Mosquitoes are a highly biodiverse group of insects with a life cycle consisting of four stages: egg, larva, pupa and adult. They are widely distributed in various ecosystems around the globe (Huang et al., 2021). They play an important role in the ecosystem, both as predators in the food chain and as vectors of many diseases. The ecosystem roles of mosquitoes are complex, with multiple environmental and biological factors involved in different stages of their life cycle. This diversity is closely related to its criticality in the ecosystem and has attracted extensive scientific attention. Different species of mosquitoes have adapted to different survival environments, making them one of the most adaptable and survivable insects on Earth. The ecological study of mosquitoes has become particularly important due to their complex role in the ecosystem and their relationship with human health. Understanding mosquitoes' ecological habits, reproductive behaviors, and their position in the food chain can help develop more scientific mosquito management strategies and reduce their potential threats to humans and ecosystems.

Microorganisms, as the basic components of ecosystems, are widely found in all corners of nature (Qiu et al., 2023). They play vital functions in soil, water bodies, and living organisms, including beneficial symbiotic relationships, treatment of environmental pollution, and disease transmission. Microorganisms also occupy an important position in the mosquito as an ecosystem participant. The microbial community in mosquitoes includes both beneficial symbiotic microorganisms and some pathogenic microorganisms that may potentially affect mosquito behavior.

Microorganisms play a variety of key roles in natural ecosystems. Ecosystems are complex ecological communities formed by the interaction of various biotic and abiotic factors, and microorganisms are an integral part of ecosystems. Microorganisms, including bacteria, fungi, viruses and protozoa, are widely found in soil, water, atmosphere and living organisms and perform important ecological functions. They are involved in nutrient cycling in living organisms, soil formation, plant growth and maintenance of animal health. In the specific ecosystem of mosquitoes, microorganisms are not only present in the gut and surface of mosquitoes, but also interact closely with their reproduction and immune system. These microbes may have a profound effect on mosquito behavior, which in turn affects the entire mosquito population and the ecosystem in which it resides.

Although past research has revealed some of the interactions between mosquitoes and microbes, there are still many uncharted areas that need to be explored in depth. The specific mechanisms by which microbes influence mosquito behavior are unknown, particularly in the reproductive and immune systems. With the global challenge of disease transmission, understanding the role of microbes in the relationship between mosquitoes and infectious diseases is crucial for the development of more effective prevention and control strategies (Yang et al., 2023). The aim of this study was to provide a systematic overview of the potential effects of microbes on mosquito behavior, with a view to providing a basis for future research and new perspectives on mosquito ecosystem management and disease prevention and control.

1 Overview of Mosquito-microbe Interactions

1.1 Overview of mosquito life cycle and behavior

Mosquitoes are a group of insects with a complex life cycle that consists of four stages: egg, larva, pupa and adult. This process usually takes place in water bodies, where mosquitoes lay eggs, hatch into larvae, and go through the pupal stage to eventually transform into adults. The reproductive behavior of mosquitoes is closely related to their ecological adaptations; females need to feed on blood for protein to support egg development, while males feed primarily on nectar.

Mosquito behavior has important implications for their interactions with microorganisms (Gao et al., 2020). For example, mosquitoes may ingest a number of microorganisms during blood feeding, and these microorganisms may colonize the mosquito to form unique microbial communities. In addition, mosquitoes' host selection, field activities and ecological location also influence the formation of their microbial communities.

1.2 Overview of microbial species and their distribution in mosquitoes

The microorganisms in mosquitoes include bacteria, fungi, viruses and many other types. These microorganisms can be present in the mosquito's mouthparts, intestines, reproductive organs and other parts of the body (Figure 1). In the larval stage of mosquitoes, microorganisms in the water body may also enter their bodies and affect the formation of microorganisms in their bodies.

Female mosquitoes may ingest microorganisms present in the host's blood while feeding on it. These microorganisms can include pathogens such as parasites, bacteria, etc. The environmental and physiological characteristics of the mosquito's body may provide suitable conditions for certain microorganisms to survive, leading to their colonization of the mosquito's body.

1.3 Overview of what established studies have learned about mosquito-microbe interactions

Past studies have revealed the complex network of interactions between mosquitoes and microorganisms (Zhang et al., 2023). Several studies have focused on identifying and characterizing the diversity of microorganisms in mosquitoes, exploring the distribution and abundance of different species of microorganisms in mosquitoes through molecular biology techniques and microbiological approaches. This provides a basis for understanding mosquito-microbe interactions.

