RT-PCR/MALDI-TOF Diagnostic Target Performance Reflects Circulating SARS-CoV-2 Variant Diversity in New York City

• Published in: The Journal of molecular diagnostics : JMD Vol. 24; no. 7; pp. 738 - 749
• Main Authors: Hernandez, Matthew M., Banu, Radhika, Gonzalez-Reiche, Ana S., Gray, Brandon, Shrestha, Paras, Cao, Liyong, Chen, Feng, Shi, Huanzhi, Hanna, Ayman, Ramírez, Juan David, van de Guchte, Adriana, Sebra, Robert, Gitman, Melissa R., Nowak, Michael D., Cordon-Cardo, Carlos, Schutzbank, Ted E., Simon, Viviana, van Bakel, Harm, Sordillo, Emilia Mia, Paniz-Mondolfi, Alberto E.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.07.2022; American Society for Investigative Pathology
• Subjects: COVID-19 - diagnosis; COVID-19 - epidemiology; Humans; New York City - epidemiology; Regular; Reverse Transcriptase Polymerase Chain Reaction; SARS-CoV-2 - genetics; Sensitivity and Specificity; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; New York City
• ISSN: 1525-1578; 1943-7811; 1943-7811
• DOI: 10.1016/j.jmoldx.2022.04.003

As severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to circulate, multiple variants of concern have emerged. New variants pose challenges for diagnostic platforms because sequence diversity can alter primer/probe-binding sites (PBSs), causing false-negative results. The MassARRAY SARS-CoV-2 Panel (Agena Bioscience) uses RT-PCR and mass spectrometry to detect five multiplex targets across N and ORF1ab genes. Herein, we use a data set of 256 SARS-CoV-2–positive specimens collected between April 11, 2021, and August 28, 2021, to evaluate target performance with paired sequencing data. During this time frame, two targets in the N gene (N2 and N3) were subject to the greatest sequence diversity. In specimens with N3 dropout, 69% harbored the Alpha-specific A28095U polymorphism that introduces a 3′-mismatch to the N3 forward PBS and increases risk of target dropout relative to specimens with 28095A (relative risk, 20.02; 95% CI, 11.36 to 35.72; P < 0.0001). Furthermore, among specimens with N2 dropout, 90% harbored the Delta-specific G28916U polymorphism that creates a 3′-mismatch to the N2 probe PBS and increases target dropout risk (relative risk, 11.92; 95% CI, 8.17 to 14.06; P < 0.0001). These findings highlight the robust capability of MassARRAY SARS-CoV-2 Panel target results to reveal circulating virus diversity, and they underscore the power of multitarget design to capture variants of concern.