Updated project metadata. Updated publication reference for PubMed record(s): 28765563. Bacterial meningitis is usually fatal without treatment. Prompt and accurate diagnosis coupled with the timely administration of parenteral antibiotics is necessary in order to save lives. We used label-free mass spectrometry based quantitative proteomics to identify specific protein signatures in cerebrospinal fluid from Malawian children with Streptococcus pneumoniae infection, the leading cause of bacterial meningitis in children, elderly and people infected with Human Immunodeficiency Virus. Samples were analysed in two independent cohorts (cases and controls, n=12 for the discovery cohort and n=16 for the verification cohort). The protein profiles clearly discriminated cases and controls. Proteins involved in the immune response and exosome cell signaling were significantly enriched in the infected samples. Several S. pneumoniae membrane proteins were identified suggesting these as potential therapeutic targets. For both cohorts, over 200 human proteins were up and down-regulated in children with confirmed S. pneumoniae infection in comparison with controls. Compiling proteins up- and down- regulated in both discovery and verification cohorts, we generated a comprehensive list of over 140 proteins from which diagnostic biomarkers could be nominated. For a panel of five selected proteins (cathelicidin, ceruloplasmin, myeloperoxidase, cystatin C and protein S100A8/S100 A9), we additionally verified the label-free results by automated quantitative western blot immunoassays (Simple Western™), with excellent agreement between the two experimental approaches. The proteins identified in our study could form the basis of development of a biomarker panel for rapid diagnosis. Our results could lead to the development of point-of-care assays to inform diagnosis, and to nominate new targets for drugs and vaccines to improve disease management and consequently to enhance the prognosis of children with bacterial meningitis.