Journal of Diagnostic Techniques and Biomedical AnalysisISSN: 2469-5653

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Sequencing the SARS-CoV-2 Genome from Stool Samples of Post-Acute Cases Implicates a Novel Mutation Associated with Reduced Antibody Neutralization

Objective: Whole genome SARS-CoV-2 sequencing tools are crucial for tracking the COVID-19 pandemic. However, current techniques require sampling of actively infectious patients following COVID-19 testing to recover enough SARS-CoV-2 RNA from the nasopharyngeal passage, which rapidly clears during the first few weeks of infection. A prospective assessment of the viral genome sourced from recovered noninfectious patients would greatly facilitate epidemiological tracking. Thus, we developed a protocol to isolate and sequence the genome of SARS-CoV-2 from stool samples of post-acute SARS-CoV-2 patients, at time points ranging from 10-120 days after onset of symptoms.

Methods: Stool samples were collected from patients at varying time points post-convalescence and viral DNA was isolated and sequenced using the QIAamp viral RNA mini kit (Qiagen Inc.) and ion Ampliseq™ library kit plus (Life Technologies Corporation). Capacity of neutralizing antibodies in patient plasma was tested using a Luminex panel (Coronavirus Ig Total Human 11-Plex ProcartaPlex™ Panel, Thermo Fisher).

Results: Out of 64 samples obtained from post-acute patients, 21 (32.8%) yielded sufficient material for whole genome sequencing. This allowed us to identify widely divergent phylogenetic relativity of the SARS-CoV-2 genome from postacute patients living in the same households and infected around the same time. Additionally, we observed that individuals who recovered from infection expressed varying degrees of antibodies against SARS-CoV-2 structural proteins that corresponded to distinct variants. Interestingly, we identified a novel point mutation in the viral genome where infected patients expressed antibodies with a significantly reduced capacity to neutralize the virus in vitro relative to that of those infected with the wild type strain.

Conclusion: Altogether, we demonstrate a protocol to successfully sequence the SARS-CoV-2 genome from stool samples from patients up to 4 months post-infection, which can be applied to studies that assess the relationship between variants and immune response post-hoc and safe monitoring of the SARS-CoV-2 genome over the course of the pandemic.

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