Respiratory Viruses
SENTINELLE Newsletter
ReViLux Bulletin hebdomadaire
03 February 2021
Semaine: 4
Résultats cliniques de la sentinelle
La surveillance SENTINELLE des infections respiratoires aiguës est une collaboration entre le Laboratoire national de santé (LNS), la Direction de la Santé et un réseau de médecins-généralistes et de pédiatres répartis à travers tout le Luxembourg. Ce rapport hebdomadaire est publié par le départment de microbiologie du LNS dans le cadre de ses missions de surveillance, de vigilance et d’alerte dans les domaines de la santé publique, l'analyse des données rapportant l’activité clinique dans la Sentinelle et des données virologiques à l’appui.
Au cours de la semaine 4 (25/01/2021 - 31/01/2021), l'activité grippale reste faible.
Le taux de consultations avec ILI1 est de 0% (seuil épidémiologique à 3%).
Le taux de consultations avec ARI2 est de 13%.
- ILI: - Influenza-Like Illness: Signes respiratoires aigus <10 jours, Fièvre 38∘C, Signes systémiques (myalgie, malaise, maux de tête, …)
- ARI: - Infection Respiratoire Aiguë: Signes respiratoires aigus (bronchite, pharyngite, rhinite, pneumonie…) avec ou sans fièvre
| Number of Patients | Percentage | |
|---|---|---|
| ARI (acute respiratory infections) | 48 | 13% |
| ILI (influenza-like illness) | 1 | 0% |
| Consultations | 361 |
Respiratory viruses circulating in Luxembourg
| results | |
|---|---|
| Cumulative number of tests performed | 179 |
| Cumulative number of positive samples | 56 |
| Positivity rate | 31% |
Patients presenting with ILI and ARI at the Covid Consultation Centre (CCC) in Luxembourg were tested using a respiratory virus panel (ADV = Adenovirus, FLU A = Influenza A, FLU B = Influenza B, HRV = Human Rhinovirus, MPV = Human metapneumovirus, PIV = Parainfluenza virus, RSV = Respiratory Syncytial Virus, SARS-CoV-2 = Severe acute respiratory syndrome coronavirus 2). SARS-CoV-2 remains the dominant circulating respiratory virus in Luxembourg, while the Human Rhinovirus (HRV) wave continue circulating in Luxembourg.
SARS-CoV-2 Sequencing
Sequenced Samples
The national genomic surveillance program had maintained its capacity of 384 samples/week and reached since March a total of 4142 SARS-CoV-2 consensus sequences from clinical samples as of week 3/2021. While the respiratory virus surveillance can address real-time data, the sequencing data have a delay of one week. This delay in reporting is represented by the technical process for sequencing samples upon reception to our premises which ranges 5-7 days. For data presentation purposes we use a window of 5 months so the plotted data are starting on 01/09/2020.
Last week, the platform sequenced 240 clinical samples. However, only 93 samples refer to week 3/2021, which represents 11% of new infections reported in Luxembourg for that week. The remaining samples are awaiting metadata (e.g. sampling date) to be integrated into the data analysis. The frequency of variants reported in ReViLux is indicative of samples referred to LNS for sequencing with potential selection bias (e.g. requests by Inspection Sanitaire, positive samples form airport testing and cluster samples), with only low numbers of samples with full metadata.
The current reported frequency of the variants should not be extrapolated to prevalence of these variants in the community, due to the emergence of new variants and to selection bias of sequenced samples. Among the 93 samples sequenced by the platform with a sampling date referring to week 3/2021 and covering 11% of new infections, more than 50% represent targeted sequencing on request by Inspection Sanitaire (e.g. contact tracing). These samples are therefore not representative for the general population.
In collaboration with Inspection Sanitaire, we are developing population surveillance to sequence 300 SARS-CoV-2 samples from different regions and age groups to offer a clear reflection of the circulation of different variants in Luxembourg on the basis of a data collection framework from partner laboratories. This will allow us to reach real-time epidemiology with accurate estimates of the frequency of circulating variants by end of February.
Detected Lineages
Lineages have been assigned based on Rambaut et al by means of pangolin (v2.1.8, pangoLEARN version 2021-01-22), allowing for a lineage (=variant) assignment of circulating viral genomes.
