Pre-treatment with Scopolamine Naturally Suppresses Japanese Encephalitis Viral Load in Embryonated Chick Through Regulation of Multiple Signaling Pathways

Abstract

Suitable recognition of invasive microorganisms is a crucial factor for evoking a strong immune response that can combat the pathogen. Toll-like receptors (TLRs) play a pivotal role in the induction of this innate immune response through stimulation of interferons (IFNs) that control viral replication in the host via distinct signaling pathways. Though the antiviral property of Atropa belladonna has been established, yet the role of one of its active components scopolamine in modulating various factors of the innate immune branch has not yet been investigated until date. Thus, the present study was conducted to assess the antiviral effects of scopolamine and its immunomodulatory role against Japanese encephalitis virus (JEV) infections in embryonated chick. Pre-treatment with scopolamine hydrobromide showed a significant decrease in the viral loads of chorioallantoic membrane (CAM) and brain tissues. Molecular docking analysis revealed that scopolamine hydrobromide binds to the active site of non-structural protein 5 (NS5) that has enzymatic activities required for replication of JEV, making it a highly promising chemical compound against the virus. The binding contributions of different amino acid residues at or near the active site suggest a potential binding of this compound. Pre-treatment with the scopolamine hydrobromide showed significant upregulation of different TLRs like TLR3, TLR7, and TLR8, interleukins like IL-4, and IL-10, as well as IFNs and their regulatory factors. However, virus-infected tissues (direct infection group) exhibited higher TLR4 expression as compared to scopolamine hydrobromide pre-treated, virus-infected tissues (medicine pre-treated group). These results indicate that scopolamine hydrobromide contributes much to launch antiviral effects by remoulding the TLR and IFN signaling pathways that are involved in sensing and initiating the much-needed anti-JEV responses.

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

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9
Fig. 10
Fig. 11

Data Availability

Not applicable.

References

  1. 1.

    Tsai, T. F. (2000). New initiatives for the control of Japanese encephalitis by vaccination: minutes of a WHO/CVI meeting, Bangkok, Thailand, 13–15 October 1998. Vaccine, 18, 1–25.

    PubMed  Article  PubMed Central  Google Scholar 

  2. 2.

    Tiwari, S., Singh, R. K., Tiwari, R., & Dhole, T. N. (2012). Japanese encephalitis: a review of the Indian perspective. The Brazilian Journal of Infectious Diseases, 16(6), 564–573.

    PubMed  Article  PubMed Central  Google Scholar 

  3. 3.

    Solomon, T., Ni, H., Beasley, D. W., Ekkelenkamp, M., Cardosa, M. J., & Barrett, A. D. (2003). Origin and evolution of Japanese encephalitis virus in southeast Asia. Journal of Virology, 77(5), 3091–3098.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  4. 4.

    Fadnis, P. R., Ravi, V., Desai, A., Turtle, L., & Solomon, T. (2013). Innate immune mechanisms in Japanese encephalitis virus infection: effect on transcription of pattern recognition receptors in mouse neuronal cells and brain tissue. Viral Immunology, 26(6), 366–377.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  5. 5.

    Li, X.-D., Shan, C., Deng, C.-L., Ye, H.-Q., Shi, P.-Y., Yuan, Z.-M., Gong, P., & Zhang, B. (2014). The interface between methyltransferase and polymerase of NS5 is essential for flavivirus replication. PLoS Neglected Tropical Diseases, 8(5), e2891.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  6. 6.

    Chen, C. J., Ou, Y. C., Lin, S. Y., Raung, S. L., Liao, S. L., Lai, C. Y., Chen, S. Y., & Chen, J. H. (2010). Glial activation involvement in neuronal death by Japanese encephalitis virus infection. Journal of General Virology, 91(4), 1028–1037.

    CAS  Article  Google Scholar 

  7. 7.

    Li, F., Wang, Y., Yu, L., Cao, S., Wang, K., Yuan, J., et al. (2015). Viral infection of the central nervous system and neuroinflammation precede blood-brain barrier disruption during Japanese encephalitis virus infection. Journal of Virology, 89(10), 5602–5614.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  8. 8.

