Male mosquitoes to be genetically engineered to poison females with semen

Image from article: Biogenetic mosquito control.

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Recombinant venom proteins in insect seminal fluid reduce female lifespan

Abstract

The emergence of insecticide resistance has increased the need for alternative pest management tools. Numerous genetic biocontrol approaches, which involve the release of genetically modified organisms to control pest populations, are in various stages of development to provide highly targeted pest control. However, all current mating-based genetic biocontrol technologies function by releasing engineered males which skew sex-ratios or reduce offspring viability in subsequent generations which leaves mated females to continue to cause harm (e.g. transmit disease). Here, we demonstrate intragenerational genetic biocontrol, wherein mating with engineered males reduces female lifespan. The toxic male technique (TMT) involves the heterologous expression of insecticidal proteins within the male reproductive tract that are transferred to females via mating. In this study, we demonstrate TMT in Drosophila melanogaster males, which reduce the median lifespan of mated females by 37 − 64% compared to controls mated to wild type males. Agent-based models of Aedes aegyptipredict that TMT could reduce rates of blood feeding by a further 40 – 60% during release periods compared to leading biocontrol technologies like fsRIDL. TMT is a promising approach for combatting outbreaks of disease vectors and agricultural pests.

Introduction

Insect pests pose a major challenge to human and environmental health. Malaria, spread by several species of Anopheles mosquitoes, causes 608,000 deaths per year1 and rates of arboviral diseases spread primarily by the Aedes aegypti mosquito, including dengue, Zika, chikungunya, and yellow fever, are reaching unprecedented levels due to increased global trade and warming climates2. Dengue virus alone causes 390 million human infections each year and is now considered the most common vector-borne viral infection worldwide3. Recent studies have calculated that the cumulative cost of biological invasions is increasing four-fold every decade4, with a conservative estimate of US$162.7 billion for damages caused in 20175. Invertebrate pests, primarily insects, are responsible for ~40% of these damages5,6, and estimates of annual losses for all major food crops ranges between 20 and 30% due to damages by pests and pathogens7. Arboviral diseases such as avian malaria also pose a significant ecological threat to many native species, and are believed to be a key driver of population decline in New Zealand and Hawaiian forest birds8,9.

Pesticides are the first line of defence against many invasive species, particularly mosquitoes10. However, over-reliance on insecticides has resulted in the widespread emergence of resistance11. Over-application of insecticides has also resulted in declining populations of non-target species, and are responsible for many other environmental and human-health concerns12. Modern integrated pest management is trending towards reducing reliance on chemical insecticides in favour of other environmentally friendly management techniques to reduce the emergence of resistances10,13.

Genetic biocontrol, defined as the release of organisms which have been genetically altered to reduce the spread and harm caused by a target species14, is one such alternative that’s been gaining broader public acceptance15. Early and well established genetic biocontrol technologies (GBT) such as the sterile insect technique (SIT) involve the release of mass numbers of males which have been radiologically sterilised16, thereby reducing the reproductive potential of the females. More modern GBT, such as the release of insects carrying a dominant lethal gene (RIDL)17 and gene drives18 function by propagating transgenes which reduce the fitness of future generations as the transgenes are inherited. Cage and field trials of these second-generation GBT have begun to demonstrate their improved efficacy19,20,21, requiring fewer released males compared to conventional approaches to achieve similar population control, albeit with a potential trade-off between enhanced efficacy and reduced control over the degree of transgene introgression14.

Currently, all GBT require a minimum of a generation to take effect on the target population and often much longer until the harm from a pest outbreak is mitigated19. While this generational lag period is less problematic for species where immature life stages are the primary cause of harm, it poses a significant challenge for many disease vectors and agricultural pests where adult females are the main concern. For example, wild female Ae. aegypti have a median adult lifespan of 2–3 weeks22,23, will typically mate within 24–48 h of emerging24, and on average will take 0.63–0.76 blood meals per day25. Females that mate with GBT males may not produce viable offspring, but they can continue to spread disease or damage crops. Although current GBT are highly effective at intergenerational population control, a more rapid response to outbreaks of disease vectors is critical to mitigate the spread of arboviruses and avert the risk of epidemics, particularly in regions with seasonal population dynamics26.

