scholarly journals Monitoring the Circulation of SARS-CoV-2 Variants by Genomic Analysis of Wastewater in Marseille, South-East France

Pathogens ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 1042
Author(s):  
Nathalie Wurtz ◽  
Océane Revol ◽  
Priscilla Jardot ◽  
Audrey Giraud-Gatineau ◽  
Linda Houhamdi ◽  
...  
Keyword(s):  
Genome Sequencing ◽  
Direct Sequencing ◽  
Genomic Analysis ◽  
Control Measures ◽  
Screening Tests ◽  
Rt Pcr ◽  
Pcr Screening ◽  
Small Areas ◽  
The Uk ◽  
Work Supports

The monitoring of SARS-CoV-2 RNA in sewage has been proposed as a simple and unbiased means of assessing epidemic evolution and the efficiency of the COVID-19 control measures. The past year has been marked by the emergence of variants that have led to a succession of epidemic waves. It thus appears that monitoring the presence of SARS-CoV-2 in wastewater alone is insufficient, and it may be important in the future to also monitor the evolution of these variants. We used a real-time RT-PCR screening test for variants in the wastewater of our city to assess the effectiveness of direct SARS-CoV-2 sequencing from the same wastewater. We compared the genome sequencing results obtained over the large RS network and the smaller B7 network with the different distributions of the variants observed by RT-PCR screening. The prevalence of the “UK variant” in the RS and B7 networks was estimated to be 70% and 8% using RT-PCR screening compared to 95% and 64% using genome sequencing, respectively. The latter values were close to the epidemiology observed in patients of the corresponding area, which were 91% and 58%, respectively. Genome sequencing in sewage identified SARS-CoV-2 of lineage B.1.525 in B7 at 27% (37% in patients), whereas it was completely missed by RT-PCR. We thus determined that direct sequencing makes it possible to observe, in wastewater, a distribution of the variants comparable to that revealed by genomic monitoring in patients and that this method is more accurate than RT-PCR. It also shows that, rather than a single large sample, it would be preferable to analyse several targeted samples if we want to more appropriately assess the geographical distribution of the different variants. In conclusion, this work supports the wider surveillance of SARS-CoV-2 variants in wastewater by genome sequencing and targeting small areas on the condition of having a sequencing capacity and, when this is not the case, to developing more precise screening tests based on the multiplexed detection of the mutations of interest.

Optimization of multiplex PCR composition to screen for SARS-CoV-2 variants of concern

2021 ◽  
Vol 4 (2) ◽  
pp. 58
Author(s):  
Maya Savira ◽  
Enikarmila Asni ◽  
Rahmat Azhari Kemal
Keyword(s):  
Whole Genome Sequencing ◽  
Multiplex Pcr ◽  
Genome Sequencing ◽  
Screening Method ◽  
Positive Control ◽  
Whole Genome ◽  
Rt Pcr ◽  
Pcr Screening ◽  
Single Target ◽  
Alpha Variant

Background: The ongoing COVID-19 pandemic has led to the emergence of several variants of concern. To rapidly identify those variants, screening samples for whole-genome sequencing (WGS) prioritization could be performed.  Objective: We optimized the polymerase chain reaction (PCR) screening method to identify the mutation in spike and ORF1a regions.  Methods: We adopted primers targeting mutation in spike and ORF1a region from another study. We optimized the PCR screening method using kits readily available in Indonesia. Firstly, we compared N1 and N2 primers as internal positive control. We also compared GoTaq® 1-Step RT-qPCR System and Indonesia TFRIC-19 BioCOV-19 for the multiplex reaction. We used the optimized composition to screen SARS-CoV-2 positive samples from April – June 2021. Samples with spike and/or ORF1a target failure were subjected to whole genome sequencing (WGS).  Results: The results demonstrated the N2 BioCOV-19 reaction as the optimized multiplex PCR composition for spike and ORF1a mutations screening. Whole-genome sequencing has shown that a sample with spike and ORF1a targets failure to be Alpha variant, while other samples with single target failure as non-variants of concern. Therefore, a multiplex RT-PCR composition has been optimized to detect mutation in spike and ORF1a regions. Conclusion: We have optimized a multiplex RT-PCR composition to detect mutation in spike and ORF1a regions.

scholarly journals Limitation of Screening of Different Variants of SARS-CoV-2 by RT-PCR

