SARS-CoV-2 shedding and evolution in patients who were immunocompromised during the omicron period: a multicentre, prospective analysis

Abstract

Background: Prolonged SARS-CoV-2 infections in people who are immunocompromised might predict or source the emergence of highly mutated variants. The types of immunosuppression placing patients at highest risk for prolonged infection have not been systematically investigated. We aimed to assess risk factors for prolonged SARS-CoV-2 infection and associated intrahost evolution.

Methods: In this multicentre, prospective analysis, participants were enrolled at five US medical centres. Eligible patients were aged 18 years or older, were SARS-CoV-2-positive in the previous 14 days, and had a moderately or severely immunocompromising condition or treatment. Nasal specimens were tested by real-time RT-PCR every 2-4 weeks until negative in consecutive specimens. Positive specimens underwent viral culture and whole genome sequencing. A Cox proportional hazards model was used to assess factors associated with duration of infection.

Findings: From April 11, 2022, to Oct 1, 2022, 156 patients began the enrolment process, of whom 150 were enrolled and included in the analyses. Participants had B-cell malignancy or anti-B-cell therapy (n=18), solid organ transplantation or haematopoietic stem-cell transplantation (HSCT; n=59), AIDS (n=5), non-B-cell malignancy (n=23), and autoimmune or autoinflammatory conditions (n=45). 38 (25%) participants were real-time RT-PCR-positive and 12 (8%) were culture-positive 21 days or longer after initial SARS-CoV-2 detection or illness onset. Compared with the group with autoimmune or autoinflammatory conditions, patients with B-cell dysfunction (adjusted hazard ratio 0·32 [95% CI 0·15-0·64]), solid organ transplantation or HSCT (0·60 [0·38-0·94]), and AIDS (0·28 [0·08-1·00]) had longer duration of infection, defined as time to last positive real-time RT-PCR test. There was no significant difference in the non-B-cell malignancy group (0·58 [0·31-1·09]). Consensus de novo spike mutations were identified in five individuals who were real-time RT-PCR-positive longer than 56 days; 14 (61%) of 23 were in the receptor-binding domain. Mutations shared by multiple individuals were rare (<5%) in global circulation.

Interpretation: In this cohort, prolonged replication-competent omicron SARS-CoV-2 infections were uncommon. Within-host evolutionary rates were similar across patients, but individuals with infections lasting longer than 56 days accumulated spike mutations, which were distinct from those seen globally. Populations at high risk should be targeted for repeated testing and treatment and monitored for the emergence of antiviral resistance.

Funding: US Centers for Disease Control and Prevention.

Figure 1:. Temporal dynamics of SARS-CoV-2 RNA viral load and culture positivity in 121 patients

(A) Cycle threshold values for total SARS-CoV-2 RNA and virus culture isolation over time in 121 patients by immunocompromised group. Open circles indicate culture negative specimens and closed circles indicate positive specimens. (B) Kaplan-Meier survival curves showing time to last positive real-time RT-PCR test by immunocompromised group. p=0·0030 for difference in time to last positive specimen across all groups. HSCT=haematopoietic stem-cell transplantation.

Figure 2:. Within-host evolution of SARS-CoV-2 in 104 patients

(A) Proportion of newly arising mutations identified at more than 2% frequency for specimens collected during the indicated time periods. Number of samples in each group is listed for each bar. (B) Genome-wide within-host divergence rate for individuals positive for SARS-CoV-2 by real-time RT-PCR for less than 21 days compared with those positive for 21 days or longer. Individuals in each group with rates of zero (eg, no mutations identified with 2% frequency or more in the final specimen) are not plotted given log transformation of y axis and are indicated in parentheses at bottom of plot; however, these individuals were included in the statistical analysis. HSCT=haematopoietic stem-cell transplantation.

Figure 3:. De novo non-synonymous SARS-CoV-2 iSNVs in 65 patients

(A) Number of patients with shared mutations, colour coded by gene. Amino acid substitutions are labelled if shared by 5% or more (n=4 or more) of patients. (B) Number of patients with iSNVs in spike, by domain. Amino acid substitutions are labelled if shared by 5% or more (n=4 or more) of patients. (C) Heatmaps of de novo non-synonymous iSNVs in SARS-CoV-2 spike and their frequencies in five individuals with infections lasting more than 56 days and with two or more sequenced samples. Patient EV138 received bebtelovimab on day 1, and patient EV022 received bebtelovimab at day 68. Mutations in the receptor binding domain are indicated in bold. Bisected squares indicate more than one codon mutation identified produced the same amino acid substitution. iSNVs=intrahost single nucleotide variants.

Figure 4:. Mutations in 15 patients who received antiviral treatment

(A) Heatmap of de novo non-synonymous mutations in SARS-CoV-2 spike among patients who were immunocompromised, received monoclonal antibody (bebtelovimab, sotrovimab, or tixagevimab–cilgavimab), and had a post-treatment sample that was sequenced (n=10). Mutations in the receptor binding domain are shown in bold. Bisected squares indicate more than one codon mutation produced the same amino acid substitution. (B) Heatmap of mutations in SARS-CoV-2 nsp12 (RNA dependent RNA polymerase) among patients who were immunocompromised, received remdesivir, and had a post-treatment sample (n=7).