Identification of the Mechanisms Causing Reversion to Virulence in an Attenuated SARS-CoV for the Design of a Genetically Stable Vaccine
Identifieur interne : 001391 ( Pmc/Curation ); précédent : 001390; suivant : 001392Identification of the Mechanisms Causing Reversion to Virulence in an Attenuated SARS-CoV for the Design of a Genetically Stable Vaccine
Auteurs : Jose M. Jimenez-Guarde O [Espagne] ; Jose A. Regla-Nava [Espagne] ; Jose L. Nieto-Torres [Espagne] ; Marta L. Dediego [Espagne] ; Carlos Casta O-Rodriguez [Espagne] ; Raul Fernandez-Delgado [Espagne] ; Stanley Perlman [États-Unis] ; Luis Enjuanes [Espagne]Source :
- PLoS Pathogens [ 1553-7366 ] ; 2015.
Abstract
A SARS-CoV lacking the full-length E gene (SARS-CoV-∆E) was attenuated and an effective vaccine. Here, we show that this mutant virus regained fitness after serial passages in cell culture or
Url:
DOI: 10.1371/journal.ppat.1005215
PubMed: 26513244
PubMed Central: 4626112
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<series><title level="j">PLoS Pathogens</title>
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<front><div type="abstract" xml:lang="en"><p>A SARS-CoV lacking the full-length E gene (SARS-CoV-∆E) was attenuated and an effective vaccine. Here, we show that this mutant virus regained fitness after serial passages in cell culture or <italic>in vivo</italic>
, resulting in the partial duplication of the membrane gene or in the insertion of a new sequence in gene 8a, respectively. The chimeric proteins generated in cell culture increased virus fitness <italic>in vitro</italic>
but remained attenuated in mice. In contrast, during SARS-CoV-∆E passage in mice, the virus incorporated a mutated variant of 8a protein, resulting in reversion to a virulent phenotype. When the full-length E protein was deleted or its PDZ-binding motif (PBM) was mutated, the revertant viruses either incorporated a novel chimeric protein with a PBM or restored the sequence of the PBM on the E protein, respectively. Similarly, after passage in mice, SARS-CoV-∆E protein 8a mutated, to now encode a PBM, and also regained virulence. These data indicated that the virus requires a PBM on a transmembrane protein to compensate for removal of this motif from the E protein. To increase the genetic stability of the vaccine candidate, we introduced small attenuating deletions in E gene that did not affect the endogenous PBM, preventing the incorporation of novel chimeric proteins in the virus genome. In addition, to increase vaccine biosafety, we introduced additional attenuating mutations into the nsp1 protein. Deletions in the carboxy-terminal region of nsp1 protein led to higher host interferon responses and virus attenuation. Recombinant viruses including attenuating mutations in E and nsp1 genes maintained their attenuation after passage <italic>in vitro</italic>
and <italic>in vivo</italic>
. Further, these viruses fully protected mice against challenge with the lethal parental virus, and are therefore safe and stable vaccine candidates for protection against SARS-CoV.</p>
</div>
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<back><div1 type="bibliography"><listBibl><biblStruct><analytic><author><name sortKey="Perlman, S" uniqKey="Perlman S">S Perlman</name>
</author>
<author><name sortKey="Netland, J" uniqKey="Netland J">J Netland</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Rota, Pa" uniqKey="Rota P">PA Rota</name>
</author>
<author><name sortKey="Oberste, Ms" uniqKey="Oberste M">MS Oberste</name>
</author>
<author><name sortKey="Monroe, Ss" uniqKey="Monroe S">SS Monroe</name>
</author>
<author><name sortKey="Nix, Wa" uniqKey="Nix W">WA Nix</name>
</author>
<author><name sortKey="Campganoli, R" uniqKey="Campganoli R">R Campganoli</name>
</author>
<author><name sortKey="Icenogle, Jp" uniqKey="Icenogle J">JP Icenogle</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Drosten, C" uniqKey="Drosten C">C Drosten</name>
</author>
<author><name sortKey="Gunther, S" uniqKey="Gunther S">S Gunther</name>
</author>
<author><name sortKey="Preiser, W" uniqKey="Preiser W">W Preiser</name>
</author>
<author><name sortKey="Van Der Werf, S" uniqKey="Van Der Werf S">S van der Werf</name>
</author>
<author><name sortKey="Brodt, Hr" uniqKey="Brodt H">HR Brodt</name>
</author>
<author><name sortKey="Becker, S" uniqKey="Becker S">S Becker</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Lau, Sk" uniqKey="Lau S">SK Lau</name>
</author>
<author><name sortKey="Woo, Pc" uniqKey="Woo P">PC Woo</name>
</author>
<author><name sortKey="Li, Ks" uniqKey="Li K">KS Li</name>
</author>
<author><name sortKey="Huang, Y" uniqKey="Huang Y">Y Huang</name>
</author>
<author><name sortKey="Tsoi, Hw" uniqKey="Tsoi H">HW Tsoi</name>
</author>
<author><name sortKey="Wong, Bh" uniqKey="Wong B">BH Wong</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Li, W" uniqKey="Li W">W Li</name>
</author>
<author><name sortKey="Shi, Z" uniqKey="Shi Z">Z Shi</name>
</author>
<author><name sortKey="Yu, M" uniqKey="Yu M">M Yu</name>
</author>
<author><name sortKey="Ren, W" uniqKey="Ren W">W Ren</name>
</author>
<author><name sortKey="Smith, C" uniqKey="Smith C">C Smith</name>
</author>
<author><name sortKey="Epstein, Jh" uniqKey="Epstein J">JH Epstein</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Woo, Pc" uniqKey="Woo P">PC Woo</name>
</author>
<author><name sortKey="Lau, Sk" uniqKey="Lau S">SK Lau</name>
</author>
<author><name sortKey="Li, Ks" uniqKey="Li K">KS Li</name>
</author>
<author><name sortKey="Poon, Rw" uniqKey="Poon R">RW Poon</name>
</author>
<author><name sortKey="Wong, Bh" uniqKey="Wong B">BH Wong</name>
</author>
<author><name sortKey="Tsoi, Hw" uniqKey="Tsoi H">HW Tsoi</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Dominguez, Sr" uniqKey="Dominguez S">SR Dominguez</name>
