Since the beginning of this pandemic, an ongoing concern is whether one person can be infected by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) more than once. Occasional reports of people retesting positive, after seemingly clearing the virus, have been published. In a retrospective article published in EBioMedicine in September, Jing Lu and colleagues (Guangdong Provincial Center for Disease Control and Prevention [CDC], Guangdong CDC, China), reported that 87 (14%) of the 619 patients who had recovered from laboratory-confirmed SARS-CoV-2 infection retested positive by real-time Reverse-Transcription Polymerase Chain Reaction (RT-PCR). All patients who tested positive a second time in the study presented with mild or moderate symptoms at initial diagnosis and the researchers were unable to isolate the infectious virus by culture or detect sequencing of full-length viral genomes at the time of the second positive test. Because all patients who tested positive a second time were socially isolated during the study, the recurrence of test positivity might have resulted from ongoing shedding of viral fragments from the original infection.
Indeed, a patient who retests positive for virus might not necessarily be experiencing a second, new SARS-CoV-2 infection. To confirm reinfection with a distinct virus as opposed to prolonged viral shedding, whole viral genome sequencing from each, potentially separate, infection is necessary. Based on this standard, true reinfection cases have now been documented in Hong Kong, the Netherlands and Belgium, and Ecuador. In October, in The Lancet Infectious Diseases, Richard Tillett and colleagues (University of Nevada, NV, USA) reported the first case of reinfection in the USA. Despite the fact that isolates from both the first infection (A) and second infection (B) belonged to Clade 20C, specimen A had four single nucleotide variants (SNVs) not seen in specimen B, while specimen B had seven distinct SNVs compared with specimen A. Considering these substantial genetic differences, the authors excluded the possibility of viral evolution within the infected individual. Unlike cases in Hong Kong and the Netherlands and Belgium, the patients in Ecuador and the USA had more severe symptoms upon secondary infection. The reasons for increased severity in these two cases are as yet unknown.
Secondary infection could result from an inadequate immune stimulation upon the first encounter with the virus, or the decline of the neutralizing antibody responses over time. In fact, the robustness and longevity of neutralizing antibody responses to SARS-CoV-2 have not yet been comprehensively defined. In the June 2020 issue of Nature Medicine, Quanxin Long and colleagues (Chongqing Medical University, China) found that, compared with the symptomatic group, patients with COVID-19 who were asymptomatic had significantly lower IgG levels in the acute phase and early convalescent phase. Moreover, 40% of patients who were asymptomatic became seronegative for IgG 8 weeks after being discharged from the hospital, compared with 12•9% who were seronegative for the symptomatic group. An insufficient immune response could be a partial reason for the reinfection case in Hong Kong, as Paul Chan and colleagues (The Chinese University of Hong Kong, Hong Kong Special Administrative Region, China) reported in Emerging Infectious Diseases (December 2020, first online now) that the patient had low or undetectable levels of neutralizing antibody against multiple viral proteins during the primary mild-symptomatic infection and acute stage of asymptomatic reinfection.
Encouragingly, however, in a larger analysis published in Science in October, Ania Wajnberg and colleagues (Icahn School of Medicine at Mount Sinai, NY, USA), found that over 90% of infected individuals with mild-to-moderate symptoms had moderate-to-high titers of antibodies against spike protein (S-protein) of SARS-CoV-2 and these titers remain relatively stable for at least a period of around 5 months. Thus, much larger, definitive, serology studies are needed across a variety of populations and disease severities to get a comprehensive look at the degrees to which durable humoral immunity is induced by this virus.
In addition to variable immune stimulation, the evolution of SARS-CoV-2 might also have a potential role in reinfection. In a Cell paper published in September, Qianqian Li and colleagues (National Institutes for Food and Drug Control, China) systematically analyzed 80 natural variants in the S-protein for their infectivity and sensitivity to neutralization by antibody or convalescent serum samples using in vitro assays. Fortunately, the strain with the D614G mutation in the viral S-protein, a major variant now circulating globally, seems to retain susceptibility to neutralization by both convalescent serum samples and a panel of tested S-protein-specific monoclonal antibodies. This result was further supported by a medRxiv preprint done in September by Drew Weissman and colleagues (University of Pennsylvania, PA, USA). Although not formally peer reviewed, this group reported data using pseudoviruses bearing either the D614 or G614 S-protein that the D614G spike mutation could increase susceptibility of the viral S-protein to antibody neutralization. That said, Qianqian Li and colleagues found that particular S-protein variants, including A475V, L452R, V483A, and F490L, were less susceptible to neutralization by a subset of antibodies tested. Although the frequencies of such variants are still very low in the population, caution is warranted, as some leading vaccine candidates are using the S-protein as the immunogen.
In addition to humoral immunity, T cells also have a crucial role in clearing viruses from the body. In a July issue of Nature, Nina Le Bert and colleagues (Duke-NUS Medical School, Singapore) reported that CD4+ and CD8+ T cells from individuals who recovered from mild-to-severe COVID-19 could recognize multiple regions of the nucleocapsid protein (N-protein) of SARS-CoV-2. Furthermore, they found that patients who recovered from the SARS outbreak 17 years ago, which was caused by SARS-CoV, possess memory T cells that have robust cross-reactivity to the N-protein of SARS-CoV-2. Thus, stimulating T-cell responses could be an important consideration in vaccine development for a long-lasting protective effect.
With only four documented cases formally reported to date, is reinfection likely to be a rare phenomenon? Considering the relatively high percentage of asymptomatic infections (up to 40% as estimated by the USA CDC), it is possible that many undocumented reinfections have occurred. However, without viral genome sequencing data, the true reinfection rate cannot be confirmed. 12 months have passed since SARS-CoV-2 was first reported, but there is still so much to learn. Until effective methods of curing and preventing COVID-19 have been found, maintaining social distance and wearing masks are still our best options for personal protection, especially with the coming winter.
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