    Ye, Q., Li, X. F., Zhao, H., Li, S. H., Deng, Y. Q., Cao, R. Y., & Zhou, X. (2012). A single nucleotide mutation in NS2A of Japanese encephalitis-live vaccine virus (SA14-14-2) ablates NS1’formation and contributes to attenuation. Journal of General Virology, 93(9), 1959–1964.

    CAS  Article  Google Scholar 

  9. 9.

    Cao, L., Fu, S., Gao, X., Li, M., Cui, S., Li, X., et al. (2016). Low protective efficacy of the current Japanese encephalitis vaccine against the emerging genotype 5 Japanese encephalitis virus. PLoS Neglected Tropical Diseases, 10(5), e0004686.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  10. 10.

    Adi, A. A. A. M., Astawa, N. M., Damayanti, P. A. A., Kardena, I. M., Erawan, I. G. M. K., Suardana, I. W., et al. (2016). Seroepidemiological evidence for the presence of Japanese encephalitis virus infection in ducks, chickens, and pigs, Bali-Indonesia. Bali Medical Journal, 5, 189.

    Article  Google Scholar 

  11. 11.

    Larena, M., Regner, M., Lee, E., & Lobigs, M. (2011). Pivotal role of antibody and subsidiary contribution of CD8+ T cells to recovery from infection in a murine model of Japanese encephalitis. Journal of Virology, 85(11), 5446–5455.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  12. 12.

    Manocha, G. D., Mishra, R., Sharma, N., Kumawat, K. L., Basu, A., & Singh, S. K. (2014). Regulatory role of TRIM21 in the type-I interferon pathway in Japanese encephalitis virus-infected human microglial cells. Journal of Neuroinflammation, 11(1), 24.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  13. 13.

    Lee, M. S., & Kim, Y. J. (2007). Pattern-recognition receptor signaling initiated from extracellular, membrane, and cytoplasmic space. Molecules and Cells, 23(1), 1–10.

  14. 14.

    Han, Y. W., Choi, J. Y., Uyangaa, E., Kim, S. B., Kim, J. H., Kim, B. S., & Eo, S. K. (2014). Distinct dictation of Japanese encephalitis virus-induced neuroinflammation and lethality via triggering TLR3 and TLR4 signal pathways. PLoS Pathogens, 10(9), e1004319.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  15. 15.

    Zhang, Y., El-Far, M., Dupuy, F. P., Abdel-Hakeem, M. S., He, Z., Procopio, F. A., & Said, E. A. (2016). HCV RNA activates APCs via TLR7/TLR8 while virus selectively stimulates macrophages without inducing antiviral responses. Scientific Reports, 6(1), 1–13.

    Article  CAS  Google Scholar 

  16. 16.

    Awais, M., Wang, K., Lin, X., Qian, W., Zhang, N., Wang, C., & Cui, M. (2017). TLR7 deficiency leads to TLR8 compensative regulation of immune response against JEV in mice. Frontiers in Immunology, 8, 160.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  17. 17.

    Jiang, R., Ye, J., Zhu, B., Song, Y., Chen, H., & Cao, S. (2014). Roles of TLR3 and RIG-I in mediating the inflammatory response in mouse microglia following Japanese encephalitis virus infection. Journal of Immunology Research, 2014, 1–11.

    Article  CAS  Google Scholar 

  18. 18.

    Nazmi, A., Mukherjee, S., Kundu, K., Dutta, K., Mahadevan, A., Shankar, S. K., & Basu, A. (2014). TLR7 is a key regulator of innate immunity against Japanese encephalitis virus infection. Neurobiology of Disease, 69, 235–247.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  19. 19.

    Beladi, I., Pusztai, R., Mucsi, I., Bakay, M., & Gabor, M. (1977). Activity of some flavonoids against viruses. Annals of the New York Academy of Sciences, 284(1), 358–364.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  20. 20.