</author>
<author><name sortKey="O Shea, Tj" uniqKey="O Shea T">TJ O'Shea</name>
</author>
<author><name sortKey="Oko, Lm" uniqKey="Oko L">LM Oko</name>
</author>
<author><name sortKey="Holmes, Kv" uniqKey="Holmes K">KV Holmes</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Zaki, Am" uniqKey="Zaki A">AM Zaki</name>
</author>
<author><name sortKey="Van Boheemen, S" uniqKey="Van Boheemen S">S van Boheemen</name>
</author>
<author><name sortKey="Bestebroer, Tm" uniqKey="Bestebroer T">TM Bestebroer</name>
</author>
<author><name sortKey="Osterhaus, Ad" uniqKey="Osterhaus A">AD Osterhaus</name>
</author>
<author><name sortKey="Fouchier, Ra" uniqKey="Fouchier R">RA Fouchier</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Bermingham, A" uniqKey="Bermingham A">A Bermingham</name>
</author>
<author><name sortKey="Chand, M" uniqKey="Chand M">M Chand</name>
</author>
<author><name sortKey="Brown, C" uniqKey="Brown C">C Brown</name>
</author>
<author><name sortKey="Aarons, E" uniqKey="Aarons E">E Aarons</name>
</author>
<author><name sortKey="Tong, C" uniqKey="Tong C">C Tong</name>
</author>
<author><name sortKey="Langrish, C" uniqKey="Langrish C">C Langrish</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Huang, C" uniqKey="Huang C">C Huang</name>
</author>
<author><name sortKey="Ito, N" uniqKey="Ito N">N Ito</name>
</author>
<author><name sortKey="Tseng, Ct" uniqKey="Tseng C">CT Tseng</name>
</author>
<author><name sortKey="Makino, S" uniqKey="Makino S">S Makino</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Huang, C" uniqKey="Huang C">C Huang</name>
</author>
<author><name sortKey="Peters, Cj" uniqKey="Peters C">CJ Peters</name>
</author>
<author><name sortKey="Makino, S" uniqKey="Makino S">S Makino</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Shen, S" uniqKey="Shen S">S Shen</name>
</author>
<author><name sortKey="Lin, Ps" uniqKey="Lin P">PS Lin</name>
</author>
<author><name sortKey="Chao, Yc" uniqKey="Chao Y">YC Chao</name>
</author>
<author><name sortKey="Zhang, A" uniqKey="Zhang A">A Zhang</name>
</author>
<author><name sortKey="Yang, X" uniqKey="Yang X">X Yang</name>
</author>
<author><name sortKey="Lim, Sg" uniqKey="Lim S">SG Lim</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Qiu, M" uniqKey="Qiu M">M Qiu</name>
</author>
<author><name sortKey="Shi, Y" uniqKey="Shi Y">Y Shi</name>
</author>
<author><name sortKey="Guo, Z" uniqKey="Guo Z">Z Guo</name>
</author>
<author><name sortKey="Chen, Z" uniqKey="Chen Z">Z Chen</name>
</author>
<author><name sortKey="He, R" uniqKey="He R">R He</name>
</author>
<author><name sortKey="Chen, R" uniqKey="Chen R">R Chen</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Saif, Lj" uniqKey="Saif L">LJ Saif</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Buchholz, Uj" uniqKey="Buchholz U">UJ Buchholz</name>
</author>
<author><name sortKey="Bukreyev, A" uniqKey="Bukreyev A">A Bukreyev</name>
</author>
<author><name sortKey="Yang, L" uniqKey="Yang L">L Yang</name>
</author>
<author><name sortKey="Lamirande, Ew" uniqKey="Lamirande E">EW Lamirande</name>
</author>
<author><name sortKey="Murphy, Br" uniqKey="Murphy B">BR Murphy</name>
</author>
<author><name sortKey="Subbarao, K" uniqKey="Subbarao K">K Subbarao</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Enjuanes, L" uniqKey="Enjuanes L">L Enjuanes</name>
</author>
<author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML DeDiego</name>
</author>
<author><name sortKey="Alvarez, E" uniqKey="Alvarez E">E Alvarez</name>
</author>
<author><name sortKey="Deming, D" uniqKey="Deming D">D Deming</name>
</author>
<author><name sortKey="Sheahan, T" uniqKey="Sheahan T">T Sheahan</name>
</author>
<author><name sortKey="Baric, R" uniqKey="Baric R">R Baric</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Huang, J" uniqKey="Huang J">J Huang</name>
</author>
<author><name sortKey="Cao, Y" uniqKey="Cao Y">Y Cao</name>
</author>
<author><name sortKey="Du, J" uniqKey="Du J">J Du</name>
</author>
<author><name sortKey="Bu, X" uniqKey="Bu X">X Bu</name>
</author>
<author><name sortKey="Ma, R" uniqKey="Ma R">R Ma</name>
</author>
<author><name sortKey="Wu, C" uniqKey="Wu C">C Wu</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Zhao, J" uniqKey="Zhao J">J Zhao</name>
</author>
<author><name sortKey="Perlman, S" uniqKey="Perlman S">S Perlman</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Channappanavar, R" uniqKey="Channappanavar R">R Channappanavar</name>
</author>
<author><name sortKey="Zhao, J" uniqKey="Zhao J">J Zhao</name>
</author>
<author><name sortKey="Perlman, S" uniqKey="Perlman S">S Perlman</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Torres, J" uniqKey="Torres J">J Torres</name>
</author>
<author><name sortKey="Parthasarathy, K" uniqKey="Parthasarathy K">K Parthasarathy</name>
</author>
<author><name sortKey="Lin, X" uniqKey="Lin X">X Lin</name>
</author>
<author><name sortKey="Saravanan, R" uniqKey="Saravanan R">R Saravanan</name>
</author>
<author><name sortKey="Liu, Dx" uniqKey="Liu D">DX Liu</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Verdia Baguena, C" uniqKey="Verdia Baguena C">C Verdia-Baguena</name>
</author>
<author><name sortKey="Nieto Torres, Jl" uniqKey="Nieto Torres J">JL Nieto-Torres</name>
</author>
<author><name sortKey="Alcaraz, A" uniqKey="Alcaraz A">A Alcaraz</name>
</author>
<author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML Dediego</name>
</author>
<author><name sortKey="Torres, J" uniqKey="Torres J">J Torres</name>
</author>
<author><name sortKey="Aguilella, Vm" uniqKey="Aguilella V">VM Aguilella</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Verdia Baguena, C" uniqKey="Verdia Baguena C">C Verdia-Baguena</name>
</author>