Including data already reported last week for the sampling time frame week 3/2021 (18th-24th January) 15 circulating SARS-CoV-2 variants have been detected within our sequencing pool. The most prevalent lineage is B.1.160 (44,1%), followed by B.1.1.7 (16,1%). Position three is shared by B.1 and B.1.177 (8,6% each). The remaining variants, not considering variants of concern, mainly represent singular cases. B.1.160, B.1 and B.1.221 represent big and widely spread European Cluster.
Variants of Concern (B.1.1.7, B.1.351, P.1)
Since the introduction of B.1.1.7 variant in Luxembourg on 19/12/2021, we have sequenced 791 clinical samples. By week 3/2021, the total number of detected B.1.1.7 cases is set to 47 (6%), while the case count for B.1.351 (South African variant B.1.351) if 4 (0.5%). No cases have been detected for the Japanese/Brazilian variant P.1 in Luxembourg by now.
Including data already reported in the previous newsletter, 15 cases of B.1.1.7 have been detected referring to week 3/2021, representing 16.1% of strains in the sequencing pool (in comparison to week 2/2020 15,4%). The earliest sampling date remains 19/12/2020 and the latest is 24/01/2021.
For the sampling time frame of week 3/2021, 2 cases of the South African variant B.1.351 have been detected, taking the case count by week 3/2021 to 4. The earliest sampling date remains 11/01/2021 and the latest is 22/01/2021.
Lineage B.1.1.7 is characterized by several spike protein mutations including S:N501Y, deletion S:H69/V70 and S:P861H. The variant seems to have a considerable epidemiological impact, as it has a higher transmissibility (more efficient and rapid).
Lineage B.1.351 holds numerous spike protein mutations, wherein three are located in the receptor binding domain (K417N, E484K and N501Y), which is relevant for antibody binding. As for B.1.1.7, a higher transmissibility and viral load is in discussion. Due to K417N and E484K an impact on vaccination and possible reinfection is subject of scientific investigation.
Lineage P.1 (descendent of B.1.1.28) initially been found in the Amazon region has a similar mutation profile as the South African variant, including E484K and N501Y. Concerns are, as for the South African variant, higher transmissibility and a decreased protection by neutralizing antibodies.
It is to note, that additional samples are in preparation for sequencing, so that numbers might change with increasing representativeness of circulating strains and proportions.
Phylogenetic Analysis
Phylogenetic analysis has been done by means of nextstrain4 and is based on genetic distance including a timeline on the x-axis. The tree is rooted versus the first sequence obtained in Luxembourg (29/02/2020). Sequences obtained before 01/11/2020 are in grey and those obtained within the last 2 weeks from most recent sampling date referring to week3/2021 (24th January) are represented in colour and by greater diameter. Due to the increasing number of sequences represented, we performed a random subsampling of 300 sequences over the complete sequencing time frame since 01/09/2020, adding on top the sequences representing B.1.1.7 and B.1.351 referring to week 3/2021.
The tree shows four visually well separated clusters. These clusters are represented by the lineages B.1.160, B.1.177, B.1.1.7 and B.1.221. Due to random selection of sequences and the explicit insertion of variants of concern, the size of clusters might be misleading and does not necessarily correlate with the prevalence of the respective variant.
Lineage B.1.1.7 (represented by green dots) forms a very well distinguished cluster, due to its high amount of accumulated mutations. B.1.1.7 subtrees might direct to different subgroups, possibly related to different sources of infection.
The cases for B.1.351 (light brownish dots, close to the middle of the tree) are as well separating well from common circulating strains.
- Nextstrain applied an additional filter on genome length > 27000, 316 sequences shown
Clinically relevant mutations
Currently genomic surveillance is - independently from lineage call - observing the occurrence of 13 different known SARS-Cov-2 mutations, being assumed to have a clinical or epidemiological relevance. The pool of observed mutations is updated continuously, depending on the development of SARS-CoV-2 variants and their prevalence.
Due to the increasing incidence of the variant B.1.1.7, the characterizing mutations N501Y, H69/V70del, Y144del and P681H are integrated in weekly reporting. Additionally, the mutation E484K is under observation, which is in combination with N501Y and K417N characteristic for the South African and Japanese/Brazilian variant.
