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Lepidoptera are the phytophagous group most at risk of impact by insect-resistant genetically modified (GM) plants since most of these crops have been developed to express toxins to specifically target lepidopteran pests. In the case of Bt-maize cultivation, pollen grains released by GM plants could dust larval host plants growing inside or nearby maize fields, thus leading to the exposure of caterpillars to the Cry toxins. This way of exposure has been studied mainly for butterflies, whereas moths, in spite of their great diversity and abundance, were rarely considered. To evaluate the potential exposure of moth species to maize pollen in absence of Cry-expressing plants, macro-moths were sampled by means of light traps in three protected areas of Northern, Central and Southern Italy where conventional maize is among the main crops. Light traps were activated in two consecutive years (2010- 11) from the beginning of June to the end of July, a period that encompasses the anthesis of most maize cultivars grown in Italy. Overall, 11446 individuals belonging to 201 species and nine families were collected. Given that larvae of several sampled species are likely to match the criteria of potential exposure to maize pollen and susceptibility to Cry toxins, macro-moths might be considered among the potential assessment endpoints for environmental risk assessment of GM plants, especially in the vicinity of protected areas. © 2017, Bulletin of Insectology. All rights reserved.

Genetically modified (GM) plants may show unintended differences compared to the original varieties, due to the modification process. Such differences might in some cases affect non-target organisms linked to the crop into an agro-ecosystem. In this paper, we aimed to study interactions of two blight-resistant GM potato clones with the aphid species Macrosiphum euphorbiae Thomas, a non-target arthropod frequently feeding on potato plants and one of the major pests of that crop. One of the potato events used in our experiments caused an increased fertility of the aphids in the first generation, and consequently, a positive effect on the growth of the aphid population was estimated. When a second generation of the aphid was reared on potato leaves of the same GM event, differences in aphid fertility were no longer observed. Behavioural studies conducted in a wind tunnel using the aphid parasitoid Aphidius ervi Haliday indicated that neither of the two tested GM varieties had a significant effect on the attractiveness of potato plants towards the parasitoid. In planta tests proved to be sensitive protocols to detect unintended effects on a non-target arthropod; experimental results, however, indicate that these effects are not expected to be biologically relevant in this tritrophic system, if these GM events become available for commercial use in the future.

Ladybird beetles (Coleoptera Coccinellidae) are important predators of aphids and other crop pests, and there is great interest in their conservation in agroecosystems. Bt-maize, genetically engineered to express insecticidal Cry proteins, is regarded as a taxon-specific way of controlling pests, but some concerns have been raised about the possibility of unintended negative effects on non-target organisms, including coccinellids. One of the possible routes of exposure of ladybird beetles to Cry toxins is by feeding on maize pollen, as for many species pollen is an important integrative food source. In this study, coccinellid adults were sampled by sweep netting in Ecological Compensation Areas (ECAs) in three sites of Northern and Central Italy, where conventional maize cultivars are grown (Bt-maize is currently forbidden for commercial cultivation in Italy). The coccinellid communities were sampled during and around the typical flowering periods of maize in order to check their diversity and their overlap with pollen shedding. A total of 11 species were recorded. Hippodamia variegata (Goeze), Coccinella septempunctata L. and the exotic Harmonia axyridis (Pallas) were the most abundant species in Northern Italy, whereas Tytthaspis sedecimpunctata (L.) and Coccinula quatuordecimpustulata (L.) were dominant in Central Italy. The potential exposure to maize pollen was different in the two areas, since in Northern Italy the maize flowering coincided with a period of high coccinellid activity in the field, whereas in Central Italy the ladybird population peaks occurred roughly one month earlier than the anthesis. The collected data might be useful for exposure characterization of ladybird beetles in a possible future scenario of Bt-maize cropping in the studied areas. © 2016, Universita degli Studi di Bologna, Department of Agroenvironmental Sciences and Technologies. All rights reserved.

Sicily (Italy) is one of the richest European regions in animal biodiversity. Butterflies offer good opportunities for studies on biodiversity, population and community ecology. Many species are strictly seasonal, preferring only a particular set of habitats. Five typical Mediterranean habitats: olive grove (conducted following organic farm management), mixed wood, ampelodesmos prairie (with dominance of Ampelodesmos mauritanicus), Mediterranean shrub and shrub at the coastal areas, were monitored for diversity and seasonal patterns in butterfly communities for 2 years. All habitats were inside protected areas indicated as Sites of Community Importance. Butterfly species richness was highest in ampelodesmos prairie and in olive grove. Pieris brassicae, Lasiommata megera, Maniola jurtina, Pararge aegeria have been reported in all habitats, although with different abundance whilst other species were observed only in one particular habitat. Melanargia pherusa, endemic species, and Hipparchia statilinus, listed in the IUCN Red List, respectively as “Least Concern” and “Near Threatened”, were observed in the ampelodesmos prairie. The morphotype Coenonympha pamphilus f. lyllus was reported in olive grove and ampelodesmos prairie. The majority of species showed abundance peaks in June and July, however many species showed a longer flight period than reported in literature with detection of specimens until November or December. The analysis of the similarity of communities among habitats shows a similarity between ampelodesmos prairie and olive grove and between Mediterranean shrub and Mediterranean shrub at the coastal areas, while the butterfly community in mixed wood is substantially different. The results of several diversity indexes suggest that ampelodesmos prairie has an important ecological role, as it supports butterfly abundance and species diversity. Copyright 2015, Universita degli Studi di Bologna, Department of Agroenvironmental Sciences and Technologies, All Rights Reserved.