    Lin, L. T., Chen, T. Y., Lin, S. C., Chung, C. Y., Lin, T. C., Wang, G. H., Anderson, R., Lin, C. C., & Richardson, C. D. (2013). Broadspectrum antiviral activity of chebulagic acid and punicalagin against viruses that use glycosaminoglycans for entry. BMC Microbiology, 13(1), 187.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  21. 21.

    Chakraborty, U., Katoch, S., Sinha, M., Nayak, D., Khurana, A., Manchanda, R. K., & Das, S. (2018). Changes in viral load in different organs of Japanese encephalitis virus-infected chick embryo under the influence of Belladonna 200C. Indian Journal of Research in Homeopathy, 12, 75–80.

  22. 22.

    Yamazaki, Z., & Tagaya, I. (1980). Antiviral effects of atropine and caffeine. Journal of General Virology, 50(2), 429–431.

    CAS  Article  Google Scholar 

  23. 23.

    Chakraborty, U., Sinha, M., Bhattacharjee, A., Nayak, D., Khurana, A., Manchanda, R. K., & Das, S. (2020). Sub-lethal dose of atropine gives protection from Japanese encephalitis virus infection in chick embryo model. In Proceedings of the Zoological Society (pp. 1–8). Springer India.

  24. 24.

    Zárate, R., Hermosin, B., Cantos, M., & Troncoso, A. (1997). Tropane alkaloid distribution in Atropabaetica plants. Journal of Chemical Ecology, 23(8), 2059–2066.

    Article  Google Scholar 

  25. 25.

    Schwahn, H. N., Kaymak, H., & Schaeffel, F. (2000). Effects of atropine on refractive development, dopamine release and slow retinal potentials in the chick. Visual Neuroscience, 17(2), 165–176.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  26. 26.

    Furey, M. L., & Drevets, W. C. (2006). Antidepressant efficacy of the antimuscarinic drug scopolamine: a randomized, placebo-controlled clinical trial. Archives of General Psychiatry, 63(10), 1121–1129.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  27. 27.

    Wagman, W. D., & Maxey, G. C. (1969). The effects of scopolamine hydrobromide and methyl scopolamine hydrobromide upon the discrimination of interoceptive and exteroceptive stimuli. Psychopharmacologia, 15(4), 280288.

    Article  Google Scholar 

  28. 28.

    de Lagarde, M., Rodrigues, N., Chevigny, M., Beauchamp, G., Albrecht, B., & Lavoie, J. P. (2014). N-Butylscopolammonium bromide causes fewer side effects than atropine when assessing bronchoconstriction reversibility in horses with heaves. Equine Veterinary Journal, 46(4), 474–478.

    PubMed  Article  PubMed Central  Google Scholar 

  29. 29.

    McBride, W. G., Vardy, P. H., & French, J. (1982). Effects of scopolamine hydrobromide on the development of the chick and rabbit embryo. Australian Journal of Biological Sciences, 35(2), 173–178.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  30. 30.

    Scher, C. D., Haudenschild, C., & Klagsbrun, M. (1976). The chick chorioallantoic membrane as a model system for the study of tissue invasion by viral transformed cells. Cell, 8(3), 373–382.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  31. 31.

    Nowak-Sliwinska, P., Segura, T., & Iruela-Arispe, M. L. (2014). The chicken chorioallantoic membrane model in biology, medicine and bioengineering. Angiogenesis, 17(4), 779–804.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  32. 32.

    Janzer, R. C. (1993). The blood-brain barrier: cellular basis. Journal of Inherited Metabolic Disease, 16(4), 639–647.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  33. 33.

    Lobrinus, J. A., Juillerat-Jeanneret, L., Darekar, P., Schlosshauer, B., & Janzer, R. C. (1992). Induction of the blood-brain barrier specific HT7 and neurothelin epitopes in endothelial cells of the chick chorioallantoic vessels by a soluble factor derived from astrocytes. Developmental Brain Research, 70(2), 207–211.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  34. 34.

    Holash, J. A., & Stewart, P. A. (1993). Chorioallantoic membrane (CAM) vessels do not respond to blood-brain barrier (BBB) induction. Advances in Experimental Medicine and Biology, 197, 223–228.

