Two-Period Study Results from a Large Italian Hospital Laboratory Attesting SARS-CoV-2 Variant PCR Assay Evolution

Liotti, Flora Marzia; Maio, Flavio De; Ippoliti, Chiara; Santarelli, Giulia; Monzo, Francesca; Sali, Michela; Santangelo, Rosaria; Ceccherini‐Silberstein, Francesca; Sanguinetti, Maurizio

doi:10.1128/spectrum.02922-22

Cited by 5 publications

(5 citation statements)

References 21 publications

(39 reference statements)

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“…Finally, RT-PCR kits focus on a select group of mutations that identify a variant using an interpretative algorithm. Additional information like lineage, genomic diversity, and genotyping must be obtained by WGS ( 21 ).…”

Section: Discussionmentioning

confidence: 99%

Frequency of SARS-CoV-2 variants identified by real-time PCR in the AUNA healthcare network, Peru

Ortiz-Gómez,

Gomez,

Chuima

et al. 2024

Front. Public Health

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IntroductionIn Peru, on 11 February 2023, the Ministry of Health registered 4 million patients infected with COVID-19 and around 219,260 deaths. In 2020, the SARS-CoV-2 virus was acquiring mutations that impacted the properties of transmissibility, infectivity, and immune evasion, leading to new lineages. In the present study, the frequency of COVID-19 variants was determined during 2021 and 2022 in patients treated in the AUNA healthcare network.MethodsThe methodology used to detect mutations and identify variants was the Allplex™ SARS-CoV-2 Variants Assay I, II, and VII kit RT-PCR. The frequency of variants was presented by epidemiological weeks.ResultsIn total, 544 positive samples were evaluated, where the Delta, Omicron, and Gamma variants were identified. The Delta variant was found in 242 (44.5%) patients between epidemiological weeks 39 and 52 in 2021. In the case of Gamma, it was observed in 8 (1.5%) patients at weeks 39, 41, 43, 45, and 46 of 2021. The Omicron variant was the most frequent with 289 (53.1%) patients during weeks 49 to 52 of 2021 and 1 to 22 of 2022. During weeks 1 through 22 of 2022, it was possible to discriminate between BA. 1 (n = 32) and BA.2 (n = 82).ConclusionThe rapid identification of COVID-19 variants through the RT-PCR methodology contributes to timely epidemiological surveillance, as well as appropriate patient management.

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Section: Discussionmentioning

confidence: 99%

Frequency of SARS-CoV-2 variants identified by real-time PCR in the AUNA healthcare network, Peru

Ortiz-Gómez,

Gomez,

Chuima

et al. 2024

Front. Public Health

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“…Although the Simplexa® SARS-CoV-2 Variants Direct assay can contribute to detecting known, circulating variants, it should be emphasized that the assay cannot totally replace the sequencing process, since such PCR-based methods are neither suitable for the detection of new variants nor able to replace the role of sequencing in detecting the emergence of new variants ( Berno et al, 2022 ). This is a weakness that generally characterizes such assays for the detection of SARS-CoV-2 variants, and as also stated by Liotti et al (2022) the original version of those assays cannot be helpful as the SARS-CoV-2 pandemic progresses. Therefore, constant updates and adjustments based on sequencing are required, either by modifying the reagents or by providing new data for the interpretation of the melting curve patterns with the appearance of new variants, so that the usage of such assays is meaningful.…”

Section: Discussionmentioning

confidence: 99%

Rapid identification of SARS-CoV-2 variants: Validation of the simplexa SARS-CoV-2 variant direct assay

Eptaminitaki,

Parakatselaki,

Petroulaki

et al. 2023

Journal of Virological Methods

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“…Our approach of using Sanger sequencing lends itself to better utility in resource limited settings. Other approaches for genomics surveillance, including the commercially available Seegene Allplex SARS-Cov_2 Variants I (E484K, N501Y and ΔH69/V70) and II (L452R, W152C, K417T, and K417N) assays, have opted to sequence the entire S-gene to identify VOC ( Liotti et al., 2022 ). Clearly, whilst the approach to identifying SNPs using qPCR assays is not unique, the context in which these were developed must be considered.…”

Section: Discussionmentioning

confidence: 99%

Development of primer-probe sets to rapidly distinguish single nucleotide polymorphisms in SARS-CoV-2 lineages

Ealand,

Gordhan,

Machowski

et al. 2023

Front. Cell. Infect. Microbiol.

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Ongoing SARS-CoV-2 infections are driven by the emergence of various variants, with differential propensities to escape immune containment. Single nucleotide polymorphisms (SNPs) in the RNA genome result in altered protein structures and when these changes occur in the S-gene, encoding the spike protein, the ability of the virus to penetrate host cells to initiate an infection can be significantly altered. As a result, vaccine efficacy and prior immunity may be diminished, potentially leading to new waves of infection. Early detection of SARS-CoV-2 variants using a rapid and scalable approach will be paramount for continued monitoring of new infections. In this study, we developed minor groove-binding (MGB) probe-based qPCR assays targeted to specific SNPs in the S-gene, which are present in variants of concern (VOC), namely the E484K, N501Y, G446S and D405N mutations. A total of 95 archived SARS-CoV-2 positive clinical specimens collected in Johannesburg, South Africa between February 2021 and March 2022 were assessed using these qPCR assays. To independently confirm SNP detection, Sanger sequencing of the relevant region in the S-gene were performed. Where a PCR product could be generated and sequenced, qPCR assays were 100% concordant highlighting the robustness of the approach. These assays, and the approach described, offer the opportunity for easy detection and scaling of targeted detection of variant-defining SNPs in the clinical setting.

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Two-Period Study Results from a Large Italian Hospital Laboratory Attesting SARS-CoV-2 Variant PCR Assay Evolution

Cited by 5 publications

References 21 publications

Frequency of SARS-CoV-2 variants identified by real-time PCR in the AUNA healthcare network, Peru

Frequency of SARS-CoV-2 variants identified by real-time PCR in the AUNA healthcare network, Peru

Rapid identification of SARS-CoV-2 variants: Validation of the simplexa SARS-CoV-2 variant direct assay

Development of primer-probe sets to rapidly distinguish single nucleotide polymorphisms in SARS-CoV-2 lineages

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