Detection of Rhopalosiphum insertum (apple-grass aphid) predation by the predatory mite Anystis baccarum using molecular gut analysis

Abstract 1 A simple, yet sensitive polymerase chain reaction based technique was developed for the detection of the apple‐grass aphid Rhopalosiphum insertum in the gut of Anystis baccarum, a predatory mite. 2 A range of conserved polymerase chain reaction primers for insect mitochondrial and ribosomal DNA were tested in order to amplify R. insertum DNA. The mitochondrial DNA primers LrRNAR2 + N1F1, amplified a region between the ND1 and large subunit RNA genes. 3 DNA sequencing of the R. insertum ND1‐LRNA polymerase chain reaction product allowed aphid‐specific polymerase chain reaction primers to be designed. These amplified a 283‐bp product from individual aphids. No polymerase chain reaction product was amplified from individual A. baccarum. 4 Using the aphid‐specific primers against A. baccarum fed on R. insertum, the diagnostic 283‐bp product was amplified. 5 Two restriction enzymes (RsaI and AluI) produced patterns that allowed unambiguous identification of R. insertum DNA from that of Macrosiphum euphorbiae and Myzus persicae.     

Determining the frequency of heteropteran predation on sweetpotato whitefly and pink bollworm using multiple ELISAs

The gut contents of field-collected, predaceous Heteroptera were assayed for the presence of eggs of the sweetpotato whitefly,Bemisia tabaci Gennadius (Homoptera: Aleyrodidae) and the pink bollworm,Pectinophora gossypiella (Saunders) (Lepidoptera: Gelechiidae) using multiple enzyme-linked immunosorbent assays (ELISAs). Of seven species examined,Geocoris species andOrius tristicolor (Say) were the most frequent predators of sweetpotato whitefly with 32–39% of the individuals tested over the whole season scoring positive for whitefly antigens. With the exception ofLygus hesperus Knight, a major insect pest as well as a predator, the frequency of predation on pink bollworm eggs was much lower (0.7–14.3% positive over the season). Relatively few predators tested positive for both antigens (0.3–12.5%).     

Testing the validity of functional response models using molecular gut content analysis for prey choice in soil predators

Analysis of predator–prey interactions is a core concept of animal ecology, explaining structure and dynamics of animal food webs. Measuring the functional response, i.e. the intake rate of a consumer as a function of prey density, is a powerful method to predict the strength of trophic links and assess motives of prey choice, particularly in arthropod communities. However, due to their reductionist set‐up, functional responses, which are based on laboratory feeding experiments, may not display field conditions, possibly leading to skewed results. Here, we tested the validity of functional responses of centipede predators and their prey by comparing them with empirical gut content data from field‐collected predators. Our predator–prey system included lithobiid and geophilomorph centipedes, abundant and widespread predators of forest soils and their soil‐dwelling prey. First, we calculated the body size‐dependent functional responses of centipedes using a published functional response model in which we included natural prey abundances and animal body masses. This allowed us to calculate relative proportions of specific prey taxa in the centipede diet. In a second step, we screened field‐collected centipedes for DNA of eight abundant soil‐living prey taxa and estimated their body size‐dependent proportion of feeding events. We subsequently compared empirical data for each of the eight prey taxa, on proportional feeding events with functional response‐derived data on prey proportions expected in the gut, showing that both approaches significantly correlate in five out of eight predator–prey links for lithobiid centipedes but only in one case for geophilomorph centipedes. Our findings suggest that purely allometric functional response models, which are based on predator–prey body size ratios are too simple to explain predator–prey interactions in a complex system such as soil. We therefore stress that specific prey traits, such as defence mechanisms, must be considered for accurate predictions.     

