
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.
Our case report presents details of the first individual in North America to have symptomatic reinfection with SARS-CoV-2. Similar to observations with the reinfection case in Ecuador,our patient showed increased symptom severity in their second infection, whereas the cases from Belgium and the Netherlands11 and Hong Kong10 did not show a difference in severity of symptoms. The mechanisms that could account for a more severe secondary infection can only be speculated. First, a very high dose of virus might have led to the second instance of infection and induced more severe disease.Second, it is possible that reinfection was caused by a version of the virus that was more virulent, or more virulent in this patient's context. Third, a mechanism of antibody-dependent enhancement might be the cause, a means by which specific Fc-bearing immune cells become infected with virus by binding to specific antibodies. This mechanism has been seen previously with the betacoronavirus causing severe acute respiratory syndrome. In that case, the patient recovered and was discharged from hospital.
The individual associated with these two SARS-CoV-2 infections had no immunological disorders that would imply facilitation of reinfection. They were not taking any immunosuppressive drugs. The individual was negative for HIV by antibody and RNA testing (data not shown) and had no obvious cell count abnormalities. The secondary positive case (reinfection) occurred simultaneously to a positive case in a cohabitant (parent), who also provided a specimen on June 5, 2020, that was positive by nucleic acid amplification testing (transcription-mediated amplification). Sequencing is underway on the co-habitant specimen to ascertain its potential role in reinfection. However, the positive specimen from the co-habitant was obtained and tested in the Hologic Aptima format, which did not align with the procedures established at our sequencing laboratory. Nevertheless, the co-habitant positive case provides a possible source for secondary exposure and reinfection of our patient.
It is possible that we have reported a case of continuous infection entailing deactivation and reactivation. However, for such a hypothesis to be true, a mutational rate of SARS-CoV-2 would be required that has not yet been recorded.Specimens A and B showed an extrapolated rate of SNV and MNV accumulation of 83·64 substitutions per year, a rate that greatly exceeds the currently observed rate of 23·12.However, even more importantly, the four substitutions noted in specimen A would have to revert to the ancestral genotype, and the odds of this reversion occurring are remote. Of course, if such an amount of base change did occur in that timeframe, the remarkable nature of specimens A and B would shift from a case of possible reinfection to one of high-rate evolution within an infected individual. Another alternative explanation for the observed differences in specimens A and B would be that of co-infection. In a co-infection hypothesis, the patient would have been infected with viruses of both genotypes at the time of sample collection. Such a hypothesis would then further require that the specimen B type virus be present, yet undetected in April, 2020, and then conversely, specimen A type virions become depleted before the June, 2020, sample collection date. Specimens A and B were both in clade 20C, which was the predominant major clade seen in northern Nevada at the time samples were obtained. Our survey of viruses in Nevada identified samples resembling each of the case genotypes. Although evidence exists that SARS-CoV-2 quasispecies exist at low and fluctuating frequencies in infected samples, whereby low-frequency (eg, 1%) SNVs could be seen in various samples from the same patient, this possible situation would not itself account for the genotype switch observed between the first infection and reinfection.
Our findings have implications for the role of vaccination in response to COVID-19. If we have truly reported a case of reinfection, initial exposure to SARS-CoV-2 might not result in a level of immunity that is 100% protective for all individuals. With respect to vaccination, this understanding is established, with influenza regularly showing the challenges of effective vaccine design.A major limitation of our case study is that we were unable to undertake any assessment of the immune response to the first episode of SARS-CoV-2 infection. We also could not assess fully the effectiveness of the immune responses (eg, neutralising antibody titres) during the second episode, when the individual was antibody-positive for total antibody assay to the SARS-CoV-2 nucleocapsid protein. If our patient is a case of natural viral evolution in vivo (although highly unlikely in view of the requirement of four reversions to reference genotypes) then the implications of these data are that SARS-CoV-2 can adapt with enough genetic dexterity to avoid a natural immune response in a manner to re-establish detectable levels of infection in an individual. If our patient is a case of reinfection, it is crucial to note that the frequency of such an occurrence is not defined by one case study: this event could be rare. The absence of comprehensive genomic sequencing of positive cases in the USA and worldwide limits the advances in public health surveillance needed to find these cases. Certainly, limitations in screening and testing availability for SARS-CoV-2 exacerbate the poor surveillance efforts being undertaken not only to diagnose COVID-19 but also to obtain actionable genetic tracking of this agent.
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