpreted in the same way.     

Evaluation of temperature gradient gel electrophoresis for the analysis of prey DNA within the guts of invertebrate predators

The utility of temperature gradient gel electrophoresis (TGGE) as a means of analysing the gut contents of predators was evaluated. Generalist predators consume multiple prey species and a species-specific primer approach may not always be a practical means of analysing predator responses to prey diversity in complex and biodiverse ecosystems. General invertebrate primers were used to amplify the gut contents of predators, generating banding patterns that identified component prey remains. There was no evidence of dominance of the polymerase chain reaction (PCR) by predator DNA. When applied to field samples of the carabid predator Pterostichus melanarius (Illiger) nine banding patterns were detected, including one for aphids. To further distinguish between species, group-specific primers were designed to separate species of earthworm and aphid. TGGE of the earthworm PCR products generated banding patterns that varied with haplotype in some species. Aphid and earthworm DNA could be detected in the guts of carabids for up to 24 h using TGGE. In P. melanarius, with low numbers of prey per insect gut (mean<3), interpretation of banding patterns proved to be tractable. Potential problems of interpretation of TGGE gels caused by multiple prey bands, cryptic bands, haplotype variation, taxonomic uncertainties (especially with regard to earthworms), secondary predation, scavenging and presence of parasites and parasitoids in the prey or the predators, are discussed. The results suggest that PCR, using combinations of general invertebrate and group-specific primers followed by TGGE, provides a potentially useful approach to the analysis of multiple uncharacterized prey in predators.     

Tracking aphid predation by lycosid spiders in spring-sown cereals using PCR-based gut-content analysis

Detection of prey DNA-remains in arthropod predators by polymerase chain reaction (PCR) is useful when investigating food webs. In this study we estimated how long after a feeding event it was possible to detect mitochondrial COII DNA (331 bp) from an important aphid pest, Rhopalosiphum padi (Homoptera: Aphididae), in spiders of the genus Pardosa (Araneae: Lycosidae). Following laboratory evaluations we tested spiders collected in spring-sown cereals for aphid predation during two seasons. Aphids were digested rapidly in laboratory-fed predators and the time point when prey DNA could be amplified from 50% of the spiders was estimated to be 3.7 h. A total of 372 field- collected predators were analyzed by PCR and despite low aphid densities many spiders (26% in 2004 and 19% in 2005) tested positive for R. padi, indicating consumption of at least one aphid within a few hours before capture. The percentage of spiders that tested positive for R. padi DNA varied considerably between fields and logistic regression analysis revealed that the probability of detecting aphid DNA was significantly influenced by location and year. We conclude that Pardosa spiders, under certain conditions, frequently feed on R. padi and deserve special attention in conservation biological control.     

Temperature and prey capture: opposite relationships in two predator taxa

Abstract 1. All other things equal, predator capture rates are expected to depend on encounter rate with prey, prey escape capability (including prey defences), and on predator agility. Ectotherm predators and their prey both respond to increasing temperature by increased activity, i.e. predators increase their search area and prey may enhance their escape capability. This means that, as temperature changes, the ability of a predator to catch prey will decrease, increase, or remain unchanged depending on the relative effect of temperature on predator and prey. Their responses may further be differentially moulded by light conditions depending on whether the predator is diurnally or nocturnally active. It was hypothesised that flying Diptera are vulnerable to carabid beetles only at low temperatures and over the full temperature range for spiders because carabids, in contrast to spiders, are not built to catch swiftly moving prey. 2. The first experiment examined the spontaneous locomotor activity of the predators and of fruit flies at different temperatures (5, 10, 15, 20, 25, and 30 °C) and light conditions (light, dark). A second experiment examined the effect of temperature and light on the predation rate of two carabid beetles (Pterostichus versicolor and Calathus fuscipes) and two spiders (Clubiona phragmitis and Pardosa prativaga) using fruit flies (Drosophila melanogaster) as prey. 3. All four predators and the fruit fly increased their locomotory activity at higher temperatures. Activity of the carabid beetles peaked at intermediate temperatures; spiders and fruit flies were most active at the highest temperatures. Predation rate of the spiders increased with temperature whereas the beetles caught flies only at low temperatures (5 and 10 °C). 4. Diurnal variation in temperature may bring different prey groups within the set of potential prey at different times of the day or at different seasons. The ability of many carabid beetles to forage at low temperatures may have nutritional benefits and increases the diversity of interactions in terrestrial food webs.     

