# Dynamics and evolution of transposable elements in mosquito genomes

Josquin Daron, Alexander Bergman, Louis Lambrechts

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Josquin Daron, Alexander Bergman, Louis Lambrechts. Dynamics and evolution of transposable elements in mosquito genomes. Current Opinion in Insect Science, 2025, 71, pp.101406. 10.1016/j.cois.2025.101406. pasteur-05232570

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ELSEVIER

Review

# Dynamics and evolution of transposable elements in mosquito genomes

Josquin Daron, Alexander Bergman and Louis Lambrechts

<figure><img src="image_3.png" /></figure>

Decoding mosquito genomes is essential for understanding their role as vectors of human pathogens. Transposable elements (TEs), once considered ‘junk DNA’, are now recognized as key drivers of genomic plasticity and evolution. These mobile DNA fragments are more abundant in culicine than anopheline mosquitoes, influencing their genome size and complexity. TEs may contribute to mosquito adaptation, influencing traits such as insecticide resistance and habitat expansion. Recent advances in long-read sequencing technologies and functional assays are uncovering TE dynamics, but challenges remain in measuring and manipulating their activity, which is critical to demonstrate their phenotypic effects. Further research into the environmental and biological conditions that activate TEs in mosquitoes, as well as the host mechanisms of TE regulation, such as small RNA pathways, will significantly enhance our understanding of mosquito biology.

**Address**

Institut Pasteur, Université Paris Cité, CNRS UMR2000, Insect-Virus Interactions Unit, Paris, France

Corresponding authors: Daron, Josquin (josquin.daron@pasteur.fr), Lambrechts, Louis (louis.lambrechts@pasteur.fr)

**Current Opinion in Insect Science** 2025, 71:101406

This review comes from a themed issue on **Insect genomics**

Edited by **Ben Matthews** and **John Soghigian**

For complete overview about the section, refer “Insect genomics (2025)”

Available online 28 June 2025

https://doi.org/10.1016/j.cois.2025.101406

2214-5745/© 2025 The Author(s). Published by Elsevier Inc. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

## Introduction

This review aims to explore the evolutionary dynamics of transposable elements (TEs) in the genomes of mosquitoes (Culicidae), highlighting their significant impact on genomic evolution and adaptive potential. TEs are mobile DNA fragments capable of duplicating or relocating within the genome. They are found in all living organisms, except for a few *Plasmodium* species, and exhibit broad variation in their abundance between

species. For instance, the TE fraction of insect genomes ranges from 1.6% to 81.5% [1,2]. TEs can be autonomous, capable of mobilizing themselves by encoding all the enzymatic machinery required for their own transposition, or non-autonomous, relying on the molecular machinery of other autonomous TEs for their movement. They are classified into two groups based on their transposition mechanism: Class I TEs (retrotransposons) use a replicative ‘copy-and-paste’ transposition mechanism, whereas Class II TEs (DNA transposons) utilize a ‘cut-and-paste’ mechanism. TE insertions can alter coding sequences, gene regulation, and genome structure, contributing significantly to genome plasticity and evolution [3]. While most TE insertions are neutral, some are detrimental or adaptive [4]. Since McClintock’s pioneering research in the 1940s, TEs have evolved from being considered ‘junk DNA’ to recognized drivers of genomic evolution, with consequences for the host ranging from harmful to beneficial [5].

Long-read sequencing technologies have revolutionized genomics by overcoming the limitations of short-read methods and enabling the analysis of large and repeat-rich genomes. Short reads often struggle to accurately sequence TEs because they do not span full-length elements, resulting in collapsed, fragmented, and incomplete assemblies [6,7]. In recent years, research in mosquito genomics has been transformed by the sequencing of multiple large mosquito genomes, with several exceeding 1 gigabase (Gb). *Aedes aegypti* is no longer the only large-genome mosquito species with a chromosome-level assembly (Figure 1). The Culicidae family comprises two main subfamilies, anophelines (Anophelinae) and culicines (Culicinae), which diverged ~179 million years ago [8]. Several mosquito species act as vectors of human pathogens, such as arboviruses and malaria parasites, fueling a long-standing interest in deciphering mosquito genomes to advance our understanding of vector biology. This article presents fresh prospects for understanding the evolutionary dynamics of TEs in mosquito genomes, providing a timely update on a two-decade-old review [9].