Other studies focus on the potential effects of microbes in mosquitoes on the ecological behavior and physiological characteristics of mosquitoes. Some microbes may directly or indirectly affect the ecological roles of mosquitoes by altering their immune system, influencing their host selection behavior, and regulating the function of the reproductive system. These studies help to reveal the mechanisms by which microbes regulate mosquito behavior and roles in the ecosystem.

There are also studies that focus on the relationship between mosquitoes and infectious diseases by examining whether the microorganisms carried by mosquitoes are potentially pathogenic and whether these microorganisms affect the ability of mosquitoes to act as disease vectors. These studies have provided new perspectives on understanding the microbial regulation of the relationship between mosquitoes and infectious diseases for disease prevention and control.

Taken together, the study of mosquito-microbe interactions involves multiple aspects of the mosquito life cycle, microbial diversity, and microbial influences on mosquito behavior and physiological characteristics. An in-depth understanding of these interactions is important for understanding the dynamic balance of the mosquito ecosystem and the mechanism of mosquito-borne diseases. Future studies should continue to dig deeper into the details of microbe-mosquito interactions to provide a more comprehensive understanding of mosquito ecology and infectious disease prevention and control.

2 Direct Effects of Microorganisms on Mosquito Behavior

2.1 Impact of mosquitoes as vectors of disease transmission

Mosquitoes have played a prominent role in human history as important vectors of disease transmission. This phenomenon is mainly attributed to the introduction of pathogens carried by mosquitoes into their hosts during their blood-sucking behavior. Pathogens such as parasites, viruses and bacteria multiply in the mosquito's body, thus making it an effective transmitter of many infectious diseases (Sun, 2019).

Microorganisms play a key role in the transmission of diseases by mosquitoes. They may be the main carriers of pathogens and are responsible for transferring pathogens from the source of infection to a new host. Microorganisms may also influence the mosquito's infective capacity and infection cycle, by modulating the mosquito's immune system or altering its host selection behavior, thus affecting the efficiency of disease transmission.

2.2 Mosquito reproduction and microbial influences

Microorganisms have a direct impact on the reproductive behavior of mosquitoes. In mosquitoes, some microorganisms may interact with their reproductive organs to influence the egg development and hatching process. This effect may manifest itself by promoting egg hatching and increasing larval survival or, conversely, by inhibiting egg development and reducing population size. These microorganisms may affect the reproductive success of mosquitoes by altering the biochemical environment within the mosquito, such as influencing hormone levels and providing specific nutrients.

Microorganisms may have a direct effect on the development of mosquitoes during their egg stage. During the process of mosquito egg laying, some microorganisms may survive with the eggs and enter a new life cycle. These microorganisms may affect egg development and hatching by altering the microenvironment of the egg, such as by providing specific nutrients or by influencing the surface properties of the egg.

The reproductive organs of mosquitoes are important settlement sites for microbial communities, including bacteria and fungi. These microorganisms may affect the reproductive health and reproductive behavior of mosquitoes. Some microorganisms may assist mosquitoes in reproduction, either by producing beneficial metabolites or by cross-fertilizing with mosquito physiological processes. Some microorganisms may adversely affect the reproductive system of mosquitoes by causing damage to reproductive organs or physiological abnormalities.

Microorganisms may influence the reproductive behavior and reproductive strategy of mosquitoes by modulating their physiological state and behavior. Some microorganisms may influence the reproductive behavior of mosquitoes by altering their immune system or endocrine system, such as the frequency and number of egg-laying and the choice of location for egg-laying. Such effects may lead to changes in mosquito populations in specific environments, thereby affecting the balance of the ecosystem.

The impact of microorganisms on the reproductive behavior of mosquitoes is a complex and multilayered process. From influencing egg development to regulating microbial communities in reproductive organs to influencing offspring over time, microbes play an important regulatory role in mosquito reproduction. An in-depth understanding of the interaction mechanisms between microbes and mosquito reproduction will help to better understand the dynamic balance in the mosquito ecosystem and provide more effective strategies for mosquito management and disease prevention and control.

2.3 Impact of changes in reproductive system microbial communities

Changes in the microbial community of the reproductive system of mosquitoes are also an important aspect of microbial influence on mosquito behavior. There are abundant microbial communities in the reproductive system of mosquitoes, including bacteria and fungi. These microbes may play different roles in mosquitoes at different reproductive stages. For example, some microorganisms may play a mediating role in the mating behavior of mosquitoes, influencing female mosquitoes to select suitable male partners.

