Corn is a dominating feature in the landscape of the Midwestern United States. Associated with this crop are a plethora of products from fertilizers to pesticides that help farmers maximize grain yield while minimizing costs. A widely accepted form of protection from major pests in the United States is genetically modified corn that has been altered so that it contains genetic material from another species, a soil bacterium, Bacillus thuringiensis Berliner (Bt corn), in order to produce proteins that are toxic to some insect pests. Additional insecticides that are commonly used to protect corn include a neonicotinoid seed coating, clothianidin, and a pyrethroid insecticide applied in-furrow, tefluthrin. The goal of my dissertation was to measure the environmental fate of the Cry1Ab Bt corn protein, clothianidin, and tefluthrin and determine if measured concentrations could be causing effects to non-target species. The Cry1Ab proteins persisted only during the corn growing season in soil, runoff water and sediment with the highest concentrations measured during pollination. Clothianidin was detected in all matrices and remained persistent throughout the year in soil pore water. Tefluthrin was consistently detected in soil, runoff water, and runoff sediment during the corn growing season, but was not found in groundwater or soil pore water. No acute toxicity was observed to any species when exposed to the Cry1Ab protein in corn plant leaf tissue, therefore little risk was anticipated. Non-target species exposed to clothianidin resulted in limited ecological risk from field exposures. Tefluthrin results indicated elevated risk for non-target species exposed to tefluthrin at concentrations that were measured in the field study. There was no increase in toxicity to tefluthrin when non-target species were exposed to a combination of all three insecticides. In summary, the genetically modified corn insecticidal proteins and clothianidin were not found at environmental concentrations exceeding benchmark values for ecological effects, but tefluthrin was consistently detected in the environment at levels that could be causing toxicity to non-target species, especially if it is able to travel off-site.

AN ABSTRACT OF THE THESIS OF Katherine E. Strain, for the Master's degree in Zoology, presented on October 21, 2014, at Southern Illinois University Carbondale. TITLE: FATE AND TRANSPORT OF CRY1AB FROM TRANSGENIC BACILLUS THURINGIENSIS CORN IN AN AGRICULTURAL FIELD AND AQUATIC MICROCOSMS MAJOR PROFESSOR: Dr. Michael Lydy, Ph.D. Genetically-modified crops expressing insecticidal crystalline proteins derived from a soil bacterium, Bacillus thuringiensis (Bt), were commercialized almost two decades ago as a means to combat agricultural pests. The Bt proteins are highly specific and only lethal upon ingestion, limiting the scope of toxicity to target insects. However, evidence for risk to non-target organisms and negative public perceptions on the use of Bt crops has caused controversy surrounding their use. The objective of this research was to monitor the fate and transport of a Bt protein, Cry1Ab, in a large-scale agricultural field and in aquatic microcosms. Quantitative methods were validated using enzyme-linked immunosorbent assay (ELISA) and then used to evaluate field and laboratory samples. The highest environmental concentrations of the Cry1Ab protein were found in runoff water and sediment, up to 130 ng/L and 143 ng/g dry weight, respectively, with the Cry1Ab protein detected in both Bt and non-Bt fields. As surface runoff and residual crop debris can transport Bt proteins to waterways adjacent to agricultural fields, a series of laboratory experiments were conducted to determine the potential risk to non-target aquatic organisms. The results showed that sediment type and temperature can influence the degradation of the Cry1Ab protein in an aquatic system and that the Cry1Ab protein can persist for two months. While Cry1Ab protein concentrations measured in the field soil indicate little risk to terrestrial organisms, the consistent input of Bt-contaminated runoff and crop debris into agricultural waterways impart chronic risk to non-target aquatic species.

