Background/Objectives Molecular epidemiology is a powerful tool to decipher the dynamics of viral transmission, quasispecies temporal evolution and origins. Little is known about the pH1N1 molecular dynamics in general population. A prospective study (CoPanFlu-RUN) was carried out in Reunion Island to characterize pH1N1 genetic variability and molecular evolution occurring in population during the pH1N1 Influenza pandemic in 2009. Methodology We directly amplified pH1N1 genomes from 28 different nasal swabs (26 individuals from 21 households). Fifteen strains were fully sequenced and 13 partially. This includes pairs of sequences from different members of 5 separate households; and two pairs from individuals, collected at different times. We assessed the molecular evolution of pH1N1 by genetic variability and phylogenetic analyses. Principal Findings We found that i) Reunion pH1N1 sequences stemmed from global “clade 7” but shaped two phylogenetic sub-clades; ii) D239E mutation was identified in the hemagglutinin protein of all Reunion sequences, a mutation which has been associated elsewhere with mild-, upper-respiratory tract pH1N1 infecting strains; iii) Date estimates from molecular phylogenies predicted clade emergence some time before the first detection of pH1N1 by the epidemiological surveillance system; iv) Phylogenetic relatedness was observed between Reunion pH1N1 viruses and those from other countries in South-western Indian Ocean area; v) Quasispecies populations were observed within households and individuals of the cohort-study. Conclusions Surveillance and/or prevention systems presently based on Influenza virus sequence variation should take into account that the majority of studies of pH1N1 Influenza generate genetic data for the HA/NA viral segments obtained from hospitalized-patients, which is potentially non-representative of the overall viral diversity within whole populations. Our observations highlight the importance of collecting unbiased data at the community level and conducting whole genome analysis to accurately understand viral dynamics.
References
[1]
Fraser C, Donnelly CA, Cauchemez S, Hanage WP, Van Kerkhove MD, et al. (2009) Pandemic potential of a strain of influenza A (H1N1): early findings. Science 324: 1557–1561.
[2]
Dawood FS, Jain S, Finelli L, Shaw MW, Lindstrom S, et al. (2009) Emergence of a novel swine-origin influenza A (H1N1) virus in humans. N Engl J Med 360: 2605–2615.
[3]
Garten RJ, Davis CT, Russell CA, Shu B, Lindstrom S, et al. (2009) Antigenic and genetic characteristics of swine-origin 2009 A(H1N1) influenza viruses circulating in humans. Science 325: 197–201.
[4]
Balcan D, Hu H, Goncalves B, Bajardi P, Poletto C, et al. (2009) Seasonal transmission potential and activity peaks of the new influenza A(H1N1): a Monte Carlo likelihood analysis based on human mobility. BMC Med 7: 45.
[5]
Nelson M, Spiro D, Wentworth D, Beck E, Fan J, et al. (2009) The early diversification of influenza A/H1N1pdm. PLoS currents 1: RRN1126.
[6]
Potdar VA, Chadha MS, Jadhav SM, Mullick J, Cherian SS, et al. (2010) Genetic characterization of the influenza A pandemic (H1N1) 2009 virus isolates from India. PLoS One 5: e9693.
[7]
Ilyicheva T, Susloparov I, Durymanov A, Romanovskaya A, Sharshov K, et al.. (2011) Influenza A/H1N1pdm virus in Russian Asia in 2009–2010. Infect Genet Evol.
[8]
Barrero PR, Viegas M, Valinotto LE, Mistchenko AS (2011) Genetic and phylogenetic analyses of influenza A H1N1pdm virus in Buenos Aires, Argentina. J Virol 85: 1058–1066.
[9]
Chan KH, To KK, Hung IF, Zhang AJ, Chan JF, et al. (2011) Differences in antibody responses of individuals with natural infection and those vaccinated against pandemic H1N1 2009 influenza. Clinical and vaccine immunology : CVI 18: 867–873.
[10]
Furuse Y, Suzuki A, Oshitani H (2010) Evolutionary analyses on the HA gene ofpandemic H1N1/09: early findings. Bioinformation 5: 7–10.
[11]
Shiino T, Okabe N, Yasui Y, Sunagawa T, Ujike M, et al. (2010) Molecular evolutionary analysis of the influenza A(H1N1)pdm, May-September, 2009: temporal and spatial spreading profile of the viruses in Japan. PLoS One 5: e11057.