One of the concerns surrounding the commercial release of genetically modified (GM) crops is the escape of transgenes into agricultural or semi-natural habitats through vertical gene flow, as this may cause environmental or economic problems. There is also the concern that GM crops may affect pollinators and the pollination services they provide. Despite the growing commercial interest of GM tomato (Solanum lycopersicum), gene flow has been assessed only sparsely in tomato. To evaluate the likelihood of gene flow from GM tomato plants to sexually compatible plants, and to assess whether bumblebee activity is affected by GM tomato, three experiments were conducted under greenhouse conditions, using a Bt-tomato expressing the insecticidal Cry3Bb1 protein as model system: (a) artificial crosses between a GM tomato line, two wild tomato relatives (Solanum hirsutum and Solanum nigrum) and a non-GM tomato variety; (b) bumblebee-mediated crosses between GM and non-GM tomato plants and (c) visual observations of bumblebees' feeding behaviour. No hybrids were obtained between the GM tomato line and S. hirsutum and S. nigrum. In an experimental design where non-GM receptor plants outnumbered GM plants by approximately 3:1, the bumblebee-mediated cross-fertilisation rate between GM and non-GM tomato plants was measured at 4.3 ± 5.47%. No significant differences in feeding behaviour of bumblebees foraging on GM and non-GM tomato plants were observed. Therefore, we conclude that: (a) the probability of transgene introgression between the GM tomato line used in this study and its wild relatives S. hirsutum and S. nigrum is negligible; (b) bumblebee activity can mediate cross-fertilisation between GM and non-GM tomato and (3) the Cry3Bb1-expressing tomato line tested does not adversely affect the feeding behaviour of bumblebees. © 2012 The Authors. Annals of Applied Biology © 2012 Association of Applied Biologists.

The tomato leafminer Tuta absoluta (Meyrick) (Lepidoptera: Gelechiidae) is a devastating pest of tomato originating from South America. After its initial detection in eastern Spain in 2006, it rapidly invaded various other European countries and spread throughout the Mediterranean basin. If no control measures are taken, then the pest can cause up to 80–100% yield losses in tomato crops in recently invaded areas and may pose a threat to both greenhouse and open-field tomato production. The exceptional speed and extent of T. absoluta invasion have called for studies documenting its biology and ecology, while indicating an urgent need for efficient and sustainable management methods. The development of approaches to manage T. absoluta would be facilitated through a detailed revision of information on this pest in its area of origin. This review combines information on the invasion by T. absoluta, its ecology, and potential management strategies, including data that may help the implementation of efficient biological control programs. These programs, together with a variety of other management tactics, may allow efficient integrated pest management of T. absoluta in Europe and Mediterranean Basin countries.

