Digital PCR Discriminates between SARS-CoV-2 Omicron Variants and Immune Escape Mutations

Holland, Steven C.; Holland, LaRinda A; Smith, Matthew F.; Lee, Mihyun B; Hu, James C; Lim, Efrem S.

doi:10.1128/spectrum.05258-22

Cited by 4 publications

(3 citation statements)

References 34 publications

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“…Similarly, its reliability has been demonstrated in the detection of both SARS-CoV-2 as a whole and of SARS-CoV-2 viral mutants that may be associated with increased antiviral resistance or atypical behaviour from the pathogenic or diagnostic point of view [6,26,27]. Thus, some studies have shown that dPCR would detect between 2.2 and 3.7 copies/reaction [27] and, compared to RT-qPCR, would increase the positive rate by 12.0% in samples from SARS-CoV-2 infected patients [27].…”

Section: Virology Applicationsmentioning

confidence: 97%

Present and Future Applications of Digital PCR in Infectious Diseases Diagnosis

Sancha Dominguez,

Cotos Suárez,

Sánchez Ledesma

et al. 2024

Diagnostics

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Infectious diseases account for about 3 million deaths per year. The advent of molecular techniques has led to an enormous improvement in their diagnosis, both in terms of sensitivity and specificity and in terms of the speed with which a clinically useful result can be obtained. Digital PCR, or 3rd generation PCR, is based on a series of technical modifications that result in more sensitive techniques, more resistant to the action of inhibitors and capable of direct quantification without the need for standard curves. This review presents the main applications that have been developed for the diagnosis of viral, bacterial, and parasitic infections and the potential prospects for the clinical use of this technology.

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Section: Virology Applicationsmentioning

confidence: 97%

Present and Future Applications of Digital PCR in Infectious Diseases Diagnosis

Sancha Dominguez,

Cotos Suárez,

Sánchez Ledesma

et al. 2024

Diagnostics

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“…Diagnostic screening nucleic acid amplification technique (NAAT)-based assays are used as an alternative method that enables the early detection and pre-screening of VOCs. Recently, PCR-based studies have been published to address the diagnosis of SARS-CoV-2 variants [ 7 , 8 , 9 , 10 , 11 ]. Holand et al showed that the use of digital PCR will enable the need for a personalized diagnosis aimed at individual patient treatment [ 8 ], and Vogels et al aimed to reduce false negative results by multiplexed RT-PCR, developing a set of dedicated primers for variants sharing common mutations [ 12 ].…”

Section: Introductionmentioning

confidence: 99%

“…Recently, PCR-based studies have been published to address the diagnosis of SARS-CoV-2 variants [7][8][9][10][11]. Holand et al showed that the use of digital PCR will enable the need for a personalized diagnosis aimed at individual patient treatment [8], and Vogels et al aimed to reduce false negative results by multiplexed RT-PCR, developing a set of dedicated primers for variants sharing common mutations [12].…”

Section: Introductionmentioning

confidence: 99%

Discriminative Identification of SARS-CoV-2 Variants Based on Mass-Spectrometry Analysis

Feldberg,

Zvi,

Yahalom-Ronen

et al. 2023

Biomedicines

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The spread of SARS-CoV-2 variants of concern (VOCs) is of great importance since genetic changes may increase transmissibility, disease severity and reduce vaccine effectiveness. Moreover, these changes may lead to failure of diagnostic measures. Therefore, variant-specific diagnostic methods are essential. To date, genetic sequencing is the gold-standard method to discriminate between variants. However, it is time-consuming (taking several days) and expensive. Therefore, the development of rapid diagnostic methods for SARS-CoV-2 in accordance with its genetic modification is of great importance. In this study we introduce a Mass Spectrometry (MS)-based methodology for the diagnosis of SARS-CoV-2 in propagated in cell-culture. This methodology enables the universal identification of SARS-CoV-2, as well as variant-specific discrimination. The universal identification of SARS-CoV-2 is based on conserved markers shared by all variants, while the identification of specific variants relies on variant-specific markers. Determining a specific set of peptides for a given variant consists of a multistep procedure, starting with an in-silico search for variant-specific tryptic peptides, followed by a tryptic digest of a cell-cultured SARS-CoV-2 variant, and identification of these markers by HR-LC-MS/MS analysis. As a proof of concept, this approach was demonstrated for four representative VOCs compared to the wild-type Wuhan reference strain. For each variant, at least two unique markers, derived mainly from the spike (S) and nucleocapsid (N) viral proteins, were identified. This methodology is specific, rapid, easy to perform and inexpensive. Therefore, it can be applied as a diagnostic tool for pathogenic variants.

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