An alternative approach to GBT that has been speculated on is utilising seminal fluid proteins (SFP) as a target to affect the fitness or fertility of mated females27,28,29,30. SFPs are produced within the male accessory glands (MAG) and stored within the lumen until they are transferred to females along with sperm and other compounds. SFPs such as ovulin (Acp26Aa) and sex peptides are known to generate various post-mating responses and physiological changes in females, such as reduced mating receptivity and median lifespan31,32. It has been demonstrated in Drosophila melanogaster that several low molecular weight SFP can pass through the female reproductive tract and enter the circulatory system (haemolymph), and even to act on receptors in the central nervous system (CNS)33,34. Evidence is mounting that mosquito SFP also act on receptors found within the CNS, with 30% of radiolabelled Culex SFP localising to the female head and thorax35, and intra-thoracic but not intra-abdominal SFP injections resulting in mating refractoriness in Anopheline and Aedes females36,37,38,39.

Here we describe the toxic male technique (TMT), which is—to the best of our knowledge—the first example of an intragenerational GBT. By genetically engineering pest species to express toxic, low molecular weight compounds within the MAG, the survival of mated females can be reduced (Fig. 1). Using D. melanogaster as a proof of concept, we engineer males to express one of seven insect-specific venom proteins in a MAG-specific pattern. Of the >1600 characterised venom compounds, 60% of these are cysteine-knotted mini-proteins40, which range from 2.6 to 14.8 kDa and are highly stable, taxonomically selective and less susceptible to pore region mutations associated with insecticide resistances41. Like endogenous SFP, the recombinant venom peptides should remain safely sequestered within the lumen of the MAG and not enter the male’s haemolymph. If the recombinant venom peptides pass through the mated female’s reproductive tract to affect the target ion channel receptors in her CNS, the neurotoxic effects are likely to be fast-acting and dose-dependent.

Fig. 1: Intergenerational vs intragenerational genetic biocontrol of pest insects.
figure 1

Current mating-based methods of genetic biocontrol (A) function by affecting the viability or sex ratio of the offspring of subsequent generations. However, mated females persist in the target area and can continue to cause harm (e.g. spread disease). Intragenerational biocontrol (B), such as TMT, directly affects the fitness of mated females, thereby rapidly reducing the harm caused by the target population. Created in BioRender. Maselko, M. (2024) https://BioRender.com/r25q201.

The median lifespans of wild-type females mated to two of the TMT strains are reduced by 37–64% after initial exposure compared to control females mated to wild-type males. Single-pair mating assays find that TMT males are able to court wild-type females as effectively as wild-type males. MAG-specific expression of a 5.2 kDa venom peptide has no significant effect on the lifespan of adult TMT males, but expression of a 2.9 kDa venom peptide reduces lifespan by 59% compared to wildtype and transgenic controls. To determine the theoretical capability of TMT to suppress a pest population in comparison to female-specific RIDL (fsRIDL), the current state-of-the-art GBT, we develop an agent-based model to simulate an idealised Ae. aegypti release programme19. Sensitivity analysis of the model found that the improved performance of TMT compared to fsRIDL is consistent across a broad range of values of polyandry, density-dependent mortality, and release ratios, with TMT reducing blood feeding rates by a further 40–60% in most simulations. These results demonstrate the potential of TMT as the next generation of genetic biocontrol, which is especially suited to rapidly respond to outbreaks of disease vectors and agricultural pests.

References

  1. World Health Organization. World malaria report 2023. (2023)

  2. Roiz, D. et al. Integrated Aedes management for the control of Aedes-borne diseases. PLoS Negl. Trop. Dis. 12, e0006845 (2018).

    Article PubMed PubMed Central MATH Google Scholar

  3. Wilder-Smith, A. et al. Epidemic arboviral diseases: priorities for research and public health. Lancet Infect. Dis. 17, e101–e106 (2017).