Changes in the microbial community of the reproductive system may also be related to the mosquito's immune system, which plays a key role in maintaining the health of the reproductive system and the proper development of eggs. Microorganisms may influence the reproductive success and reproductive strategy of mosquitoes by modulating the activity of the immune system. This microbial influence on the reproductive system may be realized through direct action on the reproductive organs of mosquitoes or through modulation of the endocrine system of mosquitoes, for example.

By delving into the direct effects of microbes on mosquito reproductive behavior, the role of mosquitoes in the ecosystem and the mechanisms by which they act as disease vectors can be better understood. These studies not only provide new perspectives on the ecology and behavior of mosquitoes, but also provide theoretical support for the development of more effective mosquito control and infectious disease prevention and control strategies. In future studies, further insights into the specific mechanisms of microbial contributions to mosquito reproductive behavior are needed for a more comprehensive understanding of the complex interactions between mosquitoes and microbes.

3 Indirect Effects of Microorganisms on Mosquito Behavior

3.1 Interaction between microorganisms and mosquito immune system

There are complex and subtle interactions between microorganisms and the mosquito immune system (Parhander, 2023). The mosquito immune system is a complex network that includes both intrinsic and adaptive immunity components. Microorganisms may trigger the mosquito's intrinsic immune response, triggering a series of immune responses such as the production of antimicrobial peptides and activation of immune-related pathways.

The mosquito's immune system may also influence the survival and reproduction of microbes. For example, the mosquito immune system may inhibit microbial proliferation by producing anti-microbial molecules, such as antimicrobial peptides. This interaction creates a state of equilibrium between microbes and the mosquito immune system.

Microbes indirectly influence mosquito behavior by modulating the mosquito's immune system. Activation of the immune system may lead to a stress response in mosquitoes that affects their feeding behavior, host selection, and egg-laying behavior. Immune system regulation may affect the life cycle of mosquitoes, including developmental rate, lifespan, etc., which further affects mosquito behavioral patterns (Figure 2).

Research suggests that certain microorganisms may influence the blood-sucking behavior of mosquitoes towards their hosts by modulating their immune system. Activation of the immune system may lead mosquitoes to engage in more frequent blood-sucking in order to obtain sufficient nutrients to maintain the activity of their immune system. This change in behavior may have profound effects on mosquito-host interactions and the efficiency of disease transmission.

3.2 Microbial impacts on mosquito populations and ecosystems

Microorganisms affect mosquito populations and entire ecosystems by influencing their survival and reproduction and, in turn, their impact on mosquito populations. Mosquitoes are keystone species in many ecosystems, and their population size and distribution are important for the balance of the entire ecosystem.

Microorganisms may have an indirect effect on the size and structure of mosquito populations by influencing ecological parameters such as mosquito life cycle and reproductive success. This effect may be transmitted to the whole ecosystem by changing the interrelationships between mosquitoes and other organisms, the structure of the food chain, and so on.

3.3 Influence of microorganisms on mosquito competition and adaptation

Microorganisms may also have an effect on mosquito competition and adaptation. In the same ecosystem, different species of mosquitoes may compete with other mosquito populations for resources, including food, breeding sites, and so on. Microorganisms may affect the competitiveness of mosquitoes in an ecosystem by influencing their ecological traits, such as feeding behavior and egg-laying habits.

Microorganisms may also affect mosquito adaptations. When faced with stresses such as environmental change and climate fluctuations, microorganisms may increase the adaptability of mosquitoes to environmental change by regulating their physiological state, gene expression, and other aspects. This increased adaptability may have profound effects on mosquito survival, reproduction and population evolution.

4 Applications of Microorganisms in Mosquito Control

4.1 Application of mosquito ecology management using microorganisms

Eco-friendly mosquito control methods have become a hotspot for research and practice, and microorganisms, as a natural means of control, show great potential to realize mosquito control in the ecosystem.

Biological control is a method that utilizes natural biological factors such as natural enemies, parasites, and pathogens to control pests (Qin et al., 2022). In the mosquito ecosystem, microorganisms play a key role in this approach. By selectively introducing or increasing certain microbial species, regulation of mosquito populations can be achieved, reducing their numbers and slowing down the rate of disease transmission.

Entomopathogenic microorganisms, such as Bacillus thuringiensis and Bacillus sphaericus, are microorganisms that are widely used in eco-friendly mosquito control. The insecticidal crystal proteins produced by these microorganisms are highly selective for mosquito larvae and have a low impact on other non-target organisms, which is in line with the concept of eco-friendly control.

