S Gene Target Failure (SGTF) in Commercial Multiplex RT-PCR assay as indicator to detect SARS-CoV-2 VOC B.1.1.7 lineage in Tamil Nadu, India

Emergence of Severe Acute Respiratory Syndrome Corona Virus-2 (SARS-CoV-2) Variants of Concern (VOC) possessing improved virulence, transmissibility and/or immune-escape capabilities has raised significant public health concerns. In order to identify VOCs, WHO recommends Whole-Genome Sequencing approach, which is costly and involves longer completion time. Hence, potential role of commercial multiplex RT-PCR kit to screen variants rapidly is being attempted in this study. A total of 1200 suspected COVID samples from different districts of Tamil Nadu State (India) were screened with Thermo TaqPath RT-PCR kit and Altona Realstar RT-PCR Assay kit. Among 1200 screened, S-gene target failure (SGTF) phenomenon were identified in 112 samples while testing with TaqPath RT-PCR Kit. 100% concordant results were observed between SGTF phenomenon and whole-genome sequencing (WGS) results in detecting SARS-CoV-2 VOC B.1.1.7. TaqPath RT-PCR assay testing can be utilized by laboratories to screen rapidly the VOC B.1.1.7 variants, thus enabling early detection of B.1.1.7 variant infection and transmission in population. This in turn will pave way to implement suitable preventive measures by appropriate authorities to control the transmission of the viral variant.

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

10.1101/2021.07.03.21257942 ◽

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.

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Two-step strategy for the identification of SARS-CoV-2 variant of concern 202012/01 and other variants with spike deletion H69–V70, France, August to December 2020

Eurosurveillance ◽

10.2807/1560-7917.es.2021.26.3.2100008 ◽

2021 ◽

Vol 26 (3) ◽

Author(s):

Antonin Bal ◽

Gregory Destras ◽

Alexandre Gaymard ◽

Karl Stefic ◽

Julien Marlet ◽

...

Keyword(s):

Whole Genome ◽

Rt Pcr ◽

Pcr Assay ◽

Gene Target ◽

S Gene

We report the strategy leading to the first detection of variant of concern 202012/01 (VOC) in France (21 December 2020). First, the spike (S) deletion H69–V70 (ΔH69/ΔV70), identified in certain SARS-CoV-2 variants including VOC, is screened for. This deletion is associated with a S-gene target failure (SGTF) in the three-target RT-PCR assay (TaqPath kit). Subsequently, SGTF samples are whole genome sequenced. This approach revealed mutations co-occurring with ΔH69/ΔV70 including S:N501Y in the VOC.

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Two-step strategy for the identification of SARS-CoV-2 variants co-occurring with spike deletion H69-V70, Lyon, France, August to December 2020

10.1101/2020.11.10.20228528 ◽

2020 ◽

Cited By ~ 2

Author(s):

Antonin Bal ◽

Gregory Destras ◽

Alexandre Gaymard ◽

Hadrien Regue ◽

Quentin Semanas ◽

...

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Large Scale ◽

False Negative ◽

False Negative Result ◽

Whole Genome ◽

Rt Pcr ◽

Pcr Assay ◽

Large Scale Screening

AbstractThe spike deletion H69-V70 (ΔH69/ΔV70) has been recently detected in a SARS-CoV-2 variant under investigation in England (VUI 202012/01) as well as in cluter-5 variant detected both in minks and humans in Denmark. Herein we report the implementation of a two-step strategy enabling to detect SARS-CoV-2 variants carrying H69-V70 deletion. We found that this deletion resulted in a false negative result for the spike target of a three-target RT-PCR assay (TaqPath kit). From August 3rd to December 20th, 59/9,266 (0.6%) of positive tests displayed a S negative profile (negative for S target and positive for N & ORF1ab targets). Among the 59 samples without detection of the S target, 36 were available for whole genome sequencing (WGS). The most frequent S mutations co-occurring with ΔH69/ΔV70 were S477N & D614G (21/36 samples). The co-occurrence of N439K and D614G mutations was found in 10/36 samples. The complete combination of S mutations detected in VUI 202012/01 or in cluster-5 variant was not found. The data presented herein emphasize that the TaqPath RT-PCR assay enables a rapid, large-scale screening of ΔH69/ΔV70 variants. Samples with S negative profiles should be further addressed to national referral laboratories for SARS-CoV-2 WGS. This 2-step strategy is currently being reinforced in France as national diagnostic platforms have mainly implemented the TaqPath RT-PCR kit.