    Article PubMed MATH Google Scholar

  4. Diagne, C. et al. High and rising economic costs of biological invasions worldwide. Nature592, 571–576 (2021).

    Article ADS CAS PubMed MATH Google Scholar

  5. Bacher, S. et al. Impacts of invasive alien species on nature, nature’s contributions to people, and good quality of life. In Thematic Assessment Report on Invasive Alien Species and their Control of the Intergovernmental Science-Policy Platform on Biodiversity and Ecosystem Services. (eds. Roy, H. E., Pauchard, A., Stoett, P. & Renard Truong, T.) Ch. 4 (Zenodo, 2023). https://doi.org/10.5281/zenodo.7430731.

  6. Leroy, B. et al. Analysing economic costs of invasive alien species with the invacost r package. Methods Ecol. Evol. 13, 1930–1937 (2022).

    Article MATH Google Scholar

  7. Savary, S. et al. The global burden of pathogens and pests on major food crops. Nat. Ecol. Evol. 3, 430–439 (2019).

    Article PubMed MATH Google Scholar

  8. Howe, L. et al. Malaria parasites (Plasmodium spp.) infecting introduced, native and endemic New Zealand birds. Parasitol. Res. 110, 913–923 (2012).

    Article PubMed Google Scholar

  9. van Riper, III C., van Riper, S. G., Goff, M. L. & Laird, M. The epizootiology and ecological significance of malaria in Hawaiian land birds. Ecol. Monogr. 56, 327–344 (1986).

    Article MATH Google Scholar

  10. World Health Organization. Test Procedures for Insecticide Resistance Monitoring in Malaria Vector Mosquitoes (World Health Organization, 2013).

  11. Liu, N. Insecticide resistance in mosquitoes: impact, mechanisms, and research directions. Annu. Rev. Entomol. 60, 537–559 (2015).

    Article CAS PubMed MATH Google Scholar

  12. Ansari, M. S., Moraiet, M. A. & Ahmad, S. In Environmental Deterioration and Human Health(eds. Malik, A., Grohmann, E. & Akhtar, R.) Ch. 6 (Springer, 2014).

  13. Bueno, A. F. et al. Challenges for adoption of integrated pest management (IPM): the soybean example. Neotrop. Entomol. 50, 5–20 (2021).

    Article CAS PubMed MATH Google Scholar

  14. Teem, J. L. et al. Genetic biocontrol for invasive species. Front. Bioeng. Biotechnol. 8, 452 (2020).

    Article PubMed PubMed Central MATH Google Scholar

  15. Winneg, K. M., Stryker, J. E., Romer, D. & Jamieson, K. H. Differences between Florida and the rest of the United States in response to local transmission of the zika virus: implications for future communication campaigns. Risk Anal. 38, 2546–2560 (2018).

    Article PubMed Google Scholar

  16. Knipling, E. F. Possibilities of insect control or eradication through the use of sexually sterile males. J. Econ. Entomol. 48, 459–462 (1955).

    Article Google Scholar

  17. Fu, G. et al. Female-specific insect lethality engineered using alternative splicing. Nat. Biotechnol. 25, 353–357 (2007).

    Article CAS PubMed Google Scholar

  18. Esvelt, K. M., Smidler, A. L., Catteruccia, F. & Church, G. M. Concerning RNA-guided gene drives for the alteration of wild populations. eLife 3, e03401 (2014).

    Article PubMed PubMed Central Google Scholar

  19. Carvalho, D. O. et al. Suppression of a field population of Aedes aegypti in Brazil by sustained release of transgenic male mosquitoes. PLoS Negl. Trop. Dis. 9, e0003864 (2015).

    Article PubMed PubMed Central Google Scholar

  20. Shelton, A. M. et al. First field release of a genetically engineered, self-limiting agricultural pest insect: evaluating its potential for future crop protection. Front. Bioeng. Biotechnol. 7, 482 (2020).

    Article PubMed PubMed Central MATH Google Scholar

  21. Hammond, A. et al. Gene-drive suppression of mosquito populations in large cages as a bridge between lab and field. Nat. Commun. 12, 4589 (2021).

    Article ADS CAS PubMed PubMed Central MATH Google Scholar

  22. Sheppard, P. M., Macdonald, W. W., Tonn, R. J. & Grab, B. The dynamics of an adult population of Aedes aegypti in relation to dengue haemorrhagic fever in Bangkok. J. Anim. Ecol. 38, 661–702 (1969).