    Article  Google Scholar 

  35. 35.

    Yuan, Y. J., Xu, K., Wu, W., Luo, Q., & Yu, J. L. (2014). Application of the chick embryo chorioallantoic membrane in neurosurgery disease. International Journal of Medical Sciences, 11(12), 1275–1281.

    PubMed  PubMed Central  Article  Google Scholar 

  36. 36.

    Toriniwa, H., & Komiya, T. (2007). Japanese encephalitis virus production in Vero cells with serum-free medium using a novel oscillating bioreactor. Biologicals, 35(4), 221–226.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  37. 37.

    Nawa, M. (1997). Japanese encephalitis virus infection in Vero cells: the involvement of intracellular acidic vesicles in the early phase of viral infection was observed with the treatment of a specific vacuolar type H+-ATPase inhibitor, bafilomycin A1. Microbiology and Immunology, 41(7), 537–543.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  38. 38.

    Nawa, M., Takasaki, T., Yamada, K. I., Kurane, I., & Akatsuka, T. (2003). Interference in Japanese encephalitis virus infection of Vero cells by a cationic amphiphilic drug, chlorpromazine. Journal of General Virology, 84(7), 1737–1741.

    CAS  Article  Google Scholar 

  39. 39.

    Wu, S. C., & Huang, G. Y. (2002). Stationary and microcarrier cell culture processes for propagating Japanese encephalitis virus. Biotechnology Progress, 18(1), 124–128.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  40. 40.

    Srivastava, A. K., Putnak, J. R., Lee, S. H., Hong, S. P., Moon, S. B., Barvir, D. A., Zhao, B., Olson, R. A., Kim, S. O., Yoo, W. D., Towle, A. C., Vaughn, D. W., Innis, B. L., & Eckels, K. H. (2001). A purified inactivated Japanese encephalitis virus vaccine made in Vero cells. Vaccine, 19(31), 4557–4565.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  41. 41.

    Trott, O., & Olson, A. J. (2010). AutoDockVina: improving the speed and accuracy of docking with a new scoring function, efficient optimization, and multithreading. Journal of Computational Chemistry, 31(2), 455–461.

    CAS  PubMed  PubMed Central  Google Scholar 

  42. 42.

    Seeliger, D., & de Groot, B. L. (2010). Ligand docking and binding site analysis with PyMOL and Autodock/Vina. Journal of Computer-Aided Molecular Design, 24(5), 417–422.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  43. 43.

    Biovia, D. S. (2017). Discovery Studio Modeling Environment, Release 2017. San Diego, CA: Dassault Systèmes

  44. 44.

    Reed, L. J., & Muench, H. (1938). A simple method of estimating fifty per cent endpoints. American Journal of Epidemiology, 27(3), 493–497.

    Article  Google Scholar 

  45. 45.

    Livak, K. J., & Schmittgen, T. D. (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2− ΔΔCT method. Methods, 25(4), 402–408.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  46. 46.

    Mayta, H., Talley, A., Gilman, R. H., Jimenez, J., Verastegui, M., Ruiz, M., et al. (2000). Differentiating Taenia solium and Taenia saginata infections by simple hematoxylin-eosin staining and PCR-restriction enzyme analysis. Journal of Clinical Microbiology, 38(1), 133–137.

    CAS  PubMed  PubMed Central  Google Scholar 

  47. 47.

    Pieper, J., Methner, U., & Berndt, A. (2008). Heterogeneity of avian γδ T cells. Veterinary Immunology and Immunopathology, 124(3-4), 241–252.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  48. 48.

    Jie, H., Lian, L., Qu, L. J., Zheng, J. X., Hou, Z. C., Xu, G. Y., Song, J. Z., & Yang, N. (2013). Differential expression of Tolllike receptor genes in lymphoid tissues between Marek’s disease virus-infected and noninfected chickens. Poultry Science, 92(3), 645–654.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  49. 49.

