Sensitivity of infectious SARS-CoV-2 B.1.1.7 and B.1.351 variants to neutralizing antibodies

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ARTICLES https://doi.org/10.1038/s41591-021-01318-5 Sensitivity of infectious SARS-CoV-2 B.1.1.7 and B.1.351 variants to neutralizing antibodies 1,2,3,22 1,2,3,221,2,3,4 1,2,3 Delphine Planas, Timothée Bruel, Ludivine Grzelak, Florence Guivel-Benhassine, 1,2,3 1,2,35 1,2,3 Isabelle Staropoli, Françoise Porrot, Cyril Planchais, Julian Buchrieser, 1,2,3,4 1,2,3,46,7 6,78 Maaran Michael Rajah, Elodie Bishop, Mélanie Albert, Flora Donati, Matthieu Prot, 6,7 6,79 1010 Sylvie Behillil, Vincent Enouf, Marianne Maquart, Mounira Smati-Lafarge, Emmanuelle Varon, 11 1213 14,1516 Frédérique Schortgen, Layla Yahyaoui, Maria Gonzalez, Jérôme De Sèze, Hélène Péré, 16,17 188 19,209,21 David Veyer, Aymeric Sève, Etienne Simon-Lorière, Samira Fafi-Kremer, Karl Stefic, 5 186,7,23 18,23 Hugo Mouquet, Laurent Hocqueloux, Sylvie van der Werf, Thierry Prazuckand ᅒ 1,2,3,23 Olivier Schwartz Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) B.1.1.7 and B.1.351 variants were first identified in the United Kingdom and South Africa, respectively, and have since spread to many countries. These variants harboring diverse mutations in the gene encoding the spike protein raise important concerns about their immune evasion potential. Here, we isolated infectious B.1.1.7 and B.1.351 strains from acutely infected individuals.

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• Coronavirus

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• Publié par: france3centre
• Publié le: 07 avril 2021
• Nombre de lectures: 4 674
• Langue: English
• Poids de l'ouvrage: 4 Mo

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ARTICLES https://doi.org/10.1038/s41591021013185

Sensitivity of infectioUs SARS-CoV-2 B.1.1.7 and B.1.351 variants to neUtralizing antibodies

1,2,3,22 1,2,3,22 1,2,3,4 1,2,3 Delphine Planas , Timothée BrUel , LUdivine Grzelak , Florence GUivel-Benhassine , 1,2,3 1,2,3 5 1,2,3 Isabelle Staropoli , Françoise Porrot , Cyril Planchais , JUlian BUchrieser , 1,2,3,4 1,2,3,4 6,7 6,7 8 Maaran Michael Rajah , Elodie Bishop , Mélanie Albert , Flora Donati , MatthieU Prot , 6,7 6,7 9 10 10 Sylvie Behillil , Vincent EnoUf , Marianne MaqUart , MoUnira Smati-Lafarge , EmmanUelle Varon , 11 12 13 14,15 16 FrédériqUe Schortgen , Layla YahyaoUi , Maria Gonzalez , Jérôme De Sèze , Hélène Péré , 16,17 18 8 19,20 9,21 David Veyer , Aymeric Sève , Etienne Simon-Lorière , Samira Fafi-Kremer , Karl Stefic , 5 18 6,7,23 18,23 HUgo MoUqUet , LaUrent HocqUeloUx , Sylvie van der Werf , Thierry PrazUck and ✉ 1,2,3,23 Olivier Schwartz

