Since its first description in December 20201, the SARS-CoV-2 variant of concern VOC-202112/01 (also known as lineage B.1.1.7, 20I/501Y.V1 or, recently, according to the WHO, simply alpha2) has been spreading all over the world. In our geographical area, it became predominant from the beginning of March 2021, accounting for more than 90% of new infections in June 20213. This is mainly due to the mutations that this variant accumulates in the gene that encodes the spike (S gene), especially in the receptor-binding domain (RBD). These mutations, especially the N501Y mutation that it shares with the beta (B.1.351 or 20H/501Y.V2) and gamma (P.1 20J/501Y.V3) variants of concern, among others, are related to an increased binding affinity of the spike with angiotensin-converting enzyme II and an increase in transmissibility4. These last two variants also share the E484K mutation, also in the RBD, which could be related to a certain degree of escape from the action of the vaccines5. Therefore, when the first sequences of the B.1.1.7 lineage with the E484K mutation emerged, the British authorities declared them variants of concern VOC-202102/026. However, the cluster in which these variants were framed does not seem to have been as successful and represents only 0.225% of the sequences of the B.1.1.7 lineage included in the GISAID (Global Initiative on Sharing All Influenza Data).
In accordance with our centre’s variant screening protocol, all SARS-CoV-2 positive samples with Ct <32 are tested using the Allplex™ SARS-CoV-2 Variants I Assay kit (Seegene, Korea), which simultaneously detects H69/V70, E484K and N501Y mutations. At the end of April, we identified a sample which was positive for all three targets studied. In addition to the sample from the patient who had all three mutations, samples from the other three positive cases from the family cluster of which the patient had been a close contact were sequenced, presenting a profile compatible with the B.1.1.7 lineage but without the E484K mutation. For the sequencing of the viral genome, the Ion AmpliSeq SARS-CoV-2 Research Panel (Thermo Fisher Scientific, USA) was used7. Libraries were prepared following the manufacturer’s instructions and loaded onto a 540 chip and the Ion GeneStudio™ S5 (Thermo Fisher Scientific, USA) platform. The genome was assembled using the IRMA plugin8 and its consistency was checked using the Integrative Genomics Viewer (IGV) program9. The Nextstrain webApp10 was also used for both clade assignment and visualisation of mutations. The sequences obtained by next-generation sequencing confirmed the results of the PCR variant screening techniques. The test sample presented the G23012A mutation that conditions the amino acid change in the E484K S gene. None of the other three samples had that mutation in the assembly, and the A readings at position 23012 accounted for less than 1% of the readings in each of the samples. Except for the G23012A mutation, the four strains of the family cluster were identical and had both the characteristic mutations of the B.1.1.7 lineage and some of their own (see Table 1).
Coverage of the mutations present in the family cluster.
Test sample | Relative 1 | Relative 2 | Relative 3 | |
---|---|---|---|---|
C241T | 2935 | 7356 | 4892 | 3110 |
C913T | 5499 | 10533 | 7634 | 5110 |
C3037T | 6899 | 10817 | 8752 | 8597 |
C3267T | 1870 | 6997 | 4489 | 3645 |
C4464T | 7318 | 13962 | 8950 | 5797 |
A5041G | 3407 | 6980 | 4543 | 2567 |
C5388A | 4309 | 8448 | 5309 | 3474 |
G5763A | 7673 | 12951 | 9983 | 10079 |
C5986T | 2074 | 3458 | 1213 | 449 |
T6954C | 2822 | 7202 | 3136 | 933 |
11288-11297 | 6911 | 11107 | 8345 | 7157 |
C11668T | 5339 | 7078 | 3973 | 2795 |
C12439T | 6903 | 11349 | 7622 | 6920 |
C14407T | 4004 | 6209 | 3489 | 2521 |
C14408T | 4006 | 6230 | 3496 | 2525 |
C14676T | 5486 | 8911 | 6115 | 4265 |
T15096C | 3185 | 5267 | 3462 | 3199 |
C15279T | 1811 | 6655 | 4096 | 2838 |
T16176C | 6050 | 11721 | 8117 | 9837 |
C18647T | 2354 | 6501 | 4085 | 3055 |
21766-21772 | 5587 | 6261 | 5807 | 5072 |
21994-21997 | 7153 | 7985 | 6569 | 5075 |
G23012A | 8771 | |||
A23063T | 2137 | 5422 | 4300 | 1691 |
C23271A | 4375 | 7485 | 6071 | 5999 |
A23403G | 13281 | 16004 | 13387 | 13398 |
C23604A | 9736 | 12985 | 11754 | 14791 |
C23709T | 7735 | 11037 | 9594 | 7600 |
T24506G | 14234 | 17157 | 15581 | 13242 |
G24914C | 9398 | 14695 | 11772 | 7804 |
C27972T | 25717 | 28812 | 39283 | 27683 |
G28048T | 25829 | 28835 | 39267 | 27589 |
A28095T | 12218 | 16000 | 19763 | 6728 |
A28111G | 12192 | 15990 | 19727 | 6687 |
28274 | 9517 | 11135 | 18433 | 8315 |
G28280C | 10624 | 12112 | 19790 | 8916 |
A28281T | 10686 | 12175 | 19865 | 8933 |
T28282A | 10689 | 12181 | 19875 | 8936 |
C28320T | 10853 | 12294 | 20071 | 9036 |
G28881A | 6831 | 10521 | 12661 | 5721 |
8882A | 6840 | 10531 | 12673 | 5728 |
G28883C | 6841 | 10531 | 12673 | 5728 |
C28977T | 6715 | 10284 | 12336 | 5378 |
Despite belonging to the B.1.1.7 lineage and also exhibiting the E484K mutation, the sequence of our sample did not share the rest of the characteristic mutations of VOC-202102/02. The appearance of this mutation was therefore probably independent to those found in February 2021 in the United Kingdom, similar to other synchronous appearances of this mutation. In this case, the epidemiological chain of transmission is quite clear, since the infection of the patient with the E484K mutation appeared later: one week after the rest, and when the patient had been in isolation for six days for being a close contact of a confirmed case, so this mutation probably emerged de novo, either in the patient himself or in any of the other three members of the cluster after taking their respective samples. Fortunately, the patient had no subsequent close contacts and no new variants belonging to the B.1.1.7 lineage with the E484K mutation have been observed in the daily screenings carried out in our health organisation.
Please cite this article as: Urrutikoetxea-Gutierrez M, Ugalde Zarraga E, Gallego Rodrigo M, Díaz de Tuesta del Arco JL. Aparición de novo de la mutación E484K en una variante del linaje B.1.1.7 de SARS-CoV-2. Enferm Infecc Microbiol Clin. 2022;40:520–522.