1 Introduction

Mosquitoes are vectors for numerous pathogens that cause significant human diseases, including malaria, dengue, Zika, chikungunya, and yellow fever. The accurate identification of mosquito species is crucial for understanding and controlling the spread of these diseases. Different mosquito species have varying capacities to transmit specific pathogens, making species identification essential for targeted vector control strategies. For instance, Aedes aegypti and Aedes albopictus are primary vectors for dengue, Zika, and chikungunya viruses, while Culex species are significant vectors for West Nile virus and Japanese encephalitis (Weaver et al., 2018; Guarner and Hale, 2019; Jones et al., 2019; Kain et al., 2022b). The global distribution of these vectors and their associated diseases is influenced by factors such as climate change, urbanization, and increased human travel, which have led to the emergence and re-emergence of mosquito-borne diseases in new regions (Colmant et al., 2018; Rückert and Ebel, 2018; Brugueras et al., 2020).

Phylogenetic studies provide insights into the evolutionary relationships among mosquito species and their associated pathogens. These studies are essential for understanding the genetic diversity and evolutionary history of mosquitoes, which can inform vector control strategies and predict potential disease outbreaks. Phylogenetic analyses have revealed the complex interactions between mosquitoes and the viruses they transmit, highlighting the role of specific mosquito-virus interactions in the emergence of global pathogens (Rückert and Ebel, 2018; Parry et al., 2021; Coşgun et al., 2023). For example, the recently identified Bamaga virus, transmitted by Culex mosquitoes, has been shown to interfere with the replication of West Nile virus, demonstrating the importance of understanding these interactions at a molecular level (Colmant et al., 2018). Additionally, the virome diversity within mosquito species, such as Aedes aegypti and Aedes albopictus, has been explored through meta-analyses, uncovering novel viruses and providing a resource for further studies on mosquito-virus interactions (Parry et al., 2021).

This study summarizes current methods and technologies utilized for identifying mosquito species, emphasizes the importance of accurate identification in disease control and prevention efforts, reviews phylogenetic studies that shed light on mosquito-virus interactions and disease transmission dynamics, and pinpoint existing research gaps while proposing future research directions. By synthesizing existing literature, this study endeavors to advance the knowledge of mosquito species identification and phylogenetics, ultimately enhancing the development of more effective vector control strategies and bolstering efforts to mitigate mosquito-borne diseases.

2 Methods of Mosquito Species Identification

2.1 Morphological identification techniques

Morphological identification of mosquito species has traditionally been the cornerstone of entomological studies and vector control programs. This method relies on the examination of physical characteristics such as wing patterns, leg markings, and body size. However, it is often challenged by the need for expert knowledge and the potential for misidentification, especially when dealing with morphologically similar species or damaged specimens. For instance, the identification of mosquitoes from Mexico State using morphology was complemented by DNA barcoding to overcome these limitations, highlighting the necessity of integrating molecular techniques for accurate species identification (Adeniran et al., 2020).

2.2 Molecular identification methods

DNA barcoding, particularly using the mitochondrial cytochrome c oxidase subunit I (COI) gene, has become a widely adopted method for mosquito species identification. This technique involves sequencing a short, standardized region of the genome, which provides a unique genetic fingerprint for each species. Studies have demonstrated the efficacy of COI barcoding in identifying mosquito species across various regions, including Mexico (Adeniran et al., 2020), the UK (Hernández-Triana et al., 2019), and Thailand (Chaiphongpachara et al., 2022). The method has proven effective in distinguishing species with high accuracy, even in cases where morphological identification is challenging due to damaged specimens or cryptic species complexes.

Polymerase Chain Reaction (PCR) and Single Nucleotide Polymorphism (SNP) genotyping are other molecular techniques used for mosquito identification. PCR-based methods can amplify specific DNA regions to detect and identify mosquito species and their associated pathogens. For example, a targeted amplicon sequencing panel was developed to simultaneously identify mosquito species and Plasmodium presence across the Anopheles genus, demonstrating the utility of multiplex PCR in large-scale genetic surveillance (Makunin et al., 2021). SNP genotyping, on the other hand, focuses on identifying genetic variations at single nucleotide positions, which can be used to generate species-specific barcodes. A decision tree-based barcoding algorithm utilizing SNPs has been proposed for the rapid and reliable identification of Anopheles species (Swain et al., 2019).

2.3 Advancements in high-throughput sequencing technologies

High-throughput sequencing technologies have revolutionized mosquito species identification and phylogenetic studies. These technologies allow for the simultaneous sequencing of multiple genetic loci or entire genomes, providing comprehensive insights into genetic diversity and evolutionary relationships. For instance, a new method of DNA metabarcoding was developed to detect microsporidian infections and identify host mosquito species using next-generation sequencing, showcasing the potential of high-throughput approaches in uncovering hidden biodiversity (Trzebny et al., 2020). Additionally, the use of multilocus amplicon sequencing has enabled the identification of mosquito species and the detection of malaria parasites in a single sequencing run, highlighting the efficiency and scalability of these methods for vector control programs (Makunin et al., 2021).

