Delgado, SoledadSomovilla, PilarFerrer-Orta, CristinaMartínez-González, BrendaVázquez-Monteagudo, SergiMuñoz-Flores, JavierSoria, Maria EugeniaGarcía-Crespo, Carlosde Ávila, Ana IsabelDurán-Pastor, AntoniGadea, IgnacioLopez-Galindez, Luis CecilioMorán, FedericoLorenzo-Redondo, RamonVerdaguer, NuriaPerales, CeliaDomingo, Esteban2024-03-202024-03-202024-03-05Proc Natl Acad Sci USA. 2024 Mar 5;121(10):e2317851121.http://hdl.handle.net/20.500.12105/19015Since its introduction in the human population, SARS-CoV-2 has evolved into multiple clades, but the events in its intrahost diversification are not well understood. Here, we compare three-dimensional (3D) self-organized neural haplotype maps (SOMs) of SARS-CoV-2 from thirty individual nasopharyngeal diagnostic samples obtained within a 19-day interval in Madrid (Spain), at the time of transition between clades 19 and 20. SOMs have been trained with the haplotype repertoire present in the mutant spectra of the nsp12- and spike (S)-coding regions. Each SOM consisted of a dominant neuron (displaying the maximum frequency), surrounded by a low-frequency neuron cloud. The sequence of the master (dominant) neuron was either identical to that of the reference Wuhan-Hu-1 genome or differed from it at one nucleotide position. Six different deviant haplotype sequences were identified among the master neurons. Some of the substitutions in the neural clouds affected critical sites of the nsp12-nsp8-nsp7 polymerase complex and resulted in altered kinetics of RNA synthesis in an in vitro primer extension assay. Thus, the analysis has identified mutations that are relevant to modification of viral RNA synthesis, present in the mutant clouds of SARS-CoV-2 quasispecies. These mutations most likely occurred during intrahost diversification in several COVID-19 patients, during an initial stage of the pandemic, and within a brief time period.engVoRhttp://creativecommons.org/licenses/by-nc-nd/4.0/COVID-19Clade transitionViral quasispeciesSelf-organized mapsIntrahost evolutionSARS-CoV-2COVID-19HumansHaplotypesViral Nonstructural ProteinsRNA, ViralAttribution-NonCommercial-NoDerivatives 4.0 Internacional3841668412110e231785112110.1073/pnas.23178511211091-6490Proceedings of the National Academy of Sciences of the United States of Americaopen access