Potential cotton aphid,Aphis gossypii,population suppression by arthropod predators in upland cotton

The cotton aphid, Aphis gossypii Glover, predation rate of convergent lady beetle, Hippodamia convergens Guerin‐Meneville, was determined by assigning a single predator randomly to each of four prey density treatments in the laboratory. Prey densities included 25, 50, 100, and 200 aphids per Petri dish arena. Predation response was recorded at 1, 4, 8, 16, 24, and 48 h after assigning predators to their prey treatments. Rate of consumption increased through time, with all 25 aphids consumed during the first 4 h of the experiment. At the highest density, adult lady beetle consumed on average 49, 99, 131, 163, 183, and 200 aphids within 1, 4, 8, 16, 24 and 48 h, respectively. Predators showed a curvilinear feeding response in relation to total available time, indicating that convergent lady beetles have the potential to suppress larger populations of aphids through continuous feeding by regulating their predation efficiency during feeding. The analysis of age‐specific mortality in absence of prey revealed that lady beetles could survive for an extended period of time (more than 2 weeks) without prey. The ability of a predator to survive without prey delays or prevents the rebound of pest populations that is a significant factor in natural biological control. A two‐year field sampling of 10 cotton arthropod predator species showed that spiders (27%) were the most dominant foliage dwelling predators in the Texas High Plains cotton followed by convergent lady beetles (23.5%), hooded beetles (13.5%), minute pirate bugs (11%), green lacewings (9.5%), bigeyed bugs (7.5%), scymnus beetles (3%), soft‐winged flower beetles (2%), damsel bugs (1.5%), and assassin bugs (1.5%). A field cage study showed that one H. convergens adult per plant released at prey density of one aphid per leaf kept the aphid population below economic threshold for the entire growing season.     

A field experiment on the effectiveness of spiders and carabid beetles as biocontrol agents in soybean

1 Spiders and carabid beetles are abundant generalist predators that prey upon insect pests of soybean. A field experiment was conducted to determine the impact of spiders and carabids on soybean yield. Prior to planting, three 7 × 7 m plots were fenced in order to reduce spider and carabid immigration. Carabids that emerged within the plots were not removed, but spiders that ballooned into these predator‐reduction plots or that entered by climbing the fence were removed by pitfall trapping and searching the vegetation. Three unmanipulated, unfenced plots served as the control treatment. 2 Densities of spiders on soybean vegetation, and activity‐densities of spiders and carabids determined by pitfall trapping, were c. 75% lower in the spider‐carabid reduction treatment than in control plots. Despite clear differences between treatments in numbers and activity of these major generalist predators, the weight of soybeans harvested did not differ between control and spider‐carabid reduction plots. 3 Paralleling the absence of an effect of predator reduction on soybean yield was the absence of any significant difference between treatments in densities of whiteflies (Aleyrodidae), leafhoppers (Cicadellidae), thrips (Thysanoptera), Lepidoptera larvae and herbivorous Coleoptera. 4 Our experiment provides no evidence that spiders and carabid beetles at ambient densities affect soybean yield. Low populations of pest species or low predation pressure on soybean pests by spiders and carabids at the ambient densities of this experiment could be responsible for this result.     

Collembola as alternative prey sustaining spiders in arable ecosystems: prey detection within predators using molecular markers

Collembola comprise a major source of alternative prey to linyphiid spiders in arable fields, helping to sustain and retain these predators as aphid control agents within the crop. Polymerase chain reaction primers were developed for the amplification, from spider gut samples, of DNA from three of the most abundant species of Collembola in wheat crops in Europe, namely Isotoma anglicana, Lepidocyrtus cyaneus and Entomobrya multifasciata. The primers amplified fragments of the mitochondrial cytochrome oxidase subunit I (COI) gene and were designed following alignment of comparable sequences for a range of predator and prey species. Each of the primer pairs proved to be species-specific to a Collembola species, amplifying DNA fragments from 211 to 276 base pairs in length. Following consumption of a single collembolan, prey DNA was detectable in 100% of spiders after 24 h of digestion. We report the first use of DNA-based techniques to detect predation by arthropods on natural populations of prey in the field. All three species of Collembola were consumed by the spiders. By comparing the ratios of the Collembola species in the field with the numbers of spiders that gave positive results for each of those species, it was possible to demonstrate that the spiders were exercising prey choice. Overall, a single target species of Collembola was eaten by 48% of spiders while a further 16% of spiders contained DNA from two different species of Collembola. Preference was particularly evident for I. anglicana, the species most frequently found in spider guts yet the least numerous of the three target species in the field.     