    Article Google Scholar

  23. Sowilem, M. M., Kamal, H. A. & Khater, E. I. Life table characteristics of Aedes aegypti(Diptera: Culicidae) from Saudi Arabia. Trop. Biomed. 30, 301–314 (2013).

  24. Ramírez-Sánchez, L. F., Hernández, B. J., Guzmán, P. A., Alfonso-Parra, C. & Avila, F. W. The effects of female age on blood-feeding, insemination, sperm storage, and fertility in the dengue vector mosquito Aedes aegypti (Diptera: Culicidae). J. Insect Physiol. 150, 104570 (2023).

    Article PubMed Google Scholar

  25. Scott, T. W. et al. Longitudinal Studies of Aedes aegypti (Diptera: Culicidae) in Thailand and Puerto Rico: blood feeding frequency. J. Med. Entomol. 37, 89–101 (2000).

    Article ADS CAS PubMed MATH Google Scholar

  26. Weaver, S. C. Prediction and prevention of urban arbovirus epidemics: a challenge for the global virology community. Antivir. Res. 156, 80–84 (2018).

    Article CAS PubMed MATH Google Scholar

  27. Alfonso-Parra, C. et al. Synthesis, depletion and cell-type expression of a protein from the male accessory glands of the dengue vector mosquito Aedes aegypti. J. Insect Physiol. 70, 117–124 (2014).

    Article CAS PubMed PubMed Central MATH Google Scholar

  28. Marshall, J. M., Pittman, G. W., Buchman, A. B. & Hay, B. A. Semele: a killer-male, rescue-female system for suppression and replacement of insect disease vector populations. Genetics 187, 535–551 (2011).

    Article CAS PubMed PubMed Central Google Scholar

  29. Hurtado, J., Revale, S. & Matzkin, L. M. Propagation of seminal toxins through binary expression gene drives could suppress populations. Sci. Rep. 12, 6332 (2022).

    Article ADS CAS PubMed PubMed Central MATH Google Scholar

  30. Sirot, L. K. et al. Identity and transfer of male reproductive gland proteins of the dengue vector mosquito, Aedes aegypti: potential tools for control of female feeding and reproduction. Insect Biochem. Mol. Biol. 38, 176–189 (2008).

    Article CAS PubMed MATH Google Scholar

  31. Chapman, T., Liddle, L. F., Kalb, J. M., Wolfner, M. F. & Partridge, L. Cost of mating in Drosophila melanogaster females is mediated by male accessory gland products. Nature373, 241–244 (1995).

    Article ADS CAS PubMed Google Scholar

  32. Avila, F. W., Sirot, L. K., LaFlamme, B. A., Rubinstein, C. D. & Wolfner, M. F. Insect seminal fluid proteins: identification and function. Annu. Rev. Entomol. 56, 21–40 (2011).

    Article CAS PubMed PubMed Central Google Scholar

  33. Lung, O. & Wolfner, M. F. Drosophila seminal fluid proteins enter the circulatory system of the mated female fly by crossing the posterior vaginal wall. Insect Biochem. Mol. Biol. 29, 1043–1052 (1999).

    Article CAS PubMed MATH Google Scholar

  34. Yapici, N., Kim, Y.-J., Ribeiro, C. & Dickson, B. J. A receptor that mediates the post-mating switch in Drosophila reproductive behaviour. Nature 451, 33–37 (2008).

    Article PubMed Google Scholar

  35. Young, A. D. M. & Downe, A. E. R. Male accessory gland substances and the control of sexual receptivity in female Culex tarsalis. Physiol. Entomol. 12, 233–239 (1987).

    Article Google Scholar

  36. Klowden, M. J. Sexual receptivity in Anopheles gambiae mosquitoes: absence of control by male accessory gland substances. J. Insect Physiol. 47, 661–666 (2001).

    Article CAS PubMed Google Scholar

  37. Shutt, B., Stables, L., Aboagye-Antwi, F., Moran, J. & Tripet, F. Male accessory gland proteins induce female monogamy in anopheline mosquitoes. Med. Vet. Entomol. 24, 91–94 (2010).

