The aim of the “Sentinel” national surveillance program is to monitor the circulating respiratory viruses, including SARS-CoV-2 variants, and hence underpin public health actions.
In Week 8/2021, the overall frequency of the SARS-CoV-2 B.1.1.7 variant, in all sequenced samples, increased to 64.4% (CI 59,4% – 69,4%, p<0,05). For the SARS-CoV-2 B.1.351 variant, we found an overall frequency of 16.4% (CI 12,5% – 20,3%, p<0,05).
The representative sample size was estimated, based on the number of positive cases in Luxembourg for week 8 (1315). The minimum sample size required to detect prevalence of B.1.1.7 (56%) with an error margin of 5% was estimated to be 295 specimens. This number corresponds to a coverage of 22.4%, which exceeds the minimum coverage recommended by ECDC (10%). Hence our sequencing results this week are representative of the circulating variants in Luxembourg.
The total number of sequences performed this week was 570, with 354 samples collected in the time frame of week 8/2021. The sequencing coverage this week was 27% of all positive cases in Luxembourg.
At present, the scope of the ReViLux report is to provide (i) surveillance and descriptive epidemiology data, including data on molecular phylogenies (identification of importation events, changes in outbreak size over time), and (ii) phylogenetic interpretation of regional virus spread and circulation in Luxembourg, and cluster investigation. The scope of the ReViLux report is not to attempt phylogeographical reconstructions in Luxembourg, as a whole.
The “Sentinel” surveillance network reported 210 consultations in week 8 (22/FEB/2021 – 28/FEB/2021). There was one case of ILI1, representing 0.47% of the consultations (epidemiological threshold is 2 new cases per week), as shown in Figure 1. The percentage of consultations for ARI2 was 7,6%.
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). The SAR-COV-2 was the most prevalent respiratopry virus detected in the “Sentinel” network, with 39 positive cases (25%). The wave of Human Rhinovirus (HRV) continued in week 8/2021, with 19 positive cases in 156 tests (12.2%). Three cases of PIV were identified in week 8. No cases of Influenza A/B were detected, indicating absence of circulation of influenza viruses in Luxembourg, as shown in Figure 2.
In Luxembourg, we have tested 156 samples from the “Sentinel” surveillance network, as compared to 1045 samples tested in Europe, in the week 08/2021. Three of these 1045 specimens tested for influenza viruses were positive. The influenza epidemic in the European Region has usually reached its peak by this point of the year but, despite widespread and regular testing for influenza, reported influenza activity still remains at a very low level, likely due to the impact of the various public health and social measures, implemented to reduce transmission of SARS-CoV-2 (Source: FluNews Europe).
The National Reference Laboratory for Acute Respiratory Infections at LNS continues to improve the representativeness of the pool of sequenced specimens to reach real-time epidemiology, by implementing the following weekly sequencing activities:
The representative sequencing sample size calculation was based on the minimum number of specimens required to extrapolate prevalence of VOC variants with error rate of 5%. The representative sample size was estimated based on the number of positive cases in Luxembourg for week 8 (1315). The minimum sample size required to detect prevalence of B.1.1.7 (56%) with an error margin of 5% was estimated to be 295 specimens. The calculation was based on a sample size calculation tool that uses the expected prevalence of the variant in the population of interest. (Population Proportion – Sample Size – Select Statistical Consultants (select-statistics.co.uk). This number represented a coverage of 22.4% which exceeds the minimum coverage recommended by ECDC (10%). Hence our sequencing results this week are representative of the circulating variants in Luxembourg.
The starting material used for sequencing is respiratory specimens (nasopharyngeal or oropharyngeal swabs) that have already been tested positive by RT PCR.
The LNS sequencing data sharing strategy includes sharing of the sequencing data with GSAID EpiCov database (www.gsaid.org ) on a periodic basis.
Last week the microbial genomics platform at the LNS sequenced 570 specimens. Out of these, 354 specimens were collected in week 8/2021. This represents 27% of the new infections reported in Luxembourg in week 8/2021. Among the 354 specimens, no specimens were reported to be part of a cluster investigation.
The population sequencing coverage in week 8/2021 was 27%. Based on statistical inference, the frequency of the reported variants in Week 8/2021 is representative of the circulating variants in Luxembourg.
In the weekly population sequencing pool from representative regions, 17 specimens were collected from non-residents. This leads to 337 specimens, collected in Week 8, and being the representative population sequencing sample.