Biocides often contain components of microbial origin, such as mosquito-specific insect biocides. These microbial sources can be applied to mosquito breeding sites by injection or spraying to provide control. Compared with chemical pesticides, these biopesticides have less environmental impact and do not produce excessive negative effects on non-target organisms.

4.2 Application of genetic modification and mosquito tolerance

Through genetic modification technology, scientists can introduce microbial resistance genes that are harmful to mosquitoes into their genomes (Demirak et al., 2022). For example, introducing genes that are resistant to pathogens that transmit diseases in mosquitoes into mosquitoes gives them a higher level of resistance when infected with the pathogens, thus slowing down the spread of the diseases.

Genetic modification also allows for the regulation of interactions between mosquitoes and microbes. Scientists can adjust gene expression in the mosquito's immune system to make it more likely to establish symbiotic relationships with certain microbes, or to increase the mosquito's tolerance of certain beneficial microbes. This helps maintain a state of equilibrium in the ecosystem.

When conducting research on genetically engineered mosquitoes, scientists must fully consider their potential impact on the ecosystem. Microorganisms, as an important part of the mosquito ecosystem, and their interactions with genetically engineered mosquitoes need to be studied in depth to ensure that the release of genetically engineered mosquitoes has no unforeseen negative impacts on the ecological balance and microbial communities.

When conducting research on genetically engineered mosquitoes, scientists must fully consider their potential impact on the ecosystem. Microorganisms, as an important part of the mosquito ecosystem, and their interactions with genetically engineered mosquitoes need to be studied in depth to ensure that the release of genetically engineered mosquitoes has no unforeseen negative impacts on the ecological balance and microbial communities.

The application of microorganisms in mosquito control exhibits a multilayered character, including both applications in eco-friendly mosquito control methods and potential applications in the study of genetically engineered mosquitoes. By utilizing microorganisms for mosquito ecological management, precise control of mosquito populations can be achieved, slowing down the rate of disease transmission. Meanwhile, genetic engineering technology provides new means for mosquito control, and the potential application of microorganisms in this field can help realize more precise and environmentally friendly mosquito control strategies. Future research should continue to dig deeper into the mechanism of microorganisms in mosquito control and actively promote the application of microorganisms in mosquito control in practice.

5 Research Methods and Techniques

5.1 Microbial identification techniques

Microorganisms play a key role in mosquito behavioral studies, and advanced microbial identification techniques need to be applied in order to gain insights into the effects of microorganisms on mosquito behavior (Chen et al., 2022). Some commonly used microbial identification techniques will be described below to provide a viable approach in mosquito behavioral studies.

DNA sequencing technique is one of the important tools for microbial identification. By extracting DNA from microbial samples and applying high-throughput sequencing technology, the entire genome sequence information of the microorganism can be obtained. This enables researchers to accurately identify microbial species, subspecies, strains, and other detailed information, which provides a basis for further research on the relationship between microbes and mosquito behavior.

Polymerase chain reaction (PCR) is a rapid and sensitive technique for amplifying microbial DNA. With selective primers, PCR amplifies specific segments of microbial DNA, providing a rapid and effective means of identifying microorganisms. In mosquito behavioral studies, PCR can be used to detect the presence and diversity of microorganisms by collecting samples from different parts of the mosquito or the environment.

16S rRNA sequence analysis is a technique commonly used to identify bacteria and archaea. Since 16S rRNA is widely present in microorganisms, variation in its sequence can be used to identify microorganisms at taxonomic levels such as genus and species. Sequencing and analyzing the 16S rRNA of microorganisms in mosquitoes or in breeding sites can provide insight into the diversity and abundance of microorganisms.

5.2 Behavioral experimental design

Studying the effects of microorganisms on mosquito behavior requires the design of rational behavioral experiments to obtain reliable data and conclusions. Some commonly used behavioral experimental design methods are described below.

Selection experiments are an effective method to study the host selection preference of mosquitoes for different microorganisms. By setting up selection devices containing different microorganisms and observing the selection behavior of mosquitoes, the effect of microorganisms on mosquito host selection can be assessed. This experimental design can be carried out in a simulated natural environment, and to increase the reliability of the experiment, the use of field-caught mosquitoes can be considered for the experiment.

