Globally, mosquitoes pose a serious threat to human health as vectors of many diseases. These diseases include, but are not limited to, dengue fever, yellow fever, and Zika virus disease, which cause a large disease burden globally, especially in tropical and subtropical regions (Iosr et al., 2015). In addition to direct threats to human health, the widespread presence of mosquitoes in aquaculture environments may also have negative impacts on the aquaculture industry, including disrupting ecological balance and reducing water quality, thereby affecting breeding efficiency and production (Rudd et al., 2023).

Given the environmental pollution and ecological risks that chemical control methods may bring, it is particularly important to find environmentally friendly mosquito control strategies. Bacillus thuringiensis var. israelensis (Bti) is a biological pesticide widely used to control mosquito larvae. Due to its high efficiency, specificity and environmental friendliness, it has become one of the preferred methods for controlling mosquito larvae. The use of Bti can not only effectively reduce mosquito populations and reduce the risk of disease transmission, but also has less impact on non-target organisms and the overall ecosystem, providing the possibility of achieving sustainable mosquito management (Mo-on et al., 2022).

The purpose of this study was to comprehensively evaluate the effectiveness and impact of Bacillus thuringiensis var. israelensis in controlling (Aedes spp.) mosquito larvae in aquaculture environments, and to explore its practical significance and potential value as an ecological control strategy. By analyzing Bti's application strategies, ecological and environmental impacts, as well as challenges and management strategies, this article aims to provide a new mosquito control strategy for the aquaculture industry that reduces the use of chemical pesticides and protects the ecological environment, and also provides public Health protection provides support to reduce the health burden caused by mosquito-borne diseases.

1 Bti Application Strategies in Aquaculture

1.1 Application methods of Bti in aquaculture environment

Bacillus thuringiensis var. israelensis (Bti) in aquaculture environments requires careful planning of its application time, concentration and frequency to achieve optimal mosquito larvae control. Generally, Bti applications should be timed to match the mosquito's life cycle, especially during peak larval hatching. In addition, the application concentration should be sufficient to kill most mosquito larvae but not have a negative impact on aquaculture organisms or the environment. The frequency of application needs to be determined based on the mosquito's breeding cycle and the continued activity of Bti in a specific water environment. In most cases, periodic application of Bti can effectively interrupt the mosquito life cycle, thereby reducing the number of adult mosquitoes.

Ângelo et al. (2015) used response surface methodology to optimize the culture medium for Bti in their study. This optimization process not only increased the production of Bti, but also provided a basis for the development of effective application strategies. The optimized culture medium can support Bti to produce higher concentrations of toxins, thereby achieving lower LC50 values in practical applications, that is, a lower concentration of toxin required to kill half of the target organisms.

Schneider et al. (2015) developed a specific PCR method for Bti, providing an effective tool for monitoring the distribution and concentration of Bti in aquaculture environments. This method can help researchers and farm managers accurately determine the presence of Bti and evaluate the effectiveness of its application methods, ensuring that the application concentration and frequency achieve the desired mosquito larval control effect while avoiding potential negative impacts on the environment and non-target organisms.

Melo et al. (2018) further emphasized the importance of cultivating Bti in bioreactors to improve its application in aquaculture. By cultivating Bti on a large scale under controlled conditions, researchers were able to obtain high-density, high-activity Bti cultures, which is essential for achieving effective mosquito larvae control. In addition, bioreactor cultivation also provides more precise experimental conditions for studying the optimal application time, concentration and frequency of Bti, making the application strategy of Bti more scientific and effective.

1.2 Effect of different Bti formulations on the effect

Different formulations of Bti, such as suspended particles and solutions, have a significant impact on its effectiveness. Planktonic formulations are particularly suitable for use in static or slow-moving water bodies due to their longer duration and ability to form a coating on the surface of the water. This formula effectively attracts and kills surface-feeding mosquito larvae. In contrast, solution formulas are more suitable for fast action and are suitable for fast-moving water bodies or situations where mosquito larvae populations need to be reduced immediately. The adaptability of different formulas to specific water types and mosquito larvae densities determines their effectiveness and economy in practical applications.

Iors et al. (2015) emphasized the importance of formulation to improve Bti activity by using multiple carbon sources to develop a new Bti feed batch method. This study shows that by optimizing the carbon source, Bti production and toxicity can be significantly improved, thereby reducing the required Bti concentration in practical applications and achieving increased cost-effectiveness. This is particularly important for large-scale application of Bti for mosquito larvae control, especially in resource-constrained aquaculture sites.

