Tick-borne encephalitis virus (TBEV) can cause life-threatening CNS infection. Changes in cerebrospinal fluid (CSF) metabolites may reflect critical aspects of host responses and end-organ damage in neuro infection and neuroinflammation. In this study, we applied an untargeted metabolomics screen of CSF samples to investigate the metabolites profile and explore biomarkers for TBEV infection. By analyzing CSF samples from 77 patients with TBEV infection and 23 without TBEV infection, tryptophan metabolism and Citrate cycle were found to be the top important metabolic pathways in differentiating the control and case groups; acetoacetate, 5’-deoxy-5’-(methylthio)-adenosine, 3-methyl-2-oxobutanoic acid,etc. were identified to be metabolic biomarkers（| log 2 FC|>1,VIP>1,FDR<0.05）in CSF and clearly separated the TBEV infection from the non-infected samples. Moreover, four metabolites were identified to be associated with fatal outcome, including kynurenic acid, 5-hydroxyindole-3-acetic acid, DL-tryptophan, indole-3-acrylic acid, demonstrating the potential predictive biomarkers for severe TBEV infection. This study explored the metabolic profile of TBEV infection both in CSF samples and identified candidate biomarkers for TBEV infection, which might be useful in target screening for differential diagnosis and therapeutic inter-vention.