key: cord-0850311-1n5750hi authors: Laiton-Donato, K.; Franco-Munoz, C.; Alvarez-Diaz, D. A.; Ruiz-Moreno, H.; Usme-Ciro, J.; Prada, D.; Reales, J.; Corchuelo, S.; Herrera-sepulveda, M.; Naizaque, J.; Santamaria, G.; Rivera, J.; Rojas, P.; Cardona Rios, A.; Hernandez-Ortiz, J.; Malo, D.; Prieto, F.; Ruiz-Gomez, F.; Wiesner, M.; Ospina-Martinez, M. L.; Mercado-Reyes, M. title: Characterization of the emerging B.1.621 variant of interest of SARS-CoV-2 date: 2021-05-10 journal: nan DOI: 10.1101/2021.05.08.21256619 sha: 5c40241edfdec1cb9a5a75097c53adaa00d6e089 doc_id: 850311 cord_uid: 1n5750hi The SARS-CoV-2 genetic diversification has a potential impact in the virus escape from natural infection- or vaccine-elicited neutralizing antibodies and higher transmissibility. Here we report the emergence of novel B.1.621 variant of interest with the insertion 145N in the N-terminal domain and amino acid change N501Y, E484K, and P681H in the Receptor Binding Domain of the Spike protein. Further studies in vitro biological assays and epidemiologic analysis will allow evaluating the public health impact of B.1.621 variant. In September 2020, SARS-CoV-2 variants of concern (VOC) and variants of interest (VOI) started to be reported, with more distinctive mutations than expected from the characteristic clock-like molecular evolution of this virus evidenced during the first year pandemic (1,2). Despite mutations spanning the whole genome, an interesting feature of these emerging variants has been the presence of several amino acid substitutions falling in the Spike protein, the viral protein responsible for receptor binding and membrane fusion and also the main target for neutralizing antibodies (3) . Monitoring the emergence of new variants of SARS-CoV-2 is a priority worldwide, as the presence of certain non-synonymous substitutions and INDELs could be related to biological properties, such as altering the ligand-receptor affinity, the efficiency of neutralization by naturally acquired polyclonal immunity or post-vaccination antibodies and transmission capacity (4) (5) (6) . In Colombia, the National Genomic Characterization Program led by the Instituto Nacional de Salud has carried out real-time monitoring of the SARS-CoV-2 lineages since the beginning of the pandemic (7, 8) . Until December 2020, over thirty lineages were circulating inside the country without evidence of VOC and VOI importation. However, a lineage turnover accompanied the third epidemic peak during March and April 2021, involving the emergence of B.1 lineage descendants with high mutation accumulation (B.1.621 and the provisionally assigned B.1+L249S+E484K) (9), as well as the introduction of the B.1.1.7, P.1 and VOI in some cities. In this study, we reported the emergence and spread of the novel B.1.621 lineage of SARS-CoV-2, a new VOI with the insertion 146N and several amino acid substitutions in the Spike protein (Y144T, Y145S, R346K, E484K, N501Y and P681H). Samples were selected from routine surveillance in all departments based on representativeness and virologic criteria (10) . Respiratory samples were used for automated RNA extraction with magnetic beads and the RNA extracts were processed by using the amplicon sequencing protocol v3 (https://www.protocols.io/view/ncov-2019-sequencing-protocol-v3-locost-bh42j8ye). The assembly of raw NGS data was performed by following the pipeline described for Oxford Nanopore Technologies (ONT) platform (https://artic.network/ncov-2019/ncov2019bioinformatics-sop.html). Lineage assignment started by filing a new issue in the pango-designation repository (https://github.com/cov-lineages/pango-designation/issues/57) followed by designation as B.1.621 lineage by the Pangolin curation team and PangoLEARN model training for subsequent automatic lineage assignment. Maximum likelihood tree reconstruction was performed with the GTR+F+I+G4 