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False COVID-19 cases due to contamination by inactivated virus vaccine

Clinical Infectious Diseases ◽

10.1093/cid/ciab684 ◽

2021 ◽

Author(s):

Kelvin Kai-Wang To ◽

Xin Li ◽

David Christopher Lung ◽

Jonathan Daniel Ip ◽

Wan-Mui Chan ◽

...

Keyword(s):

Public Health ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Virus Vaccine ◽

False Positive ◽

Spike Protein ◽

Whole Genome ◽

Rt Pcr ◽

Health Measures ◽

Respiratory Specimens

Abstract A false-positive SARS-CoV-2 RT-PCR result can lead to unnecessary public-health measures. We report two individuals whose respiratory specimens were contaminated by inactivated SARS-CoV-2 vaccine strain(CoronaVac), likely at vaccination premises. Incidentally, whole-genome sequencing of CoronaVac showed adaptive deletions on the spike protein, which do not result in observable changes of antigenicity.

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RARE-26. WHOLE GENOME SEQUENCING OF AN OSSEOUS METASTASIS DURING CHECKPOINT-CONTROLLED INTRACRANIAL GLIOBLASTOMA REVEALS NEW INSIGHTS INTO POTENTIAL MECHANISMS OF IMMUNE ESCAPE

Neuro-Oncology ◽

10.1093/neuonc/noz175.949 ◽

2019 ◽

Vol 21 (Supplement_6) ◽

pp. vi227-vi227

Author(s):

Malte Mohme ◽

Cecile Maire ◽

Simon Schliffke ◽

Simon Joosse ◽

Malik Alawi ◽

...

Keyword(s):

T Cell ◽

Bone Metastasis ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Cell Response ◽

Immune Escape ◽

T Cell Response ◽

Molecular Profile ◽

Whole Genome ◽

Checkpoint Inhibition

Abstract Glioblastoma (GBM) has a devastating prognosis and recent advances in the treatment of a variety of cancer entities, e.g. through checkpoint inhibition, could so far not be translated into improved outcome in newly-diagnosed GBM. Characterizing rare cases of peripheral metastases which succeeded in overcoming immune control, can help to understand the mechanisms of immune escape. Here we describe the first reported case of a detailed genetic and immunological characterization of a peripheral bone metastasis from a GBM which was controlled intracranially by anti-PD1 checkpoint inhibition We performed whole genome sequencing (WGS) of the primary- and recurrent tumor, as well as the bone metastasis. Genomic data was analyzed for copy number variations and mutational profiles. In addition, immune monitoring with flow cytometric phenotyping and next-generation sequencing of the peripheral T-cell repertoire was used. A 70-year old patient developed multiple osseous metastases in the spine, while his IDHwt GBM recurrence was immunologically controlled with checkpoint inhibition. Biopsy confirmed peripheral GBM metastases. Immunophenotyping reflected the effective activation of the peripheral T-cell response, with, however, simultaneous upregulation of regulatory T-cells during disease progression. WGS sequencing demonstrated a distinct molecular profile of the GBM metastasis, with amplifications in chromosome 3 and 9, as well as genomic loss on chromosomes 4, 10 and 11. The peripheral metastasis was distinguished by mutations in mismatch repair genes, such as MSH4 and MLH1, associated with a hypermutated phenotype. Among the mutated genes we found potential candidates involved in immune escape of circulating tumor cells. This case represents a unique opportunity to analyze potential mechanisms of GBM-mediated immune escape during immune activation with anti-PD1 checkpoint therapy. It highlights the fact, that although an effective, disinhibited immune response can control the cranial GBM disease, hypermutated tumor clones can evade the tumor-specific T-cell response and disseminate to distant organs.