Diagnostics ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 1241
Author(s):  
Agathe Boudet ◽  
Robin Stephan ◽  
Sophie Bravo ◽  
Milène Sasso ◽  
Jean-Philippe Lavigne
Keyword(s):  
Next Generation Sequencing ◽  
Virus Evolution ◽  
Screening Strategy ◽  
Next Generation ◽  
Rt Pcr ◽  
Screening Assay ◽  
Pcr Screening ◽  
S Gene ◽  
The Uk ◽  
Generation Sequencing

Since January 2021, the diffusion of the most propagated SARS-CoV-2 variants in France (UK variant 20I/501Y.V1 (lineage B.1.1.7), 20H/H501Y.V2 (lineage B.1.351) and 20J/H501Y.V3 (lineage P.1)) were urgently screened, needing a surveillance with an RT-PCR screening assay. In this study, we evaluated one RT-PCR kit for this screening (ID SARS-CoV-2/UK/SA Variant Triplex®, ID Solutions, Grabels, France) on 2207 nasopharyngeal samples that were positive for SARS-CoV-2. Using ID Solutions kit, 4.1% (92/2207) of samples were suspected to belonged to B.1.351 or P.1 variants. Next-generation sequencing that was performed on 67.4% (62/92) of these samples confirmed the presence of a B.1.351 variant in only 75.8% of the samples (47/62). Thirteen samples belonged to the UK variant (B.1.1.7), and two to A.27 with N501Y mutation. The thirteen with the UK variant presented one mutation in the S-gene, near the ΔH69/ΔV70 deletion (S71F or A67S), which impacted the detection of ΔH69/ΔV70 deletion. Using another screening kit (PKampVariantDetect SARS-CoV-2 RT-PCR combination 1 and 3® PerkinElmer, Waltham, MA, USA) on the misidentified samples, we observed that the two mutations, S71F or A67S, did not impact the detection of the UK variant. In conclusion, this study highlights the limitations of the screening strategy based on the detection of few mutations/deletions as well as it not being able to follow the virus evolution.

scholarly journals Whole-genome sequencing of SARS-COV-2 reveals a substantially lower frequency of the UK variant than previously announced in Libya

2021 ◽  
Author(s):  
Inas M Alhudiri ◽  
Ahmad M Ramadan ◽  
Khaled Ibrahim Ibrahim ◽  
Mouna Eljilani ◽  
Adel Abdalla Aboud ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Drop Out ◽  
Whole Genome ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Gene Target ◽  
Specific Identification ◽  
S Gene ◽  
The Uk

A cluster-5 variant was detected in September 2020 in minks and humans in Denmark and currently classified as Alpha or B.1.1.7 strain. This variant presents several mutations in the spike region (S) which could increase the transmissibility of the virus 43-90% over previously circulating variants. The national center for disease control (NCDC) announced on 24th February 2021 the discovery of B.1.1.7 strain in Libya using a reverse-transcriptase PCR assay for S-gene target failure (SGTF) and reported that 25% of the tested samples were UK variant. This assay relies on the specific identification of the H69-V70 deletion in S gene which causes S gene drop out in RT-PCR; characteristic of the UK variant (B.1.1.7). This letter discusses our whole genome sequencing results of positive SARS-COV-2 samples with SGTF collected between 25th February - 4th March 2021 in Libya.

scholarly journals Surveillance testing for SARS-COV-2 infection in an asymptomatic athlete population: a prospective cohort study with 123 362 tests and 23 463 paired RT-PCR/antigen samples

2021 ◽  
Vol 7 (2) ◽  
pp. e001137
Author(s):  
Kimberly Harmon ◽  
Anabelle M de St Maurice ◽  
Adam C Brady ◽  
Sankar Swaminathan ◽  
Doug F Aukerman ◽  
...  
Keyword(s):  
Screening Programme ◽  
Sports Participation ◽  
High Specificity ◽  
Turnaround Time ◽  
Screening Tests ◽  
Rt Pcr ◽  
Pcr Screening ◽  
Testing Approach ◽  
Antigen Tests ◽  
Antigen Testing

