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Control of dengue virus in the midgut of Aedes aegypti by ectopic expression of the dsRNA-binding protein Loqs2


Dengue virus (DENV) is an arbovirus transmitted to humans by Aedes mosquitoes1. In the insect vector, the small interfering RNA (siRNA) pathway is an important antiviral mechanism against DENV2,3,4,5. However, it remains unclear when and where the siRNA pathway acts during the virus cycle. Here, we show that the siRNA pathway fails to efficiently silence DENV in the midgut of Aedes aegypti although it is essential to restrict systemic replication. Accumulation of DENV-derived siRNAs in the midgut reveals that impaired silencing results from a defect downstream of small RNA biogenesis. Notably, silencing triggered by endogenous and exogenous dsRNAs remained effective in the midgut where known components of the siRNA pathway, including the double-stranded RNA (dsRNA)-binding proteins Loquacious and r2d2, had normal expression levels. We identified an Aedes-specific paralogue of loquacious and r2d2, hereafter named loqs2, which is not expressed in the midgut. Loqs2 interacts with Loquacious and r2d2 and is required to control systemic replication of DENV and also Zika virus. Furthermore, ectopic expression of Loqs2 in the midgut of transgenic mosquitoes is sufficient to restrict DENV replication and dissemination. Together, our data reveal a mechanism of tissue-specific regulation of the mosquito siRNA pathway controlled by Loqs2.

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Fig. 1: Production of DENV-derived siRNAs in infected mosquitoes.
Fig. 2: The antiviral siRNA pathway does not control DENV infection in the mosquito midgut but inhibits systemic dissemination and replication.
Fig. 3: The siRNA pathway triggered by endogenous and exogenous sources of dsRNA is functional in the midgut.
Fig. 4: The Aedes-specific dsRBP Loqs2 regulates the antiviral arm of the mosquito siRNA pathway.

Data availability

Small RNA libraries from this study have been deposited in the Sequence Read Archive (SRA) at NCBI. Other publicly available RNA–seq data sets were obtained from SRA. Accession numbers and references are provided in Supplementary Table 4.


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The authors thank C.N.D. dos Santos for providing 4G2 monoclonal antibodies. This work was supported with funding from Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) and Fundação de Amparo a Pesquisa do Estado de Minas Gerais (FAPEMIG) to J.T.M., Agence Nationale de la Recherche (ANR-11-ASV3-002) and Investissement d’Avenir Programs (ANR-10-LABX-36; ANR-11-EQPX-0022) to J.T.M and J.-L.I., and Inserm, CNRS and the University of Strasbourg to E.M. and J.-L.I. Mass spectrometry instrumentation was funded by the University of Strasbourg, IdEx Equipement mi-lourd 2015 to L.K. and P.H. National and/or international fellowships from CAPES were granted to R.P.O., T.C.I.-T., A.G.A.F., E.R.G.R.A., K.P.R.S., K.P.O. and C.D.O., and national and/or international fellowships from CNPq were granted to I.J.S.F., L.A.M. and J.T.M.

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Authors and Affiliations



R.P.O. and J.T.M. designed the project. R.P.O., A.G.A.F., T.C.I.-T., I.J.S.F., K.P.R.S., K.P.O., P.H., E.G.A., Y.M.T, M.N.R., T.H.J.F.L., S.C.G.A., J.N.A. and S.P. performed experiments. R.P.O., E.R.G.R.A. and L.K. performed bioinformatics analysis. C.D.O., F.D.C., L.A.M and E.M. contributed to mosquito experiments. R.P.O., E.R.G.R.A., J.-L.I. and J.T.M. analysed the data. R.P.O., J.-L.I. and J.T.M. wrote the paper. All authors discussed the results and commented on the manuscript.

Corresponding author

Correspondence to João T. Marques.

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Supplementary information

Supplementary Information

Supplementary Figures 1–4, Supplementary Tables 2–4.

Reporting Summary

Supplementary Table 1

Mass spectrometry analysis of Loqs2 interacting proteins.

Supplementary File 1

Plasmid sequence containing the cassette (attP loxP CP::3×FLAG-Loqs2-Sv40pA, PUb::mTurquoise2-Sv40pA loxP) flanked by piggyBac repeats.

Supplementary File 2

Plasmid sequence containing the cassette (attP loxP PUb::3×FLAG-eGFPSv40pA, OpIE2::Puromicin-Sv40pA loxP) flanked by piggyBac repeats.

Supplementary File 3

Plasmid sequence containing the cassette (attP loxP PUb::3×FLAG-Loqs2-Sv40pA, OpIE2::Puromicin-Sv40pA loxP) flanked by piggyBac repeats.

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Olmo, R.P., Ferreira, A.G.A., Izidoro-Toledo, T.C. et al. Control of dengue virus in the midgut of Aedes aegypti by ectopic expression of the dsRNA-binding protein Loqs2. Nat Microbiol 3, 1385–1393 (2018).

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