Lineages (variants) have been assigned based on Rambaut et al by means of Phylogenetic Assignment of Named Global Outbeak LINeages (pangolin) software (v2.3.3, pangoLEARN version 2021-02-21).
The lineage nomenclature system that we use is the one proposed by Rambaut et al. that focuses on actively circulating virus lineages (https://cov-lineages.org ).
In the sampling period of week 8/2021, 17 circulating SARS-CoV-2 variants were detected within our representative sequencing pool, as shown in Figure 3. The most prevalent lineage was B.1.1.7 (65.5%, CI 60,4% – 70,6%), followed by B.1.351 (16%, CI 12,1% – 19,9%) and B.1.1.29 (6%, CI 3,4% – 8,4%).
Among specimens collected within the week 8/2021, 221 cases of the B.1.1.7 variant have been detected, representing 65.5% of the specimens in the week’s representative sample (by comparison, the week 7/2021 had shown 53.7% of this variant). The total case count of sequenced variant B.1.1.7 was 813 by week 8/2021. The earliest collection date for this variant remains 19/DEC/2020 and the latest is 28/FEB/2021.
In the collection period of week 8/2021, 54 cases of the South African variant B.1.351 have been detected, representing 16% of the specimens in the week’s sequencing pool (by comparison, the week 7/2021 had shown 14.8% of this variant). The total case count of sequenced variant B.1.351 was 159 by week 8/2021. The earliest collection date for this variant remains 11/JAN/2021 and the latest is 28/FEB/2021.
In week 8/2021, one case of the Brazilian variant P.1 was detected, therefore the total case count of variant P.1 was 2 by week 8/2021 (collection date 02/FEB/2021 and 26/FEB/2021).
The pangolin group, which provides a global report on novel coronavirus haplotypes, published two additional variants of concern, assigned as A.23.1 and B.1.525. Both variants have already been reported by neighboring countries (Belgium, France). Retrospectively, one case of the B.1.525 variant has been detected for the week 7/2021 (sampling date 20/FEB/2021).
Lineage B.1.1.7 is characterized by several spike protein mutations, including N501Y, H69/V70del and P861H. The variant seems to have a considerable epidemiological impact, as it has a higher transmissibility rate.
Lineage B.1.351 holds numerous spike protein mutations, of which three are located in the receptor binding domain (K417N, E484K and N501Y), and are therefore relevant for antibody binding. As for B.1.1.7, a higher transmissibility rate and viral loads seem to be associated with this variant. Due to the K417N and E484K mutations, an impact on vaccination efficacy and possibility of reinfection is subject to scientific investigation.
Lineage P.1 (descendent of B.1.1.28), initially 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.
Lineage B.1.525 carries several mutations of biological significance, including E484K, Q677H and F888L. It does not carry N501Y, but a set of deletions similar to the B.1.1.7 variant.
Of note, additional specimens are in the pipeline for sequencing, so that numbers may change with increasing representativeness of circulating variants and proportions.
Currently the LNS genomic surveillance program – independently from lineage calling – notes the occurrence of 13 different known SARS-CoV-2 mutations, assumed to have clinical and epidemiological relevance. The list of observed mutations is being updated continually, based on the appearance and prevalence of SARS-CoV-2 variants.
The following table provides the overall frequencies of these mutations, detected in the lineage-assignable genome sequences, analyzed between 01/SEP/2020 and 28/FEB/2021 (N=5098), as well as the mutation frequencies in week 8/2021.
The ReViLux data are communicated as support to the understanding of respiratory virus transmission dynamics, including introduction of new variants, to the evaluation of the impact of response measures, to the contact tracing and the investigation of clusters.
Genomic sequencing of SARS-CoV-2. A guide to implementation for maximum impact on public health. WHO, 8 January 2021.
COVID-19 data portal. 2020 (https://www.covid19dataportal.org/sequences )
J Hadfield et al. Nextstrain: real-time tracking of pathogen evolution. Bioinformatics 2018;34:4121-4123
A Rambaut et al. A dynamic nomenclature proposal for SARS-CoV-2 lineages to assist genomic epidemiology. Nat Microbiol 2020;5:1403-1407
Y Guangchuang et al. ggtree: an R package for visualization and annotation of phylogenetic trees with their covariates and other associated data. Methods in Ecology and Evolution 2017;8:28-36
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