    Article CAS PubMed Google Scholar

  38. Helinski, M. E. H., Deewatthanawong, P., Sirot, L. K., Wolfner, M. F. & Harrington, L. C. Duration and dose-dependency of female sexual receptivity responses to seminal fluid proteins in Aedes albopictus and Ae. aegypti mosquitoes. J. Insect Physiol. 58, 1307–1313 (2012).

    Article CAS PubMed PubMed Central Google Scholar

  39. Duvall, L. B., Basrur, N. S., Molina, H., McMeniman, C. J. & Vosshall, L. B. A peptide signaling system that rapidly enforces paternity in the Aedes aegypti mosquito. Curr. Biol. 27, 3734–3742.e5 (2017).

    Article CAS PubMed PubMed Central Google Scholar

  40. Kuhn-Nentwig, L., Stöcklin, R. & Nentwig, W. Venom composition and strategies in spiders. Adv. Insect Physiol. 40, 1–86 (2011).

  41. King, G. F. Tying pest insects in knots: the deployment of spider‐venom‐derived knottins as bioinsecticides. Pest Manag. Sci. 75, 2437–2445 (2019).

    Article CAS PubMed MATH Google Scholar

  42. Leader, D. P., Krause, S. A., Pandit, A., Davies, S. A. & Dow, J. A. T. FlyAtlas 2: a new version of the Drosophila melanogaster expression atlas with RNA-Seq, miRNA-Seq and sex-specific data. Nucleic Acids Res. 46, D809–D815 (2018).

    Article CAS PubMed Google Scholar

  43. Ishikawa, T. et al. DCRY is a Drosophila photoreceptor protein implicated in light entrainment of circadian rhythm. Genes Cells 4, 57–65 (1999).

    Article CAS PubMed MATH Google Scholar

  44. Lee, P.-T. et al. A gene-specific T2A-GAL4 library for Drosophila. eLife 7, e35574 (2018).

    Article PubMed PubMed Central Google Scholar

  45. Yoffe, K. B., Manoukian, A. S., Wilder, E. L., Brand, A. H. & Perrimon, N. Evidence for engrailed-independent wingless autoregulation in Drosophila. Dev. Biol. 170, 636–650 (1995).

    Article CAS PubMed Google Scholar

  46. Chapman, T. et al. The sex peptide of Drosophila melanogaster: female post-mating responses analyzed by using RNA interference. Proc. Natl Acad. Sci. USA 100, 9923–9928 (2003).

    Article ADS CAS PubMed PubMed Central MATH Google Scholar

  47. Shearin, H. K., Macdonald, I. S., Spector, L. P. & Stowers, R. S. Hexameric GFP and mCherry reporters for the Drosophila GAL4, Q, and LexA transcription systems. Genetics 196, 951–960 (2014).

    Article CAS PubMed PubMed Central Google Scholar

  48. Manning, A. The control of sexual receptivity in female Drosophila. Anim. Behav. 15, 239–250 (1967).

    Article CAS PubMed MATH Google Scholar

  49. Manning, A. A sperm factor affecting the receptivity of Drosophila melanogaster females. Nature 194, 252–253 (1962).

    Article ADS MATH Google Scholar

  50. Proverbs, M. D., Newton, J. R. & Campbell, C. J. Codling moth: a pilot program of control by sterile insect release in British Columbia. Can. Entomol. 114, 363–376 (1982).

    Article Google Scholar

  51. Wee, S. L. et al. Effects of substerilizing doses of gamma radiation on adult longevity and level of inherited sterility in Teia anartoides (Lepidoptera: Lymantriidae). J. Econ. Entomol.98, 732–738 (2005).

    Article CAS PubMed Google Scholar

  52. Edward, D. A., Fricke, C., Gerrard, D. T. & Chapman, T. Quantifying the life‐history response to increased male exposure in female Drosophila melanogaster. Evolution 65, 564–573 (2011).

    Article PubMed MATH Google Scholar

  53. Von Philipsborn, A. C., Shohat-Ophir, G. & Rezaval, C. Single-pair courtship and competition assays in Drosophila. Cold Spring Harb. Protoc. 2023, pdb.prot108105 (2023).

