Uhlalutyo lwedatha yoluntu
Idatha ngemigca ye-SARS-CoV-2 kunye nomhla owathathwa isampulu kulandelelwano olukhoyo eChile yafunyanwa kwindawo yeConsorcio Genomas CoV2 efumanekayo https://auspice.cov2.cl/ncov/chile-global. Idatha yogonyo ifunyenwe kwidatha yoluntu evela kwi-Ofisi yoMphathiswa wezeNzululwazi, iTekhnoloji, uLwazi kunye noTshintsho olukhoyo https://github.com/MinCiencia/Datos-COVID19 (Imveliso 83).
Uvavanyo losulelo
Iintsholongwane ze-pseudotyped ezithwele iiproteni ezahlukeneyo ze-SARS-CoV-2 zalungiswa njengoko besichazile ngaphambili.12. Ngokufutshane, iipseudotypes ezise-HIV-1 ezisekwe kwi-SARS-CoV-2 zaveliswa kwiiseli ze-HEK293T ngokugqithisela i-pNL4.3-ΔEnv-Luc kunye ne-pCDNA-SARS-CoV-2 Spike coding vector ehambelanayo kwi-1: 1 molar ratio. IiPlasmids zikhowuda i-codon-optimized spike eswele i-19 amino acids yokugqibela ye-C-terminal end (SΔ19) eyaziwayo ukunqanda ukugcinwa kwi-endoplasmic reticulum.12 zifunyenwe nge-gene synthesis okanye i-customized site-directed mutagenesis (GeneScript) kwaye iqulethe ezi nguqu zilandelayo: umgca A (ulandelelwano lwereferensi), umgca B (D614G), umgca B.1.1.7 (Δ69-70, Δ144, N501Y, A570D, D614G, P681H, T716I, S982A, D1118H), umnombo weP.1 (L18F, T20N, P26S, D138Y, R190S, K417T, E484K, N501Y, D614G, H655Y, T1027N, P37S, D75Y, R76S, K246T, E252K, N452Y, D490G, H614Y, T859N, T3,000G1, T24, T50, T80, T1, T24, T2, T48, T96, TXNUMX, TXNUMX, TXNUMX, TXNUMX, TXNUMX, TXNUMX, TXNUMX, TXNUMX, TXNUMX, XNUMX- XNUMX, LXNUMXQ, FXNUMXS, DXNUMXG, TXNUMXN). Ulungiselelo ngalunye lwe-pseudotype lwacocwa nge-centrifugation kwi-XNUMX rpm kwiqondo lobushushu begumbi, libalwe kusetyenziswa i-HIV-XNUMX Gag pXNUMX Quantikine ELISA Kit (R&D Systems), ecatshulwe kwi-XNUMX% ye-fetus bovine serum (Sigma-Aldrich) kwaye igcinwe ku -XNUMX °C de sebenzisa. Amanani ahlukeneyo eentsholongwane ze-pseudotyped virus (njengoko kuchongwe ngamanqanaba e-HIV-XNUMX pXNUMX protein) yayisetyenziselwa ukuchaphazela iiseli ze-HEK-ACEXNUMX kunye neeyure ze-XNUMX kamva, umsebenzi we-firefly luciferase walinganiswa usebenzisa i-Luciferase Assay Reagent (Promega) kwi-Glomax XNUMX Microplate. i-luminometer (iPromega).
Uvavanyo lokungathathi hlangothi
Uvavanyo lwe-Pseudotyped virus neutralization lwenziwa ngokuyimfuneko njengoko besichazile ngaphambili12. Ngokufutshane, ukuhlanjululwa okulandelelanayo kweesampulu zeplasma (1: 4 ukuya ku-1: 8748) zalungiswa kwiDMEM kunye ne-10% ye-fetal bovine serum kwaye incubated nge-5 ng ye-p24 yentsholongwane nganye ye-pseudotyped ngexesha le-1h ku-37 °C kwaye emva koko, 1×10.4 yeeseli ze-HEK-ACE2 zongezwa kwiqula ngalinye. Iiseli ze-HEK293T (ezingayichaziyo i-ACE2) ezifukanywe ngentsholongwane eyipseudotyped (umnombo A) zasetyenziswa njengolawulo olubi. Iiseli zahlanjululwa kwiiyure ze-48 kamva, kwaye umsebenzi we-firefly luciferase walinganiswa usebenzisa i-Luciferase Assay Reagent (Promega) kwi-Glomax 96 Microplate luminometer (Promega). Ipesenti yokungathathi hlangothi kwi-dilution nganye ibalwe kwaye i-ID50 yesampuli nganye ibalwe kusetyenziswa i-GraphPad Prism version 9.0.1.
