Abstract: Biodiversity auditing represents a critical institutional instrument for advancing the green transformation of economic and social development and fostering ecological civilization, as well as a pivotal supervisory mechanism for implementing green strategic missions and safeguarding the functional integrity of ecosystems. Grounded in the macro-context of green strategic missions, this study employs sustainable development theory, the weakest-link effect theory, and ecological niche theory as its analytical framework, drawing on systematic literature review and case study methodologies to examine the current development landscape of biodiversity auditing in China. The study conducts an in-depth analysis of four core challenges confronting audit practice: evidentiary difficulties arising from the dynamic complexity and concealment of audit objects; ambiguous delineation of audit boundaries; the absence of applicable performance evaluation standards; and inadequate long-term audit mechanisms. On this basis, the study constructs a comprehensive optimization pathway encompassing the full audit cycle of “pre-audit planning, in-audit execution, and post-audit management” proposing a series of systematic countermeasures, including an ecological risk-based project initiation mechanism, an interdisciplinary collaborative audit model, a multi-source integrated evidence system, and an adaptive long-term closed-loop tracking management approach. The mbedding biodiversity auditing within the green strategic mission framework facilitates a fundamental transition in audit evaluation from an economic compliance orientation toward an ecological function orientation and drives a transformative shift in audit supervision from ex post review to ex ante risk prevention, thereby providing a quantifiable and traceable institutional guarantee for the Beautiful China strategy.