The following table provides the overall frequency of these mutations in lineage assignable genomes between 01/09/2020 and 24/01/2021 (N=3323), as well as a close-up of the last 14 days. As mentioned above, due to increased targeted sequencing of samples on request of health authorities, the frequency of mutations within the last 14 days is not representative for the general population.
| Mutation | Gene | Genomic position in reference | Frequency overall | Frequency recent 14days | Characteristics | Reference |
|---|---|---|---|---|---|---|
| D614G | S gene | 23402 | 98.0% | 96.0% | Higher infectivity, higher case fatality rate, higher transmission; replaced original Wuhan strain, became globally dominant form of the virus | Eaaswarkhanth, al Madhoun, and Al-Mulla 2020, Becerra-Flores and Cardozo 2020, Hu et al. 2020, Plante et al. 2020 |
| E484K | S gene | 23012 | 0.0% | 6.0% | 501Y.V2 / possible impact on antibody neutralization activity (escape mutation), improved ACE2 binding affinity | Greaney et al, 2020 |
| G204R | N gene | 28883 | 8.0% | 27.0% | Fitness advantage for the virus | Leary et al. 2020 |
| H69/V70del | S gene | 21765-21770 | 4.0% | 16.0% | possible impact on antibody neutralization activity and reinfection; included in "mink" mutation | Kemp et al 2020, Kemp et al 2020 |
| K417N | S gene | 22813 | 0.0% | 3.0% | 501Y.V2 / possible impact on antibody binding affinity (escape mutation) | Kemp et al 2020 |
| L37F | Nsp6 | 11081 | 8.0% | 7.0% | Favored viral infection, higher severity | Aiewsakun et al. 2020 |
| N439K | S gene | 26143 | 3.0% | 0.0% | Improved ACE2 binding affinity | Zhou et al, 2020 |
| N501Y | S gene | 23063 | 2.0% | 19.0% | 501Y.V1/V2; Improved ACE2 binding affinity/higher transmissibility | Filip Fratev 2020 COVID-19 Genomics Consortium UK, 2020 |
| P323L | ORF1ab (RdRP gene, nsp12) | 14407 | 94.0% | 88.0% | Higher severity | Biswas and Mudi 2020 |
| P681H | S gene | 23604 | 5.0% | 20.0% | immediately adjacent to the furin cleavage site, a known location of biological significance | COVID-19 Genomics Consortium UK, 2020 |
| Q57H | ORF3a | 25561 | 45.0% | 55.0% | Higher severity | Biswas and Mudi 2020 |
| R203K | N gene | 28880 | 8.0% | 27.0% | Fitness advantage for the virus | Leary et al. 2020 |
| Y144del | S gene | 21991-21993 | 2.0% | 18.0% | possible impact on antibody binding affinity | Dawood et al 2020 |
Clinically relevant mutations frequencies over time
The D614G and P323L mutations have been and are continuously present in a vast majority of sequences. They mainly occur together.
In comparison to the previous week the frequency within the last 14 days increased for S:N501Y from 15% to 19%, for S:H69/V70 deletion from 13% to 16%, for S:P681H from 19% to 20%, for S:Y144del from 14% to 18%, for K417N from 2% to 3% and for E484K from 3% to 6%.
Additional Information
For more information on lineages visit: https://cov-lineages.org
For more information and statistics on Covid-19 infections in Luxembourg visit: https://covid19.public.lu/en.html
References
Rambaut, A., Holmes, E.C., O’Toole, Á. et al. A dynamic nomenclature proposal for SARS-CoV-2 lineages to assist genomic epidemiology. Nat Microbiol 5, 1403–1407 (2020). https://doi.org/10.1038/s41564-020-0770-5
Pangolin (pangolin assigns lineages to query sequences as described in Rambaut et al 2020); Áine O’Toole, JT McCrone, Emily Scher; github.com/cov-lineages/pangolin
Hadfield et al., Nextstrain: real-time tracking of pathogen evolution, Bioinformatics (2018)
Guangchuang Yu, David Smith, Huachen Zhu, Yi Guan, Tommy Tsan-Yuk Lam. ggtree: an R package for visualization and annotation of phylogenetic trees with their covariates and other associated data. Methods in Ecology and Evolution 8(1) 28-36 (2017). https://doi.org/10.1111/2041-210X.12628