BACKGROUND: The outbreak of Zika virus (ZIKV) in the Americas has transformed a previously obscure mosquito-transmitted arbovirus of the Flaviviridae family into a major public health concern. Little is currently known about the evolution and biology of ZIKV and the factors that contribute to the associated pathogenesis. Determining genomic sequences of clinical viral isolates and characterization of elements within these are an important prerequisite to advance our understanding of viral replicative processes and virus-host interactions. METHODOLOGY/PRINCIPAL FINDINGS: We obtained a ZIKV isolate from a patient who presented with classical ZIKV-associated symptoms, and used high throughput sequencing and other molecular biology approaches to determine its full genome sequence, including non-coding regions. Genome regions were characterized and compared to the sequences of other isolates where available. Furthermore, we identified a subgenomic flavivirus RNA (sfRNA) in ZIKV-infected cells that has antagonist activity against RIG-I induced type I interferon induction, with a lesser effect on MDA-5 mediated action. CONCLUSIONS/SIGNIFICANCE: The full-length genome sequence including non-coding regions of a South American ZIKV isolate from a patient with classical symptoms will support efforts to develop genetic tools for this virus. Detection of sfRNA that counteracts interferon responses is likely to be important for further understanding of pathogenesis and virus-host interactions.

Boyd Orr Centre for Population and Ecosystem Health Institute of Biodiversity Animal Health and Comparative Medicine College of Medical Veterinary and Life Sciences University of Glasgow Glasgow Scotland United Kingdom

Department of Microbial Pathogenesis Yale University New Haven Connecticut United States of America

Faculty of Science University of South Bohemia České Budějovice Czech Republic

Fundação Oswaldo Cruz PE Centro de Pesquisas Aggeu Magalhães Departamento de Virologia Campus da UFPE Cidade Universitária Recife PE Brasil

Institute of Parasitology Biology Centre of the Academy of Sciences of the Czech Republic České Budějovice Czech Republic

Medical Research Council Human Immunology Unit Weatherall Institute of Molecular Medicine and Radcliffe Department of Medicine University of Oxford Oxford England United Kingdom

MRC University of Glasgow Centre for Virus Research Glasgow Scotland United Kingdom

Pole de Virologie Unité des arbovirus et virus des fièvres hémorragiques Institut Pasteur de Dakar Dakar Senegal

Zobrazit více v PubMed

Hancock WT, Marfel M, Bel M. Zika virus, French Polynesia, South Pacific, 2013. Emerg Infect Dis. 2014;20(11):1960 10.3201/eid2011.141380 PubMed DOI PMC

Cao-Lormeau VM, Musso D. Emerging arboviruses in the Pacific. Lancet. 2014;384(9954):1571–2. 10.1016/S0140-6736(14)61977-2 . PubMed DOI

Dupont-Rouzeyrol M, O'Connor O, Calvez E, Daures M, John M, Grangeon JP, et al. Co-infection with Zika and dengue viruses in 2 patients, New Caledonia, 2014. Emerg Infect Dis. 2015;21(2):381–2. 10.3201/eid2102.141553 PubMed DOI PMC

Musso D, Nilles EJ, Cao-Lormeau VM. Rapid spread of emerging Zika virus in the Pacific area. Clin Microbiol Infect. 2014;20(10):O595–6. 10.1111/1469-0691.12707 . PubMed DOI

Tognarelli J, Ulloa S, Villagra E, Lagos J, Aguayo C, Fasce R, et al. A report on the outbreak of Zika virus on Easter Island, South Pacific, 2014. Arch Virol. 2015. 10.1007/s00705-015-2695-5 . PubMed DOI

Campos GS, Bandeira AC, Sardi SI. Zika Virus Outbreak, Bahia, Brazil. Emerg Infect Dis. 2015;21(10):1885–6. 10.3201/eid2110.150847 PubMed DOI PMC

Zanluca C, Melo VC, Mosimann AL, Santos GI, Santos CN, Luz K. First report of autochthonous transmission of Zika virus in Brazil. Mem Inst Oswaldo Cruz. 2015;110(4):569–72. 10.1590/0074-02760150192 PubMed DOI PMC

de Oliveira CS, da Costa Vasconcelos PF. Microcephaly and Zika virus. J Pediatr (Rio J). 2016;92(2):103–5. 10.1016/j.jped.2016.02.003 . PubMed DOI