Monoclonal antibodies reveal the potential of the tetragnathid spider Pachygnatha degeeri (Araneae: Tetragnathidae) as an aphid predator

The drive towards a more sustainable and integrated approach to pest management has engendered a renewed interest in conservation biological control, the role of natural enemy communities and their interactions with prey. Monoclonal antibodies have provided significant advances in enhancing our knowledge of trophic interactions and can be employed to help quantify predation on target species. The tetragnathid spider Pachygnatha degeeri Sundevall was collected from fields of winter wheat in the UK and assayed by ELISA for aphid proteins. It was demonstrated that this spider did not simply consume greater quantities of aphids because it was bigger. In addition, P. degeeri contained significantly greater concentrations of aphid in their guts than other spiders, showing that aphids comprised a greater proportion of their diet. Although P. degeeri constituted only 6% of the spider population numerically, females and males respectively contained 16% and 37% of total aphid proteins within all spiders screened, significantly more than their density would predict. These spiders also preyed upon aphids at a disproportionately high rate in June, during the aphid establishment phase, theoretically the best time for limiting growth in the aphid population. Although less abundant than other generalist predators, the capability of these hunting spiders to consume large numbers of aphids highlights them as a more significant component of the predator complex than had previously been realized. Limitation of aphid numbers early in the year by generalist predators provides more time for the specialist aphid predators and parasitoids to move in.     

Recolonisation and distribution of spiders and carabids in cereal fields after spring sowing

Generalist predators are important for pest suppression during pest establishment because they may occur in the crop before and during pest arrival. However, different crop management practices can have a negative effect on predator populations. If so, there is a need for recolonisation by the predators to the crop fields. An important pest in Sweden is the bird cherry‐oat aphid, Rhopalosiphum padi, which migrates to cereal fields in spring. In turn, many cereal fields are spring sown and thus are disturbed by harrowing and sowing a short time before aphid arrival. In this study, three different questions about the populations of spiders and carabids in spring cereal crops were asked. First, does sowing in spring have a negative effect on the predators present in the fields? Second, if sowing has a negative effect on predator populations, do they recolonise the fields before pest arrival? Third, how are the predators distributed in the fields? We found that most carabids and most lycosid spiders were not affected by sowing and were distributed uniformly in the fields after sowing. Most linyphiid spiders were negatively affected by sowing, but then they recolonised the fields and were uniformly distributed in the fields after recolonisation. Thus, many spiders and carabids are present in the fields after sowing in spring and have the opportunity to suppress aphids during their establishment phase, which in turn may prevent a possible outbreak.     

Habitat complexity dampens selection on prey activity level

Conspecific prey individuals often exhibit persistent differences in behavior (i.e., animal personality) and consequently vary in their susceptibility to predation. How this form of selection varies across environmental contexts is essential to predicting ecological and evolutionary dynamics, yet remains currently unresolved. Here, we use three separate predator–prey systems (sea star–snail, wolf spider–cricket, and jumping spider–cricket) to independently examine how habitat structural complexity influences the selection that predators impose on prey behavioral types. Prior to conducting staged predator–prey interaction encounters, we ran prey individuals through multiple behavioral assays to determine their average activity level. We then allowed individual predators to interact with groups of prey in either open or structurally complex habitats and recorded the number and individual identity of prey that were eaten. Habitat complexity had no effect on overall predation rates in any of the three predator–prey systems. Despite this, we detected a pervasive interaction between habitat structure and individual prey activity level in determining individual prey survival. In open habitats, all predators imposed strong selection on prey behavioral types: sea stars preferentially consumed sedentary snails, while spiders preferentially consumed active crickets. Habitat complexity dampened selection within all three systems, equalizing the predation risk that active and sedentary prey faced. These findings suggest a general effect of habitat complexity that reduces the importance of prey activity level in determining individual predation risk. We reason this occurs because activity level (i.e., movement) is paramount in determining risk within open environments, whereas in complex habitats, other behavioral traits (e.g., escape ability to a refuge) may take precedence.     

Generalist predators disrupt parasitoid aphid control by direct and coincidental intraguild predation

Generalist predators and parasitoids are considered to be important regulators of aphids. The former not only feed on these pests, but might also consume parasitoids at all stages of development. This direct or coincidental interference affects the natural control of aphids, the scale of which is largely unknown, and it has rarely been examined under natural conditions. Here, molecular diagnostics were used to track trophic interactions in an aphid-parasitoid-generalist predator community during the build-up of a cereal aphid population. We found that generalist predators, principally carabid and staphylinid beetles as well as linyphiid spiders, had strong trophic links to both parasitoids and aphids. Remarkably, more than 50% of the parasitoid DNA detected in predators stems from direct predation on adult parasitoids. The data also suggest that coincidental intraguild predation is common too. Generalist predators, hence, disrupt parasitoid aphid control, although the levels at which the predators feed on pests and parasitoids seem to vary significantly between predator taxa. Our results suggest that taxon-specific trophic interactions between natural enemies need to be considered to obtain a more complete understanding of the route to effective conservation biological control.     