High-throughput genomic sequencing has also facilitated detailed phylogenetic analyses, as demonstrated in studies on the Culex pipiens complex and the Neotropical mosquito fauna. These studies have provided robust support for the monophyly of various mosquito subfamilies and tribes, offering valuable insights into the evolutionary history and diversification of mosquitoes (Adeniran et al., 2020; Lorenz et al., 2021). The integration of high-throughput sequencing with advanced analytical tools continues to enhance our understanding of mosquito phylogenetics and species identification, paving the way for more effective vector control strategies.

3 Phylogenetic Analysis in Mosquitoes

3.1 Importance of phylogenetics in mosquito research

Phylogenetic analysis is crucial in mosquito research as it helps elucidate the evolutionary relationships and divergence times among mosquito species, which are essential for understanding their role in disease transmission and developing control strategies. For instance, the study of the mitochondrial genomes of 102 mosquito species revealed significant insights into the evolutionary history of the Culicidae family, including the divergence of the Anophelinae and Culicinae subfamilies during the early Jurassic period (Lorenz et al., 2021). Additionally, phylogenetic studies can inform public health strategies by identifying species complexes and cryptic species that may have different vectorial capacities, as seen in the analysis of the Anopheles neivai species (López-Rubio et al., 2019).

3.2 Common phylogenetic methods used

Several phylogenetic methods are commonly employed in mosquito research, including Maximum Likelihood (ML) and Bayesian Inference (BI). These methods provide robust frameworks for constructing phylogenetic trees and estimating divergence times. For example, Bayesian relaxed clock methods were used to estimate divergence times in a study of Neotropical mosquitoes, providing robust support for the monophyly of several mosquito tribes (Lorenz et al., 2021). Similarly, ML and BI analyses were utilized to investigate the phylogenetic relationships within the Culex pipiens complex, revealing intricate patterns of genetic differentiation and historical admixture (Figure 1) (Aardema et al., 2020). Other methods, such as phylogenetic networks and SNP barcoding, have also been applied to uncover evolutionary patterns and species identification (López-Rubio et al., 2019; Swain et al., 2019).

3.3 Challenges and limitations in phylogenetic studies

Phylogenetic studies in mosquitoes face several challenges and limitations. One major challenge is the resolution of phylogenetic relationships among closely related species or within species complexes, as seen in the Culex pipiens complex, where ongoing genetic exchange and historical admixture obscure clear evolutionary histories (Aardema et al., 2020). Additionally, the choice of genetic markers and the quality of sequence data can significantly impact the results. For instance, the COI gene has been widely used for DNA barcoding, but it may not always provide sufficient resolution to distinguish between closely related species, as observed in British mosquitoes (Anoopkumar et al., 2019). Furthermore, the variability in phylogenetic relationships depending on the method used (e.g., concatenated vs. partitioned analyses) and the number of taxa sampled can complicate the interpretation of results (Lorenz et al., 2021). Finally, the integration of morphological, behavioral, and ecological data with molecular phylogenetics is essential for a comprehensive understanding of mosquito evolution and vectorial capacity (Anoopkumar et al., 2019; López-Rubio et al., 2019).

4 Global Diversity of Mosquito Species

4.1 Regional distribution and diversity patterns

Mosquito species exhibit significant regional diversity and distribution patterns influenced by various ecological and environmental factors. In the Caatinga biome of Brazil, a study identified 82 morphospecies, with 47 species unique to specific areas, highlighting the region's high mosquito diversity and the importance of environmental heterogeneity in shaping these patterns (Andrade et al., 2020). Similarly, in Rio de Janeiro, Brazil, mosquito diversity was assessed across urban, periurban, and rural landscapes, revealing higher species richness in rural areas and significant habitat segregation along an urban-forest gradient (Câmara et al., 2020). In Saudi Arabia, the phylogenetic analysis of Anopheles species from different zoogeographic zones demonstrated the region's diverse mosquito fauna, influenced by its unique geographic position (Munawar et al., 2020). In Honduras, the distribution and genetic diversity of Anopheles species were studied, with Anopheles albimanus being the most widespread, indicating the importance of both taxonomic and molecular approaches in understanding regional mosquito diversity (Escobar et al., 2020).

4.2 Endemic and invasive species

Endemic and invasive mosquito species play crucial roles in local and global ecosystems. The Culex pipiens complex, which includes several taxa of medical importance, presents phylogenetic challenges due to historical and contemporary admixture, highlighting the complexity of endemic species' evolutionary trajectories (Figure 2) (Aardema et al., 2021). Invasive species such as Aedes albopictus, the Asian tiger mosquito, have shown distinct genetic clusters in Reunion Island and Europe, suggesting independent invasion histories and significant genetic diversity in long-established populations (Sherpa et al., 2018). The global evaluation of the Culex pipiens complex further underscores the intricate taxonomic relationships and admixture within this group, with implications for understanding disease transmission dynamics (Aardema et al., 2020). In the UK, DNA barcoding has been employed to identify mosquito species, discover cryptic genetic diversity, and monitor invasive species, demonstrating the utility of molecular tools in managing mosquito populations (Hernández-Triana et al., 2019).