Severe acute respiratory syndrome coronavirus 2 (SARSCoV2) B.1.1.7 and B.1.351 variants were first identified in the United Kingdom and South Africa, respectively, and have since spread to many countries. These variants harboring diverse mutations in the gene encoding the spike protein raise important concerns about their immune evasion potential. Here, we isolated infec tious B.1.1.7 and B.1.351 strains from acutely infected individuals. We examined sensitivity of the two variants to SARSCoV2 antibodies present in sera and nasal swabs from individuals infected with previously circulating strains or who were recently vaccinated, in comparison with a D614G reference virus. We utilized a new rapid neutralization assay, based on reporter cells that become positive for GFP after overnight infection. Sera from 58 convalescent individuals collected up to 9 months after symptoms, similarly neutralized B.1.1.7 and D614G. In contrast, after 9 months, convalescent sera had a mean sixfold reduction in neutralizing titers, and 40% of the samples lacked any activity against B.1.351. Sera from 19 individuals vaccinated twice with Pfizer Cominarty, longitudinally tested up to 6 weeks after vaccination, were similarly potent against B.1.1.7 but less efficacious against B.1.351, when compared to D614G. Neutralizing titers increased after the second vaccine dose, but remained 14fold lower against B.1.351. In contrast, sera from convalescent or vaccinated individuals similarly bound the three spike proteins in a flow cytometrybased serological assay. Neutralizing antibodies were rarely detected in nasal swabs from vaccinees. Thus, fasterspreading SARSCoV2 variants acquired a partial resistance to neutralizing antibodies generated by natural infection or vaccination, which was most frequently detected in individuals with low antibody levels. Our results indicate that B1.351, but not B.1.1.7, may increase the risk of infection in immunized individuals.

ARS-CoV-2 varîans ave rapîdly emerged în umans and 1–5 îo S n advanage a e populaîon level. One o e îrs îdenîîed supplaned ancesral sraîns . Teîr proposed încreased raes o înerîndîvîdual ransmîssîon conerred a replîca- varîans încludes e D614G muaîon în e gene encodîng e spîke (S) proeîn, wîc enances vîral înecîvîy and sîs S pro- eîn conormaîon oward an angîoensîn-converîng enzyme 2 (ACE2)-bîndîng usîon-compeen sae, wîou sîgnîîcanly mod- 1,6–8 îyîng sensîîvîy o anîbody neuralîzaîon . More recenly, novel varîans ave appeared în mulîple counrîes, wî combînaîons

o muaîons and deleîons în e recepor-bîndîng domaîn (RBD) and N-ermînal domaîn o S proeîn, as well as în oer proeîns. Te B.1.1.7 varîan emerged în e Unîed Kîngdom, e B.1.351 varîan (also ermed 501Y.V2) în Sou Arîca and e P.1 and 2,3,5,9–12 P.2 lîneages în Brazîl . Aloug dîsînc, e varîans sare common caracerîsîcs, încludîng known escape muaîons a were prevîously îdenîîed under anîbody pressure selecîon 2,3,13–17 în vîro . Some o e muaîons or deleîons were also îdenî- îed în îmmunocompromîsed îndîvîduals wî prolonged înecîous vîral seddîng and reaed wî convalescen plasma or S-proeîn

1 2 3 Virus & Immunity Unit, Department of Virology, Institut Pasteur, Paris, France. CNRS UMR 3569, Paris, France. Vaccine Research Institute, Créteil, 4 5 France. Université de Paris, Sorbonne Paris Cité, Paris, France. Laboratory of Humoral Immunology, Department of Immunology, Institut Pasteur, 6 INSERM U1222, Paris, France. Molecular Genetics of RNA Viruses, Department of Virology, Institut Pasteur CNRS UMR 3569, Université de Paris, Paris, 7 8 France. National Reference Center for Respiratory Viruses, Institut Pasteur, Paris, France. G5 Evolutionary Genomics of RNA Viruses, Institut Pasteur, 9 10 11 Paris, France. INSERM U1259, Université de Tours, Tours, France. CHI de Créteil, Service de Biologie Médicale, Créteil, France. CHI de Créteil, Service 12 13 de Réanimation, Créteil, France. CHI de Créteil, Service des Urgences, Créteil, France. CHU de Strasbourg, Service de Pathologie Professionnelle et 14 15 Médecine du Travail, Strasbourg, France. Centre d’investigation Clinique INSERM 1434, CHU Strasbourg, France. CHU de Strasbourg, Service de 16 Neurologie, Strasbourg, France. INSERM, Functional Genomics of Solid Tumors (FunGeST), Centre de Recherche des Cordeliers, Université de Paris 17 18 and Sorbonne Université, Paris, France. Hôpital Européen Georges Pompidou, Service de Virologie, Paris, France. CHR d’Orléans, Service de maladies 19 20 infectieuses, Orléans, France. CHU de Strasbourg, Laboratoire de Virologie, Strasbourg, France. Université de Strasbourg, INSERM, IRM UMR_S 1109, 21 22 Strasbourg, France. CHRU de Tours, National Reference Center for HIV-Associated laboratory, Tours, France. These authors contributed equally: 23 Delphine Planas, Timothée Bruel. These authors jointly supervised this work: Sylvie van der Werf, Thierry Prazuck, Olivier Schwartz. ✉ e-mail:[e-mail]

