Viral evolution sustains a dengue outbreak of enhanced severity

Catherine Inizan, Marine Minier, Matthieu Prot, Olivia O'Connor, Carole Forfait, Sylvie Laumond, Ingrid Marois, Antoine Biron, Ann-Claire Gourinat, Marie-Amélie Goujart, Elodie Descloux, Anavaj Sakuntabhai, Arnaud Tarantola, Etienne Simon-Lorière, Myrielle Dupont-Rouzeyrol, Catherine Inizan, Marine Minier, Matthieu Prot, Olivia O'Connor, Carole Forfait, Sylvie Laumond, Ingrid Marois, Antoine Biron, Ann-Claire Gourinat, Marie-Amélie Goujart, Elodie Descloux, Anavaj Sakuntabhai, Arnaud Tarantola, Etienne Simon-Lorière, Myrielle Dupont-Rouzeyrol

Abstract

Compared to the previous 2013-2014 outbreak, dengue 2016-2017 outbreak in New Caledonia was characterized by an increased number of severe forms associated with hepatic presentations. In this study, we assessed the virological factors associated with this enhanced severity. Whole-genome sequences were retrieved from dengue virus (DENV)-1 strains collected in 2013-2014 and from severe and non-severe patients in 2016-2017. Fitness, hepatic tropism and cytopathogenicity of DENV 2016-2017 strains were compared to those of 2013-2014 strains using replication kinetics in the human hepatic cell line HuH7. Whole-genome sequencing identified four amino acid substitutions specific to 2016-2017 strains and absent from 2013-2014 strains. Three of these mutations occurred in predicted T cell epitopes, among which one was also a B cell epitope. Strains retrieved from severe forms did not exhibit specific genetic features. DENV strains from 2016-2017 exhibited a trend towards reduced replicative fitness and cytopathogenicity in vitro compared to strains from 2013-2014. Overall, the 2016-2017 dengue outbreak in New Caledonia was associated with a viral genetic evolution which had limited impact on DENV hepatic tropism and cytopathogenicity. These mutations, however, may have modified DENV strains antigenicity, altering the anti-DENV immune response in some patients, in turn favoring the development of severe forms.Trial registration: ClinicalTrials.gov identifier: NCT04615364.

Keywords: Severe dengue; hepatitis; viral fitness; whole-genome sequencing.

Conflict of interest statement

No potential conflict of interest was reported by the author(s).

Figures

Figure 1.
Figure 1.
Phylogenetic tree of DENV-1 whole-genome sequences retrieved from dengue patients sera from 2013–2014 and 2016–2017. Whole-genome sequences retrieved from severe forms are highlighted in blue. The tree is midpoint rooted and bootstrap values above 80% are indicated. Two DENV-1 genomes from strains retrieved in NC in 2014 are displayed on the tree. The four amino acid substitutions detected in strains from 2016–2017 in comparison to strains from 2013–2014 are indicated.
Figure 2.
Figure 2.
Replication kinetics of DENV-1 viral isolates from 2013–2014 and 2016–2017 in the human hepatic cell line HuH7. A DENV strain isolated in NC in 2014 passaged several times in C6/36 cells and yielding a high infection rate at day 3 post-infection was used as positive control. Human hepatic HuH7 cells were infected at a MOI of 1 with DENV-1 from 2013–2014 and 2016–2017. A. Percentages of HuH7 cells staining positive for 2H2 antibody measured by flow cytometry at days 1, 2 and 3 post-infection. Percentages of infected cells are normalized by the percentage of infected cells of the positive control at day 3 post-infection. B. Productivity of the infection measured as the viral titer in the supernatant of HuH7 at day 2 post-infection. C. Percentage of dead HuH7 cells staining positive for Viability dye measured by flow cytometry at days 1, 2 and 3 post-infection. NI: non-infected. Each dot is the mean of three independent replicates. Medians and interquartile ranges are indicated. ns: non significant difference in Wilcoxon-Mann-Whitney ranksum test.