O uso de culturas GM (geneticamente modificadas) tem sido questionado quanto ao destino dos produtos derivados da transgenia no ambiente. Com a liberação de exsudatos das raízes das plantas e a decomposição dos resíduos culturais, aumenta-se a quantidade de DNA transgênico no ambiente, que pode ser adsorvido à superfície ativa das partículas do solo e/ou degradado pela ação de enzimas microbianas. A comunidade microbiana do solo pode entrar em contato direto com estes produtos, aumentando a probabilidade de transferência horizontal de fragmentos de DNA transgênico para os microrganismos. Também, alterações na composição dos exsudatos das plantas GM e mudanças em função das práticas de manejo, podem resultar em alterações na composição funcional e estrutural da comunidade microbiana. Assim, faz-se necessário avaliar a persistência dos produtos derivados da transgenia no solo e seus possíveis efeitos sobre a comunidade microbiana. Os objetivos deste estudo foram: avaliar a persistência dos fragmentos 35S-hsp70, hsp70-cry1Ab e cry1Ab-planta da construção gênica do milho Bt (evento MON810) em diferentes tipos de solo e temperaturas, em condições de microcosmo e de campo; e determinar a abundância do número de cópias dos gene 16S rRNA de Bacteria, Firmicutes, Verrucomicrobria e Archaea, e 18S rRNA de Fungo nas mesmas condições, e avaliar a estrutura da comunidade bacteriana em áreas agrícolas de cultivo de milho Bt. No primeiro estudo, o DNA do milho Bt MON810 foi adicionado em solos arenoso e argiloso. Como controle negativo, apenas água estéril foi misturada ao solo. Amostras de solo foram incubadas a 15 e 25ºC. Em campo, os solos foram amostrados em três áreas agrícolas em Fátima do Sul, MS, em dois anos consecutivos. Após extração de DNA, os fragmentos foram quantificados por qPCR. No segundo estudo, foram determinadas a abundância dos genes 16S rRNA de Bacteria, Firmicutes, Verrucomicrobria e Archaea e 18S rRNA de Fungo e avaliada a estrutura da comunidade bacteriana por T-RLFP. Os resultados mostraram que em condições de microcosmo, os fragmentos hsp70-cry1Ab e cry1Ab-planta persistiram até 291 dias, e o fragmento 35S-hsp70 até 180 dias. A temperatura e o tipo de solo não afetaram a persistência dos fragmentos. Em campo, o número de cópias desses fragmentos foi maior na segunda coleta. No segundo estudo, o número de cópias do gene 16S rRNA de Bacteria aumentou com adição de DNA do milho Bt nos microcosmos, e uma redução do número de cópias foi verificada para Archaea, Verrucomicrobia e Fungo. Para Firmicutes, os resultados não foram consistentes. As temperaturas não resultaram em efeito na abundância dos genes, enquanto o tipo de solo teve efeito apenas para Archaea e Verrucomicrobia. Áreas agrícolas com cinco anos de cultivo de milho Bt apresentaram variações na estrutura da comunidade bacteriana em nível de filo, e maior abundância de Fungos no segundo ano de amostragem, enquanto em área com um ano de cultivo, observouse uma redução da população de Firmicutes e Verrucomicrobia. Os maiores efeitos na comunidade microbiana foram verificados entre os anos de amostragem / The use of GM (genetically modified) crops has been questioned about the fate of transgenes is derived products on the environment. With the release of exudates from roots of GM plants and the decomposition of its residues, the amount of transgenic DNA in the environment increases, which can be adsorbed to the active surface of soil particles and/or be degraded by the action of microbial enzymes. Soil microbial communities can come into direct contact with these products, raising the probability of horizontal transfer of transgenic DNA fragments to soil microorganisms. Moreover, changes in exudates composition of GM plants and changes depending on the management practices may result in structural and functional alterations in the microbial community. Thus, it is necessary to evaluate the persistence of transgenes is derivatives in the soil and effects on microbial community. The objectives of this study were to assess the persistence of fragments 35S-hsp70, hsp70-cry1Ab and cry1Abplant from the genetic construct of Bt corn (event MON810) in different soil types and temperatures, in microcosm and field conditions; and to determine the abundance of 16S rRNA copy number of Bacteria, Firmicutes, Verrucomicrobria and Archaea and 18S rRNA of Fungi under the same conditions, and to evaluate the structure of bacterial communities in agricultural areas of Bt corn cultivation. In