Two tapeworm specimens collected in northeast China in 2009 and 2011 were identified as Diphyllobothrium latum based on morphological criteria. Molecular methods were used to confirm their identity and analyze genetic variations compared with published data for this species. Species identity was confirmed by molecular characterization of the 18S rDNA partial sequence, complete sequences of internal transcribed spacers (ITSs) and 5.8S rDNA, and partial sequences of mitochondrial cytochrome c oxidase subunit 1 (cox1) and mitochondrial NADH dehydrogenase subunit 5 (nad5). PCR amplification and sequence analysis of 18S rDNA (1472 bp), ITS regions (1218 bp), cox1 (885 bp), and nad5 (1028 bp) revealed that these four sequences showed more than 99% identity to reference sequences for D. latum, confirming that this species is D. latum. To date, a total of 12 diphyllobothriosis cases have been documented in China. This study represents the first molecular characterization of D. latum in China, providing molecular evidence of human diphyllobothriosis in China.