References

    1. Horstick O, Farrar J, Lum L, et al. . Reviewing the development, evidence base, and application of the revised dengue case classification. Pathog Glob Health. 2012;106(2):94–101. doi:10.1179/2047773212y.0000000017.
    1. WHO . Dengue: guidelines for diagnosis, treatment, prevention and control. New Edition; Geneva: WHO Press; 2009.
    1. Brady OJ, Gething PW, Bhatt S, et al. . Refining the global spatial limits of dengue virus transmission by evidence-based consensus. PLoS Negl Trop Dis. 2012;6(8):e1760. doi:10.1371/journal.pntd.0001760.
    1. Inizan C, Tarantola A, O'Connor O, et al. . Dengue in New Caledonia: knowledge and gaps. Trop Med Infect Dis. 2019; 4(2). doi:10.3390/tropicalmed4020095.
    1. Cao-Lormeau VM, Musso D.. Emerging arboviruses in the pacific. Lancet. 2014;384(9954):1571–1572. doi:10.1016/s0140-6736(14)61977-2.
    1. DASS-NC . Documents, rapports, études - La situation sanitaire de la Nouvelle-Calédonie.
    1. Marois I, Forfait C, Inizan C, et al. . Development of a bedside score to predict dengue severity. medRxiv. 2020: 2020. doi:10.1101/2020.11.25.20238972.
    1. Warrilow D, Northill JA, Pyke A, et al. . Single rapid TaqMan fluorogenic probe based PCR assay that detects all four dengue serotypes. J Med Virol. 2002;66(4):524–528.
    1. CDC Trioplex Real-time RT-PCR Assay; 2017.
    1. Johnson BW, Russell BJ, Lanciotti RS.. Serotype-specific detection of dengue viruses in a fourplex real-time reverse transcriptase PCR assay. J Clin Microbiol. 2005;43(10):4977–4983. doi:10.1128/jcm.43.10.4977-4983.2005.
    1. Simon-Loriere E, Faye O, Koivogui L, et al. . Distinct lineages of Ebola virus in Guinea during the 2014 west african epidemic. Nature. 2015;524(7563):102–104. doi:10.1038/nature14612.
    1. Bolger AM, Lohse M, Usadel B.. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics. 2014;30(15):2114–2120. doi:10.1093/bioinformatics/btu170.
    1. Nurk S, Meleshko D, Korobeynikov A, et al. . metaSPAdes: a new versatile metagenomic assembler. Genome Res. 2017;27(5):824–834. doi:10.1101/gr.213959.116.
    1. National Center for Biotechnology Information (NCBI)[Internet]. Bethesda (MD): National Library of Medicine (US) NCfBI; 1988.
    1. Buchfink B, Xie C, Huson DH.. Fast and sensitive protein alignment using DIAMOND. Nat Methods. 2015;12(1):59–60. doi:10.1038/nmeth.3176.
    1. Pickett BE, Sadat EL, Zhang Y, et al. . ViPR: an open bioinformatics database and analysis resource for virology research. Nucleic Acids Res. 2012;40(Database issue):D593–D598. doi:10.1093/nar/gkr859.
    1. Katoh K, Standley DM.. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol. 2013;30(4):772–780. doi:10.1093/molbev/mst010.
    1. Nguyen LT, Schmidt HA, von Haeseler A, et al. . IQ-TREE: a fast and effective stochastic algorithm for estimating maximum-likelihood phylogenies. Mol Biol Evol. 2015;32(1):268–274. doi:10.1093/molbev/msu300.
    1. Hoang DT, Chernomor O, von Haeseler A, et al. . UFBoot2: improving the ultrafast bootstrap approximation. Mol Biol Evol. 2018;35(2):518–522. DOI:10.1093/molbev/msx281.
    1. Kalyaanamoorthy S, Minh BQ, Wong TKF, et al. . Modelfinder: fast model selection for accurate phylogenetic estimates. Nat Methods. 2017;14(6):587–589. DOI:10.1038/nmeth.4285.
    1. Vu TT, Holmes EC, Duong V, et al. . Emergence of the Asian 1 genotype of dengue virus serotype 2 in viet nam: in vivo fitness advantage and lineage replacement in South-East Asia. PLoS Negl Trop Dis. 2010;4(7):e757), doi:10.1371/journal.pntd.0000757.
    1. Lee KS, Lo S, Tan SS, et al. . Dengue virus surveillance in Singapore reveals high viral diversity through multiple introductions and in situ evolution. Infect Genet Evol. 2012;12(1):77–85. doi:10.1016/j.meegid.2011.10.012.
    1. Thu H M, Lowry K, Jiang L, et al. . Lineage extinction and replacement in dengue type 1 virus populations are due to stochastic events rather than to natural selection. Virology. 2005;336(2):163–172. doi:10.1016/j.virol.2005.03.018.