Zghal et al. (2018) improved the yield and effectiveness of Bti biopesticides by optimizing culture media based on agricultural by-products. Their study not only demonstrated the impact of different formulations on the effectiveness of Bti, but also highlighted the impact of environmental factors, such as culture medium composition, on the expression of Bti toxicity. By optimizing the composition of the culture medium, a significant increase in Bti toxin production was achieved, which is of great significance for achieving effective mosquito larvae control.

Gopinathan and Shalini (2022) further demonstrated the potential of formulation improvement to enhance Bti activity. By chemically pretreating rice straw as a cost-effective Bti culture medium, they found that the pretreated rice straw not only provided a rich source of nutrients but also enhanced the growth and toxicity expression of Bti by improving the physical and chemical properties of the culture medium. This approach provides a feasible strategy for developing new, efficient, and low-cost Bti formulations.

1.3 Influence of environmental conditions

The activity of Bti is affected by a variety of environmental conditions, including water type, temperature, and pH. The type of water, such as ponds, ditches, or rice fields, affects the uniformity and duration of Bti distribution. Temperature is another key factor. Too high or too low temperatures may affect the activity of Bti toxins. Generally, warm environments help Bti to be active, but extreme high temperatures may reduce its effectiveness. pH also has an important impact on Bti. Overly acidic or alkaline water environments can destroy the stability of Bti spores, thereby affecting their insecticidal effects. Therefore, when planning a Bti application strategy, these environmental conditions must be taken into account, and the application plan must be adjusted as much as possible to adapt to these conditions to ensure that Bti is optimally effective.

Allgeier et al. (2018) revealed the important impact of temperature on Bti activity through research on European common frog larvae. This study found that although Bti exposure did not significantly affect the survival rate and metamorphosis completion time of larvae, it caused a significant increase in the activities of detoxifying enzymes and antioxidant enzymes, suggesting that environmental temperature may regulate the effects of Bti by affecting metabolic activities.

Brühl et al. (2020) shows that Bti, as an environmentally friendly and effective biopesticide for specific targets, may behave differently in different types of water bodies depending on the complexity of the ecosystem. For example, static or slow-flowing water bodies, such as ponds and rice fields, are more suitable for Bti's floating particle formula, because this formula can form a stable covering layer on the water surface, effectively targeting surface-feeding mosquito larvae. However, characteristics of this ecosystem may also lead to impacts at the food web level, and further assessment of the long-term ecological effects of Bti use is needed.

Gutierrez-Villagomez et al. (2021) indirectly reflected the possible impact of pH value on Bti activity by evaluating the impact of Bti commercial formulations on tadpoles. Research shows that although Bti has little effect on the survival rate, body length, weight and other indicators of tadpoles, it causes significant changes in the composition of the intestinal microbial community. This change may reflect the potential impact of Bti on interactions among ecosystem members under different pH conditions.

2 Ecological and Environmental Impacts of Bti

2.1 The control effect of Bti on target mosquito larvae populations

Bacillus thuringiensis var. israelensis (Bti) has been widely recognized in controlling target mosquito larval populations, especially against important disease-transmitting mosquito species such as Aedes aegypti. Begum et al. (2015) showed that the use of Bti aqueous suspension for the third and fourth instar larvae of Aedes aegypti reared in the laboratory can achieve the highest lethality rate of 96.66%, with an LC50 of 1.0 μl/ml. The value is 0.0097. This result highlights that Bti has a significant control effect on the target mosquito larvae population at a specific concentration (Figure 1).

Additionally, Bti-CECIF is a biopesticide designed and developed in solid tablet form for the control of disease vector mosquitoes. An evaluation of the effectiveness and residual activity of Aedes aegypti larvae by Wilber Gómez-Vargas et al. (2018) in semi-field and field conditions in two Colombian municipalities showed that the highest tested dose showed the greatest residual activity after 15 days, with Postlarval mortality is 80%.

