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Thirteen exceptional mutations that explain certain properties of Omicron…

Some virological data concerning Omicron Article
published in French on February 3, 2022 in the Vidal site under the title
Bons baisers d’Omicron : quelles nouvelles de ce variant très atypique ?

Contributor: Dr Daniel Maller 


Published date: 06 February 2022


Thirteen exceptional mutations that explain certain properties of Omicron…

The previously mentioned Omicron mutation interaction study by Martin DP et al.(1) , deserves further investigation. This team focused its attention on 13 of the 30 mutations present in the gene for the Spike protein of Omicron, mutations hitherto exceptionally encountered in other sarbecoviruses, even in the numerous ones present in bats. Some of these thirteen mutations had never been observed before among the millions of SARS-CoV-2 genomes sequenced during the pandemic.


These 13 mutations are not scattered randomly across Spike. They form three groups (clusters), each modifying a small part of the protein and playing an important role in what makes Omicron unique.


  • Clusters 1 and 2 modify the end of the S1 subunit of Spike and would increase its affinity with the ACE2 receptor, thus improving the contagiousness of Omicron. In addition, these modifications make class 1, 2 and 4 neutralizing antibodies resulting from infection or vaccination less effective, which contributes to the relative immunoresistance of Omicron.

  • Cluster 3 modifies Spike near its base at its "fusion domain", i.e. the part of the S2 subunit that allows the virus to deliver its genes inside the target cell. This part of Spike is usually conserved (unmutated) in coronaviruses of the SARS-CoV-2 family, at least those that use ACE2 to penetrate target cells. Cluster 3 mutations dramatically change the way Omicron infects target cells (see below).


Taken separately, these mutations, when observed, represented a hindrance to the vitality of SARS-CoV-2 and were rapidly eliminated by selective pressure. But together, through a phenomenon of synergy operating both within and between the clusters (called “positive epistasis”), they have created a series of advantages for Omicron, including its greater contagiousness and its relative immunoresistance. (1)


...and sign an unusual origin for this new variant


The authors of this study(1) are also trying to better understand the conditions of appearance of Omicron. Indeed, there are now three hypotheses for this sudden appearance of a variant whose closest ancestor had been observed more than a year earlier (see article of 2 December 2021)(2):


  1. Insufficient genomic surveillance of variants in an isolated region where Omicron could have been built gradually in a stealthy manner;

  2. a slow evolution within an immunocompromised and chronically infected patient, as has been sometimes reported(3), including recently(4);

  3. an evolution in an animal intermediate host infected by humans (a “reverse zoonosis” towards a “reservoir species”), in which the mutations would have accumulated and which would then have infected humans with this novel variant.


Martin DP et al.(1) , the simultaneous appearance, from the first Omicron sequencing, of three subvariants (BA.1, BA.2 and BA.3) seems to favor the hypothesis of insufficient local genomic surveillance, at least in the months preceding the identification of Omicron. Nevertheless, they recognize that the modifications of clusters 1 and 2, by significantly increasing the affinity of Omicron for the ACE2 receptors of various animal species (mouse, rat, hen, turkey or horseshoe bat)(5) pours water on the mill of the inverted zoonosis hypothesis.


To decide between the hypotheses, it would be necessary, for example, to identify the three sub-variants in an animal or a patient, thus making possible the last two hypotheses. A mixture of hypotheses is also possible (for example, appearance of the ancestor of the sub-variants in a patient or an animal, then appearance of BA.1, BA.2 and BA.3 in various patients infected by this ancestor). The Scientific Advisory Group on the Origins of Novel Pathogens (SAGO), recently created by the World Health Organization, is expected to publish a report in early February 2022 on the origins of Omicron.


A mode of cellular invasion modified compared to the previous variants…


Structural analysis of Omicron's Spike protein, as well as various culture studies of this variant in vitro, show that Omicron is fundamentally different from other known variants in its mode of infection of target cells. The modifications of the S2 subunit by cluster 3 explain this radical change, with strong clinical consequences. As a reminder, there are two modes of intracellular penetration for SARS-CoV:

  • the cleavage of S1 and S2 by the transmembrane serine protease 2 (TMPRSS2, on the membrane of the target cell), followed by the fusion of the viral membrane with the cell membrane, thus releasing the viral RNA into the cytoplasm. This mode of infection is the one favored by all the dominant variants of SARSCoV-2 previously described.