the first study, DNA from Bt corn MON810 was added to sandy and clay soils. As negative control, only sterile water was mixed with soil. Soil samples were incubated at 15 and 25°C. At the field, soils were sampled in three agricultural areas in Fátima do Sul, MS, in two consecutive years. After DNA extraction, fragments were quantified by qPCR. In the second study, the abundance of 16S rRNA of Bacteria, Firmicutes, Verrucomicrobria and Archaea and 18S rRNA of Fungi were determined and the structure of bacterial communities was evaluated by T-RFLP. The results showed that in microcosm conditions, hsp70-cry1Ab and cry1Ab-plants fragments persisted until 291 days and the 35S-hsp70 up to 180 days. The temperature and the type of soil did not affect the persistence of fragments. In field, the copy number of these fragments was greater in the second sampling. In the second study, the copy number of 16S rRNA of Bacteria increased with the addition of DNA from Bt corn in microcosm, and a reduction in copy number was observed for Archaea, Verrucomicrobia and Fungi. The results were not consistent for Firmicutes. Temperatures resulted in no effect in gene abundance, while the soil was effective only for Archaea and Verrucomicrobia. Agricultural areas with five years of Bt corn cultivation showed variations in bacterial community structure at the phylum level, and greater abundance of fungi in the second year of sampling, while in the area with a year of cultivation, a reduction in population of Firmicutes and Verrucomicrobia was observed. The largest effects on the microbial community were observed between the sampled years

O uso de culturas GM (geneticamente modificadas) tem sido questionado quanto ao destino dos produtos derivados da transgenia no ambiente. Com a liberação de exsudatos das raízes das plantas e a decomposição dos resíduos culturais, aumenta-se a quantidade de DNA transgênico no ambiente, que pode ser adsorvido à superfície ativa das partículas do solo e/ou degradado pela ação de enzimas microbianas. A comunidade microbiana do solo pode entrar em contato direto com estes produtos, aumentando a probabilidade de transferência horizontal de fragmentos de DNA transgênico para os microrganismos. Também, alterações na composição dos exsudatos das plantas GM e mudanças em função das práticas de manejo, podem resultar em alterações na composição funcional e estrutural da comunidade microbiana. Assim, faz-se necessário avaliar a persistência dos produtos derivados da transgenia no solo e seus possíveis efeitos sobre a comunidade microbiana. Os objetivos deste estudo foram: avaliar a persistência dos fragmentos 35S-hsp70, hsp70-cry1Ab e cry1Ab-planta da construção gênica do milho Bt (evento MON810) em diferentes tipos de solo e temperaturas, em condições de microcosmo e de campo; e determinar a abundância do número de cópias dos gene 16S rRNA de Bacteria, Firmicutes, Verrucomicrobria e Archaea, e 18S rRNA de Fungo nas mesmas condições, e avaliar a estrutura da comunidade bacteriana em áreas agrícolas de cultivo de milho Bt. No primeiro estudo, o DNA do milho Bt MON810 foi adicionado em solos arenoso e argiloso. Como controle negativo, apenas água estéril foi misturada ao solo. Amostras de solo foram incubadas a 15 e 25ºC. Em campo, os solos foram amostrados em três áreas agrícolas em Fátima do Sul, MS, em dois anos consecutivos. Após extração de DNA, os fragmentos foram quantificados por qPCR. No segundo estudo, foram determinadas a abundância dos genes 16S rRNA de Bacteria, Firmicutes, Verrucomicrobria e Archaea e 18S rRNA de Fungo e avaliada a estrutura da comunidade bacteriana por T-RLFP. Os resultados mostraram que em condições de microcosmo, os fragmentos hsp70-cry1Ab e cry1Ab-planta persistiram até 291 dias, e o fragmento 35S-hsp70 até 180 dias. A temperatura e o tipo de solo não afetaram a persistência dos fragmentos. Em campo, o número de cópias desses fragmentos foi maior na segunda coleta. No segundo estudo, o número de cópias do gene 16S rRNA de Bacteria aumentou com adição de DNA do milho Bt nos microcosmos, e uma redução do número de cópias foi verificada para Archaea, Verrucomicrobia e Fungo. Para Firmicutes, os resultados não foram consistentes. As temperaturas não resultaram em efeito na abundância dos genes, enquanto o tipo de solo teve efeito apenas para Archaea e Verrucomicrobia. Áreas agrícolas com cinco anos de cultivo de milho Bt apresentaram variações na estrutura da comunidade bacteriana em nível de filo, e