Ucatshulwa
Uhlalutyo lweenkcukacha-manani lwenziwa kusetyenziswa isoftware ye-GraphPad Prism version 9.1.2. Uthelekiso lwamaqela amaninzi lokunciphisa i-antibody titers (NAbTs) ngokuchasene nephaneli ye-SARS-CoV-2 pseudotyped virus kunye nothelekiso lweempendulo ze-NAbs ngokwesondo kunye nobume bomsi lwenziwa kusetyenziswa uvavanyo oludityanisiweyo lweWilcoxon olusayiniweyo. Ukutshintsha kwezinto kubalwa njengomahluko we-geometric mean titer kwi-ID50 xa kuthelekiswa naleyo Wild uhlobo pseudotyped virus. Uhlalutyo lokunxibelelana phakathi kwe-NAbTs kunye nobudala okanye i-BMI lwenziwa kusetyenziswa uvavanyo lukaSpearman. Uvavanyo lwendlela enye ye-ANOVA kunye neTukey yothelekiso oluninzi lwenziwa kuhlalutyo lwamanani osulelo. Ixabiso le-p ≤0.05 lithathwe njengelibalulekile ngokweenkcukacha-manani.
Ukuvunywa kokuziphatha
Iprothokholi yokufunda ivunyiwe yiKomiti yezokuziphatha ye-Faculty of Medicine e-Universidad de Chile (IiProjekthi N ° 0361-2021 kunye ne-N ° 096-2020) kunye ne-Clínica Santa Maria (iProjekthi N ° 132604-21). Bonke abaxhasi basayine imvume enolwazi, kwaye iisampulu zabo azichazwanga.
Impembelelo yoguqulo lwe-spike kukwahluka kweLambda ekosulelekeni kunye nokungathathi hlangothi kweempendulo ze-antibodies
Uhlalutyo lwe-3695 olulandelelanayo olusuka eChile lufakwe kwi-GISAID ngokwe-24 kaJunith Uwama-2021 ubonisa ukongamela okucacileyo kwee-SARS-CoV-2 ezahlukeneyo zeGamma kunye neLambda ngexesha lekota yokugqibela ye-accounting, kunye, kuma-79% azo zonke izilandelelwano.
Okubangel 'umdla kukuba, eli xesha liphawulwe liphulo elikhulu lokugonya apho ama-65.6% abantu ekujoliswe kubo (abaneminyaka eyi-18 nabadala) baye bafumana iskimu sokugonya esipheleleyo ngokomhla wama-27 kuJuni.th 2021
Ngenxa yokuba ama-78.2% abantu abatofelwe ngeskimu esipheleleyo bafumana isitofu sokugonya sentsholongwane esingasetyenziswayo i-CoronaVac evela kwi-Sinovac Biotech, siye sazama ukuphanda ifuthe lotshintsho lwe-spike olukhoyo kulwahlulo lwe-Lambda kumandla okunciphisa amajoni omzimba afunyanwe sesi sitofu sokugonya.
Kule nto, senze iintsholongwane ezisekelwe kwi-SARS-CoV-1 ezisekwe kwi-HIV-2-pseudotyped virus ezithwele iprotein ye-spike ukusuka kuluhlu lwesalathiso lwe-Wuhan-1 (Uhlobo lwasendle; umnombo A), ukuguqulwa kwe-D614G (umnombo B), kunye no-Alpha (umnombo B). .1.1.7), Gamma (umnombo P.1) kunye Lambda (umnombo C.37) ezahlukeneyo.
Ngexesha lokulungiselela intsholongwane, siye saqaphela ukuba iiseli ezosulelwe yintsholongwane eyipseudotyped ethwele i-Lambda spike zivelisa amaxabiso aphezulu e-bioluminescence xa kuthelekiswa ne-D614G eguquguqukayo okanye ukwahluka kwe-Alpha kunye ne-Gamma ebonisa ukosuleleka kokonyuka okuqhutywa yi-Lambda spike protein.
Umzobo 1.Ukosuleleka kulamlwa ziiproteni ezahlukeneyo zespike.
(A) Ukumelwa okucwangcisiweyo kwe-SARS-CoV-2 spike protein kunye nokwahluka okusetyenziswe kolu phononongo. Imigca yomgca ibonakaliswe kwizibiyeli. I-RBD, i-domain-binding domain, i-CM; umsila wecytoplasmic.