## Transposable element landscape in mosquito genomes

The recent rise in sequenced Culicidae genomes allows a macro-evolutionary exploration of their TE content (Figure 2). A clear dichotomy exists between the Culicinae and Anophelinae subfamilies: Anophelinae
<figure><img src="image_4.png" /><figcaption>Figure 1</figcaption></figure>

Timeline of chromosome-level genome assemblies in Culicidae. Release dates of chromosome-level genome assemblies for Culicidae species are shown, with their cumulative number overlaid in light gray. The wide adoption of third-generation (long-read) sequencing technologies since 2020 coincides with an exponential increase in the production of high-quality genome assemblies for Culicidae species with large genomes.

genomes contain 0.13–19.55% TEs, while Culicinae genomes often exceed 40% in TE content [10–13]. Despite this dichotomy, genome size tends to positively correlate with TE content across Culicidae (Figure 2), as generally observed in both plant [14] and animal [15] genomes. Despite more than five decades of research demonstrating that genome size does not correlate with organismal complexity or gene count in eukaryotes — a concept known as the ‘C-value enigma’ — the evolutionary forces driving genome size remain poorly understood [15]. The drift–barrier hypothesis posits that a small effective population size ($N_e$) allows TEs to accumulate, increasing genome size [16]. When $N_e$ is small, genetic drift can cause large changes in allele frequencies that counteract the effect of purifying selection. Consequently, deleterious TE insertions that would be eliminated by purifying selection in larger populations can reach fixation in small populations. However, empirical support to the drift–barrier hypothesis is mixed. While some studies report a negative correlation between $N_e$ and genome size [17], others fail to detect such an association [18]. In Culicidae, large-genome species like *Ae. aegypti* tend to have smaller $N_e$ values ($10^5–10^6$) [19–21] compared to smaller-genome species like *Anopheles coluzzii* ($10^6–10^7$) [22]. These differences support the hypothesis that reduced $N_e$ in the common ancestor of Culicinae may have led to increased genome size in this subfamily, but this hypothesis remains to be demonstrated.

Studies of intraspecific TE variation across Culicidae remain sparse, despite early calls for more population-

level genetic data [23]. In *Aedes albopictus*, genome size estimates vary from 0.62 to 1.66 picograms (606 Mb to 1.5 Gb) [13,24]. Invasive *Ae. albopictus* populations, which presumably underwent demographic bottlenecks that reduced $N_e$, tend to have larger genomes [13,24], providing support for the drift–barrier hypothesis. More fine-scale research has revealed a positive correlation between TE content and genome size in *An. coluzzii* populations across cities of central Africa [25]; however, the causes of these variations remain unclear. Large population genomics data sets, such as the 1000 Genomes projects for *Anopheles gambiae* and *Ae. aegypti*, are ideal resources for exploring the evolutionary factors shaping intraspecific variation in TE content.

Culicidae genomes display a highly diverse TE composition, with up to 24 TE superfamilies identified across different species [10]. Retrotransposons are the predominant elements in Culicidae, with long interspersed nuclear elements, such as *jockey*, *RTE*, and *I* superfamilies being especially abundant in Culicinae [10]. In addition, long-terminal repeat (LTR) retrotransposons, including *gypsy*, *copia*, and *belpao*, represent about a quarter of the repeat fraction in the most up-to-date Anophelinae genome assemblies [25,26] and account for ~17% of the TE content in both *Ae. aegypti* and *Ae. albopictus* genomes [10,11]. In contrast, DNA transposons are more heterogeneously distributed among genomes, with some superfamilies like *Sola* and *Zator* found exclusively in Culicinae [10]. Non-autonomous miniature inverted-repeat TEs (MITEs) have proliferated in *Ae. aegypti* and *Ae.*
<figure><img src="image_5.png" /><figcaption>Figure 2</figcaption></figure>

