Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

An Anopheles transgenic sexing strain for vector control

Abstract

Genetic manipulation of mosquito species that serve as vectors for human malaria is a prerequisite to the implementation of gene transfer technologies for the control of vector-borne diseases. Here we report on the development of transgenic sexing lines for the mosquito Anopheles stephensi, the principal vector of human malaria in Asia. Male mosquitoes, expressing enhanced green fluorescent protein (EGFP) under the control of the β2-tubulin promoter, are identified by their fluorescent gonads in as early as their 3rd instar larval stage, and can be efficiently separated from females using both manual methods and automated sorting machines. Importantly, β2-EGFP males are not impaired in their mating ability and viable fluorescent spermatozoa are also detected in spermathecae of wild-type females mated with transgenic males. The transgenic mosquito lines described here combine most of the features desired and required for a safe application of transgenic methodologies to malaria-control programs.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Buy article

Get time limited or full article access on ReadCube.

$32.00

All prices are NET prices.

Figure 1: Fluorescent microphotographs showing β2-EGFP expression at different developmental stages.
Figure 2: Automated sorting of transgenic male larvae.
Figure 3: Dissected spermathecae of wild-type A. stephensi females mated to transgenic heterozygous β2-EGFP males from line β2-egfp2, containing multiple insertions.
Figure 4: Mate choice experiment between transgenic β2-egfp2 and wild-type A. stephensi males.

References

  1. Catteruccia, F. et al. Stable germline transformation of the malaria mosquito Anopheles stephensi. Nature 405, 959–962 (2000).

    Article  CAS  Google Scholar 

  2. Grossman, G.L. et al. Germline transformation of the malaria vector Anopheles gambiae, with the piggyBac transposable element. Insect Mol. Biol. 10, 597–604 (2001).

    Article  CAS  Google Scholar 

  3. Perera, O.P., Harrell, I.R. & Handler, A.M. Germ-line transformation of the South American malaria vector, Anopheles albimanus, with a piggyBac/EGFP transposon vector is routine and highly efficient. Insect Mol. Biol. 11, 291–297 (2002).

    Article  CAS  Google Scholar 

  4. Alphey, L. et al. Malaria control with genetically manipulated insect vectors. Science 298, 119–121 (2002).

    Article  CAS  Google Scholar 

  5. Benedict, M.Q. & Robinson, A.S. The first releases of transgenic mosquitoes: an argument for the sterile insect technique. Trends Parasitol. 19, 349–355 (2003).

    Article  Google Scholar 

  6. Knipling, E.F. Sterile-male method of population control. Science 130, 902–904 (1959).

    Article  CAS  Google Scholar 

  7. Knipling, E.F. et al. Genetic control of insects of public health importance. Bull. World Health Organ. 38, 421–438 (1968).

    CAS  PubMed  PubMed Central  Google Scholar 

  8. Alphey, L. & Andreasen, M. Dominant lethality and insect population control. Mol. Biochem. Parasitol. 121, 173–178 (2002).

    Article  CAS  Google Scholar 

  9. Krafsur, E.S., Whitten, C.J. & Novj, J.E. Screwworm eradication in North and Central America. Parasitol. Today 3, 131–137 (1987).

    Article  CAS  Google Scholar 

  10. Lindquist, D.A., Abusowa, M. & Hall, J.R. The New World screwworm fly in Lybia: a review of its introduction and eradication. Med. Vet. Entomol. 6, 2–8 (1992).

    Article  CAS  Google Scholar 

  11. Robinson, A.S. Genetic sexing strains in medfly, Ceratitis capitata, sterile insect technique programmes. Genetica 116, 5–13 (2002).

    Article  CAS  Google Scholar 

  12. Vreysen, M.J. et al. Glossina austeni (Diptera: Glossinidae) eradicated on the island of Unguja, Zanzibar, using the sterile insect technique. J. Econ. Entomol. 93, 123–135 (2000).

    Article  CAS  Google Scholar 

  13. Rendon, P., McInnis, D., Lance, D. & Stewart, J. Medfly (Diptera: Tephritidae) genetic sexing: large-scale field comparison of males-only and bisexual sterile fly releases in Guatemala. J. Econ. Entomol. 97, 1547–1553 (2004).