<author><name sortKey="Nieto Torres, Jl" uniqKey="Nieto Torres J">JL Nieto-Torres</name>
</author>
<author><name sortKey="Alcaraz, A" uniqKey="Alcaraz A">A Alcaraz</name>
</author>
<author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML Dediego</name>
</author>
<author><name sortKey="Enjuanes, L" uniqKey="Enjuanes L">L Enjuanes</name>
</author>
<author><name sortKey="Aguilella, Vm" uniqKey="Aguilella V">VM Aguilella</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Nieto Torres, Jl" uniqKey="Nieto Torres J">JL Nieto-Torres</name>
</author>
<author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML Dediego</name>
</author>
<author><name sortKey="Verdia Baguena, C" uniqKey="Verdia Baguena C">C Verdia-Baguena</name>
</author>
<author><name sortKey="Jimenez Guarde O, Jm" uniqKey="Jimenez Guarde O J">JM Jimenez-Guardeño</name>
</author>
<author><name sortKey="Regla Nava, Ja" uniqKey="Regla Nava J">JA Regla-Nava</name>
</author>
<author><name sortKey="Fernandez Delgado, R" uniqKey="Fernandez Delgado R">R Fernandez-Delgado</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Jimenez Guarde O, Jm" uniqKey="Jimenez Guarde O J">JM Jimenez-Guardeño</name>
</author>
<author><name sortKey="Nieto Torres, Jl" uniqKey="Nieto Torres J">JL Nieto-Torres</name>
</author>
<author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML DeDiego</name>
</author>
<author><name sortKey="Regla Nava, Ja" uniqKey="Regla Nava J">JA Regla-Nava</name>
</author>
<author><name sortKey="Fernandez Delgado, R" uniqKey="Fernandez Delgado R">R Fernandez-Delgado</name>
</author>
<author><name sortKey="Casta O Rodriguez, C" uniqKey="Casta O Rodriguez C">C Castaño-Rodriguez</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Teoh, Kt" uniqKey="Teoh K">KT Teoh</name>
</author>
<author><name sortKey="Siu, Yl" uniqKey="Siu Y">YL Siu</name>
</author>
<author><name sortKey="Chan, Wl" uniqKey="Chan W">WL Chan</name>
</author>
<author><name sortKey="Schluter, Ma" uniqKey="Schluter M">MA Schluter</name>
</author>
<author><name sortKey="Liu, Cj" uniqKey="Liu C">CJ Liu</name>
</author>
<author><name sortKey="Peiris, Js" uniqKey="Peiris J">JS Peiris</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Hung, Ay" uniqKey="Hung A">AY Hung</name>
</author>
<author><name sortKey="Sheng, M" uniqKey="Sheng M">M Sheng</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Munz, M" uniqKey="Munz M">M Munz</name>
</author>
<author><name sortKey="Hein, J" uniqKey="Hein J">J Hein</name>
</author>
<author><name sortKey="Biggin, Pc" uniqKey="Biggin P">PC Biggin</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Gerek, Zn" uniqKey="Gerek Z">ZN Gerek</name>
</author>
<author><name sortKey="Keskin, O" uniqKey="Keskin O">O Keskin</name>
</author>
<author><name sortKey="Ozkan, Sb" uniqKey="Ozkan S">SB Ozkan</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Javier, Rt" uniqKey="Javier R">RT Javier</name>
</author>
<author><name sortKey="Rice, Ap" uniqKey="Rice A">AP Rice</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Lamirande, Ew" uniqKey="Lamirande E">EW Lamirande</name>
</author>
<author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML DeDiego</name>
</author>
<author><name sortKey="Roberts, A" uniqKey="Roberts A">A Roberts</name>
</author>
<author><name sortKey="Jackson, Jp" uniqKey="Jackson J">JP Jackson</name>
</author>
<author><name sortKey="Alvarez, E" uniqKey="Alvarez E">E Alvarez</name>
</author>
<author><name sortKey="Sheahan, T" uniqKey="Sheahan T">T Sheahan</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Netland, J" uniqKey="Netland J">J Netland</name>
</author>
<author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML DeDiego</name>
</author>
<author><name sortKey="Zhao, J" uniqKey="Zhao J">J Zhao</name>
</author>
<author><name sortKey="Fett, C" uniqKey="Fett C">C Fett</name>
</author>
<author><name sortKey="Alvarez, E" uniqKey="Alvarez E">E Alvarez</name>
</author>
<author><name sortKey="Nieto Torres, Jl" uniqKey="Nieto Torres J">JL Nieto-Torres</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Fett, C" uniqKey="Fett C">C Fett</name>
</author>
<author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML DeDiego</name>
</author>
<author><name sortKey="Regla Nava, Ja" uniqKey="Regla Nava J">JA Regla-Nava</name>
</author>
<author><name sortKey="Enjuanes, L" uniqKey="Enjuanes L">L Enjuanes</name>
</author>
<author><name sortKey="Perlman, S" uniqKey="Perlman S">S Perlman</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML DeDiego</name>
</author>
<author><name sortKey="Alvarez, E" uniqKey="Alvarez E">E Alvarez</name>
</author>
<author><name sortKey="Almazan, F" uniqKey="Almazan F">F Almazan</name>
</author>
<author><name sortKey="Rejas, Mt" uniqKey="Rejas M">MT Rejas</name>
</author>
<author><name sortKey="Lamirande, E" uniqKey="Lamirande E">E Lamirande</name>
</author>
<author><name sortKey="Roberts, A" uniqKey="Roberts A">A Roberts</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML DeDiego</name>
</author>
<author><name sortKey="Pewe, L" uniqKey="Pewe L">L Pewe</name>
</author>
<author><name sortKey="Alvarez, E" uniqKey="Alvarez E">E Alvarez</name>
</author>
<author><name sortKey="Rejas, Mt" uniqKey="Rejas M">MT Rejas</name>
</author>
<author><name sortKey="Perlman, S" uniqKey="Perlman S">S Perlman</name>
</author>
<author><name sortKey="Enjuanes, L" uniqKey="Enjuanes L">L Enjuanes</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Graham, Rl" uniqKey="Graham R">RL Graham</name>
</author>
<author><name sortKey="Donaldson, Ef" uniqKey="Donaldson E">EF Donaldson</name>
</author>
<author><name sortKey="Baric, Rs" uniqKey="Baric R">RS Baric</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Lambert, Ph" uniqKey="Lambert P">PH Lambert</name>
</author>
<author><name sortKey="Liu, M" uniqKey="Liu M">M Liu</name>
