Marine ecosystems are threatened by a suite of anthropogenic stressors. Mitigating multiple threats is a daunting task, particularly when funding constraints limit the number of threats that can be addressed. Threats are typically assessed and prioritized via expert opinion workshops that often leave no record of the rationale for decisions, making it difficult to update recommendations with new information. We devised a transparent, repeatable, and modifiable method for collecting expert opinion that describes and documents how threats affect marine ecosystems. Experts were asked to assess the functional impact, scale, and frequency of a threat to an ecosystem; the resistance and recovery time of an ecosystem to a threat; and the certainty of these estimates. To quantify impacts of 38 distinct anthropogenic threats on 23 marine ecosystems, we surveyed 135 experts from 19 different countries. Survey results showed that all ecosystems are threatened by at least nine threats and that nine ecosystems are threatened by >90% of existing threats. The greatest threats (highest impact scores) were increasing sea temperature, demersal destructive fishing, and point-source organic pollution. Rocky reef, coral reef, hard-shelf, mangrove, and offshore epipelagic ecosystems were identified as the most threatened. These general results, however, may be partly influenced by the specific expertise and geography of respondents, and should be interpreted with caution. This approach to threat analysis can identify the greatest threats (globally or locally), most widespread threats, most (or least) sensitive ecosystems, most (or least) threatened ecosystems, and other metrics of conservation value. Additionally, it can be easily modified, updated as new data become available, and scaled to local or regional settings, which would facilitate informed and transparent conservation priority setting.