Bt Research 2024, Vol.15, No.1, 65-75 http://microbescipublisher.com/index.php/bt 67 Additionally, CRISPR/Cas9 has been employed in plant genome editing to develop crops with improved traits, including enhanced resistance to pests, which complements the use of Bt biopesticides (Chen et al., 2019). 3.3 Development of multi-trait Bt strains Developing multi-trait Bt strains involves combining multiple insecticidal genes to create a single strain capable of targeting a broader spectrum of pests or overcoming resistance in specific insect populations. This strategy is exemplified by the isolation and characterization of novel Bt strains, such as the AB1 strain from Sri Lanka, which exhibits toxicity towards Dipel-resistant Plutella xylostella (Shanmugam et al., 2020). The AB1 strain produces a variety of Cry proteins, including Cry1Ca and Cry1Da, which contribute to its effectiveness against resistant pest populations (Baragamaarachchi et al., 2019). Such multi-trait strains are crucial for maintaining the long-term sustainability and effectiveness of Bt biopesticides in agricultural pest management. By leveraging advanced genetic engineering techniques, including gene cloning, CRISPR/Cas9 genome editing, and the development of multi-trait Bt strains, researchers are making significant strides in enhancing the efficacy and sustainability of Bt biopesticides. These innovations not only improve pest control but also contribute to the broader goals of sustainable agriculture and environmental protection. 4 Environmental Impact and Safety Assessment 4.1 Impact on non-target organisms The environmental impact of Bt biopesticides on non-target organisms has been a significant area of research. Studies have shown that while Bt biopesticides are generally considered safe, they can have varying effects on different non-target species. For instance, the ecotoxicological effects of Lavandula luisieri hydrolate on soil organisms revealed acute toxicity to the plant Allium cepa and slight toxicity to the earthworm Eisenia fetida, along with significant impacts on soil bacterial communities (Pino-Otín et al., 2019). Similarly, the hydrolate from Artemisia absinthiumshowed acute toxicity to non-target soil organisms, including earthworms and plants, and altered microbial community functions (Fournier et al., 2020). Additionally, the impact of Bt crops on non-target plant-associated insects and soil organisms has been extensively reviewed, indicating no significant harmful effects in approved GM events, although continuous risk assessments are necessary (Yaqoob et al., 2016). 4.2 Soil and water health The application of Bt biopesticides and Bt crops can influence soil and water health through their interactions with soil microorganisms and aquatic organisms. Bt proteins released into the soil from transgenic plants or biopesticides can affect soil microbial diversity and activity. For example, Bt proteins have been shown to alter soil microbial communities, impacting nutrient cycling and soil health (Figure 1) (Li et al., 2022). Furthermore, the ecotoxicity of Artemisia absinthium hydrolate on aquatic organisms demonstrated acute toxicity to species such as Daphnia magna and Vibrio fisheri, highlighting potential risks to water ecosystems (Pino-Otín et al., 2019). The impact of biopesticides on soil microbial endpoints compared to conventional crops has also been systematically reviewed, emphasizing the need for comprehensive assessments to understand their long-term effects on soil biodiversity and ecosystem services (Kostov et al., 2014). 4.3 Ecotoxicological studies Ecotoxicological studies are crucial for understanding the broader environmental implications of Bt biopesticides. Research on the environmental behaviors of Bt insecticidal proteins has provided insights into their adsorption, retention, and degradation in soils, as well as their potential ecological risks. Additionally, studies on the impact of commercial Bt products on marine organisms, such as marine bivalves, have shown significant physiological and ecological responses, including altered feeding rates and energy budgets, indicating the need for careful consideration of Bt biopesticides' effects beyond terrestrial ecosystems (Manachini et al., 2013). The comparative impact of synthetic pesticides and biopesticides on soil microbial communities has also been investigated, revealing that both types of pesticides can decrease the complexity of soil microbial networks, although their specific effects on microbial community composition and function may differ (Fournier et al., 2020).
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