    Article Google Scholar

  54. Ling, J. et al. A nanobody that recognizes a 14-residue peptide epitope in the E2 ubiquitin-conjugating enzyme UBC6e modulates its activity. Mol. Immunol. 114, 513–523 (2019).

    Article CAS PubMed PubMed Central MATH Google Scholar

  55. Degner, E. C. & Harrington, L. C. Polyandry depends on postmating time interval in the dengue vector Aedes aegypti. Am. J. Trop. Med. Hyg. 94, 780–785 (2016).

    Article PubMed PubMed Central MATH Google Scholar

  56. Harrington, L. C. et al. Heterogeneous feeding patterns of the dengue vector, Aedes aegypti, on individual human hosts in rural Thailand. PLoS Negl. Trop. Dis. 8, e3048 (2014).

    Article PubMed PubMed Central Google Scholar

  57. Chan, M. & Johansson, M. A. The incubation periods of dengue viruses. PLoS ONE 7, e50972 (2012).

    Article ADS CAS PubMed PubMed Central MATH Google Scholar

  58. ten Broeke, G., van Voorn, G. & Ligtenberg, A. Which sensitivity analysis method should I use for my agent-based model? JASSS 19, 5 (2016).

    Article MATH Google Scholar

  59. Quistad, G. B., Dennis, P. A. & Skinner, W. S. Insecticidal activity of spider (Araneae), centipede (Chilopoda), scorpion (Scorpionida), and snake (Serpentes) venoms. J. Econ. Entomol. 85, 33–39 (1992).

    Article CAS Google Scholar

  60. Schwartz, E. F., Mourão, C. B. F., Moreira, K. G., Camargos, T. S. & Mortari, M. R. Arthropod venoms: a vast arsenal of insecticidal neuropeptides. Pept. Sci. 98, 385–405 (2012).

    Article CAS Google Scholar

  61. Guo, S., Herzig, V. & King, G. F. Dipteran toxicity assays for determining the oral insecticidal activity of venoms and toxins. Toxicon 150, 297–303 (2018).

    Article CAS PubMed Google Scholar

  62. Oh, R. J. Repeated copulation in the brown planthopper, Nilaparvata lugens Stál (Homoptera; Delphacidae). Ecol. Entomol. 4, 345–353 (1979).

    Article MATH Google Scholar

  63. Cabauatan, P. Q., Cabunagan, R. C. & Choi, I.-R. In Planthoppers: New Threats to the Sustainability of Intensive Rice Production Systems in Asia (eds. Heong, K. L. & Hardy, B.) 357–368 (International Rice Research Institute, 2009).

  64. Denecke, S., Swevers, L., Douris, V. & Vontas, J. How do oral insecticidal compounds cross the insect midgut epithelium. Insect Biochem. Mol. Biol. 103, 22–35 (2018).

    Article CAS PubMed Google Scholar

  65. Kandul, N. P. et al. Transforming insect population control with precision guided sterile males with demonstration in flies. Nat. Commun. 10, 84 (2019).

    Article ADS CAS PubMed PubMed Central MATH Google Scholar

  66. Massonnet-Bruneel, B. et al. Fitness of transgenic mosquito Aedes aegypti males carrying a dominant lethal genetic system. PLoS ONE 8, e62711 (2013).

    Article ADS CAS PubMed PubMed Central Google Scholar

  67. Upadhyay, A. et al. Genetically engineered insects with sex-selection and genetic incompatibility enable population suppression. eLife 11, e71230 (2022).

    Article CAS PubMed PubMed Central Google Scholar

  68. Ejima, A. & Griffith, L. C. Measurement of courtship behavior in Drosophila melanogaster. Cold Spring Harb. Protoc. 2007, pdb.prot4847 (2007).

  69. Mueller, J. L., Linklater, J. R., Ravi Ram, K., Chapman, T. & Wolfner, M. F. Targeted gene deletion and phenotypic analysis of the Drosophila melanogaster seminal fluid protease inhibitor Acp62F. Genetics 178, 1605–1614 (2008).

    Article CAS PubMed PubMed Central Google Scholar

  70. Zhang, S. D. & Odenwald, W. F. Misexpression of the white (w) gene triggers male-male courtship in Drosophila. Proc. Natl Acad. Sci. USA 92, 5525–5529 (1995).