    Article  CAS  Google Scholar 

  14. Caceres, C. Mass rearing of temperature sensitive genetic sexing strains in the Mediterranean fruit fly (Ceratitis capitata). Genetica 116, 107–116 (2002).

    Article  CAS  Google Scholar 

  15. Lofgren, C.S. et al. Release of chemosterilized males for the control of Anopheles albimanus in El Salvador. 3. Field methods and population control. Am. J. Trop. Med. Hyg. 23, 288–297 (1974).

    Article  CAS  Google Scholar 

  16. Patterson, R.S., Weidhaas, D.E., Ford, H.R. & Lofgren, C.S. Suppression and elimination of an island population of Culex pipiens quinquefasciatus with sterile males. Science 168, 1368–1370 (1970).

    Article  CAS  Google Scholar 

  17. Curtis, C.F. Genetic sex separation in Anopheles arabiensis and the production of sterile hybrids. Bull. World Health Organ. 56, 453–454 (1978).

    CAS  PubMed  PubMed Central  Google Scholar 

  18. Lines, J.D. & Curtis, C.F. Genetic sexing systems in Anopheles arabiensis Patton (Diptera: Culicidae). J. Econ. Entomol. 78, 848–851 (1985).

    Article  CAS  Google Scholar 

  19. Seawright, J.A., Kaiser, P.E., Dame, D.A. & Lofgren, C.S. Genetic method for the preferential elimination of females of Anopheles albimanus. Science 200, 1303–1304 (1978).

    Article  CAS  Google Scholar 

  20. Sharma, V.P., Patterson, R.S. & Ford, H.R. A device for the rapid separation of male and female mosquito pupae. Bull. World Health Organ. 47, 429–432 (1972).

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Kemphues, K.J., Raff, E.C., Raff, R.A. & Kaufman, T.C. Mutation in a testis-specific beta-tubulin in Drosophila: analysis of its effects on meiosis and map location of the gene. Cell 21, 445–451 (1980).

    Article  CAS  Google Scholar 

  22. Horn, C. & Wimmer, E.A. A versatile vector set for animal transgenesis. Dev. Genes Evol. 210, 630–637 (2000).

    Article  CAS  Google Scholar 

  23. Handler, A.M. & Harrell, R.A., II Germline transformation of Drosophila melanogaster with the piggyBac transposon vector. Insect Mol. Biol. 8, 449–457 (1999).

    Article  CAS  Google Scholar 

  24. Furlong, E.E., Profitt, D. & Scott, M.P. Automated sorting of live transgenic embryos. Nat. Biotechnol. 19, 153–156 (2001).

    Article  CAS  Google Scholar 

  25. Okanda, F.M. et al. Behavioural determinants of gene flow in malaria vector populations: Anopheles gambiae males select large females as mates. Malar. J. 1, 10 (2002).

    Article  CAS  Google Scholar 

  26. Handler, A.M., Zimowska, G.J. & Horn, C. Post-integration stabilization of a transposon vector by terminal sequence deletion in Drosophila melanogaster. Nat. Biotechnol. 22, 1150–1154 (2004).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We wish to thank Charles Godfray and Chris Curtis for helpful suggestions, Elisa Petris and Ann Cronin for help with the injections, and Oliver Billker for help with the microphotographs. We also wish to thank Rico Boongarts from Union Biometrica for assisting us in the automated sorting experiments. The project was supported by the Wellcome Trust and BBSRC. F.C. was sponsored by the Wellcome Trust, J.P.B. by the FP6-EU BioMolPar program.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Andrea Crisanti.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Schematic representation of transformation vector pPB[DsRed]β2EGFP. (PDF 91 kb)

Supplementary Table 1

β2-EGFP expression as a marker to predict sex. (PDF 64 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Catteruccia, F., Benton, J. & Crisanti, A. An Anopheles transgenic sexing strain for vector control. Nat Biotechnol 23, 1414–1417 (2005). https://doi.org/10.1038/nbt1152

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nbt1152

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing