Roads force wildlife to navigate degraded and fragmented habitats across the globe, creating barriers to movement and increasing the risk of mortality. Recent studies have highlighted the negative impact that roads have on the activity and movement of insect-eating bats because roads subject roosting and foraging habitats to widespread disturbance, alteration, and reduced availability. Bats have import ecosystem roles, and there is growing concern about their conservation status in North America. Washington Department of Transportation (WSDOT) has created multiple wildlife crossings along Interstate-90 near Snoqualmie Pass to increase ecosystem connectivity. WSDOT and its partners, including Central Washington University, are monitoring various wildlife species to evaluate their use of these structures. Wildlife crossings structures (WCS) are a common mitigation strategy to increase connectivity, but investigation into bat activity in or around these structures is limited. Our goal was to test whether highway locations with WCS had more bat species and higher activity levels than locations without underpasses. We predicted that species detected along the highway (i.e., underpass and unmitigated sites) would be a subset of those in the habitats adjacent to I-90 and bat activity would be higher at the location of underpasses than at areas without them, but the activity levels would be dependent on guild-specific characteristics. We used acoustic monitors to record and identify echolocation calls at six locations along Interstate-90 during summer 2020. Three locations were at wildlife underpasses and three were at unmitigated locations (no underpasses). For each highway location, we also recorded calls 100 to 300 m inside the forest. Calls were analyzed using general linear mixed models across all species and separated into 4 guilds based on species-specific frequency ranges (e.g., 20-,30-,40- and 50-kilohertz guild). Species richness was the same at all locations: the 8 species detected in the forest were also detected at all highway locations. Total bat activity was higher along the highway than the adjacent forest but did not differ between locations with vs. without underpasses. Guild activity followed similar trends, with some exceptions. The 40-kHz guild, that has relatively high-frequency calls and more maneuverable flight characteristics, showed significantly higher activity at underpasses than at locations without them. Confounding variables make interpretation challenging, but this study provides important information on bat activity along an interstate highway in Washington State. We highlight the need for more intensive monitoring efforts to better understand the effectiveness of WCS in reducing the impacts roads have on bats in North America, and suggest some study design considerations.