Behavioral observation experiments are an effective way to study the survival behavior of microorganisms on mosquitoes. The direct influence of microorganisms on the behavior of mosquitoes can be inferred by observing their flight, blood-sucking, egg-laying and other behaviors. Such experiments require meticulous observation and can be combined with video recordings and image analysis to quantify various aspects of mosquito behavior.

Studying the effects of microbes on mosquito reproduction usually requires reproduction experiments. By regulating the presence and number of microorganisms and observing reproductive parameters such as mosquito egg laying and egg hatching rates, the mechanisms by which microorganisms affect mosquito reproductive behavior can be revealed. Such experiments need to be conducted under controlled conditions to ensure reproducibility of the experiment.

5.3 Data analysis methods

In mosquito behavior research, reasonable analysis of experimental data is an important step to obtain scientific conclusions. Some commonly used data analysis methods are introduced below.

Statistical analysis is the basis of behavioral experimental data analysis. By using statistical methods such as t-test and ANOVA, the differences between different experimental groups can be compared. For large-scale datasets, machine learning algorithms can be considered to identify the influence patterns of different microorganisms on mosquito behavior by training models.

For spatially distributed data such as flight behavior of mosquitoes, spatial analysis methods can be used. Through tools such as geographic information system (GIS), the distribution of mosquitoes under different conditions can be visualized and quantified to reveal the influence of microorganisms on the spatial behavior of mosquitoes.

Ecological modeling is an advanced method to analyze the effect of microorganisms on mosquito behavior. By developing mathematical models that take into account the interactions between microbes, mosquitoes, and the environment, it is possible to simulate changes in mosquito behavior under different conditions. This approach helps to gain a deeper understanding of the complex relationship between microbes and mosquito behavior and provides a scientific basis for mosquito control in ecosystems.

Methods and techniques for studying microbial behavior towards mosquitoes involve a variety of fields such as microbial identification, behavioral experimental design, and data analysis (Raji and Potter, 2021). A deeper and more comprehensive understanding of the interactions between microbes and mosquito behavior can be achieved through the rational selection and application of these methods. Future research should continue to introduce new technological tools and incorporate interdisciplinary approaches to promote the continuous advancement of microbial research on mosquito behavior.

6 Case Analysis

6.1 Case one: the impact of gut microbiota on mosquito host seeking behavior

Mosquitoes, as carriers of pathogens, pose a huge threat to human health. In recent years, scientists have found that the gut microbiota of mosquitoes may have a significant impact on their behavior in finding and selecting hosts. This discovery provides new possibilities for controlling disease transmission by regulating the microbial community of mosquitoes.

Wang's research team discovered in 2011 that the gut microbiome of mosquitoes affects their adaptability and immunity (Figure 3), with different microbial communities observed at different developmental stages and influenced by diet, such as blood meal (Wang et al., 2011). This graph shows a sparse curve, with the x-axis representing the "Number of Reads Samples" and the y-axis representing the "Number of OTUs at 5% Distance". OTUs represent Operational Taxonomic Units, and in microbial ecology, OTUs are commonly used to refer to biological taxonomic units (such as species or populations). There are significant differences in the number of OTUs among different samples. The number of OTUs in habitat samples is the highest, indicating the highest microbial diversity. The number of OTUs in adult samples is generally lower than that in pupa samples, followed by larvae. The impact of food sources on diversity: Adult insect samples fed on sugar (1 day, 3 days, and 7 days) showed different OTUs, indicating that food sources have a significant impact on microbial diversity. As the adult age increases, the number of OTUs gradually increases. The impact of blood sucking on diversity: The number of OTUs in adults who have just consumed blood for 2 days is lower than that in sugar group adults, but as the time of blood sucking increases, the number of OTUs in adult samples of PBM at 4 and 7 days gradually increases. This may be related to physiological changes or microbial colonization after blood sucking. This sparse curve graph indicates significant differences in microbial diversity among samples from different developmental stages and food sources, particularly in habitat and developmental stages. Understanding these differences is of great value for in-depth research on the ecological and biological significance of insect microbial communities.

