Background: Invasive fungal infections, including hepatosplenic fungal infections (HSFI), cause significant morbidity and mortality in children with leukemia. There are not enough data to support for the best approach to diagnosis of HSFI in children, nor for the best treatment. Procedure: In this multicentric study, we assessed the demographic data, clinical and radiologic features, treatment, and outcome of 40 children with leukemia and HSFI from 12 centers. Results: All cases were radiologically diagnosed with abdominal ultrasound, which was performed at a median of 7 days, of the febrile neutropenic episode. Mucor was identified by histopathology in 1, and Candida was identified in blood cultures in 8 patients. Twenty-two had fungal infection in additional sites, mostly lungs. Nine patients died. Four received a single agent, and 36 a combination of antifungals. Conclusions: Early diagnosis of HSFI is challenging because signs and symptoms are usually nonspecific. In neutropenic children, persistent fever, back pain extending to the shoulder, widespread muscle pain, and increased serum galactomannan levels should alert clinicians. Abdominal imaging, particularly an abdominal ultrasound, which is easy to perform and available even in most resource-limited countries, should be recommended in children with prolonged neutropenic fever, even in the absence of localizing signs and symptoms.