</author>
<author><name sortKey="Siegrist, Ca" uniqKey="Siegrist C">CA Siegrist</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Liao, Cl" uniqKey="Liao C">CL Liao</name>
</author>
<author><name sortKey="Lai, Mmc" uniqKey="Lai M">MMC Lai</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Regla Nava, Ja" uniqKey="Regla Nava J">JA Regla-Nava</name>
</author>
<author><name sortKey="Jimenez Guarde O, Jm" uniqKey="Jimenez Guarde O J">JM Jimenez-Guardeño</name>
</author>
<author><name sortKey="Nieto Torres, Jl" uniqKey="Nieto Torres J">JL Nieto-Torres</name>
</author>
<author><name sortKey="Gallagher, Tm" uniqKey="Gallagher T">TM Gallagher</name>
</author>
<author><name sortKey="Enjuanes, L" uniqKey="Enjuanes L">L Enjuanes</name>
</author>
<author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML DeDiego</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Kamitani, W" uniqKey="Kamitani W">W Kamitani</name>
</author>
<author><name sortKey="Narayanan, K" uniqKey="Narayanan K">K Narayanan</name>
</author>
<author><name sortKey="Huang, C" uniqKey="Huang C">C Huang</name>
</author>
<author><name sortKey="Lokugamage, K" uniqKey="Lokugamage K">K Lokugamage</name>
</author>
<author><name sortKey="Ikegami, T" uniqKey="Ikegami T">T Ikegami</name>
</author>
<author><name sortKey="Ito, N" uniqKey="Ito N">N Ito</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Kamitani, W" uniqKey="Kamitani W">W Kamitani</name>
</author>
<author><name sortKey="Huang, C" uniqKey="Huang C">C Huang</name>
</author>
<author><name sortKey="Narayanan, K" uniqKey="Narayanan K">K Narayanan</name>
</author>
<author><name sortKey="Lokugamage, Kg" uniqKey="Lokugamage K">KG Lokugamage</name>
</author>
<author><name sortKey="Makino, S" uniqKey="Makino S">S Makino</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Huang, C" uniqKey="Huang C">C Huang</name>
</author>
<author><name sortKey="Lokugamage, Kg" uniqKey="Lokugamage K">KG Lokugamage</name>
</author>
<author><name sortKey="Rozovics, Jm" uniqKey="Rozovics J">JM Rozovics</name>
</author>
<author><name sortKey="Narayanan, K" uniqKey="Narayanan K">K Narayanan</name>
</author>
<author><name sortKey="Semler, Bl" uniqKey="Semler B">BL Semler</name>
</author>
<author><name sortKey="Makino, S" uniqKey="Makino S">S Makino</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Tanaka, T" uniqKey="Tanaka T">T Tanaka</name>
</author>
<author><name sortKey="Kamitani, W" uniqKey="Kamitani W">W Kamitani</name>
</author>
<author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML DeDiego</name>
</author>
<author><name sortKey="Enjuanes, L" uniqKey="Enjuanes L">L Enjuanes</name>
</author>
<author><name sortKey="Matsuura, Y" uniqKey="Matsuura Y">Y Matsuura</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Tohya, Y" uniqKey="Tohya Y">Y Tohya</name>
</author>
<author><name sortKey="Narayanan, K" uniqKey="Narayanan K">K Narayanan</name>
</author>
<author><name sortKey="Kamitani, W" uniqKey="Kamitani W">W Kamitani</name>
</author>
<author><name sortKey="Huang, C" uniqKey="Huang C">C Huang</name>
</author>
<author><name sortKey="Lokugamage, K" uniqKey="Lokugamage K">K Lokugamage</name>
</author>
<author><name sortKey="Makino, S" uniqKey="Makino S">S Makino</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Narayanan, K" uniqKey="Narayanan K">K Narayanan</name>
</author>
<author><name sortKey="Ramirez, Si" uniqKey="Ramirez S">SI Ramirez</name>
</author>
<author><name sortKey="Lokugamage, Kg" uniqKey="Lokugamage K">KG Lokugamage</name>
</author>
<author><name sortKey="Makino, S" uniqKey="Makino S">S Makino</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Narayanan, K" uniqKey="Narayanan K">K Narayanan</name>
</author>
<author><name sortKey="Huang, C" uniqKey="Huang C">C Huang</name>
</author>
<author><name sortKey="Lokugamage, K" uniqKey="Lokugamage K">K Lokugamage</name>
</author>
<author><name sortKey="Kamitani, W" uniqKey="Kamitani W">W Kamitani</name>
</author>
<author><name sortKey="Ikegami, T" uniqKey="Ikegami T">T Ikegami</name>
</author>
<author><name sortKey="Tseng, Ct" uniqKey="Tseng C">CT Tseng</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Wathelet, Mg" uniqKey="Wathelet M">MG Wathelet</name>
</author>
<author><name sortKey="Orr, M" uniqKey="Orr M">M Orr</name>
</author>
<author><name sortKey="Frieman, Mb" uniqKey="Frieman M">MB Frieman</name>
</author>
<author><name sortKey="Baric, Rs" uniqKey="Baric R">RS Baric</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Yang, X" uniqKey="Yang X">X Yang</name>
</author>
<author><name sortKey="Chen, X" uniqKey="Chen X">X Chen</name>
</author>
<author><name sortKey="Bian, G" uniqKey="Bian G">G Bian</name>
</author>
<author><name sortKey="Tu, J" uniqKey="Tu J">J Tu</name>
</author>
<author><name sortKey="Xing, Y" uniqKey="Xing Y">Y Xing</name>
</author>
<author><name sortKey="Wang, Y" uniqKey="Wang Y">Y Wang</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Zust, R" uniqKey="Zust R">R Zust</name>
</author>
<author><name sortKey="Cervantes Barragan, L" uniqKey="Cervantes Barragan L">L Cervantes-Barragan</name>
</author>
<author><name sortKey="Kuri, T" uniqKey="Kuri T">T Kuri</name>
</author>
<author><name sortKey="Blakqori, G" uniqKey="Blakqori G">G Blakqori</name>
</author>
<author><name sortKey="Weber, F" uniqKey="Weber F">F Weber</name>
</author>
<author><name sortKey="Ludewig, B" uniqKey="Ludewig B">B Ludewig</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Lei, L" uniqKey="Lei L">L Lei</name>
</author>
<author><name sortKey="Ying, S" uniqKey="Ying S">S Ying</name>
</author>
<author><name sortKey="Baojun, L" uniqKey="Baojun L">L Baojun</name>
</author>
<author><name sortKey="Yi, Y" uniqKey="Yi Y">Y Yi</name>
</author>
<author><name sortKey="Xiang, H" uniqKey="Xiang H">H Xiang</name>