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ARTICLES

3,17–19 monoclonal anîbodîes , îndîcaîng a anîbody-escape mua- îons are seleced în vîvo. Te sensîîvîy o anîbody neuralîza- îon varîes wî e vîral varîan. B.1.1.7 seems o be more sensîîve o neuralîzaîon an B.1.351. Te RBD muaîon N501Y, wîc încreases aînîy o ACE2 and îs presen în B.1.1.7 and B.1.351 20 (re. ), does no împaîr on îs own pos-vaccîne serum neural- 21 îzaîon . ï as been suggesed a e oer muaîons în B.1.1.7 22 do no resul în îmmune evasîon o lînear epîopes . Muaîons în e B.1.351 and P.1 sraîns, încludîng E484K and K417N/T, are o îg concern, sînce ey parly compromîse neuralîza- 23–25 îon generaed by prevîous înecîon or vaccînaîon or may încrease îneren vîral îness. Te Pîzer Comînary (also ermed BNT162b2) vaccîne-elîcîed uman sera neuralîzes SARS-CoV-2 lîneage B.1.1.7 pseudovîrus, wî slîgly reduced îers în some 26,27 vaccînees, wen compared o e Wuan reerence sraîn . Te Moderna mRNA-1273 vaccîne also înduces neuralîzîng anîbodîes o SARS-CoV-2 varîans wî, owever, a îve- o en-old reduc- îon în eîcacy agaîns e B.1.351 S proeîn, wen compared o 27,28 pseudovîrus bearîng e D614G muaîon . Neuralîzaîon eîcacy as so ar mosly been assessed usîng vesîcular somaîs vîrus (VSV)-derîved or lenîvîrus-derîved pseu- dovîrus assays, or wî înecîous SARS-CoV-2 carryîng poîn muaîons în S proeîn. A recen repor usîng înecîous B.1.351 vîrus sowed a plasma samples rom sîx convalescen donors were srongly aenuaed agaîns îs sraîn, wî al-maxîmal înîbîory concenraîon (ïC 50 ) values 6- o 200-old îger an 29 ancesral vîrus . ï îs us o umos împorance o use auenîc varîan sraîns în addîîon o pseudovîrus parîcles, sînce muaîons ousîde o e S proeîn may împac îneren vîral îness and/or sensîîvîy o anîbodîes. Here, we compared e sensîîvîy o ree auenîc SARS-CoV-2 sraîns, e preexîsîng D614G vîrus and e B.1.1.7 and B.1.351 varîans, o anîbody neuralîzaîon.

