<p>HIV enters the central nervous system (CNS) early, disrupting neurometabolism and inducing chronic neuroinflammation that results in the development of neurocognitive symptoms and neurodevelopmental delay in children infected with HIV. In children, neurocognitive presentation is often more severe than in adults, due to early and prolonged exposure to the virus. Tuberculous meningitis (TBM), another major pediatric CNS infection and often an opportunistic co-infection in HIV, carries high morbidity and mortality but remains difficult to diagnose with existing tests. Metabolomics provides a promising complementary diagnostic approach. We used ¹H-NMR and GCxGC-TOF/MS to compare cerebrospinal fluid (CSF) metabolic profiles from children with HIV, TBM, HIV-TBM co-infection, viral meningitis, and controls that were uninfected but presented with neurological symptoms (e.g., seizures, raised intracranial pressure, brainstem dysfunction, infarction, etc.). Profiles were analyzed using univariate (ANOVA, Wilcoxon with FDR correction) and multivariate (PCA) methods. The discriminatory performance of candidate metabolites and biomarker panels was evaluated using receiver operating characteristic (ROC) analysis, which included the area under the curve (AUC), sensitivity, specificity, likelihood ratios, diagnostic odds ratios, and Youden’s index to determine the optimal cut-offs. PCA demonstrated clear separation of viral and bacterial CNS infections. HIV was characterized by sugar alcohols, ketone bodies, and guanidinoacetate, whereas TBM showed a strong glycolytic and amino acid catabolic signature dominated by lactate, alanine, creatine, and fatty acid derivatives. Distinct CSF biosignatures can differentiate HIV and TBM in children, supporting their potential for future diagnostic applications and for advancing understanding of CNS pathogenesis in these infections.</p>