NOD2 (the nucleotide-binding oligomerization domain containing protein 2) is known to be involved in host recognition of bacteria, although its role in the host response to Staphylococcus aureus infection is unknown. NOD2-deficient (Nod2^−/−) mice and wild-type (WT) littermate controls were injected intraperitoneally with S. aureus suspension (10⁷ bacteria/g of body weight), and their survival was monitored. Cultured bone marrow-derived neutrophils were harvested from Nod2^−/− and WT mice and tested for cytokine production and phagocytosis. Compared to WT mice, Nod2^−/− mice were significantly more susceptible to S. aureus infection (median survival of 1.5 days versus >5 days; P = 0.003) and had a significantly higher bacterial tissue burden. Cultured bone marrow-derived neutrophils from Nod2^−/− and WT mice had similar levels of peritoneal neutrophil recruitment and intracellular killing, but bone marrow-derived neutrophils from Nod2^−/− mice had significantly reduced ability to internalize fluorescein-labeled S. aureus. Nod2^−/− mice had significantly higher levels of Th1-derived cytokines in serum (tumor necrosis factor alpha, gamma interferon, and interleukin-2 [IL-2]) compared to WT mice, whereas the levels of Th2-derived cytokines (IL-1β, IL-4, IL-6, and IL-10) were similar in Nod2^−/− and WT mice. Thus, mice deficient in NOD2 are more susceptible to S. aureus. Increased susceptibility is due in part to defective neutrophil phagocytosis, elevated serum levels of Th1 cytokines, and a higher bacterial tissue burden.