</author>
<author><name sortKey="Wenli, S" uniqKey="Wenli S">S Wenli</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Snijder, Ej" uniqKey="Snijder E">EJ Snijder</name>
</author>
<author><name sortKey="Bredenbeek, Pj" uniqKey="Bredenbeek P">PJ Bredenbeek</name>
</author>
<author><name sortKey="Dobbe, Jc" uniqKey="Dobbe J">JC Dobbe</name>
</author>
<author><name sortKey="Thiel, V" uniqKey="Thiel V">V Thiel</name>
</author>
<author><name sortKey="Ziebuhr, J" uniqKey="Ziebuhr J">J Ziebuhr</name>
</author>
<author><name sortKey="Poon, Llm" uniqKey="Poon L">LLM Poon</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Van Den Born, E" uniqKey="Van Den Born E">E van den Born</name>
</author>
<author><name sortKey="Gultyaev, Ap" uniqKey="Gultyaev A">AP Gultyaev</name>
</author>
<author><name sortKey="Snijder, Ej" uniqKey="Snijder E">EJ Snijder</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Zu Iga, S" uniqKey="Zu Iga S">S Zuñiga</name>
</author>
<author><name sortKey="Sola, I" uniqKey="Sola I">I Sola</name>
</author>
<author><name sortKey="Alonso, S" uniqKey="Alonso S">S Alonso</name>
</author>
<author><name sortKey="Enjuanes, L" uniqKey="Enjuanes L">L Enjuanes</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Regla Nava, Ja" uniqKey="Regla Nava J">JA Regla-Nava</name>
</author>
<author><name sortKey="Nieto Torres, Jl" uniqKey="Nieto Torres J">JL Nieto-Torres</name>
</author>
<author><name sortKey="Jimenez Guarde O, Jm" uniqKey="Jimenez Guarde O J">JM Jimenez-Guardeño</name>
</author>
<author><name sortKey="Fernandez Delgado, R" uniqKey="Fernandez Delgado R">R Fernandez-Delgado</name>
</author>
<author><name sortKey="Fett, C" uniqKey="Fett C">C Fett</name>
</author>
<author><name sortKey="Casta O Rodriguez, C" uniqKey="Casta O Rodriguez C">C Castaño-Rodriguez</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Tang, Nl" uniqKey="Tang N">NL Tang</name>
</author>
<author><name sortKey="Chan, Pk" uniqKey="Chan P">PK Chan</name>
</author>
<author><name sortKey="Wong, Ck" uniqKey="Wong C">CK Wong</name>
</author>
<author><name sortKey="To, Kf" uniqKey="To K">KF To</name>
</author>
<author><name sortKey="Wu, Ak" uniqKey="Wu A">AK Wu</name>
</author>
<author><name sortKey="Sung, Ym" uniqKey="Sung Y">YM Sung</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Smits, Sl" uniqKey="Smits S">SL Smits</name>
</author>
<author><name sortKey="De Lang, A" uniqKey="De Lang A">A de Lang</name>
</author>
<author><name sortKey="Van Den Brand, Jm" uniqKey="Van Den Brand J">JM van den Brand</name>
</author>
<author><name sortKey="Leijten, Lm" uniqKey="Leijten L">LM Leijten</name>
</author>
<author><name sortKey="Van, Iwf" uniqKey="Van I">IWF van</name>
</author>
<author><name sortKey="Eijkemans, Mj" uniqKey="Eijkemans M">MJ Eijkemans</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Peiris, Jsm" uniqKey="Peiris J">JSM Peiris</name>
</author>
<author><name sortKey="Lai, St" uniqKey="Lai S">ST Lai</name>
</author>
<author><name sortKey="Poon, Llm" uniqKey="Poon L">LLM Poon</name>
</author>
<author><name sortKey="Guan, Y" uniqKey="Guan Y">Y Guan</name>
</author>
<author><name sortKey="Yam, Lyc" uniqKey="Yam L">LYC Yam</name>
</author>
<author><name sortKey="Lim, W" uniqKey="Lim W">W Lim</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML DeDiego</name>
</author>
<author><name sortKey="Nieto Torres, Jl" uniqKey="Nieto Torres J">JL Nieto-Torres</name>
</author>
<author><name sortKey="Regla Nava, Ja" uniqKey="Regla Nava J">JA Regla-Nava</name>
</author>
<author><name sortKey="Jimenez Guarde O, Jm" uniqKey="Jimenez Guarde O J">JM Jimenez-Guardeño</name>
</author>
<author><name sortKey="Fernandez Delgado, R" uniqKey="Fernandez Delgado R">R Fernandez-Delgado</name>
</author>
<author><name sortKey="Fett, C" uniqKey="Fett C">C Fett</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Frieman, Mb" uniqKey="Frieman M">MB Frieman</name>
</author>
<author><name sortKey="Chen, J" uniqKey="Chen J">J Chen</name>
</author>
<author><name sortKey="Morrison, Te" uniqKey="Morrison T">TE Morrison</name>
</author>
<author><name sortKey="Whitmore, A" uniqKey="Whitmore A">A Whitmore</name>
</author>
<author><name sortKey="Funkhouser, W" uniqKey="Funkhouser W">W Funkhouser</name>
</author>
<author><name sortKey="Ward, Jm" uniqKey="Ward J">JM Ward</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Sheahan, T" uniqKey="Sheahan T">T Sheahan</name>
</author>
<author><name sortKey="Morrison, Te" uniqKey="Morrison T">TE Morrison</name>
</author>
<author><name sortKey="Funkhouser, W" uniqKey="Funkhouser W">W Funkhouser</name>
</author>
<author><name sortKey="Uematsu, S" uniqKey="Uematsu S">S Uematsu</name>
</author>
<author><name sortKey="Akira, S" uniqKey="Akira S">S Akira</name>
</author>
<author><name sortKey="Baric, Rs" uniqKey="Baric R">RS Baric</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Kuo, L" uniqKey="Kuo L">L Kuo</name>
</author>
<author><name sortKey="Masters, Ps" uniqKey="Masters P">PS Masters</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Kuo, L" uniqKey="Kuo L">L Kuo</name>
</author>
<author><name sortKey="Masters, Ps" uniqKey="Masters P">PS Masters</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Fu, Ks" uniqKey="Fu K">KS Fu</name>
</author>
<author><name sortKey="Baric, Rs" uniqKey="Baric R">RS Baric</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Ballesteros, Ml" uniqKey="Ballesteros M">ML Ballesteros</name>
</author>
<author><name sortKey="Sanchez, Cm" uniqKey="Sanchez C">CM Sanchez</name>
</author>
<author><name sortKey="Enjuanes, L" uniqKey="Enjuanes L">L Enjuanes</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Eckerle, Ld" uniqKey="Eckerle L">LD Eckerle</name>
</author>
<author><name sortKey="Becker, Mm" uniqKey="Becker M">MM Becker</name>
</author>
