The operation of a large-scale comprehensive port is characterized by multi-cooperative units, multi-cargo types and multi-application scenarios. In view of this, the analysis and research on port-centered ecosystem can promote the understanding of port operation mechanism and provide theoretical basis for the construction of digital twin system of large-scale comprehensive port. To clarify the boundary of digital twin system and the digital twin content, this paper analyzes and determines the external ecological participants of large-scale comprehensive port, classifies and stratifies the ecological participants of port operation, and refines the basic characteristics of the operation ecosystem model. Ecological participants are organically linked by the ecological relationship of port operation, and the construction logic of port operation ecosystem model is combed, which enriches the model base, knowledge base and rule base of port digital twin.