Development of sequence amplified characterized region (SCAR) markers of helicoverpa armigera: a new polymerase chain reaction-based technique for predator gut analysis

A method is described for the development of DNA markers for detection of Helicoverpa armigera (Hubner) (Lepidoptera: Noctuidae) in predator gut analysis, based on sequence characterized amplified regions (SCARs) derived from a randomly amplified polymorphic DNA (RAPD) band. A 1200-bp DNA fragment of H. armigera, absent in the predator band pattern and in other closely related prey species, was identified by RAPD analysis. This fragment was cloned and its extremes sequenced to design extended strand-specific 20-mer oligonucleotide primers. Three pairs of SCAR primers, which amplified three different DNA fragments, were used to study the effect of fragment length on detection of prey in the predator gut. Using the pair of primers that amplified the longest fragment of H. armigera DNA, a single band of 1100 bp was obtained, but its detection was not possible in the predator gut. Detection of the ingested prey was possible with the other two pairs of SCAR primers, obtaining bands of 600 and 254 bp, respectively. Detection of H. armigera DNA in the gut of the predator Dicyphus tamaninii was evaluated immediately after ingestion (t = 0) and after 4 h. Detection of H. armigera DNA after 4 h was only possible using the pair of primers that amplified the shortest fragment (254 bp). The test for specificity, using these last pair of primers, showed that H. armigera was the only species detected. The detection threshold was defined at a 1:8192 dilution of a H. armigera whole egg in all samples.     

Application of machine learning to identify predators of stocked fish in Lake Ontario: using acoustic telemetry predation tags to inform management

Understanding predator–prey interactions and food web dynamics is important for ecosystem-based management in aquatic environments, as they experience increasing rates of human-induced changes, such as the addition and removal of fishes. To quantify the post-stocking survival and predation of a prey fish in Lake Ontario, 48 bloater Coregonus hoyi were tagged with acoustic telemetry predation tags and were tracked on an array of 105 acoustic receivers from November 2018 to June 2019. Putative predators of tagged bloater were identified by comparing movement patterns of six species of salmonids (i.e., predators) in Lake Ontario with the post-predated movements of bloater (i.e., prey) using a random forests algorithm, a type of supervised machine learning. A total of 25 bloater (53% of all detected) were consumed by predators on average (± S.D. ) 3.1 ± 2.1 days after release. Post-predation detections of predators occurred for an average (± S.D. ) of 78.9 ± 76.9 days, providing sufficient detection data to classify movement patterns. Tagged lake trout Salvelinus namaycush provided the most reliable classification from behavioural predictor variables (89% success rate) and was identified as the main consumer of bloater (consumed 50%). Movement networks between predicted and tagged lake trout were significantly correlated over a 6 month period, supporting the classification of lake trout as a common bloater predator. This study demonstrated the ability of supervised learning techniques to provide greater insight into the fate of stocked fishes and predator–prey dynamics, and this technique is widely applicable to inform future stocking and other management efforts.     

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.     

Knowing your enemies: Integrating molecular and ecological methods to assess the impact of arthropod predators on crop pests

The importance of natural enemies as the foundation of integrated pest management (IPM) is widely accepted, but few studies conduct the manipulative field experiments necessary to directly quantify their impact on pest populations in this context. This is particularly true for predators. Studying arthropod predator–prey interactions is inherently difficult: prey items are often completely consumed, individual predator–prey interactions are ephemeral (rendering their detection difficult) and the typically fluid or soft‐bodied meals cannot be easily identified visually within predator guts. Serological techniques have long been used in arthropod predator gut‐contents analysis, and current enzyme linked immunosorbent assays (ELISA) are highly specific and sensitive. Recently, polymerase chain reaction (PCR) methods for gut‐contents analysis have developed rapidly and they now dominate the diagnostic methods used for gut‐contents analysis in field‐based research. This work has identified trophic linkages within food webs, determined predator diet breadth and preference, demonstrated the importance of cannibalism and intraguild predation within and between certain taxa, and confirmed the benefits (predator persistence) and potential disadvantages (reduced feeding on pest species) of the availability of alternative nonpest prey. Despite considerable efforts to calibrate gut‐contents assays, these methods remain qualitative. Available techniques for predator gut‐contents analysis can provide rapid, accurate, cost‐effective identification of predation events. As such, they perfectly compliment the ecological methods developed to directly assess predator impacts on prey populations but which are imperfect at identifying the key predators. These diagnostic methods for gut‐contents analysis are underexploited in agricultural research and they are almost never applied in unison with the critical field experiments to measure predator impact. This paper stresses the need for a combined approach and suggests a framework that would make this possible, so that appropriate natural enemies can be targeted in conservation biological control.