Suction sampling as a significant source of error in molecular analysis of predator diets

The molecular detection of predation is a fast growing field, allowing highly specific and sensitive detection of prey DNA within the gut contents or faeces of a predator. Like all molecular methods, this technique is prone to potential sources of error that can result in both false positive and false negative results. Here, we test the hypothesis that the use of suction samplers to collect predators from the field for later molecular analysis of predation will lead to high numbers of false positive results. We show that, contrary to previous published work, the use of suction samplers resulted in previously starved predators testing positive for aphid and collembolan DNA, either as a results of ectopic contamination or active predation in the collecting cup/bag. The contradictory evidence for false positive results, across different sampling protocols, sampling devices and different predator-prey systems, highlights the need for experimentation prior to mass field collections of predators to find techniques that minimise the risk of false positives.     

Sacrificial males: the potential role of copulation and predation in contributing to copepod sex‐skewed ratios

Predation is thought to play a selective role in the emergence of behavioural traits in prey. Differences in behaviour between prey demographics may, therefore, be driven by predation with select components of the population being less vulnerable to predators. While under controlled conditions prey demography has been shown to have consequences for predation success, investigations linking these implications to natural prey population demographics are scarce. Here we assess predator–prey dynamics between notonectid predators (backswimmers) and Lovenula raynerae (Copepoda), key faunal groups in temperate ephemeral pond ecosystems. Using a combination of field and experimental approaches we test for the development and mechanism of predation‐induced sex‐skewed ratios. A natural population of L. raynerae was tracked over time in relation to their predator (notonectid) and prey (Cladocera) numbers. In the laboratory, L. raynerae sex ratios were also assessed over time but in the absence of predation pressure. Predation success and prey performance experiments evaluating differences between L. raynerae male, female, gravid female and copulating pairs exposed to notonectid predation were then examined. Under natural conditions, a female dominated copepod population developed over time and was correlated to predation pressure, while under predator‐free conditions non sex‐skewed prey population demographics persisted. Predator–prey laboratory trials showed no difference in vulnerability and escape performance for male, female and gravid female copepods, but pairs in copula were significantly more vulnerable to predation. This vulnerability was not shared by both sexes, with only female copepods ultimately escaping from successful predation on a mating pair. These results suggest that contact periods during copula may contribute to the development of sex‐skewed copepod ratios over time in ecosystems dominated by hexapod predators. This is discussed within the context of vertebrate and invertebrate predation and how these dissimilar types of predation are likely to have acted as selective pressures for copepod mating systems.     

Can polyphagous predators control the bird cherry-oat aphid (Rhopalosiphum padi) in spring cereals?: A simulation study

Predation by two carabid species,Bembidion lampros (Herbst.) andPterostichus cupreus L., on the cereal aphid,Rhopalsiphum padi L., was described using a discrete event simulation model. The results were incorporated into an aphid population growth model. The aphid model was run using actual temperatures,R. padi emigration rates, field densities and distributions from three years with distinctly different aphid population levels. Predation was incorporated at different time periods. Predation during the arrival of emigrants and establishment of aphid populations had approximately the same effect as predation over the entire aphid period, whereas predation during the 20 days prior to peak had very little effect on resulting peak populations. We conclude that a) yearly levels ofR. padi infestation are mainly determined by the volume and intensity of emigrants; b) peak levels are sensitive to changes in predator density; and c) in some cases an outbreak ofR. padi may be prevented by the action of spring active polyphagous predators alone.     

Optimizing methods for PCR-based analysis of predation

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.     

Spider predation on a mirid pest in Japanese rice fields

Spiders are common generalist predators, and understanding their potential in biological control is important for the development of integrated pest management programs. In this study, predation by three groups of spiders on the mirid bug Stenotus rubrovittatus (Hemiptera: Miridae) in rice paddies was investigated using DNA-based gut-content analysis. A laboratory feeding study revealed that the detection half-lives of bug DNA in the spider gut at 25 °C was 3.4 days for Lycosidae and 1.5 days for Tetragnathidae. Individual spider predation on the mirid bug was investigated by detecting DNA of prey in field-collected spiders. In total, 1199 spiders were assayed from three spider groups: Pirata subpiraticus (Lycosidae), Tetragnatha spp. (Tetra-gnathidae), and Pachygnatha clercki (Tetra-gnathidae), which each differ in their preferred microhabitat as well as their predatory habits. Detection rates of prey DNA in spiders increased significantly with the density of prey across all spider groups. P. subpiraticus and Tetragnatha spp. predation showed a better fit to a saturated response curve to increasing prey density, while P. clercki showed a simple linear relationship with prey density. Densities of alternative prey species did not affect the detection rates of mirids. These results suggest that predation on pests by generalist predators in an agroecosystem is affected not only by prey abundance but also by predator preference for specific prey. Predator preference is therefore an important factor to consider when estimating the role of natural enemies as biological control agents.