<author><name sortKey="Halpin, Ra" uniqKey="Halpin R">RA Halpin</name>
</author>
<author><name sortKey="Li, K" uniqKey="Li K">K Li</name>
</author>
<author><name sortKey="Venter, E" uniqKey="Venter E">E Venter</name>
</author>
<author><name sortKey="Lu, X" uniqKey="Lu X">X Lu</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Kuo, L" uniqKey="Kuo L">L Kuo</name>
</author>
<author><name sortKey="Masters, Ps" uniqKey="Masters P">PS Masters</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Song, Ds" uniqKey="Song D">DS Song</name>
</author>
<author><name sortKey="Yang, Js" uniqKey="Yang J">JS Yang</name>
</author>
<author><name sortKey="Oh, Js" uniqKey="Oh J">JS Oh</name>
</author>
<author><name sortKey="Han, Jh" uniqKey="Han J">JH Han</name>
</author>
<author><name sortKey="Park, Bk" uniqKey="Park B">BK Park</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Woods, Rd" uniqKey="Woods R">RD Woods</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Sanchez, Cm" uniqKey="Sanchez C">CM Sanchez</name>
</author>
<author><name sortKey="Izeta, A" uniqKey="Izeta A">A Izeta</name>
</author>
<author><name sortKey="Sanchez Morgado, Jm" uniqKey="Sanchez Morgado J">JM Sanchez-Morgado</name>
</author>
<author><name sortKey="Alonso, S" uniqKey="Alonso S">S Alonso</name>
</author>
<author><name sortKey="Sola, I" uniqKey="Sola I">I Sola</name>
</author>
<author><name sortKey="Balasch, M" uniqKey="Balasch M">M Balasch</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Ye, F" uniqKey="Ye F">F Ye</name>
</author>
<author><name sortKey="Zhang, M" uniqKey="Zhang M">M Zhang</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Tonikian, R" uniqKey="Tonikian R">R Tonikian</name>
</author>
<author><name sortKey="Zhang, Y" uniqKey="Zhang Y">Y Zhang</name>
</author>
<author><name sortKey="Sazinsky, Sl" uniqKey="Sazinsky S">SL Sazinsky</name>
</author>
<author><name sortKey="Currell, B" uniqKey="Currell B">B Currell</name>
</author>
<author><name sortKey="Yeh, Jh" uniqKey="Yeh J">JH Yeh</name>
</author>
<author><name sortKey="Reva, B" uniqKey="Reva B">B Reva</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Roberts, A" uniqKey="Roberts A">A Roberts</name>
</author>
<author><name sortKey="Deming, D" uniqKey="Deming D">D Deming</name>
</author>
<author><name sortKey="Paddock, Cd" uniqKey="Paddock C">CD Paddock</name>
</author>
<author><name sortKey="Cheng, A" uniqKey="Cheng A">A Cheng</name>
</author>
<author><name sortKey="Yount, B" uniqKey="Yount B">B Yount</name>
</author>
<author><name sortKey="Vogel, L" uniqKey="Vogel L">L Vogel</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Chen, Cc" uniqKey="Chen C">CC Chen</name>
</author>
<author><name sortKey="Kruger, J" uniqKey="Kruger J">J Kruger</name>
</author>
<author><name sortKey="Sramala, I" uniqKey="Sramala I">I Sramala</name>
</author>
<author><name sortKey="Hsu, Hj" uniqKey="Hsu H">HJ Hsu</name>
</author>
<author><name sortKey="Henklein, P" uniqKey="Henklein P">P Henklein</name>
</author>
<author><name sortKey="Chen, Ym" uniqKey="Chen Y">YM Chen</name>
</author>
</analytic>
</biblStruct>
<biblStruct></biblStruct>
<biblStruct><analytic><author><name sortKey="Chiu, Rw" uniqKey="Chiu R">RW Chiu</name>
</author>
<author><name sortKey="Chim, Ss" uniqKey="Chim S">SS Chim</name>
</author>
<author><name sortKey="Tong, Yk" uniqKey="Tong Y">YK Tong</name>
</author>
<author><name sortKey="Fung, Ks" uniqKey="Fung K">KS Fung</name>
</author>
<author><name sortKey="Chan, Pk" uniqKey="Chan P">PK Chan</name>
</author>
<author><name sortKey="Zhao, Gp" uniqKey="Zhao G">GP Zhao</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Melik, W" uniqKey="Melik W">W Melik</name>
</author>
<author><name sortKey="Ellencrona, K" uniqKey="Ellencrona K">K Ellencrona</name>
</author>
<author><name sortKey="Wigerius, M" uniqKey="Wigerius M">M Wigerius</name>
</author>
<author><name sortKey="Hedstrom, C" uniqKey="Hedstrom C">C Hedstrom</name>
</author>
<author><name sortKey="Elvang, A" uniqKey="Elvang A">A Elvang</name>
</author>
<author><name sortKey="Johansson, M" uniqKey="Johansson M">M Johansson</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Ellencrona, K" uniqKey="Ellencrona K">K Ellencrona</name>
</author>
<author><name sortKey="Syed, A" uniqKey="Syed A">A Syed</name>
</author>
<author><name sortKey="Johansson, M" uniqKey="Johansson M">M Johansson</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Spaller, Mr" uniqKey="Spaller M">MR Spaller</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Bisht, H" uniqKey="Bisht H">H Bisht</name>
</author>
<author><name sortKey="Roberts, A" uniqKey="Roberts A">A Roberts</name>
</author>
<author><name sortKey="Vogel, L" uniqKey="Vogel L">L Vogel</name>
</author>
<author><name sortKey="Bukreyev, A" uniqKey="Bukreyev A">A Bukreyev</name>
</author>
<author><name sortKey="Collins, Pl" uniqKey="Collins P">PL Collins</name>
</author>
<author><name sortKey="Murphy, Br" uniqKey="Murphy B">BR Murphy</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Subbarao, K" uniqKey="Subbarao K">K Subbarao</name>
</author>
<author><name sortKey="Mcauliffe, J" uniqKey="Mcauliffe J">J McAuliffe</name>
</author>
<author><name sortKey="Vogel, L" uniqKey="Vogel L">L Vogel</name>
</author>
<author><name sortKey="Fahle, G" uniqKey="Fahle G">G Fahle</name>
</author>
<author><name sortKey="Fischer, S" uniqKey="Fischer S">S Fischer</name>
</author>
<author><name sortKey="Tatti, K" uniqKey="Tatti K">K Tatti</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Almazan, F" uniqKey="Almazan F">F Almazan</name>
</author>
<author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML DeDiego</name>
</author>
<author><name sortKey="Galan, C" uniqKey="Galan C">C Galan</name>
</author>
<author><name sortKey="Escors, D" uniqKey="Escors D">D Escors</name>