ResUlts Isolation and characterization of SARSCoV2 B.1.1.7 and B.1.351 variants.îsolaed e wo varîans rom nasal swabs We o îndîvîduals wî înecîon dîagnosed by quanîaîve PCR wî reverse ranscrîpîon (RT–qPCR) and sequence-dîagnosed înec- îon. Te vîruses were amplîîed by one o wo passages on Vero cells. Sequences o e ougrown vîruses conîrmed e îdenîy o B.1.1.7 and B.1.351 varîans. Vîral socks were îraed usîng S-Fuse 30 reporer cells . Tese cells are derîved rom a uman îmmoralîzed + lîne, ermed U2OS-ACE2 cells. Tey carry e GFP-splî comple- menaîon sysem, în wîc wo cells separaely produce al o e reporer proeîn, producîng GFP only upon usîon. Followîng înecîon, e cells produce S proeîn a eîr surace and use wî neîgborîng cells, generaîng a GFP sîgnal as soon as 6  aer înec- 30+ îon (Fîg.1a). Te number o GFP cells correlaed wî e vîral înoculum (Fîg.1b). Neuralîzîng SARS-CoV-2 monoclonal anî- bodîes (mAbs) argeîng e RBD can be classîîed îno our maîn 31,32 caegorîes . Usîng e S-Fuse assay, we esed e sensîîvîy o e ree vîral sraîns o wo RBD anîbodîes, mAb102 and mAb48, derîved rom convalescen îndîvîduals. Te anîbodîes belong o e îrs caegory and ac by blockîng bîndîng o e ‘up’ conor- 32 maîon o RBD o ACE2 (re. ). MAb102 eîcîenly and sîmîlarly −1 neuralîzed e ree vîral sraîns, wî an ïC 50  o≃0.01µg ml −1 (Fîg.1c,d). MAb48 neuralîzed D614G (ïC 50  o 0.1µg ml ) bu was înacîve agaîns B.1.1.7 and B.1.351 varîans (Fîg.1c,d). Tese resuls conîrm a e wo varîans can selecîvely dîsplay reduced sensîîvîy o ceraîn anîbodîes. O noe, e acîvîy o e mono- clonal anîbodîes was sîmîlar a dîeren vîral înocula, wîîn a + range o 50 o 200 GFP syncyîa per well (Fîg.1e), îndîcaîng a poenîal varîaîons în e number o îneced S-Fuse cells do no împac e calculaîon o ïC 50 . Tus, or urer sudîes, we seleced a mulîplîcîy o înecîon leadîng o abou 150 syncyîa per well or eac vîral sraîn.

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Sensitivity of D614G, B.1.1.7 and B.1.351 variants to sera from convalescent individuals.We assessed e neuralîzaîon abîlîy o sera rom convalescen îndîvîduals. We seleced samples rom 28 donors în a longîudînal coor o îndîvîduals wî SARS-CoV-2 rom e Frenc cîy o Orléans (Supplemenary Table 1). All îndî- vîduals were dîagnosed wî SARS-CoV-2 înecîon by RT–qPCR or serology and încludîng ose wo ad crîîcal, severe and mîld-o-moderae coronavîrus dîsease 2019 (COVïD-19). Tey were no vaccînaed. A oal o 25 îndîvîduals were sampled wîce: îrs, a a medîan o 89 d (range 77–105 d) pos onse o symp- oms (POS; 3-mon samples (M3)), and second, a a medîan o 179 d (range 168–197 d) POS (6-mon samples (M6). We încu- baed serîally dîlued sera wî D614G, B.1.1.7 or B.1.351 sraîns, + added e mîxure o S-Fuse cells, and scored e GFP cells aer overnîg înecîon. We en calculaed e medîan eecîve dose (ED 50 ) or eac combînaîon o serum and vîrus. A represena- îve example wî e same donor îs depîced în Fîg.2a, and e resuls wî all donors appear în Fîg.2b. Te D614G and B.1.1.7 sraîns were sîmîlarly sensîîve o sera. A M3, e ED 50 values were 3 3 1.5×10 and 1×10 or D614G and B.1.1.7 varîans, respecîvely, 2 4 wî uge varîaîons beween îndîvîduals (rom 10 o 2×10 ), and e values dîd no srongly declîne a M6. As expeced, îndîvîdu- als wî crîîcal dîsease dîsplayed îger neuralîzîng acîvîîes an ose wî severe or mîld-o-moderae sympoms (Exended Daa Fîg. 1). Agaîn, ere was no sîgnîîcan dîerence beween e wo vîral sraîns în eac caegory o sympoms. For B1.351, e neural- îzaîon îers were sîgnîîcanly decreased by îve and en-old a e wo îme poîns, wen compared wî D614G and B.1.1.7 sraîns (Fîg.2band Exended Daa Fîg. 1). We conîrmed ese resuls în sera rom 30 îndîvîduals rom anoer coor o RT–qPCR-conîrmed unvaccînaed eal-care workers rom Srasbourg Unîversîy Hospîal wo experîenced mîld 33,34 dîsease . Te samples were colleced a a laer îme poîn (M9), wî a medîan o 233 d (range 206–258 d) POS (Supplemenary Table 1). Overall, e neuralîzaîon acîvîy was lower (one represenaîve example îs sown în Fîg.2a and e resuls o all donors are avaîl- able în Fîg.2b). Tere was no sîgnîîcan dîerence în neuralîzaîon 2 beween D614G and B.1.1.7 sraîns, wî a sîmîlar ED 50 o 2×10 . ïn sarp conras, e neuralîzîng acîvîy agaîns B.1.351 was parîcu- larly low a îs lae îme poîn, wî a medîan ED 50 o 50, represen- îng a ourold decrease wen compared o D614G (Fîg.2b). We en arbîrarîly classîîed e îndîvîduals as neuralîzers (wî neuralîzîng anîbodîes deecable a e îrs serum dîluîon o 1:30) and non-neuralîzers, or e ree vîral sraîns and e wo coors (Fîg.2c). Mos îndîvîduals neuralîzed e ree sraîns a M3. Te racîon o neuralîzers sared o declîne a M6, a penomenon wîc was more marked or B.1.351. Te racîon o neuralîzers was îger în e second coor, wî 93% o îndîvîduals neural- îzîng eîer D614G or B.1.1.7 sraîns a M9. ïn conras, only 63% o îndîvîduals neuralîzed e B.1.351 sraîn (Fîg.2c).