The subject of this thesis is the use of transgenic plant resistance as a method to control the Colorado potato beetle, Leptinotarsa decemlineata Say in eggplant. The gene conferring resistance is coding for a Cry3B toxin and it is a synthetic version of a wild-type gene originally obtained from the soil bacterium Bacillus thuringiensis Berl.Eggplant cultivations are constantly attacked by a number of serious pests (e.g. the fruit and shoot borer, the Colorado potato beetle, soil-borne fungi). In spite of the heavy losses they may cause, breeding for resistance on this crop has been very limited because of the lack of desirable traits in the eggplant genome or sexual incompatibility with the resistant wild related species.The first chapter reviews Colorado potato beetle biology and its control, with a special emphasis on beetle-eggplant relationships. L. decemlineata has become a major problem for eggplant cultivation and sometimes its control on this crop is even more problematic compared to potato cultivation. The longer life cycle of the plant and the intensive regime under which horticultural crops are cultivated have contributed to the increasing importance of this pest.In the second chapter, a review of the source of resistant genes available in both the eggplant gene pool and wild Solanum relatives is presented. Considering the genetic basis of resistant traits, the possible strategies for eggplant breeding are discussed with emphasis on approaches based on the integration of classical breeding methods (crosses and selection) with biotechnological ones (anther culture, genetic transformation, protoplast fusion and marker-assisted selection).In the third chapter, the results of the study in which protein extracts of Escherichia coli expressing the toxin gene from B. thuringiensis were tested for effect on the behavior and development of L. decemlineata larvae using in vitro bioassays are presented. No antifeedant effect due to Cry3B toxin was found, even in concentrations which caused mortality or severely inhibited larval growth.In the fourth chapter, the production of transgenic eggplants and their evaluation is presented. A modified Bt gene of Bacillus thuringiensis var. Tolworthi, encoding a coleopteran insect-specific Cry3B toxin, was transferred via Agrobacterium tumefaciens to the female parent of the commercial F1 hybrid 'Rimina' eggplant (released by the Istituto Sperimentale per l'Orticoltura). A large number of transgenic plants were regenerated and tested by PCR and NPTII expression assays. The presence of the Cry3B toxin in leaf extracts was demonstrated by the DAS-ELISA test in 57 (62.3%) transgenic plants which contained a 74 kDa protein cross-reacting with the serum anti-Cry3B toxin. Twenty-three out of 44 S. melongena plants tested by an insect bioassay showed a significant insecticidial activity on neonate larvae of Colorado Potato Beetle (CPB). The Bt transgene and the toxic effect on CPB larvae were transmitted to progenies derived by selfing. Thus, transgenic Bt eggplants may represent a very effective means of CPB pest control.Transgenic potato clones expressing a Cry3B endotoxin were used to study the trophic interactions between newly emerged Colorado potato beetle adults and these resistant clones (Chapter 5). Adult longevity and fitness were studied for the first 3 weeks after emergence. The reproductive biology of the beetle on highly resistant clones, partly resistant clones and control potato plants was monitored by dissecting females after 7-15 days of feeding and by analyzing the haemolymph's protein content after 3 days of feeding. Feeding behavior on either highly toxin expressing or control plants was monitored individually for 36 beetles feeding on leaf-discs. Beetles feeding on transgenic or control clones as the sole source of food had similar longevity. However egg production was completely inhibited on transgenic plants.Dissection studies indicated that adult males living on transgenic plants were still able to mate and produce mobile sperms, but the females were impaired in their reproductive ability since ovarioles were not normally developed. An exam of the haemolymph revealed the protein concentration in females living on transgenic plants to be dramatically reduced (about 50% compared to the control). The feeding behavior of Colorado potato beetle adults was not affected by the different food plants. This shows that transgenic potato plants were readily accepted as host plants by the beetles. The implications of these findings for the use of transgenic plants as a means of L. decemlineata control are discussed.In the sixth chapter the most relevant results of the first year of a field experiment with the transgenic eggplant lines are presented. Two of the 3 transgenic lines used showed a high level of resistance in two separate trials, as indicated by the analyses of L. decemlineata population levels and crop yield. Fruit production was almost doubled in the resistant lines compared to a DR2 untransformed control. Only one transgenic line showed an intermediate level of resistance, giving results more similar to the control under heavy CPB attack, whereas it gave comparable results to the other transgenic lines where natural infestation was milder. No detrimental effects on non-target arthropods (including the chrysomelid beetle Altica spp.) were apparent. Field observations confirmed that Bt-expressing transgenic plants might be able to successfully control Colorado potato beetle infestations in eggplant cultivations, representing a potentially effective and environmentally safe means of pest control.In chapter 7, the relationships between L. decemlineata egg density and Coleomegillamaculata DeGeer predatory behavior is presented. Despite aggregation in areas of the highest prey density by C.maculata , egg consumption was inversely related to egg mass density at the smallest and the largest spatial scales tested. The experimental data on predation rates in high and low density field treatments, were included in a mathematical model to simulate the impact of natural enemies on the rate of L.decemlineata adaptation to Bt-toxin-expressing transgenic potato plants when Bt-expressing plants are mixed at the plot-to-plot level with normal potato plants. Results showed that C.maculata predatory behavior could decrease the rate at which L.decemlineata adapted to Bt-toxins if plot-to-plot mixed plantings were used.Finally, a simulation model to predict the possible adaptation of L. decemlineata to the Cry3 toxin expressed in transgenic eggplant is presented (Chapter 8). The use of mixed fields of transgenic and susceptible isolines at a 90:10 ratio has been simulated. Beetle movement, which is a fundamental parameter when studying plant mixtures, has been addressed with a 'two-stage' hypothesis. The biological and genetic characteristics of the beetles have been set to specifically address their possible interactions with resistant eggplant. The role of gene dominance, migration, and fitness costs associated with the resistant genotype have been examined. Using the hypothesis of partial dominance of the resistant gene, only a high level of migration (very likely, in most agricultural areas) or a considerable reduction of the fitness of resistant beetles, associated with the change in their genome, can guarantee a long-lasting efficacy of the germplasm. The simulations clearly indicate that the effect of resistance in transgenic clones expressing Bacillus thuringiensis -derived toxins can be optimized only in accordance with opportune agricultural practices.