TE abundance and composition in Culicidae genomes. The middle panel shows the proportions of TE categories in the genomes of the Culicidae family (mosquitoes), which are arranged according to their phylogeny [8] shown in the left panel. TEs were categorized into LTR retrotransposons, long interspersed nuclear element (LINE) retrotransposons, short interspersed nuclear element (SINE) retrotransposons, helitrons, other DNA transposons, and ‘unclassified’ TEs that could not be categorized by the procedure. Only species whose chromosome-level genome assemblies overlap with those included in the phylogeny [8] were selected. Genome-specific TE libraries were generated using RepeatModeler, and TEs were subsequently annotated with RepeatMasker [81]. The right panel shows the relationship between overall TE proportion and genome size (both log₁₀-transformed) for Culicinae (red) and Anophelinae (blue) subfamilies. Linear model fitting was performed using the pgls function from the R CRAN package caper, which accounts for phylogenetic nonindependence among data points to avoid phylogenetic inertia (adjusted R² = 0.297, *p*-value < 0.001). The design of the figure is inspired by Figure 1 in Ref. [1]. Data supporting this figure are available upon request.

albopictus [27]. For instance, the genome of *Ae. aegypti* contains at least 143 MITE families with a total copy number exceeding 420 000. The L-shaped age structure of TEs in Anophelinae reflects recent retrotransposon activity [25,26,28]. In contrast, the large TE content of *Ae. aegypti* does not correspond with recent amplification, as recent TE copies represent a similar repeat fraction in both *Ae. aegypti* (3.1%) and *An. coluzzii* (2.8%) [11]. Genomic TE distribution also differs sharply between Culicidae subfamilies. TEs are mostly concentrated in the pericentromeric regions of Anophelinae genomes. For instance, the proportion of TEs is 11.7–13.4% in the eu-chromatin of *An. coluzzii*, whereas it is 65.8–74.8% in heterochromatin [25], making heterochromatin a more polymorphic compartment in this species [29]. TEs are distributed more evenly along the chromosomes in *Ae. aegypti*, with their proportion ranging from 52.9% to 67.8% [11,30].

## Transposable element-mediated adaptive evolution in mosquitoes

Unlike single base-pair mutations, transposon insertion polymorphisms (TIPs) constitute a substantial source of molecular variation with notable fitness and phenotypic consequences. TEs are significant contributors to genetic diversity, providing the raw material upon which selection acts to drive adaptive evolution [4]. For example, TEs played a role in the evolution of insecticide resistance in *An. funestus* populations from across Uganda and Cameroon. A selective sweep linked to transposon-mediated duplication of the *CYP9K1* gene allowed its overexpression in response to deltamethrin exposure [31,32]. It remains unknown whether such TE-mediated adaptation can be promoted by mechanisms of TE activation, particularly under stressful conditions. Highly invasive mosquito species like *Ae. aegypti*, *Ae. albopictus*, and *Anopheles stephensi* may represent further cases of
TE-driven rapid adaptation. Despite the genetic bottlenecks due to founder effects, these species show remarkable adaptability to new environments. Preliminary work in *Ae. albopictus* supports the hypothesis that TEs could help to explain the puzzling ability of invasive species to adapt and colonize new habitats following genetic bottlenecks, referred to as the ‘genetic paradox of invasive species’ [33]. Adaptation of *Ae. albopictus* populations to temperate climate is associated with outlier TIPs linked to genes related to diapause, lipid regulation, and juvenile hormone pathways [34].

It remains unclear whether TEs contribute to large chromosomal rearrangements in mosquito genomes [35]. The ecological and behavioral plasticity of Anophelinae is often linked to high levels of polymorphic inversions, which suppress recombination and can promote species isolation [36]. TEs have been found near the breakpoints of several inversions, such as the 2Rd' inversion in *Anopheles arabiensis* [37], the 2La inversion in *An. gambiae* and *Anopheles melas* [38], and the 2Rb inversion in *An. gambiae* and *An. coluzzii* [39,40], suggesting TEs may play a role in these rearrangements and their evolutionary consequences [37]. This mirrors findings in *Drosophila melanogaster* and *Drosophila buzzatii*, where TEs likely contributed to polymorphic inversions via ectopic recombination [41,42]. However, TEs may also accumulate passively at inversion breakpoints due to the suppression of recombination, as seen in heterochromatic regions [35].