Antibody binding to cells expressing D614G, B.1.1.7 and B.1.351 S proteins.We nex examîned e bîndîng capacîy o monoclonal anîbodîes and sera rom e convalescen îndîvîduals descrîbed above o S proeîns rom dîeren lîneages. To îs aîm, we adaped e low cyomery-based S-Flow assay, wîc we prevîously esab- lîsed o measure e levels o anîbodîes bîndîng o cells expressîng 35 e Wuan S proeîn . We ransîenly ranseced uman epîe- lîal HEK293T (reerred o as 293T) cells wî plasmîds expressîng e D614G, B.1.1.7 and B.1.351 S proeîns. Sîmîlar surace levels o e vîral proeîns were deeced wî mAb10, a non-neuralîzîng pan-coronavîrus anîbody argeîng a conserved epîope în e S2 domaîn and îsolaed rom a convalescen îndîvîdual (C. Plancaîs e al., unpublîsed daa; Fîg.3aSupplemenary Fîg. 1). and Moreover, e cells used wen coculîvaed wî ACE2-expressîng cells, îndîcaîng a e ree proeîns were uncîonal (daa no

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Sensitivity of D614G, B.1.1.7 and B.1.351 variants to sera from vaccine recipients. We nex asked weer vaccîne-elîcîed anî- bodîes înîbî înecîon by e dîeren varîans. ïn France, vaccî- naîon sared în January 2020 wî e Pîzer Comînary vaccîne.

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sown). mAb48 eîcîenly bound o D614G S proeîns, slîgly less poenly o B.1.1.7, and los all bîndîng acîvîy o B.1.351. Tereore, e escape o mAb48 neuralîzaîon by e wo varîans îs due o muaîons decreasîng or abrogaîng anîbody bîndîng o îs arge. mAb102 bound sîmîlarly o e ree S proeîns, în accor- dance wî îs cross-reacîve neuralîzîng acîvîy (Fîg.3a,b). Te N501Y muaîon enances RBD aînîy o ACE2, wen esed wî 20 recombînan proeîns and yeas surace dîsplay . Usîng low cyom- ery, we assessed by e bîndîng o a labeled soluble ACE2 proeîn o cells expressîng e dîeren S proeîns. We observed a dramaîc încrease în bîndîng o soluble ACE2 o B.1.1.7 and o a lesser exen o B.1.351, wîc bo carry e N501Y muaîon, wen compared o D614G (Fîg.3a,b). We nex esed our panel o sera rom convalescen îndîvîdu- als agaîns e dîeren S proeîns. Overall, e sera bound sîmî- larly o all ree S proeîns, even oug B.1.351 dîsplayed a slîg reducîon (saîsîcally sîgnîîcan a M6) în e mean luorescence înensîy (MFï) o bîndîng (Fîg.3c). We observed a global and slîg decrease în MFï a M9, compared o samples rom M3 and M6 (Fîg.3c). Alogeer, ese resuls îndîcae a S proeîns rom B.1.351, and o a îger exen B.1.1.7, dîsplay încreased aînîy or ACE2 wîle escapîng bîndîng o some monoclonal anîbodîes and eîer o a lesser degree or no a all o polyclonal sera.

