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Clonorchiasis is a neglected tropical disease caused by the Chinese liver fluke, Clonorchis sinensis, and is often associated with a malignant form of bile duct cancer (cholangiocarcinoma). Although some aspects of the epidemiology of clonorchiasis are understood, little is known about the genetics of C. sinensis populations. Here, we conducted a comprehensive genetic exploration of C. sinensis from endemic geographic regions using complete mitochondrial protein gene sets. Genomic DNA samples from C. sinensis individuals (n = 183) collected from cats and dogs in China (provinces of Guangdong, Guangxi, Hunan, Heilongjiang and Jilin) as well as from rats infected with metacercariae from cyprinid fish from the Russian Far East (Primorsky Krai region) were deep sequenced using the BGISEQ-500 platform. Informatic analyses of mitochondrial protein gene data sets revealed marked genetic variation within C. sinensis; significant variation was identified within and among individual worms from distinct geographical locations. No clear affiliation with a particular location or host species was evident, suggesting a high rate of dispersal of the parasite across endemic regions. The present work provides a foundation for future biological, epidemiological and ecological studies using mitochondrial protein gene data sets, which could aid in elucidating associations between particular C. sinensis genotypes/haplotypes and the pathogenesis or severity of clonorchiasis and its complications (including cholangiocarcinoma) in humans. Author summary: Clonorchiasis is an important neglected tropical disease caused by the Chinese liver fluke, Clonorchis sinensis, which can induce malignant bile duct cancer (cholangiocarcinoma). Little precise information is available on the biology, epidemiology and population genetics of C. sinensis. For this reason, we explored here the genetic composition of C. sinensis populations in distinct endemic areas in China and Russia. Using a deep sequencing-informatic approach, we revealed marked mitochondrial genetic variation within and between individuals and populations of C. sinensis, with no particular affiliation with geographic or host origin. These molecular findings and the methodology established should underpin future genetic studies of C. sinensis causing human clonorchiasis and associated complications (cancer) as well as transmission patterns in endemic regions. [ABSTRACT FROM AUTHOR]