Oehler E, Watrin L, Larre P, Leparc-Goffart I, Lastere S, Valour F, et al. Zika virus infection complicated by Guillain-Barre syndrome—case report, French Polynesia, December 2013. Euro Surveill. 2014;19(9). . PubMed

Mlakar J, Korva M, Tul N, Popovic M, Poljsak-Prijatelj M, Mraz J, et al. Zika Virus Associated with Microcephaly. N Engl J Med. 2016;374(10):951–8. 10.1056/NEJMoa1600651 . PubMed DOI

Cao-Lormeau VM, Blake A, Mons S, Lastere S, Roche C, Vanhomwegen J, et al. Guillain-Barre Syndrome outbreak associated with Zika virus infection in French Polynesia: a case-control study. Lancet. 2016. 10.1016/S0140-6736(16)00562-6 . PubMed DOI PMC

Schuler-Faccini L, Ribeiro EM, Feitosa IM, Horovitz DD, Cavalcanti DP, Pessoa A, et al. Possible Association Between Zika Virus Infection and Microcephaly—Brazil, 2015. MMWR Morb Mortal Wkly Rep. 2016;65(3):59–62. 10.15585/mmwr.mm6503e2 . PubMed DOI

Rasmussen SA, Jamieson DJ, Honein MA, Petersen LR. Zika Virus and Birth Defects—Reviewing the Evidence for Causality. N Engl J Med. 2016:In press. 10.1056/NEJMsr1604338 . PubMed DOI

WHO. Situation Report: Zika virus, Microcephaly and Guillain-Barre syndrome The World Health Organisation; 2016.

Weaver SC, Costa F, Garcia-Blanco MA, Ko AI, Ribeiro GS, Saade G, et al. Zika virus: History, emergence, biology, and prospects for control. Antiviral Res. 2016;130:69–80. 10.1016/j.antiviral.2016.03.010 . PubMed DOI PMC

Weaver SC, Reisen WK. Present and future arboviral threats. Antiviral Res. 2010;85(2):328–45. 10.1016/j.antiviral.2009.10.008 PubMed DOI PMC

Chouin-Carneiro T, Vega-Rua A, Vazeille M, Yebakima A, Girod R, Goindin D, et al. Differential Susceptibilities of Aedes aegypti and Aedes albopictus from the Americas to Zika Virus. PLoS Negl Trop Dis. 2016;10(3):e0004543 10.1371/journal.pntd.0004543 PubMed DOI PMC

Wong PS, Li MZ, Chong CS, Ng LC, Tan CH. Aedes (Stegomyia) albopictus (Skuse): a potential vector of Zika virus in Singapore. PLoS Negl Trop Dis. 2013;7(8):e2348 10.1371/journal.pntd.0002348 PubMed DOI PMC

Grard G, Caron M, Mombo IM, Nkoghe D, Mboui Ondo S, Jiolle D, et al. Zika virus in Gabon (Central Africa)—2007: a new threat from Aedes albopictus? PLoS Negl Trop Dis. 2014;8(2):e2681 10.1371/journal.pntd.0002681 PubMed DOI PMC

Villamil-Gomez WE, Gonzalez-Camargo O, Rodriguez-Ayubi J, Zapata-Serpa D, Rodriguez-Morales AJ. Dengue, chikungunya and Zika co-infection in a patient from Colombia. J Infect Public Health. 2016. 10.1016/j.jiph.2015.12.002 . PubMed DOI

Lindenbach BD, Murray CL, Thiel HJ, Rice CM. Flaviviridae In: Knipe DM, Howley PM, editors. Fields Virology. 1 6th ed. Philadelphia: Lippincott Williams and W; 2013. p. 712–46.