4.3 Impact of environmental factors on mosquito diversity

Environmental factors significantly impact mosquito diversity and distribution. In the Caatinga biome, the observed dissimilarity in mosquito communities is attributed to environmental heterogeneity and anthropogenic interference, which affect species' intrinsic relationships with their habitats (Andrade et al., 2020). In Rio de Janeiro, the impact of human activities on landscape and mosquito populations was evident, with higher species richness in rural areas and habitat segregation along an urban-forest gradient (Câmara et al., 2020). The phylogenetic analysis of mosquitoes in the Neotropical region revealed that major lineages arose after the Cretaceous, coinciding with the emergence of angiosperms and the expansion of mammals and birds, suggesting that historical environmental changes have shaped mosquito diversification (Lorenz et al., 2021). In Saudi Arabia, the diverse mosquito fauna is influenced by the region's unique geographic position, bordering multiple zoogeographic zones (Munawar et al., 2020). These studies collectively highlight the complex interplay between environmental factors and mosquito diversity, emphasizing the need for comprehensive ecological and genetic analyses to inform vector control strategies.

5 Evolutionary Relationships Among Mosquito Species

5.1 Phylogenetic relationships within major genera (Anopheles, Aedes, Culex, etc.)

Phylogenetic studies have provided significant insights into the evolutionary relationships within major mosquito genera such as Anopheles, Aedes, and Culex. For instance, the phylogenetic analysis of Anopheles species in Saudi Arabia revealed distinct lineages corresponding to different zoogeographic zones, highlighting the monophyletic nature of An. stephensi and its various ecotypes (Munawar et al., 2020). Similarly, a comprehensive study on the mitochondrial genomes of 102 mosquito species, including representatives from 21 genera, confirmed the monophyly of the subfamily Anophelinae and several tribes within Culicinae, such as Aedini and Culicini (Lorenz et al., 2021). This study also suggested that the diversification of these groups coincided with significant geological and biological events, such as the emergence of angiosperms and the expansion of mammals and birds (Lorenz et al., 2021).

In the case of the Culex pipiens complex, high-throughput genomic sequencing has revealed intricate evolutionary patterns influenced by natural selection and demographic processes. This complex provides a unique system for studying speciation and taxonomic radiation, with implications for understanding the evolutionary trajectories of other cosmopolitan and invasive species (Aardema et al., 2020). Additionally, the use of the cytochrome oxidase c subunit I (COI) gene has been instrumental in elucidating the genetic divergence and evolutionary history of various mosquito species, confirming distinct clustering within genera and providing evidence for species complex formation (Anoopkumar et al., 2019).

5.2 Coevolution with hosts and pathogens

The coevolution of mosquitoes with their hosts and pathogens is a critical aspect of their evolutionary history. Wolbachia symbionts, for example, have been found in multiple mosquito species and play a significant role in their biology. Phylogenetic analysis of Wolbachia infections in mosquitoes has demonstrated multiple origins of infection and genetic links between mosquito and non-mosquito hosts, suggesting recent strain recombination and symbiont transfers (Shaikevich et al., 2019). This coevolutionary relationship can influence mosquito fitness, reproductive success, and vector competence, thereby affecting disease transmission dynamics.

Moreover, the geographic distribution and prevalence of mosquito species are influenced by their interactions with hosts and pathogens. For instance, the identification of new mosquito species in Charlotte County, Florida, underscores the impact of changing climates and increased global connectivity on mosquito distribution (Kovach et al., 2022). These changes can alter the coevolutionary dynamics between mosquitoes, their hosts, and the pathogens they transmit, potentially leading to the emergence of new vector-borne diseases.

5.3 Implications of evolutionary history on disease transmission

The evolutionary history of mosquitoes has profound implications for disease transmission. The phylogenetic relationships within mosquito genera can inform vector control strategies by identifying key species and lineages involved in disease transmission. For example, understanding the genetic divergence and evolutionary history of vector mosquitoes can help in developing targeted interventions to disrupt transmission cycles (Anoopkumar et al., 2019).

Additionally, the coevolution of mosquitoes with Wolbachia symbionts and other pathogens can influence their vector competence. The presence of Wolbachia in mosquito populations has been shown to reduce the transmission of certain pathogens, offering a potential biocontrol strategy for vector-borne diseases (Shaikevich et al., 2019). Furthermore, the phylogenetic analysis of mosquito species in different zoogeographic zones can provide insights into the spread of diseases like malaria, as seen in the study of Anopheles species in Saudi Arabia (Munawar et al., 2020).