maior abundância de Fungos no segundo ano de amostragem, enquanto em área com um ano de cultivo, observouse uma redução da população de Firmicutes e Verrucomicrobia. Os maiores efeitos na comunidade microbiana foram verificados entre os anos de amostragem / The use of GM (genetically modified) crops has been questioned about the fate of transgenes is derived products on the environment. With the release of exudates from roots of GM plants and the decomposition of its residues, the amount of transgenic DNA in the environment increases, which can be adsorbed to the active surface of soil particles and/or be degraded by the action of microbial enzymes. Soil microbial communities can come into direct contact with these products, raising the probability of horizontal transfer of transgenic DNA fragments to soil microorganisms. Moreover, changes in exudates composition of GM plants and changes depending on the management practices may result in structural and functional alterations in the microbial community. Thus, it is necessary to evaluate the persistence of transgenes is derivatives in the soil and effects on microbial community. The objectives of this study were to assess the persistence of fragments 35S-hsp70, hsp70-cry1Ab and cry1Abplant from the genetic construct of Bt corn (event MON810) in different soil types and temperatures, in microcosm and field conditions; and to determine the abundance of 16S rRNA copy number of Bacteria, Firmicutes, Verrucomicrobria and Archaea and 18S rRNA of Fungi under the same conditions, and to evaluate the structure of bacterial communities in agricultural areas of Bt corn cultivation. In the first study, DNA from Bt corn MON810 was added to sandy and clay soils. As negative control, only sterile water was mixed with soil. Soil samples were incubated at 15 and 25°C. At the field, soils were sampled in three agricultural areas in Fátima do Sul, MS, in two consecutive years. After DNA extraction, fragments were quantified by qPCR. In the second study, the abundance of 16S rRNA of Bacteria, Firmicutes, Verrucomicrobria and Archaea and 18S rRNA of Fungi were determined and the structure of bacterial communities was evaluated by T-RFLP. The results showed that in microcosm conditions, hsp70-cry1Ab and cry1Ab-plants fragments persisted until 291 days and the 35S-hsp70 up to 180 days. The temperature and the type of soil did not affect the persistence of fragments. In field, the copy number of these fragments was greater in the second sampling. In the second study, the copy number of 16S rRNA of Bacteria increased with the addition of DNA from Bt corn in microcosm, and a reduction in copy number was observed for Archaea, Verrucomicrobia and Fungi. The results were not consistent for Firmicutes. Temperatures resulted in no effect in gene abundance, while the soil was effective only for Archaea and Verrucomicrobia. Agricultural areas with five years of Bt corn cultivation showed variations in bacterial community structure at the phylum level, and greater abundance of fungi in the second year of sampling, while in the area with a year of cultivation, a reduction in population of Firmicutes and Verrucomicrobia was observed. The largest effects on the microbial community were observed between the sampled years

Conselho Nacional de Desenvolvimento Científico e Tecnológico / The objectives of this study were to evaluate the effects of Bt toxin and seeds treatment with insecticides (imidacloprid + thiodicarb) on initial growth of corn and the control of Spodoptera frugiperda (J. E. Smith) in soil; the effect of Bt corn and insecticides (seed treatment with imidacloprid + thiodicarb and spray application of novaluron and/or methomyl) for the control of S. frugiperda in crop shoots during early and late planting seasons; and the effects of Bt corn and insecticides control of S. frugiperda on the occurrences and damages from Diatraea sp. and Helicoverpa zea (Boddie) pests. Three experiments were conducted under field conditions and natural infestation of pests, in Itaara and Santiago, in Rio Grande do Sul state, Brazil, during the 2008/2009 planting seasons. The initial growth of corn was not affected by the production of Bt toxin (Cry1Ab) and insecticide seed treatment in areas without pest infestation. The toxin (Cry1Ab) present in the Bt corn as well as seeds treatment in non-transformed corn effectively protected the plants to cut-off S. frugiperda. Seeds treatment enables the reduction of damages caused by