    Article ADS CAS PubMed PubMed Central MATH Google Scholar

  71. Sirot, L. K., Buehner, N. A., Fiumera, A. C. & Wolfner, M. F. Seminal fluid protein depletion and replenishment in the fruit fly, Drosophila melanogaster: an ELISA-based method for tracking individual ejaculates. Behav. Ecol. Sociobiol. 63, 1505–1513 (2009).

    Article PubMed PubMed Central Google Scholar

  72. Maselko, M. & Beach, S. Plasmid sequences for: Recombinant venom proteins in insect seminal fluid reduces female lifespan. Zenodo https://doi.org/10.5281/zenodo.13910640(2024).

  73. Joiner, M. A. & Griffith, L. C. Visual input regulates circuit configuration in courtship conditioning of Drosophila melanogaster. Learn. Mem. 7, 32–42 (2000).

    Article CAS PubMed PubMed Central MATH Google Scholar

  74. Hegde, R., Hegde, S., Gataraddi, S., Kulkarni, S. S. & Gai, P. B. Novel and PCR ready rapid DNA isolation from Drosophila. Nucleosides Nucleotides Nucleic Acids 41, 1162–1173 (2022).

    Article CAS PubMed MATH Google Scholar

  75. Sauers, L. A., Hawes, K. E. & Juliano, S. A. Non-linear relationships between density and demographic traits in three Aedes species. Sci. Rep. 12, 8075 (2022).

    Article ADS CAS PubMed PubMed Central MATH Google Scholar

  76. Goindin, D., Delannay, C., Ramdini, C., Gustave, J. & Fouque, F. Parity and longevity of Aedes aegypti according to temperatures in controlled conditions and consequences on dengue transmission risks. PLoS ONE 10, e0135489 (2015).

    Article PubMed PubMed Central Google Scholar

  77. Christophers, A. A. The yellow fever mosquito, its life history, bionomics and structure. J. Natl Med. Assoc. 54, 132 (1960).

    Google Scholar

  78. Almeida, S. J. D., Martins Ferreira, R. P., Eiras, Á. E., Obermayr, R. P. & Geier, M. Multi-agent modeling and simulation of an Aedes aegypti mosquito population. Environ. Model. Softw.25, 1490–1507 (2010).

    Article Google Scholar

  79. Maneerat, S. & Daudé, E. A spatial agent-based simulation model of the dengue vector Aedes aegypti to explore its population dynamics in urban areas. Ecol. Model. 333, 66–78 (2016).

    Article MATH Google Scholar

  80. Pimid, M. et al. Parentage assignment using microsatellites reveals multiple mating in Aedes aegypti (Diptera: Culicidae): implications for mating dynamics. J. Med. Entomol. 59, 1525–1533 (2022).

    Article CAS PubMed MATH Google Scholar

  81. Seabold, S. & Perktold, J. Statsmodels: econometric and statistical modeling with Python. In Proc. 9th Python in Science Conference 92–96 (2010).

  82. Herman, J. & Usher, W. SALib: an open-source Python library for sensitivity analysis. J. Open Source Softw. 2, 97 (2017).

    Article ADS MATH Google Scholar

  83. Pedregosa, F. et al. Scikit-learn: machine learning in Python. J. Mach. Learn. Res. 12, 2825–2830 (2011).

    MathSciNet MATH Google Scholar

  84. Davidson-Pilon, C. lifelines: survival analysis in Python. J. Open Source Softw. 4, 1317 (2019).

    Article ADS MATH Google Scholar

  85. Hunter, J. D. Matplotlib: a 2D graphics environment. Comput. Sci. Eng. 9, 90–95 (2007).

    Article MATH Google Scholar

  86. Waskom, M. L. seaborn: statistical data visualization. J. Open Source Softw. 6, 3021 (2021).

    Article ADS MATH Google Scholar

  87. Maselko, M. & Beach, S. J. Data and code for: recombinant venom proteins in insect seminal fluid reduces female lifespan. Zenodo https://doi.org/10.5281/zenodo.13927099 (2024).

  88. MaselkoLab. TMT-Aedes-model. GitHub (2024).

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