Immune Signatures Distinguish Pure and Mixed Sepsis in Critical COVID-19: A Retrospective Cohort Study

Abstract

Background: COVID-19-associated sepsis poses unique challenges in intensive care units (ICUs), where patients are often immunocompromised and prone to secondary bacterial infections. Differentiating immune phenotypes between pure viral and viral-bacterial sepsis is essential for timely diagnosis and personalized treatment.

Objective: To compare clinical characteristics, immune profiles, and outcomes between patients with pure COVID-19 sepsis and those complicated by confirmed secondary bacterial infection, and to identify immune markers capable of differentiating sepsis phenotypes.

Methods: This retrospective cohort study enrolled ICU patients with severe COVID-19 pneumonia between July 2021 and December 2023. Patients were classified into viral sepsis (V group, n=53) and viral-bacterial sepsis (VB group, n=28) based on microbiological confirmation of secondary infection. Clinical data, inflammatory markers, cytokine levels, and neutrophil CD64 expression (via flow cytometry) were analyzed. Logistic regression, ROC curves, and Kaplan-Meier survival analysis were used to evaluate phenotypes, identify discriminative markers, and assess outcomes.

Results: The VB group exhibited significantly higher SOFA scores (median 9.5 vs 7, P<0.001), serum creatinine (103.2 vs 80.85 µmol/L, P=0.001), and LDH levels (531 vs 392 U/L, P=0.009), indicating more severe organ dysfunction. ICU stay was longer in the VB group (median 16 vs 13 days, P=0.027), and ICU mortality was slightly higher (85.7% vs 83.0%, P=0.785). Immune profiling showed significantly higher IL-2 (36.8 vs 22.1 pg/mL, P<0.001), IL-10, IFN-γ, and MYD88 in the V group, whereas TNF-α (62.3 vs 43.7 pg/mL, P=0.002) and PCT (2.51 vs 1.12 ng/mL, P=0.001) were higher in the VB group. IL-2, MYD88, and PCT independently discriminated phenotypes. Neutrophil CD64 showed strong predictive value for superinfection (AUC=0.969).

Conclusion: COVID-19 patients with secondary bacterial sepsis exhibit distinct immune and clinical profiles. Immune phenotyping may enable early recognition of bacterial superinfection and guide phenotype-driven therapy in severe viral sepsis.

Trial registration: ClinicalTrials.gov, NCT06491966. Registered 2 April 2024, https://clinicaltrials.gov/ct2/show/NCT06491966.

Keywords: COVID-19 sepsis; ICU; cytokines; immune profiling; organ dysfunction; secondary bacterial infection.

Figure 1

Kaplan-Meier survival analysis in icu patients with different sepsis types. Kaplan-Meier survival curves comparing outcomes among patients with bacterial sepsis (B group), viral sepsis (V group), and viral-bacterial sepsis (VB group). The survival patterns showed similar trends within the first 20 days of ICU stay. After day 20, patients with viral-related sepsis (V+VB groups) had significantly lower survival rates compared to bacterial sepsis (B group) (P<0.05). The 28-day survival rates were 50.5%, 28.3%, and 14.3% for B, V, and VB groups, respectively. Although VB group showed a trend of lower survival rates than V group before day 30 and higher rates thereafter, this difference between V and VB groups was not statistically significant (P=0.159). Sample sizes: B group (n=105), V group (n=106), VB group (n=65). P-values were adjusted using Bonferroni correction for multiple comparisons among three groups.

Figure 2

Detection of pathogens in bacterial vs viral related sepsis (V+VB group). The detection rates of various pathogens were compared between bacterial sepsis (B group) and viral related sepsis (V+VB group). The detection rates for blood culture positive (BCP) and bronchoalveolar lavage fluid positive (BALF+) showed no significant differences between groups. Significantly higher detection rates were found in V+VB group for cytomegalovirus (CMV, P=0.013), Epstein-Barr virus (EBV, P=0.025), Aspergillus fumigatus (AF, P=0.002), and lower respiratory aspirate culture positive (LRAC+, P<0.001). The B group showed significantly higher rates for catheter-related culture positive (CRC+, P=0.017), Gram-positive bacteria (GPB, P=0.001), Gram-negative bacteria (GNB, P=0.006), fungi (P=0.002), and Acinetobacter baumannii (AB, P<0.001). Other pathogens including Klebsiella pneumoniae (KP), Pseudomonas aeruginosa (PA), Escherichia coli (EC), Enterococci (E), Staphylococcus aureus (SA), and Candida albicans (CA) showed no significant differences between groups (all P>0.05).

Figure 3

Distribution of various pathogens in SARS-negative and SARS-positive patients based on third-generation sequencing of BALF specimens.

Figure 4

Use of antimicrobials in bacterial versus viral sepsis groups. This figure illustrated the use of various antimicrobials in patients with bacterial sepsis (B group) and viral sepsis (V group). The antibiotics were listed on the x-axis, and the number of patients using each antibiotic was shown on the y-axis. Blue bars represent the bacterial sepsis group, and green bars represented the viral sepsis group. Significant differences in the use of specific antimicrobials between the two groups were marked with an asterisk (*). The significant differences were observed in the use of Cephalosporins (2/3 generation), Beta-lactamase inhibitors, Vancomycin, Tigecycline, Fluconazole, Echinocandin, Polymyxin, and Clindamycin.