Suwito et al. (2021) examined the effectiveness of Bti H-14 biopesticide in controlling (Aedes spp.) larval density. Continuous observation of 3171 containers in household containers found that Bti H-14 was highly effective in controlling larval density over a 6-month period. These research results not only confirm the efficient killing effect of Bti on target mosquito larvae under laboratory conditions, but also demonstrate its potential for application under semi-field and field conditions. Through precise control of different doses, Bti can significantly reduce the density of mosquito larvae in specific areas, thereby reducing the number of adult mosquitoes and effectively controlling the risk of the spread of mosquito-borne diseases.

2.2 Potential effects of Bti on non-target organisms

Bacillus thuringiensis var. israelensis (Bti) is an effective biological agent for mosquito larvae control, but its potential effects on non-target organisms remain an important topic of research. The safety of Bti is generally recognized for its high specificity against mosquito larvae, but recent studies have pointed out that long-term or high-volume use of Bti may have effects on coexisting non-target organisms. Particularly when using Bti in biodiversity protected areas and sensitive wetland ecosystems, potential impacts on non-target organisms and the ecosystem as a whole should be more deeply assessed and monitored.

Derua et al. (2018) showed that although the newly developed long-lasting microbial insecticides (LLML) FourStar and LL3 can effectively control malaria vector mosquitoes, they have almost no significant impact on the population density of symbiotic non-target organisms. The study showed that after a round of standard-dose treatment, the species richness, abundance and diversity of non-target organisms did not change significantly compared with untreated controls.

Gutierrez-Villagome et al. (2021) focused on the impact of Bti on amphibians. They found that although it had little effect on the survival rate, body length, weight and other indicators of Lithobates sylvaticus and Anaxyrus americanus tadpoles, the commercial formula of Bti significantly extended the median time to the completion of metamorphosis in L. sylvaticus tadpoles and changed A. americanus, which may have implications for individual fitness, including susceptibility to parasitic infections.

Allgeier et al. (2019) further explored the impact of Bti on non-target organisms, especially non-biting midges (chironomids). Their study found that although Bti had a significant control effect on target mosquitoes, its operational dosage also significantly reduced the overall chironomid emergence rate by about half. This reduction in large numbers of non-target organisms may result in indirect negative impacts on other aquatic life such as birds and bats.

2.3 Consideration of ecosystem service functions by Bti applications

When exploring the environmental impacts of Bacillus thuringiensis var. israelensis (Bti) as a mosquito control strategy, its potential impact on ecosystem services has become an important research focus. Ecosystem services, including pollination, water purification, soil fertility maintenance, and biological control services, are the basis for ecosystem health and stability and are essential for human well-being.

Poulin and Lefebvre (2018) revealed that the impact of Bti on non-target organisms in wetland ecosystems, especially Chironomidae populations, may indirectly affect high-trophic-level organisms such as birds and bats that rely on these non-target organisms as food sources, thereby having a negative impact on ecosystem service functions. This impact is mainly reflected in the change of food web structure, which may weaken the self-purification capacity and biodiversity of wetland ecosystems.

Kästel et al. (2017) emphasized that the sensitivity of non-target organisms to Bti varies with age, such as chironomid larvae, and the changes in their population structure after Bti treatment may affect ecosystem service functions, such as the decomposition of organic matter in water bodies and nutrient cycling processes. These changes may lead to the weakening of ecosystem service functions, affecting water purification and soil fertility maintenance.

Although Bti is highly effective and target-specific as a mosquito control tool, its long-term and large-scale application needs to consider its potential impact on ecosystem services. Future research should pay more attention to the impact of Bti application on ecosystem services, especially in biodiversity-rich and ecologically sensitive areas, to promote the rational and sustainable use of Bti.

3 Challenges and Management Strategies

3.1 Risk of mosquito resistance to Bti developing

Bacillus thuringiensis var. israelensis (Bti) for mosquito control worldwide, the emergence of mosquito resistance to Bti has attracted the attention of the scientific community. Although Bti is considered a relatively safe biological control method for non-target organisms, the emergence of resistance not only reduces the control effect of Bti, but may also force the use of more chemical insecticides, thereby bringing additional environmental and health risks.

Bonin et al. (2015) found 16 QTL regions associated with Bti resistance in a study of dengue and yellow fever vector mosquitoes, Aedes aegypti, through quantitative trait loci (QTL) and mixed analysis. Four of these QTL regions were revealed in different analysis methods, explaining 29.2% and 62.2% of the phenotypic variance in the cross between the two QTLs. This study not only reveals the complexity of Bti resistance that may involve simultaneous variation in multiple genes, but also provides important genetic markers for in-depth understanding and management of resistance.