In conclusion, the evolutionary relationships among mosquito species, their coevolution with hosts and pathogens, and the implications of their evolutionary history on disease transmission are critical areas of research. These insights can inform the development of effective vector control strategies and improve our understanding of the dynamics of vector-borne diseases.

6 Case Study

6.1 Selection of study location

The selection of study locations for mosquito species identification and phylogenetic analysis is crucial to understanding the diversity and evolutionary relationships of these vectors across different ecological zones. For instance, the Neotropical region was chosen for a study that investigated the mitochondrial genomes of 102 mosquito species, emphasizing the region's rich biodiversity and its implications for mosquito evolution and public health (Lorenz et al., 2021). Similarly, the Kingdom of Saudi Arabia (KSA) was selected due to its unique position bordering the Afrotropical, Oriental, and Palaearctic zoogeographic zones, which contributes to its diverse mosquito fauna (Munawar et al., 2020). The Colombian rainforest, particularly the Sierra Nevada de Santa Marta, was another significant location due to its natural ecosystem and the presence of sylvatic mosquito species that are potential vectors for arboviruses and parasites (Muñoz-Gamba et al., 2021). These diverse locations provide a comprehensive understanding of mosquito species' phylogenetics and their role in disease transmission.

6.2 Data collection

The methodologies employed in these studies typically involve a combination of classical taxonomy and molecular techniques. In the Neotropical study, mitochondrial genomes were sequenced, and Bayesian relaxed clock methods were used to estimate divergence times, providing robust phylogenetic analyses (Lorenz et al., 2021). In KSA, mosquito larvae were collected from various regions and identified morphologically using pictorial keys. Molecular characterization was performed using single and multi-locus analysis of the internal transcribed spacer 2 (ITS2) region and cytochrome oxidase c subunit I (COI) (Munawar et al., 2020). In the Colombian rainforest study, manual capture methods were used to collect mosquitoes, which were then identified via classical taxonomy. The COI marker was used for species confirmation, and phylogenetic analysis was performed using the neighbor-joining method with the Kimura-2-Parameters model (Figure 3) (Muñoz-Gamba et al., 2021). These methodologies ensure accurate species identification and provide insights into the genetic variability and phylogenetic relationships of mosquito species.

6.3 Key findings and implications

The key findings from these studies highlight the complexity and diversity of mosquito species and their evolutionary relationships. The Neotropical study revealed that the two mosquito subfamilies, Anophelinae and Culicinae, diverged in the early Jurassic, with most major lineages arising after the Cretaceous. This diversification is linked to the emergence of angiosperms and the expansion of mammals and birds, suggesting that geographic isolation due to continental drift played a role in the worldwide distribution of Culicidae (Lorenz et al., 2021). In KSA, the phylogenetic analysis showed that An. stephensi is a monophyletic species with different ecotypes found in various geographic regions, emphasizing the need for comprehensive phylogenetics and population genetics studies to understand their role in malarial transmission (Munawar et al., 2020). The Colombian rainforest study identified several mosquito species and demonstrated the utility of combining classical and molecular taxonomy for accurate species identification, especially when morphological characteristics are not well preserved (Muñoz-Gamba et al., 2021).

These findings have significant implications for public health and vector control strategies. Understanding the phylogenetic relationships and genetic variability of mosquito species can inform the development of targeted control measures and improve the accuracy of species identification, which is essential for effective vector surveillance and disease prevention programs.

7 Implications for Public Health and Disease Control

7.1 Role of accurate species identification in vector control

Accurate identification of mosquito species is crucial for effective vector control and disease prevention. Misidentification can lead to ineffective control measures and wasted resources. For instance, the study in Spain highlighted the importance of integrating morphological and genetic analyses to accurately identify mosquito species, which is the first step in establishing a robust vector surveillance program (Ruíz-Arrondo et al., 2020). Similarly, DNA barcoding has proven to be an effective molecular approach for identifying mosquito species in Thailand, ensuring precise targeting of vector control efforts (Chaiphongpachara et al., 2022). The use of geometric morphometrics of mosquito wings also provides a reliable method for species identification, even when specimens are damaged, which is often the case in field collections (Chonephetsarath et al., 2021).

7.2 Phylogenetic insights into vector competence and disease dynamics

Phylogenetic studies offer valuable insights into the vector competence of different mosquito species and their role in disease transmission. For example, research on Australian mosquito species has reinforced canonical virus-vector groupings but also revealed significant variations within these groupings, highlighting the complexity of arbovirus transmission dynamics (Kain et al., 2022a; 2022b). Phylogenetic analysis of a novel densovirus isolated from Aedes albopictus demonstrated varied pathogenicity depending on the host species, which could inform the development of targeted biological control agents (Li et al., 2019). Additionally, the study of the virome of Aedes aegypti and Aedes albopictus has expanded our understanding of the diversity and evolution of viruses within these vectors, which can influence their susceptibility to arbovirus infections and impact disease dynamics (Parry et al., 2021).

