The evidence-based public health course equips public health professionals with skills and tools for applying evidence-based frameworks and processes in public health practice. To date, training has included participants from all the 50 U.S. states, 2 U.S. territories, and multiple other countries besides the U.S. This study pooled follow-up efforts (5 surveys, with 723 course participants, 2005–2019) to explore the benefits, application, and barriers to applying the evidence-based public health course content. All analyses were completed in 2020. The most common benefits (reported by >80% of all participants) were identifying ways to apply knowledge in their work, acquiring new knowledge, and becoming a better leader who promotes evidence-based approaches. Participants most frequently applied course content to searching the scientific literature (72.9%) and least frequently to writing grants (42.7%). Lack of funds for continued training (35.3%), not having enough time to implement evidence-based public health approaches (33.8%), and not having coworkers trained in evidence-based public health (33.1%) were common barriers to applying the content from the course. Mean scores were calculated for benefits, application, and barriers to explore subgroup differences. European participants generally reported higher benefits from the course (mean difference=0.12, 95% CI=0.00, 0.23) and higher frequency of application of the course content to their job (mean difference=0.17, 95% CI=0.06, 0.28) than U.S. participants. Participants from later cohorts (2012–2019) reported more overall barriers to applying course content in their work (mean difference=0.15, 95% CI=0.05, 0.24). The evidence-based public health course represents an important strategy for increasing the capacity (individual skills) for evidence-based processes within public health practice. Organization-level methods are also needed to scale up and sustain capacity-building efforts.