ObjectiveTo assess the diagnostic accuracy of antigen compared with reverse transcriptase (RT)-PCR testing in an asymptomatic athlete screening programme and to monitor infection in college athletes.MethodsQuidel Sofia-2 SARS-CoV-2 Antigen Tests were performed daily before sports participation for football, basketball, wrestling and water polo from 29 September 2020 to 28 February 2021. Paired RT-PCR and antigen tests were performed at least once a week. Positive antigen tests were confirmed with RT-PCR.Results81 175 antigen and 42 187 RT-PCR tests were performed, including 23 462 weekly paired antigen/RT-PCR screening tests in 1931 athletes. One hundred and seventy-two athletes had a positive screening RT-PCR (0.4%), of which 83 (48%) occurred on paired testing days. The sensitivity of antigen tests varied with the frequency of RT-PCR testing and prevalence of COVID-19. The sensitivity of antigen testing was 35.7% (95% CI: 17% to 60%) and specificity 99.8% (95% CI: 99.7% to 99.9%) with once-a-week RT-PCR testing after adjusting for school prevalence. Daily antigen testing was similar to RT-PCR testing two to three times a week in identifying infection. Antigen testing identified infection before the next scheduled PCR on 89 occasions and resulted in 234 days where potentially infectious athletes were isolated before they would have been isolated with RT-PCR testing alone. Two athletic-related outbreaks occurred; 86% of total infections were community acquired.ConclusionAntigen testing has high specificity with a short turnaround time but is not as sensitive as RT-PCR. Daily antigen testing or RT-PCR testing two to three times a week is similar. There are benefits and drawbacks to each testing approach.

scholarly journals The risk of introducing SARS-CoV-2 to the UK via international travel in August 2020

2020 ◽  
Author(s):  
Rachel Taylor ◽  
Catherine A McCarthy ◽  
Virag Patel ◽  
Ruth Moir ◽  
Louise Kelly ◽  
...  
Keyword(s):  
International Travel ◽  
Control Measures ◽  
Rt Pcr ◽  
Additional Test ◽  
Disease Occurrence ◽  
Alternative Measures ◽  
Travel Policy ◽  
The Uk ◽  
The Impact

International travel poses substantial risks for continued introduction of SARS-CoV-2. As of the 17th August 2020, travellers from 12 of the top 25 countries flying into the UK are required to self-isolate for 14 days. We estimate that 895 (CI: 834-958) infectious travellers arrive in a single week, of which 87% (779, CI: 722-837) originate from countries on the UK quarantine list. We compare alternative measures to the 14 day self-isolation (78.0% effective CI: 74.4-81.6) which could be more feasible long-term. A single RT-PCR taken upon arrival at the airport is 39.6% (CI: 35.2-43.7) effective, or equivalently, it would only detect 2 in 5 infectious passengers. Alternatively, testing four days after arrival is 64.3% (CI: 60.0-68.3) effective whereas a test at the airport plus additional test four days later is 68.9% (CI: 64.9-73.0) effective. Rapidly implementing control measures for travellers from risky countries is vital to protect public health; this methodology can be quickly updated to assess the impact of any further changes to international travel policy or disease occurrence.

Retrospective analysis of whole genome sequencing compared to prospective typing data in further informing the epidemiological investigation of an outbreak of Shigella sonnei in the UK

2013 ◽  
Vol 141 (12) ◽  
pp. 2568-2575 ◽  
Author(s):  
J. McDONNELL ◽  
T. DALLMAN ◽  
S. ATKIN ◽  
D. A. TURBITT ◽  
T. R. CONNOR ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Shigella Sonnei ◽  
Control Measures ◽  
Whole Genome ◽  
Single Strain ◽  
Local Schools ◽  
Infection Control Measures ◽  
Typing Methods ◽  
The Uk

SUMMARYThe aim of this study was to retrospectively assess the value of whole genome sequencing (WGS) compared to conventional typing methods in the investigation and control of an outbreak of Shigella sonnei in the Orthodox Jewish (OJ) community in the UK. The genome sequence analysis showed that the strains implicated in the outbreak formed three phylogenetically distinct clusters. One cluster represented cases associated with recent exposure to a single strain, whereas the other two clusters represented related but distinct strains of S. sonnei circulating in the OJ community across the UK. The WGS data challenged the conclusions drawn during the initial outbreak investigation and allowed cases of dysentery to be implicated or ruled out of the outbreak that were previously misclassified. This study showed that the resolution achieved using WGS would have clearly defined the outbreak, thus facilitating the promotion of infection control measures within local schools and the dissemination of a stronger public health message to the community.