[12]
Parks D, Macdonald N, Beiko R (2009) Tracking the evolution and geographic spread of Influenza A. PLoS currents. 1: RRN1014.
[13]
Pariani E, Piralla A, Frati E, Anselmi G, Campanini G, et al. (2011) Early co-circulation of different clades of influenza A/H1N1v pandemic virus in northern Italy. Journal of preventive medicine and hygiene 52: 17–20.
[14]
Graham M, Liang B, Van Domselaar G, Bastien N, Beaudoin C, et al. (2011) Nationwide molecular surveillance of pandemic H1N1 influenza A virus genomes: Canada, 2009. PLoS One 6: e16087.
[15]
Balcan D, Hu H, Goncalves B, Bajardi P, Poletto C, et al. (2009) Seasonal transmission potential and activity peaks of the new influenza A(H1N1): a Monte Carlo likelihood analysis based on human mobility. BMC medicine 7: 45.
[16]
Brownstein JS, Wolfe CJ, Mandl KD (2006) Empirical evidence for the effect of airline travel on inter-regional influenza spread in the United States. PLoS medicine 3: e401.
[17]
D’Ortenzio E, Renault P, Jaffar-Bandjee MC, Gauzere BA, Lagrange-Xelot M, et al. (2010) A review of the dynamics and severity of the pandemic A(H1N1) influenza virus on Reunion island, 2009. Clin Microbiol Infect 16: 309–316.
[18]
Dellagi K, Rollot O, Temmam S, Salez N, Guernier V, et al. (2011) Pandemic Influenza due to pH1N1/2009 virus: estimation of infection burden in Reunion Island through a prospective serosurvey, austral winter 2009. PloS one 6: e25738.
[19]
Koita OA, Sangare L, Poudiougou B, Aboubacar B, Samake Y, et al.. (2011) A seroepidemiological study of pandemic A/H1N1(2009) influenza in a rural population of Mali. Clinical microbiology and infection : the official publication of the European Society of Clinical Microbiology and Infectious Diseases.
[20]
Ninove L, Nougairede A, Gazin C, Thirion L, Delogu I, et al. (2011) RNA and DNA bacteriophages as molecular diagnosis controls in clinical virology: a comprehensive study of more than 45,000 routine PCR tests. PLoS One 6: e16142.
[21]
Ninove L, Gazin C, Gould EA, Nougairede A, Flahault A, et al. (2009) A simple method for molecular detection of Swine-origin and human-origin influenza a virus. Vector Borne Zoonotic Dis 10: 237–240.
[22]
Zhirnov OP, Vorobjeva IV, Saphonova OA, Poyarkov SV, Ovcharenko AV, et al. (2009) Structural and evolutionary characteristics of HA, NA, NS and M genes of clinical influenza A/H3N2 viruses passaged in human and canine cells. Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology 45: 322–333.
[23]
Drummond AJ, Ashton B, Buxton S, Cheung M, Cooper A, et al.. (2010) Geneious v5.3.
[24]
Edgar RC (2004) MUSCLE: a multiple sequence alignment method with reduced time and space complexity. BMC bioinformatics 5: 113.
[25]
Posada D (2008) jModelTest: phylogenetic model averaging. Molecular biology and evolution 25: 1253–1256.
[26]
Guindon S, Gascuel O (2003) A simple, fast, and accurate algorithm to estimate large phylogenies by maximum likelihood. Systematic biology 52: 696–704.
Tamura K, Peterson D, Peterson N, Stecher G, Nei M, et al. (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Molecular biology and evolution 28: 2731–2739.
[29]
Tajima F, Nei M (1984) Estimation of evolutionary distance between nucleotide sequences. Molecular biology and evolution 1: 269–285.
[30]
Drummond AJ, Rambaut A (2007) BEAST: Bayesian evolutionary analysis by sampling trees. BMC Evol Biol 7: 214.
[31]
Goni N, Fajardo A, Moratorio G, Colina R, Cristina J (2009) Modeling gene sequences over time in 2009 H1N1 influenza A virus populations. Virology journal 6: 215.
[32]
Nelson MI, Holmes EC (2007) The evolution of epidemic influenza. Nat Rev Genet 8: 196–205.