</author>
<author><name sortKey="Alvarez, E" uniqKey="Alvarez E">E Alvarez</name>
</author>
<author><name sortKey="Ortego, J" uniqKey="Ortego J">J Ortego</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Nieto Torres, Jl" uniqKey="Nieto Torres J">JL Nieto-Torres</name>
</author>
<author><name sortKey="Dediego, Ml" uniqKey="Dediego M">ML DeDiego</name>
</author>
<author><name sortKey="Alvarez, E" uniqKey="Alvarez E">E Alvarez</name>
</author>
<author><name sortKey="Jimenez Guarde O, Jm" uniqKey="Jimenez Guarde O J">JM Jimenez-Guardeño</name>
</author>
<author><name sortKey="Regla Nava, Ja" uniqKey="Regla Nava J">JA Regla-Nava</name>
</author>
<author><name sortKey="Llorente, M" uniqKey="Llorente M">M Llorente</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Bustin, Sa" uniqKey="Bustin S">SA Bustin</name>
</author>
<author><name sortKey="Benes, V" uniqKey="Benes V">V Benes</name>
</author>
<author><name sortKey="Garson, Ja" uniqKey="Garson J">JA Garson</name>
</author>
<author><name sortKey="Hellemans, J" uniqKey="Hellemans J">J Hellemans</name>
</author>
<author><name sortKey="Huggett, J" uniqKey="Huggett J">J Huggett</name>
</author>
<author><name sortKey="Kubista, M" uniqKey="Kubista M">M Kubista</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Kallberg, M" uniqKey="Kallberg M">M Kallberg</name>
</author>
<author><name sortKey="Wang, H" uniqKey="Wang H">H Wang</name>
</author>
<author><name sortKey="Wang, S" uniqKey="Wang S">S Wang</name>
</author>
<author><name sortKey="Peng, J" uniqKey="Peng J">J Peng</name>
</author>
<author><name sortKey="Wang, Z" uniqKey="Wang Z">Z Wang</name>
</author>
<author><name sortKey="Lu, H" uniqKey="Lu H">H Lu</name>
</author>
</analytic>
</biblStruct>
</listBibl>
</div1>
</back>
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<pmc article-type="research-article"><pmc-dir>properties open_access</pmc-dir>
<front><journal-meta><journal-id journal-id-type="nlm-ta">PLoS Pathog</journal-id>
<journal-id journal-id-type="iso-abbrev">PLoS Pathog</journal-id>
<journal-id journal-id-type="publisher-id">plos</journal-id>
<journal-id journal-id-type="pmc">plospath</journal-id>
<journal-title-group><journal-title>PLoS Pathogens</journal-title>
</journal-title-group>
<issn pub-type="ppub">1553-7366</issn>
<issn pub-type="epub">1553-7374</issn>
<publisher><publisher-name>Public Library of Science</publisher-name>
<publisher-loc>San Francisco, CA USA</publisher-loc>
</publisher>
</journal-meta>
<article-meta><article-id pub-id-type="pmid">26513244</article-id>
<article-id pub-id-type="pmc">4626112</article-id>
<article-id pub-id-type="doi">10.1371/journal.ppat.1005215</article-id>
<article-id pub-id-type="publisher-id">PPATHOGENS-D-15-01312</article-id>
<article-categories><subj-group subj-group-type="heading"><subject>Research Article</subject>
</subj-group>
</article-categories>
<title-group><article-title>Identification of the Mechanisms Causing Reversion to Virulence in an Attenuated SARS-CoV for the Design of a Genetically Stable Vaccine</article-title>
<alt-title alt-title-type="running-head">SARS-CoV Vaccine with Increased Biosafety</alt-title>
</title-group>
<contrib-group><contrib contrib-type="author" equal-contrib="yes"><name><surname>Jimenez-Guardeño</surname>
<given-names>Jose M.</given-names>
</name>
<xref ref-type="aff" rid="aff001"><sup>1</sup>
</xref>
</contrib>
<contrib contrib-type="author" equal-contrib="yes"><name><surname>Regla-Nava</surname>
<given-names>Jose A.</given-names>
</name>
<xref ref-type="aff" rid="aff001"><sup>1</sup>
</xref>
</contrib>
<contrib contrib-type="author"><name><surname>Nieto-Torres</surname>
<given-names>Jose L.</given-names>
</name>
<xref ref-type="aff" rid="aff001"><sup>1</sup>
</xref>
</contrib>
<contrib contrib-type="author"><name><surname>DeDiego</surname>
<given-names>Marta L.</given-names>
</name>
<xref ref-type="aff" rid="aff001"><sup>1</sup>
</xref>
<xref ref-type="author-notes" rid="currentaff001"><sup>¤</sup>
</xref>
</contrib>
<contrib contrib-type="author"><name><surname>Castaño-Rodriguez</surname>
<given-names>Carlos</given-names>
</name>
<xref ref-type="aff" rid="aff001"><sup>1</sup>
</xref>
</contrib>
<contrib contrib-type="author"><name><surname>Fernandez-Delgado</surname>
<given-names>Raul</given-names>
</name>
<xref ref-type="aff" rid="aff001"><sup>1</sup>
</xref>
</contrib>
<contrib contrib-type="author"><name><surname>Perlman</surname>
<given-names>Stanley</given-names>
</name>
<xref ref-type="aff" rid="aff002"><sup>2</sup>
</xref>
</contrib>
<contrib contrib-type="author"><name><surname>Enjuanes</surname>
<given-names>Luis</given-names>
</name>
<xref ref-type="aff" rid="aff001"><sup>1</sup>
</xref>
<xref rid="cor001" ref-type="corresp">*</xref>
</contrib>
</contrib-group>
<aff id="aff001"><label>1</label>
<addr-line>Department of Molecular and Cell Biology, Centro Nacional de Biotecnología (CNB-CSIC), Campus Universidad Autónoma de Madrid, Madrid, Spain</addr-line>
</aff>
<aff id="aff002"><label>2</label>
<addr-line>Department of Microbiology, University of Iowa, Iowa City, Iowa, United States of America</addr-line>
</aff>
<contrib-group><contrib contrib-type="editor"><name><surname>Drosten</surname>
<given-names>Christian</given-names>
</name>
<role>Editor</role>
<xref ref-type="aff" rid="edit1"></xref>
</contrib>
</contrib-group>
<aff id="edit1"><addr-line>University of Bonn, GERMANY</addr-line>
</aff>
<author-notes><fn fn-type="COI-statement" id="coi001"><p>The authors have declared that no competing interests exist.</p>
</fn>
<fn fn-type="con" id="contrib001"><p>Conceived and designed the experiments: JMJG JARN JLNT LE. Performed the experiments: JMJG JARN JLNT MLD RFD. Analyzed the data: JMJG JARN JLNT LE. Contributed reagents/materials/analysis tools: JMJG JARN JLNT MLD CCR RFD SP LE. Wrote the paper: JMJG JARN LE. Revised the manuscript: JMJG JARN JLNT MLD SP LE.</p>
