An 81-Nucleotide Deletion in SARS-CoV-2 ORF7a Identified from Sentinel Surveillance in Arizona (January to March 2020)

Abstract

O n 26 January 2020, the first coronavirus disease 2019 (COVID-19) case was reported in Arizona (third case in the United States) (1).Here, we report on early severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sentinel surveillance in Tempe, Arizona.Genomic characterization identified an isolate encoding a 27-amino-acid in-frame deletion in accessory protein ORF7a, the ortholog of SARS-CoV immune antagonist ORF7a/X4.In anticipation of COVID-19 spreading in Arizona, we initiated a surveillance effort for the local emergence of SARS-CoV-2 starting 24 January 2020.We leveraged an ongoing influenza surveillance project at Arizona State University (ASU) Health Services in Tempe, Arizona.Individuals presenting with respiratory symptoms (ILI) were tested for influenza A and B viruses (Alere BinaxNOW).Subsequently, we tested influenzanegative nasopharyngeal (NP) swabs for SARS-CoV-2.We extracted total nucleic acid using the bioMérieux eMAG automated platform and performed real-time quantitative reverse transcription-PCR (qRT-PCR) assays specific for SARS-CoV-2 N and E genes (2, 3).Out of 382 NP swabs collected from 24 January to 25 March 2020, we detected SARS-CoV-2 in 5 swabs from 16 to 19 March (Fig. 1).This corresponds to a prevalence of 1.31%.Given the estimated 1-to 14-day incubation period for COVID-19, the spike in cases might be related to university spring break holiday travel (8 to 15 March), as previously seen in other outbreaks (4, 5).To understand the evolutionary relationships and characterize the SARS-CoV-2 genomes, we performed next-generation sequencing (NGS; Illumina NextSeq, 2ϫ76) directly on specimen RNA, thereby avoiding cell culture passage and potentially associated mutations.This generated an NGS data set of 20.7 to 22.7 million paired-end reads per sample.We mapped quality-filtered reads to a reference SARS-CoV-2 genome (MN908947) using BBMap (version 39.64) to generate three full-length genomes: AZ-ASU2922 (376ϫ coverage), AZ-ASU2923 (50ϫ), and AZ-ASU2936 (879ϫ) (Geneious prime, version 2020.0.5).We aligned a total of 222 SARS-CoV-2 genome sequences comprising at least 5 representative sequences from phylogenetic lineages defined by Rambaut et al. (6).We performed phylogenetic reconstruction with BEAST (version 1.10.4;strict molecular clock, HKY ϩ ⌫ nucleotide substitution, exponential growth for coalescent model) (7-10).The ASU sequences were phylogenetically distinct, indicating independent transmissions (Fig. 2A).

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