Frankel Bricker et al. found in 2019 that a common gut fungus in mosquito larvae significantly affects microbial populations and host microbial interactions (Figure 4), affecting behavioral characteristics such as host seeking (Frankel Bricker et al., 2019). The figure compares the SCML Calibrated Counts of samples at different developmental stages under two different treatments. The horizontal axis represents the developmental stage, including larvae and adults, and the vertical axis represents the SCML calibrated counts. The legend lists two types of treatments: fungal treatment (Fungal) and non-fungal treatment (Non-Fungal). The red column represents fungal treatment, while the blue column represents non-fungal treatment. The SCML count of the fungal treatment group was slightly lower than that of the non-fungal treatment group, but the difference between the two was not significant (about 10000 counts). The overlapping error lines between the two groups indicate that during the larval stage, fungal and non-fungal treatments have a relatively small impact on SCML counting. Compared with the larval stage, the SCML count in the adult stage significantly decreased, indicating a lower number of microbial communities in the adult stage. Fungal treatment (red column) showed significantly lower levels than non-fungal treatment (blue column), indicating that fungal treatment has a significant inhibitory effect on the microbial community in the adult stage. The SCML count of the non-fungal treatment group was significantly higher than that of the fungal treatment group, and the difference was significant.

The number of microbial communities in the larval stage is significantly higher than that in the adult stage, indicating that the number of microbial communities will significantly decrease with changes in insect development stage. Fungal treatment significantly inhibited the number of microbial communities during the adult stage, while the effect was relatively small during the larval stage. This may be because fungi have a stronger pathogenic or inhibitory effect on the microbial community during the adult stage. An asterisk (*) is marked in the figure to indicate statistical significance: there is a significant difference in SCML counts between the larval and adult stages (indicated by the horizontal line above the asterisk). There is a significant difference between fungal and non-fungal treatments during the adult stage. This bar chart indicates significant differences in the number of microbial communities at different developmental stages and under different treatments. The number of microbial communities in the larval stage is much higher than that in the adult stage, while fungal treatment has a significant inhibitory effect on the microbial communities in the adult stage. This difference may be related to changes in insect physiological conditions and microbial colonization environment, indicating that the adaptability and resistance of microbial communities to insects are of great significance at different developmental stages and treatment conditions.

In summary, the impact of gut microbiota on mosquito host seeking behavior provides a new perspective and method for controlling mosquito populations and preventing mosquito borne diseases. Future research needs to further explore the specific mechanisms by which different microorganisms affect mosquito behavior, as well as how to effectively apply this strategy in practical environments.

6.2 Case two: microbial regulation of mosquito reproductive behavior

The reproductive behavior of mosquitoes is a crucial aspect of their life cycle, directly related to the maintenance of the population and the ability to spread diseases. In the natural environment, the reproductive behavior of mosquitoes is influenced by various biological factors, among which the role of microorganisms has received widespread attention from researchers in recent years. This study selected Aedes aegypti, a mosquito with yellow fever, as the research object. This mosquito is not only the main carrier of yellow fever, but also can transmit various viruses such as dengue fever and Zika virus. In the natural environment, the gut microbiota of these mosquitoes includes but is not limited to lactobacilli, enterobacteria, etc. These microorganisms have been proven to affect mosquito reproductive behavior by affecting their hormone levels.

Segata et al. (2016) found that the reproductive tracts of two main malaria transmission vectors, Anopheles gambiae and Culex mosquitoes An. coluzzii, contain complex microbial communities (Figure 5), including bacteria such as Spiroplasma, which can manipulate insect reproduction and potentially affect the transmission of malaria (Segata et al., 2016). This graph shows the relative abundance of four different microbial groups (Shewanella, Rhodococcaceae, Pseudomonas, Azospira) in different sample types and geographical locations. The horizontal axis indicates the sampling location (VK5, Soumousso, VK7) and sampling time point, while the vertical axis represents relative abundance. The colors in the legend represent different sample types, including ovaries, lower reproductive tract, male accessory glands, testes, as well as two mosquito species (A. gambiae and A. coluzzii). There are significant differences in the abundance of different microbial groups among the samples. For example, Pseudomonas has a higher relative abundance in male accessory glands and lower reproductive tract, while Azospira has a lower relative abundance in all samples. In addition, geographical location and sample type also have a significant impact on the relative abundance of microbial communities. For example, Shewanella has a higher relative abundance in the VK5 sample, while Rhodococcaceae is more abundant in the Soumousso sample. This chart visually presents the ecological distribution differences of different microbial groups in different geographical locations and sample types.

These results indicate that gut microbiota significantly enhances mosquito reproductive activity by influencing hormone levels or other physiological mechanisms. Possible mechanisms include metabolic products produced by microorganisms affecting the levels of sex hormones in mosquitoes or altering their behavior to stimulate hormone expression. This discovery is of great significance for understanding the biology and control strategies of mosquitoes.