7.3 Strategies for integrated mosquito management

Integrated mosquito management (IMM) strategies are essential for controlling mosquito populations and reducing the transmission of mosquito-borne diseases. The emergence of insecticide resistance poses a significant challenge to traditional vector control methods. A study on Aedes aegypti populations in Cabo Verde demonstrated the utility of targeted amplicon sequencing to monitor insecticide resistance mutations, providing a rapid and cost-effective tool for informing control strategies (Collins et al., 2022). The use of mosquito-specific entomopathogenic viruses, such as the novel densovirus AalDV-7, offers a promising alternative to chemical pesticides, with potential applications in biological control programs (Li et al., 2019). Furthermore, metatranscriptomic sequencing of individual mosquitoes can identify vectors, emerging pathogens, and reservoirs in a single assay, providing comprehensive data to support public health surveillance and intervention decisions (Batson et al., 2020).

By integrating accurate species identification, phylogenetic insights, and innovative control strategies, public health programs can enhance their effectiveness in managing mosquito populations and mitigating the impact of mosquito-borne diseases.

8 Technological Advances and Future Directions

8.1 Innovations in identification technologies (CRISPR, AI, etc.)

Recent advancements in mosquito species identification have leveraged cutting-edge technologies such as CRISPR and artificial intelligence (AI). CRISPR technology has been instrumental in gene editing, allowing for precise modifications that can help in understanding mosquito genetics and potentially controlling vector populations. AI, on the other hand, has been utilized to enhance the accuracy of species identification through image recognition and machine learning algorithms. For instance, wing geometric morphometrics combined with AI has shown high accuracy in identifying mosquito species, even in cases where traditional morphological methods fail due to damaged specimens or cryptic species complexes (Souza et al., 2020). Additionally, the use of mitochondrial DNA barcoding has proven effective in accurately identifying mosquito species, as demonstrated in studies conducted in Thailand (Chaiphongpachara et al., 2022) and India (Anoopkumar et al., 2019). These innovations are paving the way for more reliable and efficient mosquito identification methods, which are crucial for vector control and disease prevention.

8.2 Future trends in phylogenetic research

The field of phylogenetic research is evolving with the integration of advanced genomic sequencing technologies and sophisticated analytical tools. High-throughput genomic sequencing has enabled detailed analyses of evolutionary patterns in mosquito genes and genomes, providing insights into speciation and taxonomic radiation (Aardema et al., 2020). Mitogenome-based phylogenetic studies have revealed significant evolutionary relationships and divergence times within the Culicidae family, highlighting the impact of historical events such as the emergence of angiosperms and the expansion of mammals and birds on mosquito diversification (Chen et al., 2023). Future trends in phylogenetic research will likely focus on integrating multi-locus data and employing Bayesian and maximum-likelihood methods to resolve ambiguous relationships among mosquito clades (Lorenz et al., 2021; Munawar et al., 2020). These approaches will enhance our understanding of mosquito evolution and inform strategies for vector control and disease mitigation.

8.3 Potential for integrating multi-omics approaches

The integration of multi-omics approaches, including genomics, transcriptomics, proteomics, and metabolomics, holds great potential for advancing mosquito research. These approaches can provide a comprehensive understanding of mosquito biology, from gene expression and protein function to metabolic pathways. For example, the application of RNA sequencing (RNA-seq) has allowed for the simultaneous assessment of host and viral RNA in mosquito specimens, improving our understanding of mosquito-virus interactions (Koh et al., 2023). Additionally, the study of the virome of Aedes mosquitoes has uncovered a diverse array of insect-specific viruses, which can influence mosquito susceptibility to arbovirus infections (Parry et al., 2021). By combining data from various omics platforms, researchers can gain deeper insights into the molecular mechanisms underlying mosquito behavior, vector competence, and evolutionary dynamics. This holistic approach will be instrumental in developing novel vector control strategies and enhancing our ability to predict and respond to emerging mosquito-borne diseases.

In conclusion, the integration of innovative identification technologies, advanced phylogenetic methods, and multi-omics approaches is transforming mosquito research. These advancements are not only improving our understanding of mosquito biology and evolution but also providing new tools and strategies for effective vector control and disease prevention.

9 Concluding Remarks

The identification and phylogenetic analysis of mosquito species are crucial for understanding their role in disease transmission and for developing effective control strategies. Various methods, including morphological identification, DNA barcoding, and mitogenome analysis, have been employed to identify and classify mosquito species. Morphological identification remains a fundamental approach, but it is often challenged by the lack of taxonomic expertise and the difficulty in interpreting morphological characters. DNA barcoding, particularly using the mitochondrial cytochrome c oxidase subunit I (COI) gene, has proven effective in accurately identifying mosquito species, even when morphological features are damaged or indistinguishable. Mitogenome-based phylogenetic studies have provided robust support for the monophyly of major mosquito subfamilies and tribes, revealing significant evolutionary insights. Additionally, the study of the virome of mosquitoes has expanded our understanding of the interactions between insect-specific viruses and arboviruses, which could influence mosquito susceptibility to disease transmission.