  • capture of the virus in a “bubble” of cell membrane (an “endosome”) which then penetrates the cell. This endosome is degraded there by proteases, the cathepsins (and, in the case of coronaviruses, in particular the cathepsins L). This mode of infection is possible, but incidental for the SARS-CoV-2 variants studied before the appearance of Omicron.


Several culture studies of Omicron in vitro (e.g., Peacock TP et al.(6) or Willett BJ et al.)(7), using selective inhibitors of each of these two modes of infection, have shown that the route of entry preferred by Omicron is that involving the formation of endosomes and their digestion by cathepsins L.


... and which has clinical and therapeutic consequences


Why is this information on how Omicron is infected, clinically important? First of all, this mode of infection allows Omicron to overcome the presence of TMPRSS2 on the target cell: the mere presence of the ACE2 receptor is sufficient. Unlike the other variants, it does not show a selective tropism for cells carrying TMPRSS2. However, this membrane protease is particularly present on the cells of the pulmonary alveoli, taste buds, heart, digestive tract, brain,(8) etc., all organs strongly targeted by the previous variants of SARSCoV-2.


In fact, two in vitro studies and two ex vivo studies (Willett BJ et al.(7) and Meng B et al.(9) Chan MCW et al.(10) and Huy KPJ et al.(11)) have provided elements suggesting a lower infectivity of Omicron for cells pulmonary. Thus, Omicron's indifference to TMPRSS2 could contribute to explaining its lower pathogenicity, in particular regarding organs rich in this protease.


But this does not explain everything. In the mucous membranes of the nasopharynx, cells carrying only ACE2 receptors are seven times more numerous than those carrying ACE2 and TMPRSS2(12): there is therefore still a vast field of action for Omicron which, in vitro, multiplies a hundred times faster than Delta in a mixture of nasal cells(12). Under these conditions, how to explain the reduction in viral load in the nasopharynx observed in vivo with Omicron (see above)?


In fact, Omicron's preference for the endosomal pathway leads to an unfavorable consequence for its multiplication, a consequence which could counterbalance its affinity for cells bearing ACE2. Among the factors of innate immunity (non-specific, unlike acquired immunity) are cellular proteins, interferon-induced transmembrane proteins (IFITM), which have the ability to prevent endosomes from releasing their viral content. In variants like Alpha or Delta, penetration through TMPRSS2 bypasses this defense mechanism. Omicron, given its endosomal mode of infection, is itself sensitive to these immune proteins(12), which could explain its lower virulence and lower viral load in the nasopharynx.


Another possible biological basis for the lower pathogenicity of Omicron, the endosomal mode of penetration, unlike that using TMPRSS2, only weakly causes the formation of syncitia, these fusions of infected cells and viruses which favor the contamination of neighboring cells. In the context of viral respiratory infections, and in particular COVID-19, the formation of syncitia seems particularly marked during severe pulmonary forms. Two in vitro studies showed that Omicron was ten times less "fusiogenic" than Delta(12) (see also Suzuki R et al.(13)) Thus, it is possible that the switch from TMPRSS2 mode to endosomal mode is accompanied by a reduction in lesions tissue(12), especially since Omicron seems to infect alveolar cells only slightly.


  1. Selection analysis identifies unusual clustered mutational changes in Omicron lineage BA.1 that likely impact Spike function

  2. Omicron : un variant en marche ?

  3. Persistent SARS-CoV-2 infection and intra-host evolution in association with advanced HIV infection


  4. Persistent SARS-CoV-2 Infection with Accumulation of Mutations in a Patient with Poorly Controlled HIV Infection

  5. Evidence for a mouse origin of the SARS-CoV-2 Omicron variant

  6. The SARS-CoV-2 variant, Omicron, shows rapid replication in human primary nasal epithelial cultures and efficiently uses the endosomal route of entry.

  7. The hyper-transmissible SARS-CoV-2 Omicron variant exhibits significant antigenic change, vaccine escape and a switch in cell entry mechanism.

  8. ACE2, TMPRSS2 distribution and extrapulmonary organ injury in patients with COVID-19

  9. Altered TMPRSS2 usage by SARS-CoV-2 Omicron impacts tropism and fusogenicity


  10. SARS-CoV-2 Omicron variant replication in human respiratory tract ex vivo


  11. SARS-CoV-2 Omicron variant replication in human bronchus and lung ex vivo


  12. The SARS-CoV-2 variant, Omicron, shows rapid replication in human primary nasal epithelial cultures and efficiently uses the endosomal route of entry.

  13. Attenuated fusogenicity and pathogenicity of SARS-CoV-2 Omicron variant

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