6.3 Case three: potential of using microorganisms to control mosquito borne diseases

Mosquitoes, as vectors of disease transmission, cause millions of people to contract various diseases every year, such as malaria, dengue fever, yellow fever, and Zika virus infections. Especially Aedes aegypti and Anopheles mosquitoes, which are the main carriers of yellow fever and malaria, respectively. In recent years, scientists have been searching for safer and more effective ways to reduce the spread of these diseases. The potential of using microorganisms to control mosquito borne diseases has become a research hotspot, as this method relies less on chemical insecticides and can reduce environmental impact while avoiding mosquito resistance.

Gao's research team implemented intervention measures in 2019: strategies explored include the use of genetically modified symbiotes and technologies targeting mosquito microbiota. The method includes releasing bacteria carrying blocking pathogens such as Wolbachia in mosquitoes, which significantly reduces the mosquito's ability to transmit viruses (Gao et al., 2019).

A prominent example in this regard is the use of Wolbachia, an endophytic bacterium, to intervene in the transmission ability of mosquitoes. Wolbachia can be found in various insects, but it is not commonly found in Anopheles or Aedes aegypti mosquitoes under natural conditions. Researchers introduced Wolbachia into these mosquitoes through laboratory technology, hoping to block the spread of pathogens by affecting the mosquito's reproductive system. The specific experimental method includes injecting Wolbachia bacteria into mosquito embryos through microinjection in the laboratory, so that they can carry this bacteria during the adult stage.

Aedes aegypti mosquitoes treated with Wolbachia exhibit a characteristic of reducing virus transmission ability. The experiment showed that after being injected with Wolbachia, the dengue fever virus load in these mosquitoes significantly decreased, and the virus transmission ability decreased. Wolbachia also affects the reproductive ability of mosquitoes, which can prevent non bacterial mosquitoes from reproducing normally through cytoplasmic incompatibility, gradually increasing the proportion of mosquitoes carrying bacteria in natural populations.

The strategy of using microorganisms to control mosquito borne diseases provides a new approach for global public health. The microbial control methods represented by Wolbachia have achieved significant results in experiments and practices in multiple countries, demonstrating enormous potential. However, in the promotion process, attention still needs to be paid to issues such as public awareness, ecological risks, and technical details to ensure that this strategy can truly benefit people in more regions around the world and help us better cope with the challenges posed by mosquito borne diseases.

7 Research Challenges and Future Directions

7.1 Limitations of existing research

Despite some notable advances in the study of microbial-mosquito behavioral relationships, there are still some limitations in the existing research that affect the overall understanding of this complex ecosystem.

Existing studies have focused on certain mosquito species and specific environmental conditions, while relatively few studies have been conducted on microbe-mosquito relationships in different regions, seasons, and species. Mosquito-microbe interactions may vary across geographies and ecosystems, and more extensive studies are needed to reveal these differences.

Most studies of microbe-mosquito behavioral relationships are short-term laboratory studies that lack long-term, large-scale field observations. Long-term observations can help to better understand the cumulative effects, seasonal variations, and long-term adaptations of microbes on mosquito behavior. For the life cycle of mosquitoes and the dynamics of microorganisms, long-term studies can provide more comprehensive data support.

Current studies have mostly focused on the micro level and less on integrating the overall ecosystem factors. Microbial-mosquito interactions are influenced by many environmental factors, including temperature, humidity, vegetation, and other biological communities. Considering these factors in an integrated manner will help predict the relationship between microbes and mosquito behavior more accurately.

7.2 Unanswered questions about the relationship between microbes and mosquito behavior

Although many studies have revealed the direct and indirect effects of microorganisms on mosquito behavior, there are still some unanswered questions that need to be answered by future studies (Yang and Yang, 2021).

Little is known about how specific microorganisms finely regulate the life cycle of mosquitoes. For example, some microorganisms may have an impact by influencing the reproductive success, longevity, and feeding habits of mosquitoes. Understanding how these microbes regulate specific physiological processes in mosquitoes will provide the basis for more precise mosquito control.

There may be time-scale differences in the relationship between microbes and mosquito behavior, including rapid responses over short periods of time and adaptive changes over long time scales. The current understanding of these time-scale effects is relatively limited, and future research needs to more fully account for temporal factors.

In mosquito populations, there may be differences between individuals, including genetic differences and behavioral differences. These individual differences may affect how they interact with microbes, but there is currently a relative lack of individualized research on mosquito individual differences and microbial interactions. Future research needs to focus more on the effects of individual differences on the relationship between microbes and mosquito behavior.