S. frugiperda in Bt corn (Cry1Ab) to the shoots of crops in early infestations. The Bt corn (Cry1Ab) is effective in controlling S. frugiperda, especially when infestations are low to moderate. When high infestations of S. frugiperda occurred in late planting, insecticide spray on Bt corn (Cry1Ab) resulted to less damages and fewer lavae development comparatively with unsprayed plots. Insecticide treatment of nontransformed corn seeds during early infestations of Diatraea sp. and Bt corn reduced the percentage of stems attacked and injuries. However, insecticides sprayed for the control of S. frugiperda had no effect on the percentage of stems attacked and the injuries caused by Diatraea sp.. Application of insecticides to seeds or spraying for the control of S. frugiperda did not affect the percentage of corn ears attacked and the injuries caused by H. zea, while the Bt corn (Cry1Ab) reduced the percentage of ears attacked by H. zea despite not having much effect in reducing the ear-feeding injuries. / O objetivo do estudo foi avaliar: o efeito da toxina Bt e do tratamento de semente com inseticidas (imidacloprido + tiodicarbe), no crescimento inicial das plantas de milho e no controle de Spodoptera frugiperda (J. E. Smith) no solo; o efeito do milho Bt e dos inseticidas (imidacloprido + tiodicarbe em tratamento de semente e novalurom e/ou metomil em pulverizações), no controle de S. frugiperda na parte aérea da cultura, na semeadura do cedo e do tarde; e os efeitos do milho Bt e dos inseticidas para o controle de S. frugiperda, na ocorrência e na injúria da Diatraea sp. e da Helicoverpa zea (Boddie). Foram realizados três experimentos em condições de campo e com infestação natural dos insetos-praga, em Itaara e Santiago, RS, durante a safra 2008/09. O crescimento inicial das plantas de milho não é afetado pela produção da toxina Bt (Cry1Ab) e pelos inseticidas em tratamento de semente, em áreas sem infestação de insetos-praga. A toxina Cry1Ab, presente no milho Bt, assim como o tratamento de semente no milho convencional, protegem de forma eficiente as plantas do corte de S. frugiperda. O tratamento de semente, possibilita a redução da injúria na parte aérea da cultura causada por S. frugiperda em milho Bt (Cry1Ab), em infestações precoces. O milho Bt (Cry1Ab) é eficiente no controle de S. frugiperda, especialmente quando as infestações são baixas a moderadas. Em época de semeadura, na qual ocorrem altas infestações de S. frugiperda, o milho Bt (Cry1Ab) quando pulverizado com inseticida, apresenta em relação ao sem pulverização, menos injúria e menor número de lagartas grandes. O tratamento de semente com inseticidas em milho convencional, quando em infestações precoces de Diatraea sp. e o milho Bt, reduzem as injúrias e o percentual de colmos atacados, porém os inseticidas pulverizados para o controle de S. frugiperda, não têm efeito sofre as injúrias e no percentual de colmos atacados por Diatraea sp.. Os inseticidas aplicados na semente ou em pulverizações para o controle de S. frugiperda, não afetam as injúrias e o percentual de espigas atacadas por H. zea, porém o milho Bt (Cry1Ab) reduz o percentual de espigas atacadas por H. zea, apesar de não ter efeito significativo na redução das injúrias nas espigas.

No description available.

The high-dose/refuge strategy is used globally to manage insect resistance development in genetically modified crops with insecticidal properties (Bt crops). The “refuge in a bag” (RIB) strategy is also being considered for deployment against several pest species. Busseola fusca, the target pest of Bt maize in South Africa, evolved resistance to Cry1Ab proteins. The objective of this study was to determine whether migrating B. fusca larvae are effectively controlled using the RIB strategy. A field study with a single-gene event (Cry1Ab) and a “pyramid” event (Cry1A.105 + Cry2Ab2) was conducted in which the migration patterns of B. fusca larvae in plots with different seed mixture treatments were studied. The experiment consisted of five seed mixture ratios (5%, 10%, 15%, 20% non-Bt seed and 100 % non-Bt seed as control). Natural infestation was augmented by artificial inoculation with neonate larvae into the central non-Bt maize plant of each plot. Rate of larval survival and migration, measured in terms of increase in number of plants per plot that exhibited borer damage was recorded at weekly intervals until flowering. A laboratory study was conducted to determine larval growth and survival