    Yao, Y., Xu, X., Li, Y., Wang, X., Yang, H., Chen, J., Liu, S., Deng, Y., Zhao, Z., Yin, Q., Sun, M., & Shi, L. (2020). Study of the association of seventeen single nucleotide polymorphisms and their haplotypes in the TNF-α, IL-2, IL-4 and IL-10 genes with the antibody response to inactivated Japanese encephalitis vaccine. Human Vaccines & Immunotherapeutics, 16(10), 2449–2455.

    CAS  Article  Google Scholar 

  50. 50.

    Wang, T., Town, T., Alexopoulou, L., Anderson, J. F., Fikrig, E., & Flavell, R. A. (2004). Toll-like receptor 3 mediates West Nile virus entry into the brain causing lethal encephalitis. Nature Medicine, 10(12), 1366–1373.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  51. 51.

    Wang, J. P., Liu, P., Latz, E., Golenbock, D. T., Finberg, R. W., & Libraty, D. H. (2006). Flavivirus activation of plasmacytoid dendritic cells delineates key elements of TLR7 signaling beyond endosomal recognition. Immunology, 177(10), 7114–7121.

    CAS  Article  Google Scholar 

  52. 52.

    Song, P., Zheng, N., Zhang, L., Liu, Y., Chen, T., Bao, C., Li, Z., Yong, W., Zhang, Y., Wu, C., & Wu, Z. (2017). Downregulation of interferon-β and inhibition of TLR3 expression are associated with fatal outcome of severe fever with thrombocytopenia syndrome. Scientific Reports, 7(1), 6532.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  53. 53.

    Daffis, S., Samuel, M. A., Suthar, M. S., Gale, M., & Diamond, M. S. (2008). Toll-like receptor 3 has a protective role against West Nile virus infection. Journal of Virology, 82(21), 10349–10358.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  54. 54.

    Matsumoto, M., Oshiumi, H., & Seya, T. (2011). Antiviral responses induced by the TLR3 pathway. Reviews in Medical Virology, 21(2), 67–77.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  55. 55.

    Barton, G. M. (2007). Viral recognition by Toll-like receptors. In Seminars in immunology (Vol. 19, No. 1, pp. 33–40). Academic Press.

  56. 56.

    Yamamoto, M., Sato, S., Hemmi, H., Hoshino, K., Kaisho, T., Sanjo, H., et al. (2003). Role of adaptor TRIF in the MyD88-independent toll-like receptor signaling pathway. Science, 301(5633), 640–643.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  57. 57.

    Shirey, K. A., Lai, W., Scott, A. J., Lipsky, M., Mistry, P., Pletneva, L. M., et al. (2013). The TLR4 antagonist Eritoran protects mice from lethal influenza infection. Nature, 497(7450), 498–502.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  58. 58.

    Best, S. M. (2017). The many faces of the flavivirus NS5 protein in antagonism of type I interferon signaling. Journal of Virology, 91(3), e01970–e01916.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  59. 59.

    Philbin, V. J., Iqbal, M., Boyd, Y., Goodchild, M. J., Beal, R. K., Bumstead, N., et al. (2005). Identification and characterization of a functional, alternatively spliced Toll-like receptor 7 (TLR7) and genomic disruption of TLR8 in chickens. Immunology, 114(4), 507–521.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  60. 60.

    Lester, S. N., & Li, K. (2014). Toll-like receptors in antiviral innate immunity. Journal of Molecular Biology, 426(6), 1246–1264.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  61. 61.

    Kao, Y. T., Chang, B. L., Liang, J. J., Tsai, H. J., Lee, Y. L., Lin, R. J., & Lin, Y. L. (2015). Japanese encephalitis virus nonstructural protein NS5 interacts with mitochondrial trifunctional protein and impairs fatty acid β-oxidation. PLoS Pathogens, 11(3), e1004750.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  62. 62.

    Lin, R. J., Chang, B. L., Yu, H. P., Liao, C. L., & Lin, Y. L. (2006). Blocking of interferon-induced Jak-Stat signaling by Japanese encephalitis virus NS5 through a protein tyrosine phosphatase-mediated mechanism. Journal of Virology, 80(12), 5908–5918.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  63. 63.