7.3 Future research directions and potential research questions

In order to gain a deeper understanding of the complex relationship between microbes and mosquito behavior, future research can be expanded in the following directions.

Future research could focus more on microbe-mosquito relationships at the level of population ecology, exploring the effects of microbes on the entire mosquito population and the interactions of different microbes in the mosquito population. This will help to better understand the collective effects of microbes and mosquito behavior.

Using genomics technology to dig deeper into the genetic information of mosquitoes and microbes can reveal their interaction mechanisms at the molecular level. A comprehensive analysis of mosquito and microbial genomes can reveal how changes in the expression of specific genes affect mosquito behavior.

Considering the far-reaching effects of climate change on ecosystems, future studies could focus on how climate change affects microbe-mosquito interactions. Climate change may lead to changes in ecosystem structure and function, which may affect the ecological relationship between mosquitoes and microbes.

The application of emerging technologies, such as single-cell sequencing and high-throughput microscopy techniques, is expected to provide more comprehensive data support for in-depth studies of the relationship between microbes and mosquito behavior. Meanwhile, innovative research methods, such as network ecology and metabolomics, can also bring new perspectives to the study of the relationship between microbes and mosquito behavior.

Research on the relationship between microbes and mosquito behavior still faces a series of challenges and unanswered questions. Solving these problems requires the continued development of advanced research methods and the adoption of comprehensive interdisciplinary research strategies. Future research will help to reveal a deeper relationship between microbes and mosquito behavior, and provide a scientific basis for mosquito ecology, infectious disease prevention and control.

8 Summary and Outlook

The interaction between microorganisms and mosquito behavior is a complex and multilayered ecosystem, and the review of this study has provided insight into the effects of microorganisms on mosquito behavior (Dormont et al., 2021). Microorganisms play an important role in all stages of the mosquito life cycle, covering a wide range of aspects such as host selection, reproduction, and disease transmission. In mosquito host selection, microorganisms play a key role in mosquito host selection by altering the mosquito's blood-sucking behavior towards the host, guiding it to select a specific host, or by releasing chemical signals that affect its ability to find a host. Microbial influence on mosquito reproductive behavior involves selection of breeding sites, egg production, and larval development, and microbes regulate mosquito reproduction by forming symbiotic relationships with mosquitoes or by releasing inhibitory substances. The relationship between microbes and mosquito-borne diseases has also received much attention. Microbes may slow disease transmission by increasing mosquito resistance to pathogens; some microbes may also increase the probability of mosquitoes acting as disease vectors. This complex and bidirectional relationship of influence makes the relationship of microbes between mosquitoes and infectious diseases even more complex. Overall, the influence of microorganisms on mosquito behavior is multilayered and multifactorial, and different microorganisms may produce very different effects under different environmental conditions. This profound relationship of influence largely affects mosquito ecology, disease transmission, and survival strategies, with far-reaching impacts on both micro-level and macro-level ecosystems.

Despite some important advances in the relationship between microbes and mosquito behavior, there are still many unknowns that require further in-depth research. Future research can be directed in several directions: by leveraging the regulatory effects of microbes on mosquitoes, future research can go deeper to optimize eco-friendly mosquito control methods. This includes the introduction of more targeted microbes, the development of new biological control methods, and the integration of microbes with other environmentally friendly control methods to form a more comprehensive and efficient mosquito control strategy. As genetic engineering techniques continue to evolve, researchers are beginning to explore the regulation of mosquito-microbe interactions through genetic engineering. Future studies will also need to more comprehensively assess the feasibility and potential risks of genetically engineered mosquitoes and focus on their interactions with microorganisms to ensure that potential impacts on ecosystems and human health are fully considered. As genetic engineering technology continues to evolve, researchers have begun to explore the modulation of mosquito-microbe interactions through genetic engineering. Future research will also need to more fully assess the feasibility and potential risks of genetically engineering mosquitoes, focusing on their interactions with microorganisms to ensure that potential impacts on ecosystems and human health are adequately considered. As global climate change intensifies, there may be changes in ecosystem structure and function that could in turn affect microbial-mosquito interactions. Attention can also be paid to the response of climate change to microbe-mosquito relationships to better predict the behavioral and ecological effects of mosquitoes under different climatic conditions. Taken together, the study of microbe-mosquito behavioral relationships will continue to be a challenging but highly interesting area. Through continued research, it will be possible to better utilize microorganisms to control mosquitoes, slow down the spread of disease, and make more effective contributions to human health and ecological balance.

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