A recent epidemic of Zika virus in the Americas, affecting well over a million people, caused substantial mortality and morbidity, including Guillain–Barre syndrome, microcephaly and other fetal developmental defects1,2. Preventive and therapeutic measures that specifically target the virus are not readily available. The transmission of Zika virus is predominantly mosquito-borne, and Aedes aegypti mosquitoes serve as a key vector for Zika virus3. Here, to identify salivary factors that modulate mosquito-borne Zika virus infection, we focused on antigenic proteins in mice that were repeatedly bitten by mosquitoes and developed antibodies against salivary proteins. Using a yeast surface display screen, we identified five antigenic A. aegypti salivary proteins in mice. Antiserum against one of these five proteins—A. aegypti bacteria-responsive protein 1 (AgBR1)—suppressed early inflammatory responses in the skin of mice bitten by Zika-virus-infected mosquitoes. AgBR1 antiserum also partially protected mice from lethal mosquito-borne—but not needle-injected—Zika virus infection. These data suggest that AgBR1 is a target for the prevention of mosquito-transmitted Zika virus infection.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Data availability

The data that support the findings of this study are available from the corresponding authors on request. The RNA-seq data have been deposited in the Gene Expression Omnibus under the accession number GSE125194.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.


  1. 1.

    Ventura, C. V., Maia, M., Bravo-Filho, V., Gois, A. L. & Belfort, R. Jr Zika virus in Brazil and macular atrophy in a child with microcephaly. Lancet 387, 228 (2016).

  2. 2.

    Olsen, B. & Lundkvist, A. Zika virus—ancient virus gets new life in a new ecosystem. Microcephaly and Guillain–Barre syndrome are possible consequences when there is no background herd immunity in the population (in Swedish). Lakartidningen 113, DX9X (2016).

  3. 3.

    Musso, D. & Gubler, D. J. Zikavirus. Clin. Microbiol. Rev. 29, 487–524 (2016).

  4. 4.

    Coutinho-Abreu, I. V., Guimaraes-Costa, A. B. & Valenzuela, J. G. Impact of insect salivary proteins in blood feeding, host immunity, disease, and in the development of biomarkers for vector exposure. Curr. Opin. Insect Sci. 10, 98–103 (2015).

  5. 5.

    Fontaine, A. et al. Implication of haematophagous arthropod salivary proteins in host-vector interactions. Parasit. Vectors 4, 187 (2011).

  6. 6.

    Hayashi, H. et al. Anopheline anti-platelet protein from a malaria vector mosquito has anti-thrombotic effects in vivo without compromising hemostasis. Thromb. Res. 129, 169–175 (2012).

  7. 7.

    Pingen, M. et al. Host inflammatory response to mosquito bites enhances the severity of arbovirus infection. Immunity 44, 1455–1469 (2016).

  8. 8.

    Ruckert, C. & Ebel, G. D. How do virus-mosquito interactions lead to viral emergence? Trends Parasitol. 34, 310–321 (2018).

  9. 9.

    Conway, M. J. et al. Mosquito saliva serine protease enhances dissemination of dengue virus into the mammalian host. J. Virol. 88, 164–175 (2014).

  10. 10.

    Conway, M. J. et al. Aedes aegypti D7 saliva protein inhibits dengue virus infection. PLoS Negl. Trop. Dis. 10, e0004941 (2016).

  11. 11.

    Jin, L. et al. Salivary factor LTRIN from Aedes aegypti facilitates the transmission of Zika virus by interfering with the lymphotoxin-β receptor. Nat. Immunol. 19, 342–353 (2018).

  12. 12.

    Wasinpiyamongkol, L. et al. Blood-feeding and immunogenic Aedes aegypti saliva proteins. Proteomics 10, 1906–1916 (2010).

  13. 13.

    Rabilloud, T., Chevallet, M., Luche, S. & Lelong, C. Two-dimensional gel electrophoresis in proteomics: past, present and future. J. Proteomics 73, 2064–2077 (2010).

  14. 14.

