Background A chikungunya virus outbreak of unprecedented magnitude is currently ongoing in Indian Ocean territories. In Réunion Island, this alphavirus has already infected about one-third of the human population. The main clinical symptom of the disease is a painful and invalidating poly-arthralgia. Besides the arthralgic form, 123 patients with a confirmed chikungunya infection have developed severe clinical signs, i.e., neurological signs or fulminant hepatitis. Methods and Findings We report the nearly complete genome sequence of six selected viral isolates (isolated from five sera and one cerebrospinal fluid), along with partial sequences of glycoprotein E1 from a total of 127 patients from Réunion, Seychelles, Mauritius, Madagascar, and Mayotte islands. Our results indicate that the outbreak was initiated by a strain related to East-African isolates, from which viral variants have evolved following a traceable microevolution history. Unique molecular features of the outbreak isolates were identified. Notably, in the region coding for the non-structural proteins, ten amino acid changes were found, four of which were located in alphavirus-conserved positions of nsP2 (which contains helicase, protease, and RNA triphosphatase activities) and of the polymerase nsP4. The sole isolate obtained from the cerebrospinal fluid showed unique changes in nsP1 (T301I), nsP2 (Y642N), and nsP3 (E460 deletion), not obtained from isolates from sera. In the structural proteins region, two noteworthy changes (A226V and D284E) were observed in the membrane fusion glycoprotein E1. Homology 3D modelling allowed mapping of these two changes to regions that are important for membrane fusion and virion assembly. Change E1-A226V was absent in the initial strains but was observed in >90% of subsequent viral sequences from Réunion, denoting evolutionary success possibly due to adaptation to the mosquito vector. Conclusions The unique molecular features of the analyzed Indian Ocean isolates of chikungunya virus demonstrate their high evolutionary potential and suggest possible clues for understanding the atypical magnitude and virulence of this outbreak.
References
[1]
Strauss EG, Strauss JH (1986) Structure and replication of the alphavirus genome. In: Schlesinger S, Schlesinger MJ, editors. The Togaviridae and Flaviviridae. New York: Plenum Press. pp. 35–90.
[2]
Porterfield JH (1980) Antigenic characteristics and classification of the Togaviridae. In: Schlesinger R, editor. The Togaviruses. New York: Academic Press. pp. 13–46.
[3]
Ross RW (1956) The Newala epidemic. III. The virus: Isolation, pathogenic properties and relationship to the epidemic. J Hyg 54: 177–191.
[4]
Jupp PG, McIntosh BM (1988) Chikungunya disease. In: Monath TP, editor. The Arboviruses: Epidemiology and ecology. Boca Raton (Florida): CRC Press. pp. 137–157.
[5]
Johnston RE, Peters CJ (1996) Alphaviruses associated primarily with fever and polyarthritis. In: Fields BN, Knipe DM, Howley PM, editors. Fields virology. Philadelphia: Lippincott-Raven Publishers. pp. 843–898.
[6]
Pastorino B, Muyembe-Tamfum JJ, Bessaud M, Tock F, Tolou H, et al. (2004) Epidemic resurgence of Chikungunya virus in democratic Republic of the Congo: Identification of a new central African strain. J Med Virol 74: 277–282.
[7]
Laras K, Sukri NC, Larasati RP, Bangs MJ, Kosim R, et al. (2005) Tracking the re-emergence of epidemic chikungunya virus in Indonesia. Trans R Soc Trop Med Hyg 99: 128–141.
[8]
Paquet C, Quatresous I, Solet JL, Sissoko D, Renault P, et al. (2006) Chikungunya outbreak in Reunion: Epidemiology and surveillance, 2005 to early January 2006. Euro Surveill 11: 2.
[9]
Khan AH, Morita K, Parquet Md Mdel C, Hasebe F, Mathenge EG, et al. (2002) Complete nucleotide sequence of chikungunya virus and evidence for an internal polyadenylation site. J Gen Virol 83: 3075–3084.
[10]
Powers AM, Brault AC, Tesh RB, Weaver SC (2000) Re-emergence of Chikungunya and O'nyong-nyong viruses: Evidence for distinct geographical lineages and distant evolutionary relationships. J Gen Virol 81: 471–479.
[11]
Navarro-Sanchez E, Altmeyer MR, Amara A, Schwartz O, Fieschi F, et al. (2003) DC-SIGN is essential for the productive infection of human dendritic cells by mosquito-cell-derived dengue viruses. EMBO Rep 7: 1–6.
[12]
Gordon D Abajian C, Green P (1998) Consed: A graphical tool for sequence finishing. Genome Res 8: 195–202.
[13]
Rozas J, Sanchez-DelBarrio JC, Messeguer X, Rozas R (2003) DnaSP, DNA polymorphism analyses by the coalescent and other methods. Bioinformatics 19: 2496–2497.
[14]
Xia X, Xie Z (2001) DAMBE: Software package for data analysis in molecular biology and evolution. J Hered 92: 371–373.
[15]
Thompson JD, Higgins DG, Gibson TJ (1994) CLUSTAL W: Improving the sensitivity of progressive multiple sequence alignment through sequence weighting, positions-specific gap penalties and weight matrix choice. Nucleic Acids Res 22: 4673–4680.
Hofacker IL (2003) Vienna RNA secondary structure server. Nucleic Acids Res 31: 3429–3431.