Lanciotti RS, Kosoy OL, Laven JJ, Velez JO, Lambert AJ, Johnson AJ, et al. Genetic and serologic properties of Zika virus associated with an epidemic, Yap State, Micronesia, 2007. Emerg Infect Dis. 2008;14(8):1232–9. 10.3201/eid1408.080287 PubMed DOI PMC

Faye O, Freire CC, Iamarino A, Faye O, de Oliveira JV, Diallo M, et al. Molecular evolution of Zika virus during its emergence in the 20(th) century. PLoS Negl Trop Dis. 2014;8(1):e2636 10.1371/journal.pntd.0002636 PubMed DOI PMC

Haddow AD, Schuh AJ, Yasuda CY, Kasper MR, Heang V, Huy R, et al. Genetic characterization of Zika virus strains: geographic expansion of the Asian lineage. PLoS Negl Trop Dis. 2012;6(2):e1477 10.1371/journal.pntd.0001477 PubMed DOI PMC

Brinton MA, Basu M. Functions of the 3' and 5' genome RNA regions of members of the genus Flavivirus. Virus Res. 2015;206:108–19. 10.1016/j.virusres.2015.02.006 PubMed DOI PMC

Villordo SM, Carballeda JM, Filomatori CV, Gamarnik AV. RNA Structure Duplications and Flavivirus Host Adaptation. Trends Microbiol. 2016;24(4):270–83. 10.1016/j.tim.2016.01.002 PubMed DOI PMC

Roby JA, Pijlman GP, Wilusz J, Khromykh AA. Noncoding subgenomic flavivirus RNA: multiple functions in West Nile virus pathogenesis and modulation of host responses. Viruses. 2014;6(2):404–27. 10.3390/v6020404 PubMed DOI PMC

Clarke BD, Roby JA, Slonchak A, Khromykh AA. Functional non-coding RNAs derived from the flavivirus 3' untranslated region. Virus Res. 2015;206:53–61. 10.1016/j.virusres.2015.01.026 . PubMed DOI

Schnettler E, Sterken MG, Leung JY, Metz SW, Geertsema C, Goldbach RW, et al. Noncoding flavivirus RNA displays RNA interference suppressor activity in insect and Mammalian cells. J Virol. 2012;86(24):13486–500. 10.1128/JVI.01104-12 PubMed DOI PMC

Schnettler E, Tykalova H, Watson M, Sharma M, Sterken MG, Obbard DJ, et al. Induction and suppression of tick cell antiviral RNAi responses by tick-borne flaviviruses. Nucleic Acids Res. 2014;42(14):9436–46. 10.1093/nar/gku657 PubMed DOI PMC

Chang RY, Hsu TW, Chen YL, Liu SF, Tsai YJ, Lin YT, et al. Japanese encephalitis virus non-coding RNA inhibits activation of interferon by blocking nuclear translocation of interferon regulatory factor 3. Vet Microbiol. 2013;166(1–2):11–21. 10.1016/j.vetmic.2013.04.026 . PubMed DOI

Moon SL, Dodd BJ, Brackney DE, Wilusz CJ, Ebel GD, Wilusz J. Flavivirus sfRNA suppresses antiviral RNA interference in cultured cells and mosquitoes and directly interacts with the RNAi machinery. Virology. 2015;485:322–9. 10.1016/j.virol.2015.08.009 PubMed DOI PMC

Pijlman GP, Funk A, Kondratieva N, Leung J, Torres S, van der Aa L, et al. A highly structured, nuclease-resistant, noncoding RNA produced by flaviviruses is required for pathogenicity. Cell Host Microbe. 2008;4(6):579–91. 10.1016/j.chom.2008.10.007 . PubMed DOI

Manokaran G, Finol E, Wang C, Gunaratne J, Bahl J, Ong EZ, et al. Dengue subgenomic RNA binds TRIM25 to inhibit interferon expression for epidemiological fitness. Science. 2015;350(6257):217–21. 10.1126/science.aab3369 PubMed DOI PMC

Schuessler A, Funk A, Lazear HM, Cooper DA, Torres S, Daffis S, et al. West Nile virus noncoding subgenomic RNA contributes to viral evasion of the type I interferon-mediated antiviral response. J Virol. 2012;86(10):5708–18. 10.1128/JVI.00207-12 PubMed DOI PMC

