Hepatosplenic candidiasis is traditionally diagnosed by blood culture, magnetic resonance imaging (MRI), and histological analysis. The limitations of these methods include: low sensitivity (blood culture) and failure to isolate the organism (MRI/histology). This report describes a case of hepatosplenic candidiasis diagnosed by simultaneous polymerase chain reaction (PCR) analysis of liver biopsy and serum. PCR of biopsy and/or serum may offer a sensitive and specific diagnostic test for hepatosplenic candidiasis. Candida species can be identified, which helps guide antifungal chemotherapy, an important aspect in this difficult to treat infection.