    Schuijt, T. J. et al. Identification and characterization of Ixodes scapularis antigens that elicit tick immunity using yeast surface display. PLoS ONE 6, e15926 (2011).

  15. 15.

    Lee, C. G. et al. Role of chitin and chitinase/chitinase-like proteins in inflammation, tissue remodeling, and injury. Annu. Rev. Physiol. 73, 479–501 (2011).

  16. 16.

    Ribeiro, J. M. et al. An annotated catalogue of salivary gland transcripts in the adult female mosquito, Aedes aegypti. BMC Genom. 8, 6 (2007).

  17. 17.

    Beatty, P. R. et al. Dengue virus NS1 triggers endothelial permeability and vascular leak that is prevented by NS1 vaccination. Sci. Transl. Med. 7, 304ra141 (2015).

  18. 18.

    Tanaka, T. & Kishimoto, T. The biology and medical implications of interleukin-6. Cancer Immunol. Res. 2, 288–294 (2014).

  19. 19.

    Calvo, E., Mans, B. J., Andersen, J. F. & Ribeiro, J. M. Function and evolution of a mosquito salivary protein family. J. Biol. Chem. 281, 1935–1942 (2006).

  20. 20.

    Rathore, A. P. S. & St John, A. L. Immune responses to dengue virus in the skin. Open Biol. 8, 18087 (2018).

  21. 21.

    Kieny, M. P., Excler, J. L. & Girard, M. Research and development of new vaccines against infectious diseases. Am. J. Public Health 94, 1931–1935 (2004).

  22. 22.

    Sarathy, V. V., Milligan, G. N., Bourne, N. & Barrett, A. D. Mouse models of dengue virus infection for vaccine testing. Vaccine 33, 7051–7060 (2015).

  23. 23.

    Zhu, J., Huang, X. & Yang, Y. Type I IFN signaling on both B and CD4 T cells is required for protective antibody response to adenovirus. J. Immunol. 178, 3505–3510 (2007).

  24. 24.

    Fong, S. W., Kini, R. M. & Ng, L. F. P. Mosquito saliva reshapes alphavirus infection and immunopathogenesis. J. Virol. 92, e01004-17 (2018).

  25. 25.

    Shi, C. & Pamer, E. G. Monocyte recruitment during infection and inflammation. Nat. Rev. Immunol. 11, 762–774 (2011).

  26. 26.

    Osuna, C. E. et al. Zika viral dynamics and shedding in rhesus and cynomolgus macaques. Nat. Med. 22, 1448–1455 (2016).

  27. 27.

    Bai, F. et al. A paradoxical role for neutrophils in the pathogenesis of West Nile virus. J. Infect. Dis. 202, 1804–1812 (2010).

  28. 28.

    Dudley, D. M. et al. Infection via mosquito bite alters Zika virus tissue tropism and replication kinetics in rhesus macaques. Nat. Commun. 8, 2096 (2017).

  29. 29.

    Carman, W. F. et al. Vaccine-induced escape mutant of hepatitis B virus. Lancet 336, 325–329 (1990).

  30. 30.

    Uraki, R., Hastings, A. K., Gloria-Soria, A., Powell, J. R. & Fikrig, E. Altered vector competence in an experimental mosquito-mouse transmission model of Zika infection. PLoS Negl. Trop. Dis. 12, e0006350 (2018).

  31. 31.

    Chao, G. et al. Isolating and engineering human antibodies using yeast surface display. Nat. Protoc. 1, 755–768 (2006).

  32. 32.

    Uraki, R. et al. Zika virus causes testicular atrophy. Sci. Adv. 3, e1602899 (2017).

  33. 33.

    Balaban, A. E., Neuman, K., Sinnis, P. & Balaban, R. S. Robust fluorescent labelling of micropipettes for use in fluorescence microscopy: application to the observation of a mosquito borne parasite infection. J. Microsc. 269, 78–84 (2018).

  34. 34.