## Transposable element activity in mosquitoes

Estimating ongoing TE activity is a challenge, as TE diversity largely reflects past transposition events, not current activity. The gold-standard method for detecting active TE copies is the identification of *de novo* insertions using long-read sequencing technologies [43,44]. Although these methods have dramatically advanced our ability to detect TE insertions with unmatched accuracy, they are still more costly and less scalable than short-read sequencing technologies. As a result, most large-scale population genetic studies continue to rely primarily on short-read sequencing data. Alternative methods to whole-genome shotgun sequencing infer TE activity indirectly through intermediate transposition products, such as mRNAs or proteins identified in transcriptomic or proteomic data sets [45]. However, these methods often overestimate actual TE activity due to influences such as read-through transcription [46]. Cost-effective innovations such as mobilome-seq, which detects extra-chromosomal circular DNA during the life cycle of LTR retrotransposons, have proven useful [47]. The effectiveness of this approach was exemplified in *An. gambiae* by the successful identification of one-LTR circles, which serve as replication intermediates in the lifecycle of *copia* elements [48]. Recent advancements in TE display

sequencing have introduced techniques that use targeted amplification of TE extremities to detect TIPs [49]. This assay involves the digestion of genomic DNA followed by polymerase chain reaction amplification using primers specific to the target TE sequence and has been adapted for high-throughput detection via sequencing. These methods offer high versatility by enabling multiplexing to identify insertions from multiple TE families simultaneously. Recent advancements in TE display sequencing and single-cell technologies are expected to further improve the ability to assess somatic TE activity.

Whole-genome resequencing has yet to directly measure TE activity in Culicidae. However, transcriptomic and proteomic data have provided key insights. RNA-seq analyses across 12 mosquito species indicate that most TEs show constitutive but low RNA expression, typically under 3% of all mapped RNA-seq reads, except for *Ae. albopictus* (10.4%) and *An. coluzzii* (16.7%) [50]. *Ae. aegypti* shows low TE expression broadly, with ~3% of its transcriptome [50] and proteome [51] made up of TE transcripts and TE proteins, respectively. Interestingly, TE expression at the mRNA or protein level does not correlate with genomic abundance in *Ae. aegypti*. Similarly, despite constituting only ~10% of the TE fraction, LTR retrotransposons are disproportionately represented in the transcriptome of *An. funestus*, where they account for 86% of transcripts derived from Class I elements [52]. Currently, studying TE transcription offers limited insight into actual TE activity. Large, TE-rich genomes do not necessarily show higher levels of active TEs under natural conditions, a hypothesis that requires further validation through more accurate measurements of transposition rates.

Genome-wide studies of TE activity have been complemented by functional validation in heterologous systems. The Class II *Herves hAT* element shows transposition activity in *Drosophila* S2 cells and embryos [53]. Variations in *hAT* copy number and site distribution in natural populations of *An. gambiae*, *An. arabiensis*, and *Anopheles merus* suggest active transposition, particularly given the abundance of low-frequency copies [53]. In *Ae. aegypti*, functionally active *Mutator*-like elements were identified in silico and validated through yeast transposition assays, demonstrating both insertion and excision [54]. In a similar assay, a nonautonomous MITE called *Gnome*, highly abundant in *Ae. aegypti*, was shown to be mobilized by its autonomous relative *Ozma* [27]. By conducting site-directed mutagenesis, both studies revealed that mutations in conserved regions of the transposase, such as the nuclear export signal motif of *Ozma*, substantially increased transposition frequency.

Understanding how environmental and biological stressors, such as temperature shifts, bloodmeal ingestion, or
viral infections, modulate TE activity is particularly relevant in mosquito vector species. Stress-induced TE activation has been observed across various species [55,56]. For example, viral infections trigger TE expression in the somatic tissues of *Drosophila melanogaster* [57]. However, recent transcriptomic analyses using public data sets have not consistently shown widespread TE activation or repression under abiotic stresses or upon viral infections [50,58], underscoring the need for more in-depth research to clarify the conditions that influence TE activity.

## Transposable element regulation in mosquitoes

In *D. melanogaster*, the regulation of TE expression at the post-transcriptional level is primarily managed through small RNA pathways, particularly the Piwi-interacting RNA (piRNA) pathway [59] and the small interfering RNA (siRNA) pathway [60–62]. TE expression in the germline and surrounding ovarian tissues is regulated by the piRNA pathway, whereas in the somatic tissues, TE expression involves other mechanisms, including siRNAs. Detection of somatic piRNAs in many arthropod species, including mosquitoes, has challenged the notion that piRNAs exclusively specialize in controlling TEs in the germline [63].