when simulating migration between Bt and non-Bt maize plants. A feeding experiment in which larvae were reared on different types of maize (Bt and non-Bt) was conducted and larval survival and mass recorded after a 7-day feeding period. The incidence of damaged ears, stem damage and damaged internodes per stem were recorded and relationships between these variables determined by means of correlation analyses. A review was conducted in order to identify and discuss similarities and differences between the high-dose/refuge and seed mixture strategies. This was done to determine which strategy would be the most appropriate insect resistance management (IRM) strategy against B. fusca. The rate of survival and migration of B. fusca larvae was significantly higher in the plots with maize expressing Cry1Ab and control plots, than in plots with the pyramid Bt event. Older larvae exhibited improved growth and survival in the laboratory experiment when they were transferred from non-Bt to Bt plants. Positive correlations were found between early and late season damage, although some weaker than others. Plants of the “pyramid event” suffered less late-season damage than those of the single-gene event. Since the increase in number of damaged maize plants over time is associated with migration of older and larger larvae, the observed tendencies may indicate that the assumed high-dose does not kill larvae above a certain developmental stage. The high-dose refuge strategy seems to be the better option for delaying resistance development. / MSc (Environmental Sciences), North-West University, Potchefstroom Campus, 2014

Genetically modified maize expressing the Bt-protein Cry1Ab (Bt maize) is planted widely in South Africa. Crop residues of Bt maize often end up in aquatic ecosystems where aquatic organisms are exposed to Cry1Ab protein. The effect of this protein on non-target aquatic organisms has not yet been studied in South Africa. The aim of this study was to evaluate the possible effect of exposure to Bt maize on morphological development of Xenopus laevis and Amietophrynus gutturalis tadpoles. Three experiments were conducted with each of X. laevis and A. gutturalis. Five of these were conducted in the bio-secure Amphibian Biology laboratory and one with A. gutturalis in a shade-house facility where microcosms were exposed to natural conditions. In the first experiment of X. laevis and A. gutturalis, which was replicated three times, large portions of maize leaves were placed in the bottoms of microcosms. X. laevis received supplementary pulverised leaves in suspension while A. gutturalis tadpoles fed on provided leaves. For both control and experimental groups microcosms were divided in three groups receiving respectively 15, 30 and 45 g of maize leaves. In the second and third experiment tadpoles only received pulverised Bt maize leaves in suspension. Each replicate (microcosm) contained 50 one-day old tadpoles. Experiment two was conducted to determine whether the Bt-protein has adverse effects on A. gutturalis tadpoles when tadpoles are exposed to the protein in the water but not feeding on the plant material. A total of 100 tadpoles were used during the experiment and tadpoles were placed individually in 250 ml plastic cups that were filled with 100 ml water witch contained an extract of either Bt and non-Bt maize leaves. Tadpoles were fed twice a week with TetraTabimin bottom-feeding fish pellets in suspension. Experiment three was conducted to determine whether the Bt-protein will have adverse effects on A. gutturalis tadpoles when tadpoles feed on Bt maize leaves. Tadpoles were divided into a treatment in which 50 tadpoles were fed Bt maize leaves and a control treatment in which 50 tadpoles were fed non-Bt maize leaves. Tadpoles were placed individually in 250 ml plastic cups that were each filled with 100 ml borehole water. On a weekly basis 10 randomly selected tadpoles were collected, measured and staged for morphological development, using the Nieuwkoop and Faber Normal Table for X. laevis and Gosner stages for A. gutturalis tadpoles. The significant effects observed in some life history parameters of tadpoles exposed to Cry1Ab protein cannot be ascribed to the effect of the protein. Poor husbandry turned out to be the single most important confounding factor. Before follow-up studies are conducted husbandry practices should be optimized. / Thesis (MSc (Environmental Sciences))--North-West University, Potchefstroom Campus, 2013.