Despite significant advancements, several gaps remain in our understanding of mosquito identification and phylogenetics. One major challenge is the limited availability of comprehensive identification keys and taxonomic expertise, particularly in regions with high mosquito diversity. There is also a need for more extensive molecular and phylogenetic studies to resolve ambiguous relationships between mosquito clades and to identify cryptic species. The genetic variability and evolutionary dynamics of mosquito populations across different geographic regions require further investigation to understand their role in disease transmission and adaptation to changing environments. Additionally, the interactions between insect-specific viruses and arboviruses within mosquito hosts are not fully understood, necessitating more research to elucidate their impact on vector competence.

The future of mosquito identification and phylogenetics lies in the integration of multiple approaches, including morphological, molecular, and genomic methods. Advances in high-throughput genomic sequencing and bioinformatics tools will enable more detailed and comprehensive analyses of mosquito genomes, providing deeper insights into their evolutionary history and speciation mechanisms. The development of more accessible and affordable DNA barcoding techniques will facilitate accurate species identification, even in resource-limited settings. Furthermore, interdisciplinary research combining entomology, virology, and ecology will be essential to understand the complex interactions between mosquitoes, their pathogens, and the environment. By addressing the current gaps in knowledge and leveraging new technologies, we can enhance our ability to monitor and control mosquito populations, ultimately reducing the burden of mosquito-borne diseases globally.

Acknowledgments

Authors sincerely thank all the experts and scholars who reviewed the manuscript of this study. Their valuable comments and suggestions have contributed to the improvement of this study.

Conflict of Interest Disclosure

Authors affirm that this research was conducted without any commercial or financial relationships that could be construed as a potential conflict of interest.

References

Aardema M., Olatunji S., and Fonseca D., 2021, The enigmatic Culex pipiens (Diptera: Culicidae) species complex: phylogenetic challenges and opportunities from a notoriously tricky mosquito group, Annals of the Entomological Society of America, 115: 95-104.

https://doi.org/10.1093/aesa/saab038

Aardema M., Vonholdt B., Fritz M., and Davis S., 2020, Global evaluation of taxonomic relationships and admixture within the Culex pipiens complex of mosquitoes, Parasites & Vectors, 13: 8.

https://doi.org/10.1186/s13071-020-3879-8

Adeniran A., Hernández-Triana L., Ortega-Morales A., Garza-Hernández J., Cruz-Ramos J., Chan-Chable R., Vázquez-Marroquín R., Huerta-Jiménez H., Nikolova N., Fooks A., and Rodríguez‐Pérez M., 2020, Identification of mosquitoes (Diptera: Culicidae) from mexico state, mexico using morphology and COI DNA barcoding, Acta tropica, 213: 105730.

https://doi.org/10.1016/j.actatropica.2020.105730

Andrade D., Morais S., Marteis L., Gama R., Freire R., Rekowski B., Ueno H., and Corte R., 2020, Diversity of mosquitoes (Diptera: Culicidae) in the caatinga biome, brazil, from the widespread to the endemic, Insects, 11: 68.

https://doi.org/10.3390/insects11080468

Anoopkumar A., Puthur S., Rebello S., and Aneesh E., 2019, Molecular characterization of Aedes, Culex, Anopheles, and armigeres vector mosquitoes inferred by mitochondrial cytochrome oxidase i gene sequence analysis, Biologia, 24: 1-14.

https://doi.org/10.2478/s11756-019-00231-0

Batson J., Dudas G., Haas-Stapleton E., Kistler A., Li L., Logan P., Ratnasiri K., and Retallack H., 2020, Single mosquito metatranscriptomics identifies vectors, emerging pathogens and reservoirs in one assay, ELife, 10: 68353.

https://doi.org/10.7554/eLife.68353

Brugueras S., Martínez B., Puente J., Figuerola J., Porro T., Rius C., Larrauri A., and Gómez-Barroso D., 2020, Environmental drivers, climate change and emergent diseases transmitted by mosquitoes and their vectors in southern europe: a systematic review, Environmental Research, 10: 110038.

https://doi.org/10.1016/j.envres.2020.110038

Câmara D., Pinel C., Rocha G., Codeço C., and Honório N., 2020, Diversity of mosquito (Diptera: Culicidae) vectors in a heterogeneous landscape endemic for arboviruses, Acta Tropica, 212: 105715.

https://doi.org/10.1016/j.actatropica.2020.105715

Chaiphongpachara T., Changbunjong T., Laojun S., Nutepsu T., Suwandittakul N., Kuntawong K., Sumruayphol S., and Ruangsittichai J., 2022, Mitochondrial dna barcoding of mosquito species (Diptera: Culicidae) in thailand, PLOS ONE, 17(9): e0275090.

