Insufficient information on existing bridge substructures and foundations poses significant challenges for structural condition evaluation and can cause significant uncertainties for the safety and serviceability of bridges. Characterization and condition evaluation of bridges substructure and foundations will not only help to decrease the vulnerability to natural hazards but also provide opportunities for their reuse with considerable benefits. In this paper, the feasibility of leveraging structural identification techniques to characterize bridge substructures and foundations is investigated. A three-span simply supported bridge located in Mossy, West Virginia, USA, is used as a study case. Modal analysis and finite element model updating techniques are used to investigate and estimate the uncertainties and conditions of the substructure. Updated finite element model for this structure provides valuable information for bridge condition assessment and proves how structural identification is a viable tool for the case considered.