The high-dose/refuge strategy is used globally to manage insect resistance development in genetically modified crops with insecticidal properties (Bt crops). The “refuge in a bag” (RIB) strategy is also being considered for deployment against several pest species. Busseola fusca, the target pest of Bt maize in South Africa, evolved resistance to Cry1Ab proteins. The objective of this study was to determine whether migrating B. fusca larvae are effectively controlled using the RIB strategy. A field study with a single-gene event (Cry1Ab) and a “pyramid” event (Cry1A.105 + Cry2Ab2) was conducted in which the migration patterns of B. fusca larvae in plots with different seed mixture treatments were studied. The experiment consisted of five seed mixture ratios (5%, 10%, 15%, 20% non-Bt seed and 100 % non-Bt seed as control). Natural infestation was augmented by artificial inoculation with neonate larvae into the central non-Bt maize plant of each plot. Rate of larval survival and migration, measured in terms of increase in number of plants per plot that exhibited borer damage was recorded at weekly intervals until flowering. A laboratory study was conducted to determine larval growth and survival when simulating migration between Bt and non-Bt maize plants. A feeding experiment in which larvae were reared on different types of maize (Bt and non-Bt) was conducted and larval survival and mass recorded after a 7-day feeding period. The incidence of damaged ears, stem damage and damaged internodes per stem were recorded and relationships between these variables determined by means of correlation analyses. A review was conducted in order to identify and discuss similarities and differences between the high-dose/refuge and seed mixture strategies. This was done to determine which strategy would be the most appropriate insect resistance management (IRM) strategy against B. fusca. The rate of survival and migration of B. fusca larvae was significantly higher in the plots with maize expressing Cry1Ab and control plots, than in plots with the pyramid Bt event. Older larvae exhibited improved growth and survival in the laboratory experiment when they were transferred from non-Bt to Bt plants. Positive correlations were found between early and late season damage, although some weaker than others. Plants of the “pyramid event” suffered less late-season damage than those of the single-gene event. Since the increase in number of damaged maize plants over time is associated with migration of older and larger larvae, the observed tendencies may indicate that the assumed high-dose does not kill larvae above a certain developmental stage. The high-dose refuge strategy seems to be the better option for delaying resistance development. / MSc (Environmental Sciences), North-West University, Potchefstroom Campus, 2014

Genetically modified maize expressing the Bt-protein Cry1Ab (Bt maize) is planted widely in South Africa. Crop residues of Bt maize often end up in aquatic ecosystems where aquatic organisms are exposed to Cry1Ab protein. The effect of this protein on non-target aquatic organisms has not yet been studied in South Africa. The aim of this study was to evaluate the possible effect of exposure to Bt maize on morphological development of Xenopus laevis and Amietophrynus gutturalis tadpoles. Three experiments were conducted with each of X. laevis and A. gutturalis. Five of these were conducted in the bio-secure Amphibian Biology laboratory and one with A. gutturalis in a shade-house facility where microcosms were exposed to natural conditions. In the first experiment of X. laevis and A. gutturalis, which was replicated three times, large portions of maize leaves were placed in the bottoms of microcosms. X. laevis received supplementary pulverised leaves in suspension while A. gutturalis tadpoles fed on provided leaves. For both control and experimental groups microcosms were divided in three groups receiving respectively 15, 30 and 45 g of maize leaves. In the second and third experiment tadpoles only received pulverised Bt maize leaves in suspension. Each replicate (microcosm) contained 50 one-day old tadpoles. Experiment two was conducted to determine whether the Bt-protein has adverse effects on A. gutturalis tadpoles when tadpoles are exposed to the protein in the water but not feeding on the plant material. A total of 100 tadpoles were used during the experiment and tadpoles were placed individually in 250 ml plastic cups that were filled with 100 ml water witch contained an extract of either Bt and non-Bt maize leaves. Tadpoles were fed twice a week with TetraTabimin bottom-feeding fish pellets in suspension. Experiment three was conducted to determine whether the Bt-protein will have adverse effects on A. gutturalis tadpoles when tadpoles feed on Bt maize leaves. Tadpoles were divided into a treatment in which 50 tadpoles were fed Bt maize leaves and a control treatment in which 50 tadpoles were fed non-Bt maize leaves. Tadpoles were placed individually in 250 ml plastic cups that were each filled with 100 ml borehole water. On a weekly basis 10 randomly selected tadpoles were collected, measured and staged for morphological development, using the Nieuwkoop and Faber Normal Table for X. laevis and Gosner stages for A. gutturalis tadpoles. The significant effects observed in some life history parameters of tadpoles exposed to Cry1Ab protein cannot be ascribed to the effect of the protein. Poor husbandry turned out to be the single most important confounding factor. Before follow-up studies are conducted husbandry practices should be optimized. / Thesis (MSc (Environmental Sciences))--North-West University, Potchefstroom Campus, 2013.