Transbronchial lung biopsy is an indispensable method for the diagnosis of peripheral lung lesions; however, the diagnostic yield still remains unsatisfactory. Endobronchial ultrasound with guide sheath (EBUS-GS) is an excellent method for the decision of biopsy points and has contributed to improvements in diagnostic yield, but the decision of choosing the proper bronchus depends on the individual ability of each bronchoscopist. To clarify the usefulness of the virtual bronchoscopic navigation system (VBN), we evaluated the diagnostic yield and time required to determine the target lesion. Fifty-seven cases using EBUS-GS with VBN (VBN/EBUS-GS group) and 55 cases using EBUS-GS (EBUS-GS group) were compared. In the VBN/EBUS-GS group, computer software detects the air density in the bronchi from the computed tomography image and imports a detailed virtual Bronchoscopic Image. After inserting the starting position and the peripheral target lesion, the software depicts the most ideal route to the target lesion during the bronchoscopic procedure. EBUS is then used to confirm the accuracy of the route. Diagnostic yield was 84.2% for the VBN/EBUS-GS group and 80.0% for EBUS-GS group. The required time to determine the biopsy position was significantly less in the VBN/EBUS-GS group (5.54±0.57 min in VBN/EBUS-GS group vs. 9.27±0.86 min in EBUS-GS group, P<0.01). In conclusion, VBN proved useful in shortening the time needed to determine the biopsy position.