Optimizing SARS-CoV-2 molecular diagnostic using N gene target: insights about reinfection

Abstract

IntroductionMolecular diagnosis of SARS-CoV-2 is a huge challenge to many countries around the world. The cost of tests to check infected people is inaccessible since specialized teams and equipment are not disposable in remote locations. Herein, we compared the fitness of two primers sets to the SARS-CoV-2 N gene in the molecular diagnosis of COVID-19.Materials and MethodsThe 1029 patient samples were tested to presense/abscence molecular test using in house US CDC protocol. We compared the fitness of two primers sets to two different regions of N gene targets.ResultsBoth targets, N1 and N2 displayed similar fitness during testing with no differences between Ct or measurable viral genome copies. In addition, we verified security ranges Cts related to positive diagnostic with Ct above 35 value failuring in 66,6% after retesting of samples.Main conclusionOur data suggest that it is secure to use just one primer set to the N gene to identify SARS-CoV-2 in samples and the labs should be careful to set positive samples in high Ct values using high cutoffs.

Development and validation of cost-effective SYBR Green-based RT-qPCR and its evaluation in a sample pooling strategy for detecting SARS-CoV-2 infection in the Indonesian setting

“…US CDC protocol utilizes two primer sets to two different regions of N gene targets, which are tested in two separate tubes. Both targets, namely N1 and N2, have shown similar performance in the detection of the virus, with no differences between Ct or measurable viral genome copies [36,37]. Our previous study [20] also demonstrated that singleplex amplification of the N gene was equally sensitive and specific in comparison to the commercial TaqMan assay.…”

Receptor-Binding-Motif-Targeted Sanger Sequencing: a Quick and Cost-Effective Strategy for Molecular Surveillance of SARS-CoV-2 Variants

“…It could also be used for other sample types, including urine, other body fluids, and water ( 22 ). In this study, CDC N1 oligonucleotide ( 23 ) pairs with 6-carboxyfluorescein (FAM) probe were utilized in RT-qPCR to screen samples for the presence of SARS-CoV-2 (see Fig. S1 in the supplemental material).…”