Genomic evidence for reinfection with SARS-CoV-2: a case study

Abstract

Background: The degree of protective immunity conferred by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is currently unknown. As such, the possibility of reinfection with SARS-CoV-2 is not well understood. We describe an investigation of two instances of SARS-CoV-2 infection in the same individual.

Methods: A 25-year-old man who was a resident of Washoe County in the US state of Nevada presented to health authorities on two occasions with symptoms of viral infection, once at a community testing event in April, 2020, and a second time to primary care then hospital at the end of May and beginning of June, 2020. Nasopharyngeal swabs were obtained from the patient at each presentation and twice during follow-up. Nucleic acid amplification testing was done to confirm SARS-CoV-2 infection. We did next-generation sequencing of SARS-CoV-2 extracted from nasopharyngeal swabs. Sequence data were assessed by two different bioinformatic methodologies. A short tandem repeat marker was used for fragment analysis to confirm that samples from both infections came from the same individual.

Findings: The patient had two positive tests for SARS-CoV-2, the first on April 18, 2020, and the second on June 5, 2020, separated by two negative tests done during follow-up in May, 2020. Genomic analysis of SARS-CoV-2 showed genetically significant differences between each variant associated with each instance of infection. The second infection was symptomatically more severe than the first.

Interpretation: Genetic discordance of the two SARS-CoV-2 specimens was greater than could be accounted for by short-term in vivo evolution. These findings suggest that the patient was infected by SARS-CoV-2 on two separate occasions by a genetically distinct virus. Thus, previous exposure to SARS-CoV-2 might not guarantee total immunity in all cases. All individuals, whether previously diagnosed with COVID-19 or not, should take identical precautions to avoid infection with SARS-CoV-2. The implications of reinfections could be relevant for vaccine development and application.

Funding: Nevada IDEA Network of Biomedical Research, and the National Institute of General Medical Sciences (National Institutes of Health).

Figure 1

Timeline of symptom onset, molecular diagnosis, and sequencing of specimens TMA=transcription-mediated amplification. *Sequenced specimens.

Figure 2

Variant mapping of specimens A and B against the reference genome ORF1a and ORF1b encode replicase proteins. The other ORFs encode assembly proteins. ORF=open reading frame. S=spike. E=envelope. M=membrane. N=nucleocapsid. *Identifies variant 14 407 in specimen A and variants 14 407 and 14 408 in specimen B.

Figure 3

Phylogenetic placement of specimens A and B within Nevada isolates, reference genomes, and global clades 171 sequences were from Nevada. Wuhan Hu 1 was the reference genome (GenBank MN908947.3). USA WA1 was the isolate USA-WA1/2020 (Bei Resources, Manassas, VA, USA).