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We us seleced 19 vaccîne recîpîens rom a coor o vaccînaed eal-care workers esablîsed în Orléans. Te caracerîsîcs o vaccînees are depîced în Supplemenary Table 2. Sera and nasal swabs were sampled a week 2 (17 îndîvîduals), week 3 (18 îndî- vîduals), week 4 (11 îndîvîduals; correspondîng o week 1 aer e second dose), week 5 (16 îndîvîduals; correspondîng o week 2 aer e second dose) and week 6 (15 îndîvîduals; correspondîng o week 3 aer e second dose), wî a medîan o 13, 19, 28, 34 and 41 d aer e îrs dose, respecîvely. Tîs allowed us o assess e early umoral response o vaccînaîon. We îrs analyzed 16 o 19 vaccîn- ees a were no prevîously îneced wî SARS-CoV-2, as assessed by e absence o preexîsîng S-proeîn and anî-nucleoproeîn (N) anîbodîes. A represenaîve example o e evoluîon o e neu- ralîzîng response în one vaccîne recîpîen a e our îme poîns îs depîced în Fîg.4a. ïn îs însance, e serum only neuralîzed D614G a week 2, wereas B.1.1.7 sared o be neuralîzed a week 3, aloug less eîcîenly an D614G. B.1.1.7 and D614G sraîns were sîmîlarly neuralîzed a week 4. Te anî-B.1.351 response was negaîve up o week 3, and became deecable a week 4, a a level lower an e wo oer vîruses. Te ED 50  resuls rom sera o e 16 vaccîne recîpîens a e our îme poîns are presened în Fîg.4b, wereas e evoluîon o eîr neuralîzaîon îers over e 6 weeks aer vaccînaîon appears în Fîg.4c. We also arbîrarîly classîîed îndîvîduals as neuralîz- ers (wî neuralîzîng anîbodîes deecable a a serum dîluîon o 1:30) and non-neuralîzers, or e ree vîral sraîns (Fîg.4d). Two weeks aer vaccînaîon, anîbodîes neuralîzîng e D614G sraîn were deeced în sera rom 5 o 15 recîpîens (33%; wî an

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Fig. 1 |Effect of two SARS-CoV-2 monoclonal antibodies on D614G, B.1.1.7 and B1.351 strains.a, Principle of the S-Fuse reporter assay. S-Fuse cells, a mix of U20S-ACE2-GFP1–10 and U20S-ACE2-GFP11, were cocultured at a 1:1 ratio and infected with SARS-CoV-2. Cells fused upon infection and generated + a GFP signal. Infection was quantified by measuring the number of GFP syncytia at 18 h.b, Linearity of the assay with D614G (gray), B.1.1.7 (blue) and B.1.351 (green) strains. S-Fuse cells were exposed to serial dilutions of viral stocks and infection was quantified by high-content imaging.c, Representative images (from five independent experiments) of S-Fuse cells infected with D614G, B.1.1.7 and B1.351 strains in the presence or absence of two SARS-CoV-2 monoclonal antibodies (mAb48 and mAb102). Scale bar, 400µm.d, Dose-response analysis of the neutralization by mAb102 and mAb48 on the three viral strains.e, Neutralization of SARS-CoV-2 (B.1.1.7) by mAb102 at three viral inocula. Results are shown as the mean±standard deviation (s.d.) from four independent experiments.

