OBJECTIVE: To explore the value of serum microRNA-155-5p and -133a-3p (miR-155-5p and miR-133a-3p) expression for the diagnosis and prognosis evaluation of sepsis. METHODS: A prospective observational study was conducted. 105 sepsis patients admitted to emergency intensive care unit (EICU) of the First Affiliated Hospital of Zhengzhou University from January 2015 to January 2016 were enrolled. They were divided into three groups according to the severity: 35 patients with sepsis, 35 with severe sepsis, and 35 with septic shock. At the same time, 35 healthy persons were selected as the control group. According to the prognosis, the patients were divided into improved group (n = 70) and in-hospital death group (n = 35). The clinical data of all the subjects were collected. The mRNA expressions of miR-155-5p and miR-133a-3p were determined by reverse transcription-polymerase chain reaction (RT-PCR). The receiver-operating characteristic curve (ROC) was plotted to evaluate their clinical value for the diagnosis and prognosis of sepsis. The binary logistic regression was used to analyze the risk factors affecting the prognosis of sepsis patients. RESULTS: (1) The mRNA expressions of serum miR-155-5p and miR-133a-3p were gradually increased with the aggravation of sepsis. The mRNA expression of miR-155-5p (2-ΔCt) in sepsis, severe sepsis, sepsis shock groups was 1.89±0.48, 2.21±0.41, 2.79±0.73 (F = 23.737, P = 0.000), and the mRNA expression of miR-133a-3p (2(-ΔCt)) was 1.38±0.31, 1.74±0.65, 2.08±0.47, respectively (F = 27.710, P = 0.000). It was shown by ROC curve analysis that the area under the ROC curve (AUC) of serum miR-155-5p and miR-133a-3p for the diagnosis of sepsis was 0.855 [95% confidence interval (95%CI) = 0.761-0.949] and 0.769 (95%CI = 0.666-0.872) respectively. The cut-off value of miR-155-5p for the diagnosis of sepsis was 1.64, the sensitivity was 85.3%, and specificity was 80.6%. While the cut-off value of miR-133a-3p was 0.82, the sensitivity and specificity were 97.9% and 54.8% respectively. (2) Compared with improved group, the patients of in-hospital death group were more serious, and procalcitonin (PCT), C-reactive protein (CRP), D-dimer, lactic acid (Lac), sequential organ failure assessment (SOFA) score, acute physiology and chronic health evaluation II (APACHE II) score, and the mRNA expressions of miR-155-5p and miR-133a-3p were significantly increased (all P < 0.05). While there was no statistically significant difference in gender, age, white blood cells (WBC), serum creatinine (SCr) between the two groups (all P > 0.05). It was shown by binary logistic regression analysis that Lac [odds ratio (OR) = 0.514, 95%CI = 0.260-0.893, P = 0.024], sepsis severity (OR = 0.039, 95%CI = 0.023-2.955, P = 0.016), SOFA score (OR = 0.668, 95%CI = 0.474-0.825, P = 0.001), serum miR-155-5p expression (OR = 0.117, 95%CI = 0.020-0.530, P = 0.007) were the risk factors affecting the prognosis of patients with sepsis. CONCLUSIONS: The expression of serum miR-155-5p and miR-133a-3p may be used as specific indicators for the diagnosis of sepsis. And the expression of miR-155-5p can be used as independent impact factor for the estimation of sepsis prognosis.