Becker et al. (2018) conducted a study on Aedes vexans in the Upper Rhine Valley region of Germany, where Bti has been used for mosquito control for 36 years. They found that despite using almost 5,000 tonnes of the Bti formula during this long-term application, no increase in resistance to Bti was detected between Aedes vexans larvae in treated and untreated areas. This finding has important implications for assessing the sustainability of long-term Bti use and monitoring resistance development.

Stalinski et al. (2016) revealed that the impact of Bti on mosquitoes may involve changes in mosquito physiological responses. They found that the gene expression levels of 11 alkaline phosphatases (ALPs) were significantly changed in Bti-treated mosquito larvae, and that in four Bti-resistant Cry toxin or Bti-treated mosquito strains, these The activity of ALPs is reduced. This indicates that ALPs plays a key role in the toxicological effects of Bti toxins, and its changes may be related to the development of mosquito resistance to Bti.

3.2 Strategies to manage and delay the development of resistance

Managing and delaying the development of resistance to Bacillus thuringiensis var. israelensis (Bti) is key to ensuring its continued effectiveness. Multi-toxin strategy and gene deposition, bioassay and genetic analysis, application of refuge and rotation strategies, and synergistic effects of Cyt1Aa and Bin toxins can effectively delay the development of mosquito resistance to Bti through these integrated management strategies, ensuring the long-term effectiveness and sustainability of Bti as a biological control measure.

Zafar et al. (2020) showed that a strategy to manage resistance using varieties with multiple different Bt toxin genes (pyramided cotton) and RNA interference (RNAi) technology has been proposed. This approach, called multi-gene stacking and silencing (MGPS), aims to control pests by simultaneously expressing multiple Bt toxins and RNAi to knock out key genes of pests. The MGPS approach can delay or prevent the development of pest resistance to Bt cotton by delivering high doses of Bt toxins and RNAi and complying with refuge requirements.

Bonin et al. (2015) suggested regular susceptibility monitoring of mosquito populations, using bioassays and genetic analysis methods to assess the level of resistance to Bti. This includes the use of QTL (quantitative trait loci) and admixture analysis to explore genetic markers associated with Bti resistance. In this way, signs of resistance can be detected in a timely manner and control strategies can be adjusted.

Nascimento et al. (2020) showed that Cyt1Aa toxin plays a key role in promoting the entry of Bin toxin or its BinA subunit into midgut cells of mosquito larvae that lack Bin toxin receptors. This synergistic effect provides new ideas for the development of improved strategies to combat insect resistance.

3.3 Technical and practical challenges faced by Bti applications

In the process of widespread adoption of Bacillus thuringiensis var. israelensis (Bti) for mosquito control, we face multiple technical and practical challenges that affect not only the effectiveness of Bti, but also its use in public health and ecosystems. application sustainability.

The continuity and stability of Bti products is a significant technical challenge. Although Bti is widely considered to be relatively safe for non-target organisms, its long-term residue in the environment may have unknown effects on ecological balance. For example, Gutierrez-Villagomez et al. (2021) showed that the impact of Bti on amphibians may be more complex than previously thought, especially changes in their intestinal microbial communities that may affect the health and ecological functions of these organisms. In addition, the study by Johnson et al. (2020) revealed that in a mixed mangrove-salt marsh system, high mangrove coverage may significantly reduce the deposition of Bti products and reduce its lethality to mosquito larvae, indicating that the effect of Bti application is affected by Strong influence of ecosystem structure.

The practical challenges of Bti application cannot be ignored. Cost is an important factor, especially in areas with limited resources. Although Bti is a more environmentally friendly option relative to chemical pesticides, the economic cost of its production and application remains a limiting factor. Additionally, community acceptance and participation are critical to the success of Bti control programs. Effective community communication and education activities can increase public awareness of the benefits of Bti applications and increase their acceptance.

Although the application of Bti in mosquito control has faced challenges, these challenges have also prompted the development of scientific research and technological innovation. Through continued research, it will be possible to develop more efficient and sustainable Bti application strategies. For example, the research by Nascimento et al. (2020) explored the synergistic effect of Bti and other microbial toxins, providing a scientific basis for the development of new and more effective mosquito control products.

