Abstract

Over 50% of patients who survive neuroinvasive infection with West Nile virus (WNV) exhibit chronic cognitive sequelae1,2. Although thousands of cases of WNV-mediated memory dysfunction accrue annually3, the mechanisms responsible for these impairments are unknown. The classical complement cascade, a key component of innate immune pathogen defence, mediates synaptic pruning by microglia during early postnatal development4,5. Here we show that viral infection of adult hippocampal neurons induces complement-mediated elimination of presynaptic terminals in a murine WNV neuroinvasive disease model. Inoculation of WNV-NS5-E218A, a WNV with a mutant NS5(E218A) protein6,7 leads to survival rates and cognitive dysfunction that mirror human WNV neuroinvasive disease. WNV-NS5-E218A-recovered mice (recovery defined as survival after acute infection) display impaired spatial learning and persistence of phagocytic microglia without loss of hippocampal neurons or volume. Hippocampi from WNV-NS5-E218A-recovered mice with poor spatial learning show increased expression of genes that drive synaptic remodelling by microglia via complement. C1QA was upregulated and localized to microglia, infected neurons and presynaptic terminals during WNV neuroinvasive disease. Murine and human WNV neuroinvasive disease post-mortem samples exhibit loss of hippocampal CA3 presynaptic terminals, and murine studies revealed microglial engulfment of presynaptic terminals during acute infection and after recovery. Mice with fewer microglia (Il34−/− mice with a deficiency in IL-34 production) or deficiency in complement C3 or C3a receptor were protected from WNV-induced synaptic terminal loss. Our study provides a new murine model of WNV-induced spatial memory impairment, and identifies a potential mechanism underlying neurocognitive impairment in patients recovering from WNV neuroinvasive disease.

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Accessions

Primary accessions

Gene Expression Omnibus

Data deposits

Microarray data has been deposited in the Gene Expression Omnibus under the accession number GSE72139.

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Acknowledgements

Funding for this research was provided by the NIH F31 NS077640 (M.J.V.), R01 NS052632 (R.S.K.), and U19 AI083019 (R.S.K. and M.S.S.). The authors would like to thank J. Atkinson and X. Wu for reagents and M. Diamond for critical reading of the manuscript.

Author information

Affiliations

  1. Department of Medicine, Washington University School of Medicine, St Louis, Missouri 63110, USA

    • Michael J. Vasek
    • , Charise Garber
    • , Denise Dorsey
    • , Douglas M. Durrant
    • , Bryan Bollman
    • , Allison Soung
    • , Kristen Funk
    •  & Robyn S. Klein
  2. Biological Sciences Department, California State Polytechnic University, 3801 West Temple Avenue, Pomona, California 91768, USA

    • Douglas M. Durrant
  3. Department of Genetics, Washington University School of Medicine, St Louis, Missouri 63110, USA

    • Jinsheng Yu
  4. Department of Radiology, Washington University School of Medicine, St Louis, Missouri 63110, USA

    • Carlos Perez-Torres
    •  & Joel R. Garbow
  5. Department of Neurology, F. M. Kirby Neurobiology Center, Boston Children’s Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA

    • Arnaud Frouin
    • , Daniel K. Wilton
    •  & Beth Stevens
  6. Department of Neurology, University of Colorado School of Medicine, Aurora, Colorado 80045, USA

    • Bette K. DeMasters
    •  & Kenneth L. Tyler
  7. Department of Psychiatry, Washington University School of Medicine, St Louis, Missouri 63110, USA

    • Xiaoping Jiang
    •  & Steven Mennerick
  8. Department of Pediatrics and Children’s Healthcare of Atlanta, Emory Vaccine Center, Yerkes National Primate Research Center, Emory University School of Medicine, Atlanta, Georgia 30329, USA

    • James R. Bowen
    •  & Mehul S. Suthar
  9. Department of Psychology, Stony Brook University, Stony Brook, New York 11794, USA

    • John K. Robinson
  10. Department of Pathology and Immunology, Washington University School of Medicine, St Louis, Missouri 63110, USA

    • Robert E. Schmidt
    •  & Robyn S. Klein
  11. Department of Anatomy and Neurobiology, Washington University School of Medicine, St Louis, Missouri 63110, USA

    • Robyn S. Klein

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Contributions

M.J.V. and R.S.K. contributed to the study design. M.J.V., C.G., D.D., D.M.D., B.B., A.S., J.Y., C.P.-T., A.F., D.K.W., K.F., X.J., S.M., J.K.R., J.R.G., R.E.S., B.S. and R.S.K. contributed to data collection and/or interpretation. C.G., J.R.B. and M.S.S. developed single-strand PCR assays for WNV. B.K.D., K.L.T. identified, collected and provided patient samples. M.J.V. and R.S.K. wrote the paper. All authors discussed and commented on the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Robyn S. Klein.

Extended data

Supplementary information

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    Supplementary Information

    This file contains Supplementary Tables 1-6.

Videos

  1. 1.

    Video 1: Barnes Maze day 2, trial 2 in mock‐infected mouse at day 47 post‐infection

    Video of Barnes Maze day 2, trial 2 in mock‐infected mouse at day 47 post‐infection. Number of errors before finding target hole and latency to find target hole was quantified in Figure 1.

  2. 2.

    Video 2: Barnes Maze day 2, trial 2 in WNV‐NS5‐E218A‐recovered mouse at day 47 post‐infection

    Video of Barnes Maze day 2, trial 2 in WNV‐NS5‐E218A‐recovered mouse at day 47 post‐infection. Number of errors before finding target hole and latency to find target hole was quantified in Figure 1.

  3. 3.

    Video 3: 3D reconstruction from confocal Z‐stack images of immunostaining of CX3CR1‐GFP and the presynaptic marker, synaptophysin, in CX3CR1‐GFP +/‐ mice in hippocampus of mock‐infected control mouse at day 7 post‐infection.

    This video shows a 3D reconstruction from confocal Z‐stack images of immunostaining of CX3CR1‐GFP and the presynaptic marker, synaptophysin, in CX3CR1‐GFP +/‐ mice in hippocampus of mock‐infected control mouse at day 7 post‐infection.

  4. 4.

    Video 4: 3D reconstruction from confocal Z‐stack images of immunostaining of CX3CR1‐GFP and the presynaptic marker, synaptophysin, in CX3CR1‐GFP +/‐ mice in hippocampus of WNV‐NS5‐E218A‐infected mouse at day 7 post‐infection.

    This video shows a 3D reconstruction from confocal Z‐stack images of immunostaining of CX3CR1‐GFP and the presynaptic marker, synaptophysin, in CX3CR1‐GFP +/‐ mice in hippocampus of WNV‐NS5‐E218A‐infected mouse at day 7 post‐infection.

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DOI

https://doi.org/10.1038/nature18283

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