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First detection and report of SARS-CoV-2 Spike protein N501Y mutations in Oklahoma USA

10.1101/2021.01.26.21250584 ◽

2021 ◽

Author(s):

Sai Narayanan ◽

Girish Patil ◽

Sunil More ◽

Jeremiah Saliki ◽

Anil Kaul ◽

...

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Spike Protein ◽

Whole Genome ◽

S Gene ◽

Pathogen Dynamics ◽

Geographical Locations

AbstractWe describe the detection of SARS-CoV-2 (VOC)B.1.1.7 lineage in Oklahoma, USA. Various mutations in the S gene and ORF8 with similarity to the genome of B.1.1.7 lineage were detected in 4 of the 6 genomes sequenced and reported here. The sequences have been made available in GISAID. Presence of novel lineages indicate the need for frequent whole genome sequencing to better understand pathogen dynamics in different geographical locations.

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Reliability of Spike Gene Target Failure for ascertaining SARS-CoV-2 lineage B.1.1.7 prevalence in a hospital setting

10.1101/2021.04.12.21255084 ◽

2021 ◽

Author(s):

José Afonso Guerra-Assunção ◽

Paul A. Randell ◽

Florencia A. T. Boshier ◽

Michael A. Crone ◽

Juanita Pang ◽

...

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Lower Cost ◽

Hospital Setting ◽

Qpcr Assay ◽

Spike Protein ◽

Whole Genome ◽

Gene Target ◽

Spike Gene ◽

The Uk

AbstractThe appearance of the SARS-CoV-2 lineage B.1.1.7 in the UK in late 2020, associated with faster transmission, sparked the need to find effective ways to monitor its spread. The set of mutations that characterise this lineage include a deletion in position 69 and 70 of the spike protein, which is known to be associated with Spike Gene Target Failure (SGTF) in a commonly used three gene diagnostic qPCR assay. The lower cost and faster turnaround times compared to whole genome sequencing make the use of qPCR for monitoring of the variant spread an attractive proposition. However, there are several potential issues with this approach. Here we use 826 SARS-CoV-2 samples collected in a hospital setting as part of the Hospital Onset COVID Infection (HOCI) study where qPCR was used for viral detection, followed by whole genome sequencing (WGS), to identify the factors to consider when using SGTF to infer lineage B.1.1.7 prevalence in a hospital setting, with potential implications for locations where this variant has recently been introduced.

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Detection of SARS-CoV-2 from Patient Fecal Samples by Whole Genome Sequencing

10.21203/rs.3.rs-48863/v1 ◽

2020 ◽

Author(s):

Andreas Papoutsis ◽

Thomas Borody ◽

Siba Dolai ◽

Jordan Daniels ◽

Skylar Steinberg ◽

...

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Genome Analysis ◽

Nasopharyngeal Swab ◽

Whole Genome ◽

Rt Pcr ◽

Whole Genome Analysis ◽

Fecal Samples ◽

Oral Transmission ◽

Study Participants

Abstract Background SARS-CoV-2 has been detected not only in respiratory secretions, but also in stool collections. Here were sought to identify SARS-CoV-2 by enrichment NGS from fecal samples, and to utilize whole genome analysis to characterize SARS-CoV-2 mutational variations in COVID-19 patients. Results Study participants underwent testing for SARS-CoV-2 from fecal samples by whole genome enrichment NGS (n = 14), and RT-PCR nasopharyngeal swab analysis (n = 12). The concordance of SARS-CoV-2 detection by enrichment NGS from stools with RT-PCR nasopharyngeal analysis was 100%. Unique variants were identified in four patients, with a total of 33 different mutations among those in which SARS-CoV-2 was detected by whole genome enrichment NGS. Conclusion These results highlight the potential viability of SARS-CoV-2 in feces, its ongoing mutational accumulation, and its possible role in fecal-oral transmission. This study also elucidates the advantages of SARS-CoV-2 enrichment NGS, which may be a key methodology to document complete viral eradication.

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