    Gilhare, V. R., Hirpurkar, S. D., Kumar, A., Naik, S. K., & Sahu, T. (2015). Pock forming ability of fowl pox virus isolated from layer chicken and its adaptation in chicken embryo fibroblast cell culture. Veterinary World, 8(3), 245–250.

    PubMed  PubMed Central  Article  Google Scholar 

  64. 64.

    Glick, S. D., & Zimmerberg, B. (1972). Amnesic effects of scopolamine. Behavioral Biology, 7(2), 245–254.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  65. 65.

    Chang, R. Y., Hsu, T. W., Chen, Y. L., Liu, S. F., Tsai, Y. J., Lin, Y. T., Chen, Y. S., & Fan, Y. H. (2013). Japanese encephalitis virus non-coding RNA inhibits activation of interferon by blocking nuclear translocation of interferon regulatory factor 3. Veterinary Microbiology, 166(1-2), 11–21.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  66. 66.

    Ye, J., Chen, Z., Li, Y., Zhao, Z., He, W., Zohaib, A., et al. (2017). Japanese encephalitis virus NS5 inhibits type I interferon (IFN) production by blocking the nuclear translocation of IFN regulatory factor 3 and NF-κB. Journal of Virology, 91(8), e00039–e00017.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  67. 67.

    Kennedy, M. K., Torrance, D. S., Picha, K. S., & Mohler, K. M. (1992). Analysis of cytokine mRNA expression in the central nervous system of mice with experimental autoimmune encephalomyelitis reveals that IL-10 mRNA expression correlates with recovery. Journal of Immunology, 149, 2496–2505.

    CAS  Google Scholar 

  68. 68.

    Saxena, V., Mathur, A., Krishnani, N., & Dhole, T. N. (2008). An insufficient anti-inflammatory cytokine response in mouse brain is associated with increased tissue pathology and viral load during Japanese encephalitis virus infection. Archives of Virology, 153, 282–292.

    Article  CAS  Google Scholar 

Download references

Acknowledgements

The authors express their deep gratitude of thanks to the Department of Microbiology, Peerless Hospital & B.K Roy Research Centre, Kolkata, Department of Cardiology, R.G Kar Medical College & Hospital, Kolkata, and Department of Botany, Berhampur University, Odisha. We are also thankful to the National Institute of Virology, Pune, for providing the viral strains of JEV. The authors are also deeply indebted to Rev. Dr. Dominic Savio, S.J., Principal and Rector, St. Xavier’s College (Autonomous), Kolkata, Dr. Subhankar Tripathi, Principal, Sarsuna College, Kolkata, and Dr. Satadal Das, Consultant Senior Microbiologist, Peerless Hospital & B.K Roy Research Centre, for their constant support and guidance throughout.

Author information

Affiliations

Authors

Contributions

The experimental design was performed collectively by all the authors. Preparation of reagents and samples along with data collection and analysis was done by AB, LC, and SR. The manuscript was written by AB and RC with inputs from all the authors. Conceptualization: AB, MS; methodology: AB, LC, and JJD; writing—original draft preparation: AB; writing review and editing: AB, RC, and SR; resources: AB, JJD, and MS.

Corresponding author

Correspondence to Souvik Roy.

Ethics declarations

Ethics Approval

Not applicable.

Consent to Participate

Not applicable.

Consent to Publish

All the authors consent to the consideration for this manuscript for publication.

Competing Interests

The authors declare no competing interests.

Additional information

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Bhattacharjee, A., Chaudhuri, R., Dash, J.J. et al. Pre-treatment with Scopolamine Naturally Suppresses Japanese Encephalitis Viral Load in Embryonated Chick Through Regulation of Multiple Signaling Pathways. Appl Biochem Biotechnol (2021). https://doi.org/10.1007/s12010-021-03526-8

Download citation

Keywords

  • Toll-like receptors
  • Interferons
  • Japanese encephalitis virus
  • Scopolamine hydrobromide
  • Embryonated chick
  • Signaling pathways