Background: Radial endobronchial ultrasound with a guide sheath for transbronchial biopsy (EBUS-GS-TBB) can be considered for diagnosing peripheral pulmonary lesions (PPLs) with fewer complications in patients with emphysema. However, the utility and safety of bronchoscopy for PPLs in the proximity of emphysema-area lesions remain unclear. The aim of this study was to assess the efficacy and complications of the initial diagnostic procedure of bronchoscopy with EBUS-GS-TBB according to the proximity of PPLs to emphysema areas, along with factors affecting the successful diagnostic yield for PPLs, and to identify the feasibility of molecular and genetic testing using EBUS-GS-TBB-obtained tumor samples.Methods: The medical records of 278 consecutive patients with PPLs who underwent EBUS-GS-TBB without X-ray fluoroscopy guidance were screened. We compared PPLs with emphysema in such lesions. PPLs with emphysema were divided into two groups: PPLs located in non-emphysema areas and those inside or near emphysema areas.Results: This study included 84 patients with emphysema (non-emphysema area group=46; inside or near emphysema area group=38). The diagnostic yield was significantly higher for PPLs located in non-emphysema areas than for PPLs inside or near emphysema areas (82.6% vs. 52.6%, p=0.013). Multivariate analysis revealed that PPLs located in non-emphysema areas (odds ratio=5.614) and EBUS images within lesions were significant factors affecting diagnostic yield. Further, 91.7%–100% of EBUS-GS-TBB-obtained tissue samples were sufficient for molecular testing of PD-L1, EGFR, ALK, and ROS1.Conclusions: In patients with emphysema, the positional relation of PPLs to emphysema lesions and EBUS images within lesions were important factors affecting successful diagnosis using EBUS-GS-TBB.