[18]
Kumar S, Tamura K, Nei M (2004) MEGA3: Integrated software for molecular evolutionary genetics analysis and sequence alignment. Brief Bioinform 5: 150–163.
[19]
Martin DP, Williamson C, Posada D (2005) RDP2: Recombination detection and analysis from sequence alignments. Bioinformatics 21: 260–262.
[20]
Roussel A, Lescar J, Vaney MC, Wengler G, Wengler G, et al. (2006) Structure and interactions at the viral surface of the envelope protein E1 of semliki forest virus. Structure 14: 75–86.
[21]
Carson M (1987) Ribbon models of macromolecules. J Mol Graph 5: 103–106.
[22]
Strauss JH, Strauss EG (1994) The alphaviruses: Gene expression, replication, and evolution. Microbiol Rev 58: 491–562.
[23]
Lavergne A, Thoisy BD, Lacoste V, Pascalis H, Pouliquen JF, et al. (2005) Mayaro virus: Complete nucleotide sequence and phylogenetic relationships with other alphaviruses. Virus Res. In press.
[24]
Lanciotti RS, Ludwig ML, Rwaguma EB, Lutwama JJ, Kram TM, et al. (1998) Emergence of epidemic O'nyong-nyong fever in Uganda after a 35-year absence: Genetic characterization of the virus. Virology 252: 258–268.
[25]
Strauss EG, Levinson R, Rice CM, Dalrymple J, Strauss JH (1988) Nonstructural proteins nsP3 and nsP4 of Ross River and O'Nyong-nyong viruses: Sequence and comparison with those of other alphaviruses. Virology 164: 265–274.
[26]
Griffin DE (2001) Alphaviruses. In: Knipe DM, Howley PM, editors. Fields virology. Philadelphia: Lippincott Williams & Wilkins. pp. 917–962.
[27]
Vashishtha M, Phalen T, Marquardt MT, Ryu JS, Ng AC, et al. (1998) A single point mutation controls the cholesterol dependence of Semliki Forest virus entry and exit. J Cell Biol 140: 91–99.
[28]
Ahn A, Schoepp RJ, Sternberg D, Kielian M (1999) Growth and stability of a cholesterol-independent Semliki Forest virus mutant in mosquitoes. Virology 262: 452–456.
[29]
Williams MC, Woodall JP, Corbet PS, Gillett JD (1965) O'nyong-Nyong Fever: An epidemic virus disease in East Africa. 8. Virus isolations from Anopheles mosquitoes. Trans R Soc Trop Med Hyg 59: 300–306.
[30]
Weaver SC, Barrett AD (2004) Transmission cycles, host range, evolution and emergence of arboviral disease. Nat Rev Microbiol 2: 789–801.
[31]
Lu YE, Cassese T, Kielian M (1999) The cholesterol requirement for sindbis virus entry and exit and characterization of a spike protein region involved in cholesterol dependence. J Virol 73: 4272–4278.
[32]
Holland J, Spindler K, Horodyski F, Grabau E, Nichol S, et al. (1982) Rapid evolution of RNA genomes. Science 215: 1577–1585.
[33]
Domingo E, Holland JJ (1997) RNA virus mutations and fitness for survival. Annu Rev Microbiol 51: 151–178.
[34]
Vignuzzi M, Stone JK, Arnold JJ, Cameron CE, Andino R (2006) Quasispecies diversity determines pathogenesis through cooperative interactions in a viral population. Nature 439: 344–348.
[35]
Kim KH, Rumenapf T, Strauss EG, Strauss JH (2004) Regulation of Semliki Forest virus RNA replication: A model for the control of alphavirus pathogenesis in invertebrate hosts. Virology 323: 153–163.
Heise MT, White LJ, Simpson DA, Leonard C, Bernard KA, et al. (2003) An attenuating mutation in nsP1 of the Sindbis-group virus S.A.AR86 accelerates nonstructural protein processing and up-regulates viral 26S RNA synthesis. J Virol 77: 1149–1156.
[38]
Suthar MS, Shabman R, Madric K, Lambeth C, Heise MT (2005) Identification of adult mouse neurovirulence determinants of the Sindbis virus strain AR86. J Virol 79: 4219–4228.
[39]
Condon RJ, Rouse IL (1995) Acute symptoms and sequelae of Ross River virus infection in South-Western Australia: A follow-up study. Clin Diagn Virol 3: 273–284.
[40]
Selden SM, Cameron AS (1996) Changing epidemiology of Ross River virus disease in South Australia. Med J Aust 165: 313–317.
[41]
Mazaud R, Salaün JJ, Montabone H, Goube P, Bazillio R (1971) Troubles neurologiques et sensoriels aigus dans la dengue et la fièvre à Chikungunya. Bull Soc Pathol Exot 64: 22–30.
[42]
Nimmannitya S, Halstead SB, Cohen SN, Margiotta MR (1969) Dengue and chikungunya virus infection in man in Thailand, 1962–1964. I. Observations on hospitalized patients with hemorrhagic fever. Am J Trop Med Hyg 18: 954–971.
[43]
Gratz NG (2004) Critical review of the vector status of . Med Vet Entomol 18: 215–227.
[44]
Lescar J, Roussel A, Wien MW, Navaza J, Fuller SD, et al. (2001) The Fusion glycoprotein shell of Semliki Forest virus: An icosahedral assembly primed for fusogenic activation at endosomal pH. Cell 105: 137–148.