https://doi.org/10.1371/journal.pone.0275090

Chen D., He S., Fu W., Yan Z., Hu Y., Yuan H., Wang M., and, Chen, B., 2023, Mitogenome-based phylogeny of mosquitoes (Diptera: Culicidae), Insect science, 31(2): 599-612.

https://doi.org/10.1111/1744-7917.13251

Chonephetsarath S., Raksakoon C., Sumruayphol S., Dujardin J., and Potiwat R., 2021, The unequal taxonomic signal of mosquito wing cells, Insects, 12(5): 376.

https://doi.org/10.3390/insects12050376

Collins E., Phelan J., Hubner M., Spadar A., Campos M., Ward D., Acford-Palmer H., Gomes A., Silva K., Gomez L., Clark T., and Campino S., 2022, A next generation targeted amplicon sequencing method to screen for insecticide resistance mutations in Aedes aegypti populations reveals a rdl mutation in mosquitoes from cabo verde, PLOS Neglected Tropical Diseases, 16(2): e0010935.

https://doi.org/10.1371/journal.pntd.0010935

Colmant A., Hall-Mendelin S., Ritchie S., Bielefeldt-Ohmann H., Harrison J., Newton N., Brien C., Cazier C., Johansen C., Hobson-Peters J., Hall R., and Hurk A., 2018, The recently identified flavivirus bamaga virus is transmitted horizontally by Culex mosquitoes and interferes with west nile virus replication in vitro and transmission in vivo, PLoS Neglected Tropical Diseases, 12: 86.

https://doi.org/10.1371/journal.pntd.0006886

Coşgun Y., Bayrakdar F., Akiner M., Giray B., Demirci B., Bedir H., Korukluoglu G., Topluoglu S., and Kılıç S., 2023, Investigation of the presence of zika, dengue, chikungunya, and west nile virus in Aedes type mosquitoes in the eastern black sea area of turkey, Turkish Bulletin of Hygiene and Experimental Biology, 80(1): 101-108.

https://doi.org/10.5505/turkhijyen.2023.58235

Escobar D., Ascencio K., Ortiz A., Palma A., and Fontecha G., 2020, Distribution and phylogenetic diversity of Anopheles species in malaria endemic areas of honduras in an elimination setting, Parasites & Vectors, 10: 31.

https://doi.org/10.1186/s13071-020-04203-1

Guarner J., and Hale G., 2019, Four human diseases with significant public health impact caused by mosquito-borne flaviviruses: west nile, zika, dengue and yellow fever, Seminars in Diagnostic Pathology, 36(3): 170-176.

https://doi.org/10.1053/j.semdp.2019.04.009

Hernández-Triana L., Brugman V., Nikolova N., Ruiz-Arrondo I., Barrero E., Thorne L., Marco M., Krüger A., Lumley S., Johnson N., and Fooks A., 2019, DNA barcoding of british mosquitoes (diptera, culicidae) to support species identification, discovery of cryptic genetic diversity and monitoring invasive species, ZooKeys, 832: 57-76.

https://doi.org/10.3897/zookeys.832.32257

Jones R., Kulkarni M., Davidson T., and Talbot B., 2019, Arbovirus vectors of epidemiological concern in the americas: a scoping review of entomological studies on zika, dengue and chikungunya virus vectors, PLoS ONE, 15: 53.

https://doi.org/10.1371/journal.pone.0220753

Kain M., Skinner E., Athni T., Ramírez A., ErinA M., and Hurk A., 2022a, Not all mosquitoes are created equal: incriminating mosquitoes as vectors of arboviruses, medRxiv, 8: 22272101.

https://doi.org/10.1101/2022.03.08.22272101

Kain M., Skinner E., Athni T., Ramírez A., Mordecai E., and Hurk A., 2022b, Not all mosquitoes are created equal: a synthesis of vector competence experiments reinforces virus associations of australian mosquitoes, PLoS Neglected Tropical Diseases, 16: 68.

https://doi.org/10.1371/journal.pntd.0010768

Koh, C., Frangeul L., Blanc H., Ngoagouni C., Boyer S., Dussart P., Grau N., Girod R., Duchemin J., and Saleh M., 2023, Ribosomal rna (rrna) sequences from 33 globally distributed mosquito species for improved metagenomics and species identification, ELife, 12: 76.

https://doi.org/10.7554/eLife.82762

Kovach B., Reeves L., Domingo C., L'heureux S., Burger G., Schermerhorn S., and Riles M., 2022, Aedes pertinax, Anopheles perplexens, Culex declarator, and cx. interrogator: an update of mosquito species records for charlotte county, florida, Journal of the American Mosquito Control Association, 38(4): 241-249.

https://doi.org/10.2987/22-7087

Li J., Dong Y., Sun Y., Lai Z., Zhao Y., Liu P., Gao Y., Chen X., and Gu J., 2019, A novel densovirus isolated from the asian tiger mosquito displays varied pathogenicity depending on its host species, Frontiers in Microbiology, 10: 54.