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îers încreased în mos o e recîpîens and were sîmîlar beween D614G and B.1.1.7. Tîers remaîned 14-old and 53-old lower agaîns B.1.351, wen compared o D614G and B.1.117, respecîvely (Fîg.4b,c). A week 3, 63%, 38% and 0% o e samples neuralîzed D614G, B.1.1.7 and B.1.351 sraîns, respecîvely (Fîg.4d). A week 6, 92% o e samples neuralîzed D614G and B.1.1.7, wereas 77% neuralîzed B.1.351 wî a low îer (Fîg.4d). Te S-Flow assay dem- onsraed e presence o anîbodîes bîndîng o e ree S proeîns

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arbîrary resold o ED 50 >30 or neuralîzaîon posîîvîy; Fîg. 4b,d). Te îers were relaîvely modes a îs early îme poîn (ED 50 o 30). Tese low îers were less eîcîen agaîns B.1.1.7, wî 2 o 15 neuralîzers (13%), and were înacîve agaîns B.1.351 (Fîg.4b,c). A week 3, e neuralîzîng acîvîy încreased agaîns D614G and B.1.1.7. Tere was, owever, a reeold reducîon în e neural- îzaîon îers agaîns B.1.1.7 (Fîg.4b), wereas B.1.351 remaîned însensîîve. A e las îme poîn (week 3 aer e second dose),

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Fig. 2 |Sensitivity of SARS-CoV-2 D614G, B.1.1.7 and B.1.351 variants to sera from convalescent individUals.a, Examples of neutralization curves with sera from two donors (D18 and D59). The first donor, from the Orléans cohort, was sequentially sampled at months 3 (M3) and 6 (M6; left and middle, respectively). The second donor, from the Strasbourg cohort, was sampled at month 9 (M9; right). Results are shown as the mean±s.d. from three independent experiments.b, ED 50 of neutralization of the three viral isolates. Sera from the Orléans cohort were sequentially sampled at M3 (n=28) and M6 (n=25; left and middle, respectively), and 30 sera from the Strasbourg cohort were sampled at M9 (right). Data are the mean from two to four independent experiments. The dotted line indicates the limit of detection (ED 50 =30). Two-sided Friedman test with Dunn’s test for multiple comparisons was performed between each viral strain at the different time points; **P<0.01, ***P<0.001, ****P<0.0001. M3: D614G versus B.1.351,P=0.0012; B.1.1.7 versus B.1.351,P=0.0012. M6: D614G versus B.1.351,P=0.0005; B.1.1.7 versus B.1.351,P<0.0001. M9: D614G versus B.1.351,P<0.0001; B.1.1.7 versus B.1.351,P<0.0001.c, Each individual was arbitrarily defined as a ‘neutralizer’ (blue) if neutralizing activity was detected at the first serum dilution (1:30) or ‘non-neutralizer’ (gray) if no activity was detected. The numbers indicate the percentage of neutralizers.

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Fig. 3 |Antibody binding to cells expressing D614G, B.1.1.7 and B.1.351 S proteins.a, Binding of monoclonal antibodies or soluble ACE2. HEK293T cells were transiently transfected with plasmids expressing the D614G, B.1.1.7 and B.1.351 S proteins. After 24 h, cells were stained with SARS-CoV-2 antibody −1 mAb10 (a pan-coronavirus antibody), mAb48, mAb102 or soluble ACE2 (ACE2-biotin at 10μg ml revealed with fluorescent streptavidin) and analyzed by flow cytometry. One representative example of binding is shown.b, Titration binding curves of mAb48, mAb102 and ACE2 to the three S proteins. Data are the mean±s.d. of three independent experiments.c, Binding of the panel of 83 sera from 58 convalescent individuals. Sera were tested at a 1:300 dilution. Data are the mean of two independent experiments. Two-sided Friedman test with Dunn’s test for multiple comparisons was performed between each viral strain at the different time points, *P<0.05. M6: B.1.1.7 versus B.1.351,P=0.037.