Bidet K, Dadlani D, Garcia-Blanco MA. G3BP1, G3BP2 and CAPRIN1 are required for translation of interferon stimulated mRNAs and are targeted by a dengue virus non-coding RNA. PLoS Pathog. 2014;10(7):e1004242 10.1371/journal.ppat.1004242 PubMed DOI PMC

Brennan B, Welch SR, Elliott RM. The consequences of reconfiguring the ambisense S genome segment of Rift Valley fever virus on viral replication in mammalian and mosquito cells and for genome packaging. PLoS Pathog. 2014;10(2):e1003922 10.1371/journal.ppat.1003922 PubMed DOI PMC

Carlos TS, Young DF, Schneider M, Simas JP, Randall RE. Parainfluenza virus 5 genomes are located in viral cytoplasmic bodies whilst the virus dismantles the interferon-induced antiviral state of cells. J Gen Virol. 2009;90(Pt 9):2147–56. 10.1099/vir.0.012047–0 PubMed DOI PMC

Hilton L, Moganeradj K, Zhang G, Chen YH, Randall RE, McCauley JW, et al. The NPro product of bovine viral diarrhea virus inhibits DNA binding by interferon regulatory factor 3 and targets it for proteasomal degradation. J Virol. 2006;80(23):11723–32. 10.1128/JVI.01145-06 PubMed DOI PMC

Yoneyama M, Suhara W, Fukuhara Y, Fukuda M, Nishida E, Fujita T. Direct triggering of the type I interferon system by virus infection: activation of a transcription factor complex containing IRF-3 and CBP/p300. Embo J. 1998;17(4):1087–95. 10.1093/emboj/17.4.1087 . PubMed DOI PMC

Pichlmair A, Schulz O, Tan CP, Rehwinkel J, Kato H, Takeuchi O, et al. Activation of MDA5 requires higher-order RNA structures generated during virus infection. J Virol. 2009;83(20):10761–9. 10.1128/JVI.00770-09 PubMed DOI PMC

Rehwinkel J, Tan CP, Goubau D, Schulz O, Pichlmair A, Bier K, et al. RIG-I detects viral genomic RNA during negative-strand RNA virus infection. Cell. 2010;140(3):397–408. 10.1016/j.cell.2010.01.020 . PubMed DOI

Langmead B, Salzberg SL. Fast gapped-read alignment with Bowtie 2. Nat Methods. 2012;9(4):357–9. 10.1038/nmeth.1923 PubMed DOI PMC

Edgar RC. MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res. 2004;32(5):1792–7. 10.1093/nar/gkh340 PubMed DOI PMC

Kearse M, Moir R, Wilson A, Stones-Havas S, Cheung M, Sturrock S, et al. Geneious Basic: an integrated and extendable desktop software platform for the organization and analysis of sequence data. Bioinformatics. 2012;28(12):1647–9. 10.1093/bioinformatics/bts199 PubMed DOI PMC

Martin DP, Murrell B, Golden M, Khoosal A, Muhire B. RDP4: Detection and analysis of recombination patterns in virus genomes. Virus Evolution. 2015;1(1). 10.1093/ve/vev003 PubMed DOI PMC

Guindon S, Gascuel O. A simple, fast, and accurate algorithm to estimate large phylogenies by maximum likelihood. Syst Biol. 2003;52(5):696–704. . PubMed

Huelsenbeck JP, Ronquist F. MRBAYES: Bayesian inference of phylogenetic trees. Bioinformatics. 2001;17(8):754–5. . PubMed

Darriba D, Taboada GL, Doallo R, Posada D. jModelTest 2: more models, new heuristics and parallel computing. Nat Methods. 2012;9(8):772 10.1038/nmeth.2109 PubMed DOI PMC

Pond SL, Frost SD, Muse SV. HyPhy: hypothesis testing using phylogenies. Bioinformatics. 2005;21(5):676–9. 10.1093/bioinformatics/bti079 . PubMed DOI

Ladner JT, Wiley MR, Prieto K, Yasuda CY, Nagle E, Kasper MR, et al. Complete Genome Sequences of Five Zika Virus Isolates. Genome Announc. 2016;4(3). 10.1128/genomeA.00377-16 . PubMed DOI PMC

