Biodiversity underpins the fundamental elements for human well-being including food security, human health and access to clean water. In 2010, the Aichi Targets were adopted by world leaders to address the crisis of biodiversity loss. Despite conservation efforts, none of the Aichi Targets have been fully met. However, comprehensive analysis of the reasons for failure in terms of implementation mechanisms is, to date, rare and limited in scope. Here, we demonstrate that most parties did not set effective national targets in accordance with the Aichi Targets, and investments, knowledge and accountability for biodiversity conservation have been inadequate to enable effective implementation. We recommend that the new global targets under the post-2020 Global Biodiversity Framework should be adopted by parties as the minimum national targets to achieve the 2050 Vision. We propose that financial resources for biodiversity conservation are substantially increased through a variety of sources, including the deployment of new economic instruments such as payments for ecosystem services. In addition, science–policy interfaces at all levels need to be strengthened to integrate scientific, Indigenous and local knowledge to support decision-making. We suggest that a compliance and accountability mechanism, based on monitoring systems, is created to provide transparent and credible review of parties’ implementation of the new global targets.