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all îme poîns (up o 6 weeks aer vaccînaîon; Exended Daa Fîg. 2a,b). We also analyzed ree addîîonal vaccîne recîpîens wo were seroposîîve or anî-N a e îme o vaccînaîon, îndîca- îve o a prevîous înecîon. Two recîpîens were dîagnosed wî SARS-CoV-2 by PCR în Marc and Aprîl 2020 and experîenced a mîld dîsease. Te îrd recîpîen dîd no repor any prevîous sîgns remînîscen o COVïD-19. ïn ese ree îndîvîduals, e serum neuralîzîng îers were srîkîngly îg a week 2 aer e îrs dose 4 (ED 50 o abou 10 ; Exended Daa Fîg. 2c) and remaîned sîmîlarly îg a week 5 (daa no sown). Two o e ree vaccîne recîpîens wo were prevîously seroposîîve or SARS-CoV-2 dîsplayed a low neuralîzîng acîvîy în eîr nasal swabs, wen measured a week 2 aer e îrs dose (Exended Daa Fîg. 2d). Teîr nasal swabs neuralîzed D614G and B.1.117 sraîns sîmîlarly bu were înacîve agaîns B.1.351. As a conrol, we analyzed en pre-pandemîc naso- paryngeal swab specîmens (colleced by e Naîonal Reerence

a e dîeren samplîng îmes (Exended Daa Fîg. 2). Tereore, e Pîzer Comînary vaccîne generaed a neuralîzîng response a eîcîenly argeed D614G and B.1.1.7 wî a delay în e appear- ance o neuralîzîng anîbodîes o B.1.1.7 and B.1.351 relaîve o D614G. Te îers remaîned lower agaîns B.1.351, even în respond- ers. Te vaccîne dîsplays a proecîve eîcacy agaîns COVïD-19 as 36 soon as 2 weeks aer e îrs dose . Our resuls sugges a e low neuralîzîng îers (ED 50 o 50–100) în e sera may correspond o e observed proecîon agaîns severe dîsease.

Sensitivity of SARSCoV2 B.1.1.7 and B.1.351 variants to nasal swabs from vaccine recipients.Lîle îs known abou e levels and uncîon o vaccîne-elîcîed anîbodîes în mucosal samples. We us measured e neuralîzîng acîvîy and e levels o anîbodîes în nasal swabs rom e serîes o vaccîne recîpîens (Exended Daa Fîg. 2). We dîd no deec any anîvîral eec în ese samples a

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Fig. 4 |Sensitivity of SARS-CoV-2 D614G, B.1.1.7 and B.1.351 variants to sera from vaccine recipients.a, Examples of neutralization curves with sera from one donor (D9) at week 2 (W2), W3, W4 (W1 after second dose) and W6 (W3 after second dose) after vaccination. Results are shown as the mean±s.d. from three independent experiments.b, Neutralization ED 50 values of the three viral isolates. Sera from 10 to 16 vaccine recipients were sampled at W2 (n=15), W3 (n=16) and W1 and W3 after the second dose (n=10 and 15, respectively). Data are the mean from two to four independent experiments. The dotted line indicates an ED 50 of 30. Two-sided Friedman test with Dunn’s test for multiple comparisons was performed between each viral strain at the different time points; *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001. W2: D614G versus B.1.1.7,P=0.0105; D614G versus B.1.351,P=0.008. W3: D614G versus B.1.351,P=0.0002. W4: D614G versus B.1.351,P=0.011; B.1.1.7 versus B.1.351,P=0.0052. W6: D614G versus B.1.1.7,P=0.0324; D614G versus B.1.351,P=0.0324; B.1.1.7 versus B.1.351,P<0.0001.c, Evolution overtime of neutralizing antibody titers in 14 vaccine recipients. The arrows indicate the first and second doses of vaccines.d, Each individual was arbitrarily defined as a ‘neutralizer’ (blue) if neutralizing activity was detected at a 1:30 serum dilution or ‘non-neutralizer’ (gray) if no activity was detected. The numbers indicate the percentage of neutralizers.

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