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Complications of respiratory infections include pleural effusion (PE), associated with a high morbidity. Differentiating between PE caused by bacterial infections and Mycobacterium tuberculosis (TB) in endemic areas is difficult in children, impacting treatment. We investigated the aetiology of PE and features distinguishing TB from bacterial PE in children. Methods In this prospective study, children with PE admitted to a tertiary hospital in Cape Town from December 2016 to December 2019 were enrolled. Clinical information and routine laboratory investigations were compared between children with bacterial, TB or unclassified PE, categorised according to study definitions. Results A total of 91 patients were included; their median age was 31 months (IQR 11.8–102.1). Aetiology was bacterial in 37 (40%), TB in 36 (39%) and unclassified in 18 (20%) patients. Staphylococcus aureus was the most common bacterial isolate, confirmed in 24/37 (65%) patients; and Streptococcus pneumoniae confirmed in only 3/37 patients (8%). TB was microbiologically confirmed in 12/36 (33%) patients. Patients with TB were older (median age 91.6 vs 11.8 months, p< 0.001), with more weight loss (28/36 (77.8%) vs 12/37 (32%) patients, p< 0.001), and longer cough duration (10 vs 4 days, p< 0.001) than those with bacterial PE. In contrast, the latter had significantly higher median values: serum C-reactive protein (250 vs 122 mg/L, p< 0.001), procalcitonin (11 vs 0.5 mg/L, p< 0.001) pleural fluid lactate dehydrogenase (7280 vs 544 U/L, p< 0.001), and adenosine deaminase levels (162 vs 48 U/L, p< 0.001) and lower glucose levels (1.3 vs 4 mmol/L, p< 0.001). Conclusion Post-PCV, S. aureus is the dominant cause of PE in children using traditional culture methods, while TB remains a common cause of PE in our setting. Useful clinical and laboratory differences between TB and bacterial PE were identified, but the cause of PE in 20% of children was underdetermined. Molecular testing of pleural fluid for respiratory pathogens may be useful in such children.