Giovanetti M, Faria NR, Nunes MR, de Vasconcelos JM, Lourenco J, Rodrigues SG, et al. Zika virus complete genome from Salvador, Bahia, Brazil. Infect Genet Evol. 2016. 10.1016/j.meegid.2016.03.030 . PubMed DOI

Lanciotti RS, Lambert AJ, Holodniy M, Saavedra S, Signor Ldel C. Phylogeny of Zika Virus in Western Hemisphere, 2015. Emerg Infect Dis. 2016;22(5):933–5. 10.3201/eid2205.160065 PubMed DOI PMC

Faria NR, Azevedo Rdo S, Kraemer MU, Souza R, Cunha MS, Hill SC, et al. Zika virus in the Americas: Early epidemiological and genetic findings. Science. 2016;352(6283):345–9. 10.1126/science.aaf5036 . PubMed DOI PMC

Diamond MS, Gale M Jr. Cell-intrinsic innate immune control of West Nile virus infection. Trends Immunol. 2012;33(10):522–30. 10.1016/j.it.2012.05.008 PubMed DOI PMC

Lazear HM, Diamond MS. New insights into innate immune restriction of West Nile virus infection. Curr Opin Virol. 2015;11:1–6. 10.1016/j.coviro.2014.12.001 PubMed DOI PMC

Green AM, Beatty PR, Hadjilaou A, Harris E. Innate immunity to dengue virus infection and subversion of antiviral responses. J Mol Biol. 2014;426(6):1148–60. 10.1016/j.jmb.2013.11.023 PubMed DOI PMC

Castillo Ramirez JA, Urcuqui-Inchima S. Dengue Virus Control of Type I IFN Responses: A History of Manipulation and Control. J Interferon Cytokine Res. 2015;35(6):421–30. 10.1089/jir.2014.0129 PubMed DOI PMC

Lazear HM, Govero J, Smith AM, Platt DJ, Fernandez E, Miner JJ, et al. A Mouse Model of Zika Virus Pathogenesis. Cell Host Microbe. 2016;19(5):720–30. 10.1016/j.chom.2016.03.010 PubMed DOI PMC

Bayer A, Lennemann NJ, Ouyang Y, Bramley JC, Morosky S, Marques ET Jr., et al. Type III Interferons Produced by Human Placental Trophoblasts Confer Protection against Zika Virus Infection. Cell Host Microbe. 2016;19(5):705–12. 10.1016/j.chom.2016.03.008 PubMed DOI PMC

Grant A, Ponia SS, Tripathi S, Balasubramaniam V, Miorin L, Sourisseau M, et al. Zika Virus Targets Human STAT2 to Inhibit Type I Interferon Signaling. Cell Host Microbe.In press. PubMed PMC

Oshiumi H, Kouwaki T, Seya T. Accessory Factors of Cytoplasmic Viral RNA Sensors Required for Antiviral Innate Immune Response. Front Immunol. 2016;7:200 10.3389/fimmu.2016.00200 PubMed DOI PMC

Yoneyama M, Onomoto K, Jogi M, Akaboshi T, Fujita T. Viral RNA detection by RIG-I-like receptors. Curr Opin Immunol. 2015;32:48–53. 10.1016/j.coi.2014.12.012 . PubMed DOI

Freire CCdM, Iamarino A, Neto DFdL, Sall AA, Zanotto PMdA. Spread of the pandemic Zika virus lineage is associated with NS1 codon usage adaptation in humans. bioRxiv. 2015. 10.1101/032839 DOI

Shan C, Xie X, Muruato AE, Rossi SL, Roundy CM, Azar SR, et al. An Infectious cDNA Clone of Zika Virus to Study Viral Virulence, Mosquito Transmission, and Antiviral Inhibitors. Cell Host Microbe. 2016:In press. 10.1016/j.chom.2016.05.004 . PubMed DOI PMC

Tick-borne encephalitis virus capsid protein induces translational shutoff as revealed by its structural-biological analysis

Analysis of tick-borne encephalitis virus-induced host responses in human cells of neuronal origin and interferon-mediated protection