4 Discussion and Outlook

This study provides insight into the effectiveness and challenges of using Bacillus thuringiensis var. israelensis (Bti) to control mosquito larvae (Aedes spp.) in aquaculture environments. The widespread application of Bti not only significantly improves public health and safety, reducing the risk of disease transmission by effectively reducing mosquito populations, but also highlights its important role in environmental protection, especially in reducing the use of chemical pesticides and protecting non-target Biological aspects (Brühl et al., 2020).

Although Bti has gained wide acceptance and application as a biological control method, research in recent years has revealed several key challenges. The first is the development of mosquito resistance to Bti, which poses a threat to the long-term effectiveness of Bti. In addition, the effects of Bti application in different ecosystems and its potential impacts on non-target organisms and the entire ecosystem have also raised concerns. For example, recent studies have raised the need for further evaluation of the long-term ecological impacts of Bti, emphasizing the importance of continuous monitoring and adaptive management strategies to ensure the sustainability and eco-friendliness of its use (Ioannou et al., 2021).

In the aquaculture environment, the application of Bacillus thuringiensis var. israelensis (Bti) not only reflects its effectiveness in controlling (Aedes spp.) mosquito larvae, but also highlights its role in maintaining ecological balance and promoting the sustainable development of aquaculture. potential.

Continue to optimize Bti formulations and application strategies to improve its efficiency and persistence in aquaculture environments. This includes developing Bti formulations that adapt to different water conditions (such as salinity, pH and temperature), as well as exploring more precise application times and methods to reduce the impact on non-target organisms while ensuring effective control of mosquito larvae (Docile et al., 2021).

Strengthen the monitoring and assessment of the ecological effects of Bti application. Considering the complexity and diversity of the aquaculture environment, it is particularly important to conduct a comprehensive assessment of the ecological impacts of the long-term use of Bti. This includes its impact on aquatic biodiversity, non-target organisms (especially aquatic insects and amphibians), and potential risks to the health of aquaculture organisms (Becker et al., 2018).

Explore biological control methods that work synergistically with Bti to form an integrated mosquito management strategy. For example, the use of natural enemies (such as predatory fish and insects) combined with Bti can not only enhance the mosquito control effect, but also help reduce intervention in the ecosystem and promote biodiversity in aquaculture environments (Mataba et al., 2023).

Improving the public and aquaculture industry's understanding of the application value and ecological impact of Bti is the key to achieving its continuous optimization and ecological balance. Through education and training, we can enhance the understanding of Bti as an eco-friendly mosquito control method and encourage the adoption of scientific application methods and management strategies, which can effectively improve the application effect of Bti in aquaculture.

By continuously optimizing the application methods and strategies of Bti, strengthening the monitoring and evaluation of its ecological impact, and exploring synergistic effects with other biological control methods, we can effectively control mosquito populations and promote the aquaculture industry while protecting the aquaculture ecological environment. sustainable development.

With the widespread acceptance of Bacillus thuringiensis var. israelensis (Bti) for controlling mosquito larvae (Aedes spp.) in aquaculture environments, future research will face new directions and challenges. Improving the efficiency of Bti application, exploring strategies to slow the development of resistance, and comprehensively evaluating its long-term ecological impact will be the focus of research (Begum et al., 2015).

Future research needs to focus on improving the efficiency of Bti applications. This includes developing new Bti formulations and application technologies to prolong its persistence in the environment and enhance its ability to control target mosquito populations. At the same time, exploring how to improve the activity and stability of Bti without increasing the impact on non-target organisms will be the key to research (Muhammad et al., 2024).

The development of mosquito resistance to Bti has become an important factor limiting the long-term effective use of Bti. Future research needs to gain an in-depth understanding of the mechanisms of mosquito resistance to Bti and explore molecular and genetic-based strategies to slow or avoid the development of resistance. At the same time, research on comprehensive management strategies that combine the use of Bti with other non-chemical control methods may provide an effective way to slow down the development of resistance.

A comprehensive assessment of the long-term ecological impacts of Bti is particularly important given the potential increased impact of global climate change and reduced biodiversity on mosquito-borne diseases. Future research needs to assess the impact of Bti applications on biodiversity, non-target organisms, and entire ecosystem functions from an ecosystem perspective (Rahman et al., 2021). In addition, considering that climate change may change the breeding habits and distribution of mosquitoes, studying the effectiveness and applicability of Bti under different climate conditions will help develop more effective mosquito control strategies.

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