nucleotide substitution model using IQTREE. Branch support was estimated with an SH-like approximate likelihood ratio test (SH-aLRT). Recombination detection was performed using RDP4 software with RDP, GENECONV, Bottscan, Maxchi, Chimaera, SiSscan, and 3Seq tests (P-value < 0.05). Dataset 1 included Colombian SARS-CoV-2 sequences representative of the different lineages and dataset 2 included sequences previously reported as VOC or VOI. Adaptive evolution analysis at the codon level was estimated by Hyphy using stochastic evolutionary models. The detection of individual sites was performed with methods such as MEME (Mixed Effects Model of Evolution), and FEL (Fixed Effects Likelihood) (P-value <0.3). Lineage B.1.621 has become predominant in some departments of Colombia The routine genomic surveillance of SARS-CoV-2 in Colombia was reinforced in January 2021 for a higher sensitivity monitoring of the potential importation of VOC. By May 7, 2021, a total of 908 sequences from Colombia were available in the GISAID database. Lineage B.1 is the best-represented lineage (with 229 records) due to its higher frequency from the beginning of the pandemic. The recently designated B.1.621 lineage has been increasingly detected from January 11, 2021 (collection date of the first genome belonging to the lineage) to date (77 records), occupying the fifth place in frequency( Figure 1A ), rapidly becoming fixed in some departments located in the North of the country or co-circulating with others lineages in Bogotá D.C. ( Figure 1B ). The genetic background of the B.1.621 lineage includes some convergent amino acid changes previously found in several VOI and VOC. . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 10, 2021. The original assignment through the Pangolin algorithm for this monophyletic group was the B.1 lineage. However, a large number of distinctive synonymous and non-synonymous substitutions, including the following amino acid changes I95I, Y144T, Y145S in the N-terminal domain; R346K, E484K and N501Y in the Receptor Binding Domain and P681H in the S1/S2 cleavage site (Table 1) In Colombia, the current strategy for SARS-CoV-2 genomic surveillance includes sampling in principal and border cities, in special groups of interest, in patients with distinctive clinical features and severity, and finally in community transmission with an unusual increase in cases. . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 10, 2021. ; https://doi.org/10.1101/2021.05.08.21256619 doi: medRxiv preprint The high frequency of the emerging B.1.621 lineage also could be related to the strengthening of the SARS-COV-2 genomic surveillance during the third peak of the pandemic in Colombia, nevertheless, it is expected to characterize an approximate 1% of the cases and determine the adjusted frequency of the lineage in the country and to evaluate the possible predominance and the replacement of other lineages in the country. For this, intensified genomic characterization will be carried out with a multi-stage sample design throughout the national territory. During the first quarter of 2021, several convergent substitutions have been evidenced in the different lineage of SARS-CoV-2 explained by a high rate of the naive population that has explored wide options of genetic variability, the other scenario is explained by the selection pressure by monoclonal antibody therapies (11, 12) and vaccination (13, 14) . The distinctive substitutions observed in the spike protein are common, despite the characteristics attributed to some substitutions, for instance, the presence of E484K has been associated with lower neutralizing activity from convalescent plasma (12) . The 69/70 deletion spike together with the E484K and N501Y substitutions decrease the ability to neutralize antibodies (15) . The insertion N-terminal domain of S1 subunit . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 10, 2021. P681H S1/S2 cleavage region . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 10, 2021. ; https://doi.org/10.1101/2021.05.08.21256619 doi: medRxiv preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 10, 2021. ; https://doi.org/10.1101/2021.05.08.21256619 doi: medRxiv preprint red RENATA and Universidad Industrial de Santander for the workstation bioinformatic support. No conflict of interest was reported by the authors. This work was funded by the Project CEMIN-4-2020 Instituto Nacional de Salud. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. SARS-CoV-2 Colombian sequences belonging to the B.1.621 were deposited in GISAID under accession numbers: EPI_ISL_1220045, EPI_ISL_1582980, EPI_ISL_1424054, EPI_ISL_1424056, EPI_ISL_1582978, EPI_ISL_1820926, EPI_ISL_1424055, EPI_ISL_1582993, EPI_ISL_1582979, EPI_ISL_1424057, EPI_ISL_1820929, EPI_ISL_1820930, EPI_ISL_1820932, EPI_ISL_1820927, EPI_ISL_1424058, EPI_ISL_1582984, EPI_ISL_1582986, EPI_ISL_1582991, EPI_ISL_1820958, EPI_ISL_1582990, EPI_ISL_1582992, EPI_ISL_1582981, EPI_ISL_1582994, EPI_ISL_1582988, EPI_ISL_1820959, EPI_ISL_1820928, EPI_ISL_1582996, EPI_ISL_1632530, EPI_ISL_1582989, EPI_ISL_1820934, EPI_ISL_1582987, EPI_ISL_1820955, EPI_ISL_1820935, EPI_ISL_1582997, EPI_ISL_1820925, EPI_ISL_1820950, EPI_ISL_1820949, EPI_ISL_1820944, EPI_ISL_1820947, EPI_ISL_1820943, EPI_ISL_1820946, EPI_ISL_1820954, EPI_ISL_1821882, EPI_ISL_1820939, EPI_ISL_1820942, EPI_ISL_1820960, EPI_ISL_1820936, EPI_ISL_1820948, EPI_ISL_1820931, EPI_ISL_1582977, EPI_ISL_1820965, EPI_ISL_1821070, EPI_ISL_1821075, EPI_ISL_1821063, EPI_ISL_1820967, EPI_ISL_1820968, EPI_ISL_1821062, EPI_ISL_1821064, EPI_ISL_1821069, EPI_ISL_1821073, EPI_ISL_1821074, EPI_ISL_1821076, EPI_ISL_1821077, EPI_ISL_1821071, EPI_ISL_1821072, EPI_ISL_1820962, EPI_ISL_1821065, . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted May 10, 2021. . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted May 10, 2021. ; https://doi.org/10.1101/2021.05.08.21256619 doi: medRxiv preprint . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 10, 2021. ; https://doi.org/10.1101/2021.05.08.21256619 doi: medRxiv preprint New SARS-CoV-2 Variants -Clinical, Public Health, and Vaccine Implications Comprehensive mapping of mutations in the SARS-CoV-2 receptor-binding domain that affect recognition by polyclonal human plasma antibodies The impact of Spike mutations on SARS-CoV-2 neutralization 1 Immunological considerations for COVID-19 vaccine strategies Estimated transmissibility and impact of SARS-CoV-2 lineage B.1.1.7 in England Genomic epidemiology of severe acute respiratory syndrome coronavirus 2, Colombia. Emerg Infect Dis Noticias coronavirus-genoma Novel highly divergent SARS-CoV-2 lineage with the Spike substitutions L249S and E484K. medRxiv pre-print Guidance for SARS-CoV-2 samples selection for genomic characterization and surveillance -PAHO/WHO | Pan American Health Organization SARS-CoV-2 501Y.V2 escapes neutralization by South African COVID-19 donor plasma Identification of SARS-CoV-2 spike mutations that attenuate monoclonal and serum antibody neutralization mRNA vaccine-elicited antibodies to SARS-CoV-2 and circulating variants Multiple SARS-CoV-2 variants escape neutralization by vaccine-induced humoral immunity Neutralization of SARS-CoV-2 spike 69/70 deletion, E484K and N501Y variants by BNT162b2 vaccine-elicited sera Two-step strategy for the identification of SARS-CoV-2 variant of concern 202012/01 and other variants with The authors thank the National Laboratory Network for routine virologic surveillance of SARS-CoV-2 in Colombia. We also thank all researchers who deposited genomes in GISAID's EpiCoV Database contributing to genomic diversity and phylogenetic relationship of SARS-CoV-2. We thank Rotary International and Charlie Rut Castro for equipment's donation. Finally, we thank