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There are significant unmet requirements for rapid differential diagnosis of infection in patients admitted to intensive care units. Serum levels of interleukin-6 (IL-6), procalcitonin (PCT), presepsin, and C-reactive protein (CRP) are measured in clinical practice; however, their clinical utility in patients with organ dysfunction has not been tested adequately. Thus, we investigated the diagnostic and prognostic value of IL-6, PCT, presepsin, and CRP in critically ill patients who had organ dysfunction with suspicion of infection.In 100 consecutive critically ill patients with organ dysfunction and suspected infection, serum levels of IL-6, PCT, presepsin, and CRP were measured upon suspicion of infection and serially every other day up to 7 days (cohort 1). The primary outcome variable was the presence of infections. The diagnostic value of IL-6 was further tested in cohort 2 (n = 72, case-control matched). The secondary outcome variables were the sequential organ failure assessment (SOFA) score, serum creatinine levels, and 28-day mortality.Among the four biomarkers, serum IL-6 levels had the highest area under the curve (AUC) value of 0.824 (95% confidence interval [CI] 0.735–0.913) for diagnosing infection in critically ill patients with organ dysfunction and suspected infection in cohort 1 (AUC [95% CI] for the other biomarkers: PCT, 0.813 [0.714–0.911]; CRP, 0.764 [0.645–0.883]; presepsin, 0.681 [0.513–0.849]). In cohort 2, the sensitivity and specificity of IL-6 for diagnosing infection were 0.861 and 0.806, respectively. The presepsin levels were significantly correlated with the SOFA score and serum creatinine levels upon suspicion of infection (r > 0.5), especially serum creatinine levels in the patients without infection (r = 0.789). Serum IL-6 levels were significant predictors of 28-day mortality. The AUC value of serum IL-6 levels for 28-day mortality increased over time; the serum IL-6 levels on Day 7 had the highest AUC value of 0.883 (95% CI, 0.788–0.978) for 28-day mortality.Among serum IL-6, PCT, presepsin, and CRP levels, serum IL-6 levels had the highest diagnostic value for infection. They were also significant predictors of 28-day mortality. Hence, they may improve diagnosis of infection and prediction of 28-day mortality in critically ill patients with organ dysfunction.