[33]
Nelson M, Spiro D, Wentworth D, Beck E, Fan J, et al. (2009) The early diversification of influenza A/H1N1pdm. PLoS Curr 1: RRN1126.
[34]
Furuse Y, Suzuki A, Oshitani H (2010) Reassortment between swine influenza A viruses increased their adaptation to humans in pandemic H1N1/09. Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases 10: 569–574.
[35]
Ferraris O, Lina B (2008) Mutations of neuraminidase implicated in neuraminidase inhibitors resistance. Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology 41: 13–19.
[36]
Giria MT, Rebelo de Andrade H, Santos LA, Correia VM, Pedro SV, et al.. (2011) Genomic signatures and antiviral drug susceptibility profile of A(H1N1)pdm09. Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology.
[37]
Glinsky GV (2010) Genomic analysis of pandemic (H1N1) 2009 reveals association of increasing disease severity with emergence of novel hemagglutinin mutations. Cell cycle 9: 958–970.
[38]
Reid AH, Fanning TG, Hultin JV, Taubenberger JK (1999) Origin and evolution of the 1918 “Spanish” influenza virus hemagglutinin gene. Proc Natl Acad Sci U S A 96: 1651–1656.
[39]
Tse H, Kao RY, Wu WL, Lim WW, Chen H, et al. (2011) Structural basis and sequence co-evolution analysis of the hemagglutinin protein of pandemic influenza A/H1N1 (2009) virus. Experimental biology and medicine 236: 915–925.
[40]
Chen H, Wen X, To KK, Wang P, Tse H, et al. (2010) Quasispecies of the D225G substitution in the hemagglutinin of pandemic influenza A(H1N1) 2009 virus from patients with severe disease in Hong Kong, China. J Infect Dis 201: 1517–1521.
[41]
Tse H, To KK, Wen X, Chen H, Chan KH, et al. (2011) Clinical and virological factors associated with viremia in pandemic influenza A/H1N1/2009 virus infection. PLoS One 6: e22534.
[42]
Zhang AJ, To KK, Tse H, Chan KH, Guo KY, et al. (2011) High incidence of severe influenza among individuals over 50 years of age. Clinical and vaccine immunology : CVI 18: 1918–1924.
[43]
Filleul L, D’Ortenzio E, Kermarec F, Le Bot F, Renault P (2010) Pandemic influenza on Reunion Island and school closure. Lancet Infect Dis 10: 294–295.
[44]
Kelly HA, Mercer GN, Fielding JE, Dowse GK, Glass K, et al. (2010) Pandemic (H1N1) 2009 influenza community transmission was established in one Australian state when the virus was first identified in North America. PloS one 5: e11341.
[45]
Chao DY, Cheng KF, Li TC, Wu TN, Chen CY, et al. (2011) Serological evidence of subclinical transmission of the 2009 pandemic H1N1 influenza virus outside of Mexico. PloS one 6: e14555.
[46]
Gubareva LV, Novikov DV, Hayden FG (2002) Assessment of hemagglutinin sequence heterogeneity during influenza virus transmission in families. J Infect Dis 186: 1575–1581.
[47]
Poon LL, Chan KH, Chu DK, Fung CC, Cheng CK, et al. (2011) Viral genetic sequence variations in pandemic H1N1/2009 and seasonal H3N2 influenza viruses within an individual, a household and a community. J Clin Virol 52: 146–150.
[48]
Boelle PY, Ansart S, Cori A, Valleron AJ (2011) Transmission parameters of the A/H1N1 (2009) influenza virus pandemic: a review. Influenza and other respiratory viruses 5: 306–316.
[49]
Nelson MI, Holmes EC (2007) The evolution of epidemic influenza. Nature reviews Genetics 8: 196–205.
[50]
Ghedin E, Laplante J, DePasse J, Wentworth DE, Santos RP, et al. (2011) Deep sequencing reveals mixed infection with 2009 pandemic influenza A (H1N1) virus strains and the emergence of oseltamivir resistance. J Infect Dis 203: 168–174.
[51]
Greninger AL, Chen EC, Sittler T, Scheinerman A, Roubinian N, et al. (2010) A metagenomic analysis of pandemic influenza A (2009 H1N1) infection in patients from North America. PLoS One 5: e13381.
[52]
Lauring AS, Andino R (2010) Quasispecies theory and the behavior of RNA viruses. PLoS pathogens 6: e1001005.
