DNA methods are useful to identify ingested prey items from the gut of predators, but reliable detection is hampered by low amounts of degraded DNA. PCR-based methods can retrieve minute amounts of starting material but suffer from amplification biases and cross-reactions with the predator and related species genomes. Here, we use PCR-free direct shotgun sequencing of total DNA isolated from the gut of the harlequin ladybird Harmonia axyridis at five time points after feeding on a single pea aphid Acyrthosiphon pisum. Sequence reads were matched to three reference databases: Insecta mitogenomes of 587 species, including H. axyridis sequenced here; A. pisum nuclear genome scaffolds; and scaffolds and complete genomes of 13 potential bacterial symbionts. Immediately after feeding, multicopy mtDNA of A. pisum was detected in tens of reads, while hundreds of matches to nuclear scaffolds were detected. Aphid nuclear DNA and mtDNA decayed at similar rates (0.281 and 0.11 h(-1) respectively), and the detectability periods were 32.7 and 23.1 h. Metagenomic sequencing also revealed thousands of reads of the obligate Buchnera aphidicola and facultative Regiella insecticola aphid symbionts, which showed exponential decay rates significantly faster than aphid DNA (0.694 and 0.80 h(-1) , respectively). However, the facultative aphid symbionts Hamiltonella defensa, Arsenophonus spp. and Serratia symbiotica showed an unexpected temporary increase in population size by 1-2 orders of magnitude in the predator guts before declining. Metagenomics is a powerful tool that can reveal complex relationships and the dynamics of interactions among predators, prey and their symbionts.

Characterizing trophic networks is fundamental to many questions in ecology, but this typically requires painstaking efforts, especially to identify the diet of small generalist predators. Several attempts have been devoted to develop suitable molecular tools to determine predatory trophic interactions through gut content analysis, and the challenge has been to achieve simultaneously high taxonomic breadth and resolution. General and practical methods are still needed, preferably independent of PCR amplification of barcodes, to recover a broader range of interactions. Here we applied shotgun-sequencing of the DNA from arthropod predator gut contents, extracted from four common coccinellid and dermapteran predators co-occurring in an agroecosystem in Brazil. By matching unassembled reads against six DNA reference databases obtained from public databases and newly assembled mitogenomes, and filtering for high overlap length and identity, we identified prey and other foreign DNA in the predator guts. Good taxonomic breadth and resolution was achieved (93% of prey identified to species or genus), but with low recovery of matching reads. Two to nine trophic interactions were found for these predators, some of which were only inferred by the presence of parasitoids and components of the microbiome known to be associated with aphid prey. Intraguild predation was also found, including among closely related ladybird species. Uncertainty arises from the lack of comprehensive reference databases and reliance on low numbers of matching reads accentuating the risk of false positives. We discuss caveats and some future prospects that could improve the use of direct DNA shotgun-sequencing to characterize arthropod trophic networks.

Natural enemy conservation is known to be affected by ecological processes that range from local to landscape scales. At the farm scale, there are cropped and noncropped areas that differ in their management and plant diversity; these differences affect the spatiotemporal dynamics of natural enemies. We investigated how different habitat types can affect the conservation and spatial dynamics of predatory and herbivore insects in organic vegetable crops in Brazil. Insects were simultaneously sampled in two cropped (focal and neighbourhood crops) and two noncropped habitats (fallow and native forests) during five consecutive focal crop cycles. We found a higher species richness of predators and herbivores in noncropped habitats. All of the habitats shared species from both functional groups throughout the year, indicating that species could disperse among habitats. Fallow areas can serve as a source and sink for species migrating to/from cropped habitats where predators and herbivores can numerically increase their populations during the crop cycles. The spatiotemporal dynamics of herbivores and predators depend on the management and maintenance of natural, seminatural and cropped habitats within the farm.

Efeitos Ambientais de Culturas Geneticamente Modificadas Resistentes a InsetosRESUMO -Cultivares transgênicas de várias culturas estão sendo utilizadas em escala comercial em muitos países. A área dedicada ao cultivo com plantas transgênicas resistentes às pragas em todo o mundo alcançou 13 milhões de hectares em 2001. As cultivares transgênicas proporcionam benefícios, mas também apresentam riscos potenciais. As avaliações do seu impacto no ambiente são conduzidas antes da sua aprovação para uso comercial, como requerido pelas normas de biossegurança. Nesta revisão, serão discutidas as conseqüências ecológicas potenciais do uso comercial na agricultura de cultivares geneticamente modificadas que apresentam resistência aos insetos-pragas. Também serão discutidos os impactos ambientais causados pelas mudanças nas práticas agrícolas, identificando-se falhas e oportunidades de pesquisa, considerando-se essa nova ferramenta tecnológica. Os comentários e análises serão baseados no conhecimento atual que se tem dos riscos e beneficios do uso de cultivares resistentes a insetos, geneticamente modificadas, dentro do contexto dos programas de manejo integrado de pragas tradicionais. PALAVRAS-CHAVE: Planta transgênica, Bacillus thuringiensis, organismo não-alvo, fluxo gênico, biossegurança ABSTRACT -Transgenic crops are currently being cultivated on a commercial scale in many countries. The area devoted to transgenic pest resistant varieties worldwide reached 13 million hectares in 2001. These varieties offer valuable benefits but also pose potential risks. Assessments of their impact on the environment are conducted before they are approved for commercial use, as required by the regulatory biosafety frameworks. In this review, we discuss the potential ecological consequences of the commercial use in agriculture of genetically modified insect resistant crops. We also discuss the impacts caused by the change in agricultural practices, and attempt to identify gaps and possible opportunities for research, considering this new technological tool. We based our analysis and comments on the current knowledge of the risks and benefits of these genetically modified insect resistant crops, within the context of traditional insect management strategies.KEY WORDS: Transgenic crop, Bt gene, non-target organism, gene flow, biosafety affect the environment (Dale et al. 2002). The discussions on GM pest resistant crops in many parts of the world has led to questions about their potential impacts on biodiversity, particularly on their effects on non-target organisms, including insect herbivores and natural enemies, and soil microbiota. Further concern arises from the possibility of resistance development in insect pests, which could endanger important natural resources such as the bacterium Bacillus thuringiensis Berliner, a natural microbial insecticide. Another issue frequently cited as a potential risk of pest resistant GM crops is the possible movement of the inserted gene from crops to wild or weedy relatives, and the consequences of...

The Enemy Release Hypothesis (ERH) has been a factor influencing the establishment of some exotic species worldwide. We tested if the ERH could be influencing populations of the exotic predator Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae) in Brazil. During two years, three local predatory coccinellid species and Harmonia axyridis were sampled in Brassica oleracea in six organic farms in the midwest of the country. Parasitoids were detected by rearing (emerged) and qPCR Melting Curve Analysis (unemerged). Host density-dependence and spillovers were used to evaluate coccinellid population effects of parasitism. Six parasitoid species were found, including two new records for Brazil. Although Harmonia axyridis harbored fewer parasitoid species, parasitism was not lower and parasitoid densitydependence was not weaker on it compared to the local species. Spillovers among hosts occurred both onto Harmonia axyridis and from it. The ERH was rejected for these parasitoids and, therefore, it did not contribute to the invasiveness of Harmonia axyridis in Brazil.