Monitoring the marine environment for small round structured viruses (SRSVS): a new approach to combating the transmission of these viruses by molluscan shellfish

1998 ◽  
Vol 38 (12) ◽  
pp. 51-56 ◽  
Author(s):  
K. Henshilwood ◽  
J. Green ◽  
D. N. Lees
Keyword(s):  
Enteric Virus ◽  
Winter Period ◽  
Rt Pcr ◽  
Cloning And Sequencing ◽  
New Approach ◽  
Human Enteric Virus ◽  
Different Strains ◽  
The Uk ◽  
Public Health Protection

This study investigates human enteric virus contamination of a shellfish harvesting area. Samples were analysed over a 14-month period for Small Round Structured Viruses (SRSVs) using a previously developed nested RT-PCR. A clear seasonal difference was observed with the largest numbers of positive samples obtained during the winter period (October to March). This data concurs with the known winter association of gastroenteric illness due to oyster consumption in the UK and also with the majority of the outbreaks associated with shellfish harvested from this area during the study period. RT-PCR positive amplicons were further characterised by cloning and sequencing. Sequence analysis of the positive samples identified eleven SRSV strains, of both Genogroup I and Genogroup II, occurring throughout the study period. Many shellfish samples contained a mixture of strains with a few samples containing up to three different strains with both Genogroups represented. The observed common occurrence of strain mixtures may have implications for the role of shellfish as a vector for dissemination of SRSV strains. These results show that nested RT-PCR can identify SRSV contamination in shellfish harvesting areas. Virus monitoring of shellfish harvesting areas by specialist laboratories using RT-PCR is a possible approach to combating the transmission of SRSVs by molluscan shellfish and could potentially offer significantly enhanced levels of public health protection.

scholarly journals Eastern Cape Healthcare Workers Acquisition of SARS-CoV-2 (ECHAS): Cross-Sectional (Nested Cohort) Study Protocol

2021 ◽  
Vol 18 (1) ◽  
pp. 323
Author(s):  
Oladele Vincent Adeniyi ◽  
David Stead ◽  
Mandisa Singata-Madliki ◽  
Joanne Batting ◽  
Leo Hyera ◽  
...  
Keyword(s):  
Cohort Study ◽  
Infection Rate ◽  
Healthcare Workers ◽  
Health Facilities ◽  
Control Measures ◽  
Infection Prevention And Control ◽  
Eastern Cape ◽  
Rt Pcr ◽  
Cross Sectional ◽  
Increased Risk

Healthcare workers (HCWs) are at increased risk of infection by the virulent severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Though data exist on the positivity rate of the SARS-CoV-2 reverse transcription polymerase chain reaction (RT-PCR) test as well as COVID-19-related deaths amongst HCWs in South Africa, the overall infection rate remains underestimated by these indicators. It is also unclear whether the humoral immune response after SARS-CoV-2 infection offers durable protection against reinfection. This study will assess the SARS-CoV-2 seroprevalence amongst HCWs in the Eastern Cape (EC) and examine the longitudinal changes (rate of decay) in the antibody levels after infection in this cohort. Using a multi-stage cluster sampling of healthcare workers in selected health facilities in the EC, a cross-sectional study of 2250 participants will be recruited. In order to assess the community infection rate, 750 antenatal women in the same settings will be recruited. Relevant demographic and clinical characteristics will be obtained by a self-administered questionnaire. A chemiluminescent microparticle immunoassay (CMIA) will be used for the qualitative detection of IgG antibodies against SARS-CoV-2 nucleocapsid protein. A nested cohort study will be conducted by performing eight-weekly antibody assays (X2) from 201 participants who tested positive for both SARS-CoV-2 RT-PCR and serology. Logistic regression models will be fitted to identify the independent risk factors for SARS-CoV-2 infection. The cumulative SARS-CoV-2 infection rate and infection fatality rate among the frontline HCWs will be estimated. In addition, the study will highlight the overall effectiveness of infection prevention and control measures (IPC) per exposure sites/wards at the selected health facilities. Findings will inform the South African Department of Health’s policies on how to protect HCWs better as the country prepares for the second wave of the SARS-CoV pandemic.

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