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 21/2021, the overall frequency of the SARS-CoV-2 B.1.1.7 variant in all sequenced specimens decreased to 66% (CI 59,6% – 72,4%, p<0,05). For the SARS-CoV-2 B.1.351 and P.1 variant, we found for each an overall frequency of 2,8% (CI 0,6% – 5%, p<0,05) within the sequenced sample. The frequency detected for the variant B.1.617.2 doubled to 14.6% (CI 9,8% – 19,3%, p<0,05).
The representative sample was estimated, based on the number of positive cases in Luxembourg for week 21 (354). The minimum sample size required to detect prevalence of B.1.1.7 (76%) reported in week 20, with an error margin of 5%, was estimated to be 157 specimens. This number corresponds to a coverage of 44.3 %, which exceeds the minimum coverage recommended by ECDC (10%). The sequencing results of week 16 are representative of the circulating variants in Luxembourg with a margin of error of 5%.
The total number of sequences performed this week was 293, with 212 specimens having been collected in the time frame of week 21/2021. The sequencing coverage this week was 55,4% from all positive cases in Luxembourg (based on sequenced samples collected in the time frame of week 21 and corresponding to Luxembourg residents).
The “Sentinel” surveillance network reported 187 consultations in week 21 (24/MAY/2021 – 30/MAY/2021). There were 2 cases of ILI1, corresponding to 1,1% of the consultations, as shown in Figure 1. The number of consultations for ARI2 was 34, which represents 18.2% of the consultations.
Covid Consultation Centres have been closed by 16/May/2021. We are currently working on an alternative in partnership with private laboratories.
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 was based on the minimum number of specimens required to extrapolate prevalence of VOC variants with error rate of 5%. The representative sample was estimated based on the number of positive cases in Luxembourg in week 21 (354). The minimum sample size required to detect prevalence of B.1.1.7 (76%) reported in week 20 with an error margin of 5% was estimated to be 157 specimens. The calculation was based on a sample size calculation tool that uses the expected prevalence of the variant in the total population. (Population Proportion – Sample Size – Select Statistical Consultants (select-statistics.co.uk). This number represented a coverage of 44,3% which exceeds the minimum coverage recommended by ECDC (10%). The number of non-targeted specimens from Luxembourgish residents sequenced this week was 196. Therefore, 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.
In parallel, we will introduced the PCR screening for VOC on 14/06/2021, using the Allplex™ SARS-CoV-2 Variants I Assay which will allow simultaneous detection and differentiation of SARS-CoV-2 and three mutation sites in S gene for identification of SARS-CoV-2 variants to detect the Variants of Concern (VOCs. This will enable a faster investigation time of any outbreak and allow us to identify variants of concerns that do not pass the quality criteria for sequencing.
The LNS sequencing data sharing strategy includes sharing of the sequencing data with GISAID EpiCov database (www.gisaid.org ) on a periodic basis.
Last week the microbial genomics platform at the LNS sequenced 293 specimens, with 212 collected in week 21/2021. The sequencing pool referring to Luxembourgish residents represents 55,4% of new infections reported in Luxembourg in week 21/2021. Among these 212 specimens, 12 specimens were reported to be part of a cluster or outbreak investigation, and 15 specimens were from non-residents (5 specimen overlapping). This leads to 190 specimens, collected in week 21, and being the representative population sequencing sample. In the population representative sample of residents, the frequencies of B.1.1.7, B.1.617.2, B.1.351 and P.1 were 65,3%, 16,3%, 3,2% and 2,1% respectively.
The population sequencing coverage in week 21/2021 was 55,4% (Figure 2). Based on statistical inference, the frequency of the reported variants in week 21/2021 is representative of the circulating variants in Luxembourg with a margin of error of 5%.
Lineages (variants) have been assigned based on Rambaut et al by means of Phylogenetic Assignment of Named Global Outbeak LINeages (pangolin) software (v3.0.3, pangoLEARN version 2021-05-27).
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 week 21/2021, in the population representative sample, after removal of cluster specimens, and excluding specimens collected from non-residents, there were 13 variants, with the main three variants being B.1.1.7 (65,3%, CI 58,5% – 72,1%), followed by B.1.617.2 (16,3%, CI 11% – 21,5%) and P.1.1 / B.1.1 (3,7% each , CI 1% – 6,4%), as shown in Figure 3.
Among specimens collected within the week 21/2021, 140 cases of the B.1.1.7 variant have been detected, representing 66% of the specimens in the week’s sequencing pool (by comparison, the week 20/2021 pool had shown a frequency of 76,1% of this variant, including additional specimens having been sequenced from previous weeks). The total case count of sequenced variant B.1.1.7 was 6053 by week 21/2021. The earliest collection date for this variant remains 19/DEC/2020 and the latest is 30/05/2021.
In the collection period of week 21/2021, 6 cases of the South African variant B.1.351 have been detected, representing 2,8% of the specimens in the week’s sequencing pool (by comparison, the week 20/2021 pool had shown 2,5% of this variant, including additional specimens having been sequenced from previous weeks). The total case count of sequenced variant B.1.351 was 1031 by week 21/2021. The earliest collection date for this variant remains 11/JAN/2021 and the latest is 28/MAY/2021.
In week 21/2021, no new case of B.1.525 has been detected, and 6 new cases of the Brazilian variant P.1. The case count by week 21 for B.1.525 remains 45 (latest sampling date 17/MAY/2021) and for P.1 the case count increases to 111 (latest sampling date 30/MAY/2021)
In week 21/2021, 31 additional cases of the Delta variant B.1.617.2 have been detected (latest sampling date 30/MAY/2021). The case count by week 21 for B.1.617.1 remains 6, and for B.1.617.2 it raises to 85. Since May 6th, B.1.617.2 has been escalated by Public Health England from a variant under investigation to a variant of concern (Figure 4).
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.
Lineage B.1.617 is a variant first detected in India and was designated “Under Investigation” on 1st April 2021 by Public Health England. It contains a number of spike mutations associated with antigenic escape or found in other variants of concern, including L452R, E484Q and P681R. Subtype B.1.617.2 does not carry S:E484Q and seems to be at least as transmissible as B.1.1.7 (increasing confidence). Neutralization studies show reductions in cross-neutralizing activity between B.1.1.7 and B.1.351.
By week 21/2021, 86,3 % of the variants detected in the sequenced specimen pool are declared as either Variants of Concern (VOC – B.1.1.7, B.1.351, P.1, B.1.617.2) or Variants Under Investigation (VUI – B.1.525, B.1.1.318, B.1.617.1, B.1.1).
The World Health Organization (WHO) has recently announced a new nomeculature system for tracking SARS-CoV-2 variants of interest (VOI) and variants of concern (VOC). The system is based on the Greek alphabet.
According to the new system, B.1.1.7 was the first VOC designated by WHO and it is now called Alpha. The B.1.351, which originated in Brazil, is now called Beta. The two other VOCs are the P.1, the variant first identified in Brazil and now referred to as Gamma, and the B.1.617.2 that originated in India, now called Delta (Figure 5).
The ReViLux will continue to use the Rambaut et al. (“Pango”) nomenclature system to enable linking between any evolving variants and their ancestor.
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 30/MAY/2021 (N=12080), as well as the frequencies in week 21/2021.
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
https://github.com/cov-lineages/pangolin
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