In Anophelinae, the proportion of TE-derived piRNAs is lower than in *D. melanogaster* [64]. This proportion is even lower in Culicinae, despite their larger TE content [64]. Culicinae also show a reduced antisense/sense bias and include satellites in their piRNA clusters [64,65]. These and other observations suggest that the biological roles of piRNAs may also be significantly different in mosquitoes compared to *D. melanogaster*. The larger TE content of Culicinae correlates with *PIWI* gene family expansion [66], frequent use of genic transcripts as substrates for piRNA biogenesis [65,67,68], and increased abundance of nonretroviral endogenous viral elements (nrEVEs) in piRNA clusters [69–71]. nrEVEs are fragments of RNA viruses, most often insect-specific viruses, that have been reverse transcribed and integrated into the mosquito genome. When expressed, nrEVEs can contribute to antiviral defense by generating antiviral piRNAs in the presence of a cognate virus [69,72]. nrEVEs are commonly found near LTR retrotransposons [70,73], which likely play a prominent role in the piRNA activity of Culicinae. LTR retrotransposon-derived piRNAs dominate small RNA populations [64] and are overrepresented in 'core' piRNA clusters that are active in both germline and soma [71]. Abundant LTR retrotransposons are found in genomic loci containing a high density of nrEVEs [70,73]. LTR retrotransposons and their regulation by piRNAs may have shaped the adaptations of Culicinae in domains beyond TE control.

The role of siRNAs in the regulation of TE expression in mosquitoes remains less studied. In stable conditions, the siRNA pathway alone appears to play a minor role in controlling TE expression [74]. This is consistent with studies in *D. melanogaster* showing that either siRNA or piRNA pathways are sufficient to control TE expression [75]. More generally, little is known about mosquito small RNAs in their role as TE regulators, except for insights drawn from *Drosophila* research [76]. Research on small RNA pathways in mosquitoes often focuses on their role during viral infections, particularly in Culicinae [58,77–79]. Studies on their role as TE suppressors remain largely limited to descriptive analyses of sequencing data [64,71,80]. Finally, other possible TE control mechanisms, such as epigenetic regulation through histone modifications or DNA methylation, remain to be studied in mosquitoes.

## Conclusion

Studying TEs across mosquito genomes has revealed their significant impact on genomic architecture and evolution. Advanced sequencing technologies have shed light on substantial differences in TE abundance and genomic distribution among mosquito subfamilies. Large-scale population genomics and functional assays have begun to clarify TE dynamics across mosquito species, but challenges persist in effectively measuring and manipulating TE activity for functional studies. Further research is necessary to understand the environmental conditions that regulate TE activation and the host mechanisms that control TE expression.

## Data Availability

Data will be made available on request.

## Declaration of Competing Interest

Louis Lambrechts reports financial support was provided by The French National Research Agency (French Government's Investissement d'Avenir Program, grant ANR-10-LABX-62-IBEID). Alexander Bergman reports financial support was provided by Institut Pasteur (Pasteur-Paris University International PhD Program). If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

## Declaration of Generative AI and AI-assisted technologies in the writing process

During the preparation of this work, the authors used ChatGPT-4 (OpenAI) to improve the readability and language of the manuscript. After using this tool, the authors reviewed and edited the content as needed and take full responsibility for the content of the publication.
## Acknowledgements

This work was supported by the French Government's Investissement d'Avenir Program, Laboratoire d'Excellence Integrative Biology of Emerging Infectious Diseases (grant ANR-10-LABX-62-IBEID to L.L.). A.B. was supported by a stipend from the Pasteur-Paris University (PPU) International PhD Program.

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75. Beek M, van den, Silva B, da Pouch J, Chaouche MEAA, Carré C, Antoniewski C: Dual-layer transposon repression in heads of *Drosophila melanogaster*. *RNA* 2018, **24**:1749-1760.

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77. Henderson C, Brustolin M, Hegde S, Dayama G, Lau N, Hughes GL, Bergey C, Rasgon JL: Transcriptomic and small RNA response to Mayaro virus infection in *Anopheles stephensi* mosquitoes. *PLoS Negl Trop Dis* 2022, **16:e0010507**.

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