    Subramanian, A. et al. Gene set enrichment analysis: a knowledge-based approach for interpreting genome-wide expression profiles. Proc. Natl Acad. Sci. USA 102, 15545–15550 (2005).

  35. 35.

    Nakajima, S. et al. Prostaglandin I2-IP signaling promotes Th1 differentiation in a mouse model of contact hypersensitivity. J. Immunol. 184, 5595–5603 (2010).

  36. 36.

    Chang, Q., Ornatsky, O. & Hedley, D. Staining of frozen and formalin-fixed, paraffin-embedded tissues with metal-labeled antibodies for imaging mass cytometry analysis. Curr. Protoc. Cytom. 82, 12.47.1–12.47.8 (2017).

Download references


We thank S. Shresta at the La Jolla Institute for Allergy and Immunology for originally providing us with the AG129 mouse strain. We also thank S. Ren, Y.-M. Chuang, S. Householder, S. Stanley, H. Sproch and B. V. Wyk for supporting experiments and analysing data. The imaging mass cytometry was conducted at the Yale CyTOF facility and the RNA-seq service was conducted at the Yale Stem Cell Center Genomics Core facility, which was supported by the Connecticut Regenerative Medicine Research Fund and the Li Ka Shing Foundation. This work was supported by NIH grant nos AI089992 and AI127865, and the Japan Society for the Promotion of Science Overseas Research Fellowships. E.F. and A.I. are investigators with the Howard Hughes Medical Institute.

Author information


  1. Section of Infectious Diseases, Department of Internal Medicine, School of Medicine, Yale University, New Haven, CT, USA

    • Ryuta Uraki
    • , Andrew K. Hastings
    • , Alejandro Marin-Lopez
    •  & Erol Fikrig
  2. Department of Neurology, School of Medicine, Yale University, New Haven, CT, USA

    • Tomokazu Sumida
    •  & David A. Hafler
  3. Department of Immunobiology, School of Medicine, Yale University, New Haven, CT, USA

    • Tomokazu Sumida
    • , Takehiro Takahashi
    • , Akiko Iwasaki
    •  & David A. Hafler
  4. Department of Microbial Pathogenesis, School of Medicine, Yale University, New Haven, CT, USA

    • Jonathan R. Grover
  5. Howard Hughes Medical Institute, Chevy Chase, MD, USA

    • Akiko Iwasaki
    •  & Erol Fikrig
  6. Department of Internal Medicine, School of Medicine, Yale University, New Haven, CT, USA

    • Ruth R. Montgomery


  1. Search for Ryuta Uraki in:

  2. Search for Andrew K. Hastings in:

  3. Search for Alejandro Marin-Lopez in:

  4. Search for Tomokazu Sumida in:

  5. Search for Takehiro Takahashi in:

  6. Search for Jonathan R. Grover in:

  7. Search for Akiko Iwasaki in:

  8. Search for David A. Hafler in:

  9. Search for Ruth R. Montgomery in:

  10. Search for Erol Fikrig in:


R.U. and E.F. designed the experiments. R.U. performed the majority of the experiments and analysed the data. A.K.H., A.M.-L. and J.R.G. assisted in the experiments with mosquitoes. R.U. and T.S. performed the analysis of the RNA-seq. T.T. performed the histopathological analyses. A.I., D.A.H. and R.R.M. contributed experimental suggestions and strengthened the writing of the manuscript. R.U. and E.F wrote the manuscript. All authors reviewed, critiqued and provided comments on the text.

Competing interests

The authors declare no competing interests.

Corresponding author

Correspondence to Erol Fikrig.

Supplementary information

  1. Supplementary Information

    Supplementary Figures 1–12, Supplementary Tables 1–3 and Supplementary Table 6.

  2. Reporting Summary

  3. Supplementary Table 4

    List of upregulated genes at the bite site of mice treated with control (naïve) serum.

  4. Supplementary Table 5

    List of GSEA-enriched pathway at bite sites in mice treated with control serum using hallmark gene sets (http://software.broadinstitute.org/gsea/msigdb/genesets.jsp?collection=H).

About this article

Publication history