(B) I-Titration yee-pseudotypes zomnombo ngamnye kusetyenziswa izixa ezilinganayo ze-HIV-1 p24. Umsebenzi we-firefly luciferase ulinganiswe njengeeyunithi ze-luminescence ezihambelanayo (RLU) kwiiyure ze-48 emva kosulelo. I-avareji kunye ne-SD zibalwe kumfuniselo omele kathathu.
Emva koko, sasebenzisa iintsholongwane ze-pseudotyped ezikhankanywe ngasentla ukwenza iimvavanyo zokungathathi hlangothi zisebenzisa iisampuli ze-plasma ezingama-79 ezivela kubasebenzi bezempilo abasempilweni abavela e-Universidad de Chile naseClínica Santa María eSantiago, eChile.
Asibandakanyi iisampulu ezi-4 njengoko asikwazanga ukubala i-ID50 titer. Ukususela kwiisampuli ezihlalutyiweyo, i-73% ihambelana nabasetyhini, iminyaka yobudala be-34 iminyaka (IQR 29 - 43) kunye ne-index max index (BMI) ye-25 (IQR 22.7 - 27). Ama-20.5% abathathi-nxaxheba babhengeze ukuba bayatshaya ngelixa ixesha lokugonywa liqhubeka. Iisampulu zifunyenwe kumdibaniso weentsuku ezingama-95 (IQR 76 – 96) emva kwedosi yesibini yesitofu sokugonya seCoronaVac.
Siye saqaphela ukuba ukungathathi hlangothi kwentsholongwane eyipseudotyped ethwele i-Wild type spike protein ibangele i-50% inhibitory dilution (ID50) ithetha i-titer ye-191.46 (154.9 - 227.95, 95% CI,), ngelixa i-153.92 (115.68 - 192.16, 95% CI), 124.73 (86.2 - 163.2, 95% CI.104.57, 75.02% CI ) kunye ne-134.11 (95 - 78.75, 49.8% CI) kwiintsholongwane ze-pseudotyped ezithwala iprotheni ye-spike ukusuka kwi-D107.6G ye-mutant okanye i-Alpha, i-Gamma kunye ne-Lambda eyahlukileyo.
Siye saqaphela kwakhona ukuba i-geometric ithetha inqaku le-ID50 i-titers yehle nge-factor ye-3.05 (2.57 - 3.61, 95% CI) ye-virus ye-pseudotyped ephethe i-Lambda spike, 2.33 (1.95 - 2.80, 95% CI) ye-Gamma spike, 2.03 (1.71 - 2.41% 95) ye-Alpha spike kunye ne-1.37 (1.20 - 1.55, 95% CI) ye-D614G spike xa ithelekiswa ne-Wild type spike.
Akukho lunxulumano phakathi kwesondo, ubudala, isalathiso sobunzima bomzimba (BMI) okanye imeko yomsi kunye ne-antibody titers engathathi hlangothi yabonwa kwiqela lethu lophononongo.
Umzobo 2.Uvavanyo lokungathathi hlangothi kusetyenziswa iisampulu zeplasma ezivela kugonyo lweCoronaVac
(A) Utshintsho kwireciprocal 50% neutralization titer (ID50) kwiisampuli zeplasma ezivela kwi-75 abafumana isitofu sokugonya i-CoronaVac ngokuchasene ne-D614G (umnombo B), i-Alpha (umnombo B.1.1.7),
IGamma (umnombo P.1) kunye neLambda (umnombo C.37) ezahlukeneyo xa kuthelekiswa nentsholongwane yohlobo lwasendle. Iziphumo ziboniswa njengomahluko kwi-neutralization titers yeesampuli ezihambelanayo. Amaxabiso e-P okuthelekisa i-ID50 zibalwa kunye Wilcoxon sign-inqanaba uvavanyo.
(B) Iibhokisi zebhokisi zibonise uluhlu oluphakathi kunye ne-interquartile (IQR) ye-ID50 kwintsholongwane nganye echwetheziweyo. Utshintsho lwezinto luboniswa njengomahluko we-geometric mean titer kwi-ID50 xa kuthelekiswa nezo zeWild uhlobo pseudotyped virus. Uhlalutyo lweenkcukacha-manani lwenziwa kusetyenziswa uvavanyo lwe-Wilcoxon olumatanisiweyo-izibini ezisayiniweyo.
Ngokudibeneyo, idatha yethu ibonakalise ukuba iprotein ye-spike yokwahluka okusanda kuvunywa komdla weLambda, ejikeleza kakhulu kumazwe aseChile nakuMzantsi Melika, ithwala utshintsho olubonisa ukosuleleka kunye nokukwazi ukuphuncuka kwii-antibodies ezibangwe yiCoronaVac.