https://doi.org/10.3389/fmicb.2019.01549

López-Rubio A., Suaza-Vasco J., Solari S., Gutiérez-Builes L., Porter C., and Uribe S., 2019, Intraspecific phylogeny of Anopheles (kerteszia) neivai howard, dyar & knab 1913, based on mitochondrial and nuclear ribosomal genes, Infection, Genetics and Evolution: Journal of Molecular Epidemiology and Evolutionary Genetics in Infectious Diseases, 67: 183-190.

https://doi.org/10.1016/j.meegid.2018.10.013

Lorenz C., Alves J., Foster P., Suesdek L., and Sallum M., 2021, Phylogeny and temporal diversification of mosquitoes (Diptera: Culicidae) with an emphasis on the neotropical fauna, Systematic Entomology, 46: 89.

https://doi.org/10.1111/syen.12489

Makunin A., Korlević P., Park N., Goodwin S., Waterhouse R., Wyschetzki K., Jacob C., Davies R., Kwiatkowski D., Laurent B., Ayala D., and Lawniczak M., 2021, A targeted amplicon sequencing panel to simultaneously identify mosquito species and plasmodium presence across the entire Anopheles genus, Molecular ecology resources, 22: 28-44.

https://doi.org/10.1111/1755-0998.13436

Munawar K., Saleh A., Afzal M., Qasim M., Khan K., Zafar M., and Khater E., 2020, Molecular characterization and phylogenetic analysis of anopheline (Anophelinae: Culicidae) mosquitoes of the oriental and afrotropical zoogeographic zones in saudi arabia, Acta Tropica, 10: 105494.

https://doi.org/10.1016/j.actatropica.2020.105494

Muñoz-Gamba A., Laiton-Donato K., Perdomo-Balaguera E., Castro L., Usme-Ciro J., and Parra-Henao G., 2021, Molecular characterization of mosquitoes (Diptera: Culicidae) from the colombian rainforest, Revista Do Instituto De Medicina Tropical De São Paulo, 63: 24.

https://doi.org/10.1590/S1678-9946202163024

Parry R., James M., and Asgari S., 2021, Uncovering the worldwide diversity and evolution of the virome of the mosquitoes Aedes aegypti and Aedes albopictus, Microorganisms, 9: 53.

https://doi.org/10.3390/microorganisms9081653

Rückert C., and Ebel G., 2018, How do virus-mosquito interactions lead to viral emergence, Trends in Parasitology, 34(4): 310-321.

https://doi.org/10.1016/j.pt.2017.12.004

Ruíz-Arrondo I., Hernández-Triana L., Nikolova N., Fooks A., and Oteo J., 2020, Integrated approaches in support of taxonomic identification of mosquitoes (Diptera: Culicidae) in vector surveillance in spain, Vector borne and zoonotic diseases, 10: 62.

https://doi.org/10.1089/vbz.2020.2662

Shaikevich E., Bogacheva A., Rakova V., Ganushkina L., and Ilinsky Y., 2019, Wolbachia symbionts in mosquitoes: Intra and intersupergroup recombinations, horizontal transmission and evolution, Molecular phylogenetics and evolution, 134: 24-34.

https://doi.org/10.1016/j.ympev.2019.01.020

Sherpa S., Rioux D., Pougnet-Lagarde C., and Després L., 2018, Genetic diversity and distribution differ between long-established and recently introduced populations in the invasive mosquito Aedes albopictus, Infection, Genetics and Evolution : Journal of Molecular Epidemiology and Evolutionary Genetics in Infectious Diseases, 58: 145-156.

https://doi.org/10.1016/j.meegid.2017.12.018

Souza A., Multini L., Marrelli M., and Wilke A., 2020, Wing geometric morphometrics for identification of mosquito species (Diptera: Culicidae) of neglected epidemiological importance, Acta Tropica, 211: 105593.

https://doi.org/10.1016/j.actatropica.2020.105593

Swain S., Makunin A., Dóra A., and Barik T., 2019, SNP barcoding based on decision tree algorithm: a new tool for identification of mosquito species with special reference to Anopheles, Acta Tropica, 199: 105152.

https://doi.org/10.1016/j.actatropica.2019.105152

Trzebny A., Słodkowicz-Kowalska A., Becnel J., Sanscrainte N., and Dabert M., 2020, A new method of metabarcoding microsporidia and their hosts reveals high levels of microsporidian infections in mosquitoes (culicidae), Molecular Ecology Resources, 20: 1486-1504.

https://doi.org/10.1111/1755-0998.13205

Weaver S., Charlier C., Vasilakis N., and Lecuit M., 2018, Zika, chikungunya, and other emerging vector-borne viral diseases, Annual Review of Medicine, 69: 395-408.

https://doi.org/10.1146/annurev-med-050715-105122