[Background] In the past, there were few studies on the taxonomy of bifidobacteria plasmids, and the phylogenetic classification method of Bifidobacterium plasmid was missing. [Objective] In order to establish a method for phylogenetic classification and identification of natural plasmids in Bifidobacterium, and increase understanding and application of plasmids in the research of bifidobacteria biology. [Methods] We conducted to phylogenetic classification of all sequenced plasmids in Bifidobacterium by the analysis of phylogenetic relationship of replication initiation protein and genome colinearity. [Results] All known natural plasmids of Bifidobacterium were divided into six different types of plasmid families and three unique complex plasmids. Plasmids of type III and VI family were further divided into different subtype groups. Plasmids of the type III and VI family can be further divided into different subtype groups. Plasmids of the type III family were the main type and dominant family of natural plasmids in Bifidobacterium. Plasmid subtypes of type VI family were most abundantly classified. At the plasmid family level, the classification results of the two methods were completely identical. [Conclusion] These results comprehensively and accurately revealed the phylogenetic classification relationship between all the plasmids that were analyzed, and established the phylogenetic classification standard and method of natural plasmids in Bifidobacterium, which provided important theoretical references and classification basis for the future classification and identification of natural plasmids in Bifidobacterium.