Most human navigation studies in MRI rely on virtual navigation. However, the necessary supine position in MRI makes it fundamentally different from daily ecological navigation. Nonetheless, until now, no study has assessed whether differences in physical body orientation (BO) affect participants' experienced BO during virtual navigation. Here, combining an immersive virtual reality navigation task with subjective BO measures and implicit behavioral measures, we demonstrate that physical BO (either standing or supine) modulates experienced BO. Also, we show that standing upright BO is preferred during spatial navigation: participants were more likely to experience a standing BO and were better at spatial navigation when standing upright. Importantly, we report that showing a supine virtual agent reduces the conflict between the preferred BO and physical supine BO. Our study provides critical, but missing, information regarding experienced BO during virtual navigation, which should be considered cautiously when designing navigation studies, especially in MRI.