</fn>
<fn fn-type="current-aff" id="currentaff001"><label>¤</label>
<p>Current address: David H. Smith Center for Vaccine Biology and Immunology, University of Rochester Medical Center, Rochester, New York, United States of America</p>
</fn>
<corresp id="cor001">* E-mail: <email>L.Enjuanes@cnb.csic.es</email>
</corresp>
</author-notes>
<pub-date pub-type="epub"><day>29</day>
<month>10</month>
<year>2015</year>
</pub-date>
<pub-date pub-type="collection"><month>10</month>
<year>2015</year>
</pub-date>
<volume>11</volume>
<issue>10</issue>
<elocation-id>e1005215</elocation-id>
<history><date date-type="received"><day>5</day>
<month>6</month>
<year>2015</year>
</date>
<date date-type="accepted"><day>18</day>
<month>9</month>
<year>2015</year>
</date>
</history>
<permissions><copyright-statement>© 2015 Jimenez-Guardeño et al</copyright-statement>
<copyright-year>2015</copyright-year>
<copyright-holder>Jimenez-Guardeño et al</copyright-holder>
<license xlink:href="http://creativecommons.org/licenses/by/4.0/"><license-p>This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.</license-p>
</license>
</permissions>
<self-uri content-type="pdf" xlink:type="simple" xlink:href="ppat.1005215.pdf"></self-uri>
<abstract><p>A SARS-CoV lacking the full-length E gene (SARS-CoV-∆E) was attenuated and an effective vaccine. Here, we show that this mutant virus regained fitness after serial passages in cell culture or <italic>in vivo</italic>
, resulting in the partial duplication of the membrane gene or in the insertion of a new sequence in gene 8a, respectively. The chimeric proteins generated in cell culture increased virus fitness <italic>in vitro</italic>
but remained attenuated in mice. In contrast, during SARS-CoV-∆E passage in mice, the virus incorporated a mutated variant of 8a protein, resulting in reversion to a virulent phenotype. When the full-length E protein was deleted or its PDZ-binding motif (PBM) was mutated, the revertant viruses either incorporated a novel chimeric protein with a PBM or restored the sequence of the PBM on the E protein, respectively. Similarly, after passage in mice, SARS-CoV-∆E protein 8a mutated, to now encode a PBM, and also regained virulence. These data indicated that the virus requires a PBM on a transmembrane protein to compensate for removal of this motif from the E protein. To increase the genetic stability of the vaccine candidate, we introduced small attenuating deletions in E gene that did not affect the endogenous PBM, preventing the incorporation of novel chimeric proteins in the virus genome. In addition, to increase vaccine biosafety, we introduced additional attenuating mutations into the nsp1 protein. Deletions in the carboxy-terminal region of nsp1 protein led to higher host interferon responses and virus attenuation. Recombinant viruses including attenuating mutations in E and nsp1 genes maintained their attenuation after passage <italic>in vitro</italic>
and <italic>in vivo</italic>
. Further, these viruses fully protected mice against challenge with the lethal parental virus, and are therefore safe and stable vaccine candidates for protection against SARS-CoV.</p>
</abstract>
<abstract abstract-type="summary"><title>Author Summary</title>
<p>Zoonotic coronaviruses, including SARS-CoV, Middle East respiratory syndrome (MERS-CoV), porcine epidemic diarrhea virus (PEDV) and swine delta coronavirus (SDCoV) have recently emerged causing high morbidity and mortality in human or piglets. No fully protective therapy is still available for these CoVs. Therefore, the development of efficient vaccines is a high priority. Live attenuated vaccines are considered most effective compared to other types of vaccines, as they induce a long-lived, balanced immune response. However, safety is the main concern of this type of vaccines because attenuated viruses can eventually revert to a virulent phenotype. Therefore, an essential feature of any live attenuated vaccine candidate is its stability. In addition, introduction of several safety guards is advisable to increase vaccine safety. In this manuscript, we analyzed the mechanisms by which an attenuated SARS-CoV reverted to a virulent phenotype and describe the introduction of attenuating deletions that maintained virus stability. The virus, engineered with two safety guards, provided full protection against challenge with a lethal SARS-CoV. Understanding the molecular mechanisms leading to pathogenicity and the <italic>in vivo</italic>
evaluation of vaccine genetic stability contributed to a rational design of a promising SARS-CoV vaccine.</p>
</abstract>
<funding-group><funding-statement>This work was supported by grants from the government of Spain (BIO2013-42869-R), the European Community's Seventh Framework Programme under the project “EMPERIE” EC grant agreement number 223498, and a U.S. National Institutes of Health (NIH) project (5P01AI060699). JMJG, JARN and JLNT received contracts from NIH. CCR received a contract from Fundacion La Caixa. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.</funding-statement>
</funding-group>
<counts><fig-count count="16"></fig-count>
<table-count count="0"></table-count>
<page-count count="36"></page-count>
</counts>
<custom-meta-group><custom-meta id="data-availability"><meta-name>Data Availability</meta-name>
<meta-value>All relevant data are within the paper and its Supporting Information files.</meta-value>
</custom-meta>
</custom-meta-group>
</article-meta>
<notes><title>Data Availability</title>
<p>All relevant data are within the paper and its Supporting Information files.</p>
</notes>
</front>
</pmc>
</record>
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