    1. Yamanaka A, Mulyatno KC, Susilowati H, et al. . Displacement of the predominant dengue virus from type 2 to type 1 with a subsequent genotype shift from IV to I in surabaya, Indonesia 2008-2010. PloS one. 2011;6(11):e27322. doi:10.1371/journal.pone.0027322.
    1. Bennett SN, Drummond AJ, Kapan DD, et al. . Epidemic dynamics revealed in dengue evolution. Mol Biol Evol. 2010;27(4):811–818. doi:10.1093/molbev/msp285.
    1. Kukreti H, Mittal V, Chaudhary A, et al. . Continued persistence of a single genotype of dengue virus type-3 (DENV-3) in Delhi, India since its re-emergence over the last decade. J Microbiol Immunol Infect. 2010;43(1):53–61. doi:10.1016/s1684-1182(10)60008-4.
    1. Zhang C, Mammen MP Jr., Chinnawirotpisan P, et al. . Clade replacements in dengue virus serotypes 1 and 3 are associated with changing serotype prevalence. J Virol. 2005; 79(24): 15123–15130. doi:10.1128/jvi.79.24.15123-15130.2005.
    1. Tuiskunen A, Monteil V, Plumet S, et al. . Phenotypic and genotypic characterization of dengue virus isolates differentiates dengue fever and dengue hemorrhagic fever from dengue shock syndrome. Arch Virol. 2011;156(11):2023–2032. doi:10.1007/s00705-011-1100-2.
    1. Campen A, Williams RM, Brown CJ, et al. . TOP-IDP-scale: a new amino acid scale measuring propensity for intrinsic disorder. Protein Pept Lett. 2008;15(9):956–963.
    1. Lai Y-C, Chuang Y-C, Liu C-C, et al. . Antibodies against modified NS1 wing domain peptide Protect against dengue virus infection. Sci Rep. 2017;7(1):6975. doi:10.1038/s41598-017-07308-3.
    1. Lin C-F, Wan S-W, Chen M-C, et al. . Liver injury caused by antibodies against dengue virus nonstructural protein 1 in a murine model. Lab Invest. 2008;88:1079. doi:10.1038/labinvest.2008.70. .
    1. Steel A, Gubler DJ, Bennett SN.. Natural attenuation of dengue virus type-2 after a series of island outbreaks: A retrospective phylogenetic study of events in the South Pacific three decades ago. Virology. 2010;405(2):505–512. doi:10.1016/j.virol.2010.05.033.
    1. Tsetsarkin KA, Vanlandingham DL, McGee CE, et al. . A single mutation in chikungunya virus affects vector specificity and epidemic potential. PLoS Pathog. 2007;3(12):e201. doi:10.1371/journal.ppat.0030201.
    1. Wong G, He S, Leung A, et al. . Naturally occurring single mutations in Ebola virus observably impact infectivity. J Virol. 2019;93(1):e01098–18. doi:10.1128/jvi.01098-18.
    1. Pryor MJ, Carr JM, Hocking H, et al. . Replication of dengue virus type 2 in human monocyte-derived macrophages: comparisons of isolates and recombinant viruses with substitutions at amino acid 390 in the envelope glycoprotein. Am J Trop Med Hyg. 2001;65(5):427–434. doi:10.4269/ajtmh.2001.65.427.
    1. Cologna R, Rico-Hesse R.. American genotype structures decrease dengue virus output from human monocytes and dendritic cells. J Virol. 2003;77(7):3929–3938. doi:10.1128/jvi.77.7.3929-3938.2003.
    1. Inizan C, O'Connor O, Worwor G, et al. . Molecular characterization of dengue type 2 outbreak in Pacific islands countries and territories, 2017-2020. Viruses. 2020; 12(10). doi:10.3390/v12101081.
    1. Morens DM, Marchette NJ, Chu MC, et al. . Growth of dengue type 2 virus isolates in human peripheral blood leukocytes correlates with severe and mild dengue disease. Am J Trop Med Hyg. 1991;45(5):644–651. doi:10.4269/ajtmh.1991.45.644.
    1. Gutiérrez-Barbosa H, Castañeda NY, Castellanos JE.. Differential replicative fitness of the four dengue virus serotypes circulating in Colombia in human liver Huh7 cells. Braz J Infect Dis. 2020;24(1):13–24. doi:10.1016/j.bjid.2019.11.003.
    1. Payne AF, Binduga-Gajewska I, Kauffman EB, et al. . Quantitation of flaviviruses by fluorescent focus assay. J Virol Methods. 2006;134(1-2):183–189. doi:10.1016/j.jviromet.2006.01.003.

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