Genetic diversity of the Chinese liver fluke Clonorchis sinensis from Russia and Vietnam Galina N. Chelomina a,⇑ , Yulia V. Tatonova a , Nguyen Manh Hung b , Ha Duy Ngo b a Institute of Biology and Soil Science, Far Eastern Branch, Russian Academy of Sciences, 100-letiya Street, 159, Vladivostok 690022, Russia b Institute of Ecology and Biological Resources, Vietnam Academy of Science and Technology, 18 Hoang Quoc Viet Street, Cau Giay Dist., Hanoi, Viet Nam article info Article history: Received 19 March 2014 Received in revised form 1 June 2014 Accepted 2 June 2014 Available online 12 August 2014 Keywords: Clonorchis sinensis cox1 gene COX1 Genetic diversity Intraspecific phylogeny abstract Clonorchiasis is a parasitic disease of high public health importance in many countries in southeastern Asia and is caused by the Chinese liver fluke Clonorchis sinensis. However, the genetic structure and demo- graphic history of its populations has not been sufficiently studied throughout the geographic range of the species and available data are based mainly on partial gene sequencing. In this study, we explored the genetic diversity of the complete 1560 bp cytochrome c oxidase subunit 1 (cox1) gene sequence for geographically isolated C. sinensis populations in Russia and Vietnam, to our knowledge for the first time. The results demonstrated low nucleotide and high haplotype differentiation within and between the two compared regions and a clear geographical vector for the distribution of genetic diversity patterns among the studied populations. These results suggest a deep local adaptation of the parasite to its environment including intermediate hosts and the existence of gene flow across the species’ range. Additionally, we have predicted an amino acid substitution in the functional site of the COX1 protein among the Vietnam- ese populations, which were reported to be difficult to treat with praziquantel. The haplotype networks consisted of several region-specific phylogenetic lineages, the formation of which could have occurred during the most extensive penultimate glaciations in the Pleistocene Epoch. The patterns of genetic diver- sity and demographics are consistent with population growth of the liver fluke in the late Pleistocene fol- lowing the Last Glacial Maximum, indicating the lack of a population bottleneck during the recent past in the species’ history. The data obtained have important implications for understanding the phylogeogra- phy of C. sinensis, its host-parasite interactions, the ability of this parasite to evolve drug resistance, and the epidemiology of clonorchiasis under global climate change. Ó 2014 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved. 1. Introduction Clonorchis sinensis is a parasite from the class Trematoda that infects both people and some species of animals. This liver fluke is one of the most important fish-borne zoonotic parasites in east Asian countries such as China, Korea, Thailand and Vietnam, and the southern Far East of Russia (Mas-Coma and Bargues, 1997; Chai et al., 2005; Keiser and Utzinger, 2009; Besprozvannykh et al., 2012; Tatonova et al., 2012, 2013). More than 15 million peo- ple globally are infected with clonorchiasis, and more than 200 million are at risk of this infection (Fürst et al., 2012; Hong and Fang, 2012). Although a carcinogenic effect of C. sinensis was revealed in the early 1990s (Lee et al., 1993, 1994), this parasite was officially listed as a biological carcinogen to humans (Group 1) quite recently (Bouvard et al., 2009). As a tissue parasite, this fluke contributes to human diseases, disability and death (Kruglyakova et al., 1987; Posokhov et al., 1987; Mas-Coma and Bargues, 1997) due to pathological changes in the liver and bile ducts. For all of these reasons, C. sinensis is of great scientific, med- ical and epidemiological significance. Until recently, it was believed that there were no true clonor- chiasis nidi in the Primorye region of southern Far East Russia. This conclusion was based on an examination of more than 200 wild animals, belonging to different species of carnivores (Posokhov, 2004). However, over approximately the last 20 years, surveillance from the Russian Federal Service on Customers’ Rights Protection and Human Well-being has registered an increase in human clo- norchiasis cases in Russia (Besprozvannykh et al., 2012). Clonor- chiasis nidi were revealed in the native range of freshwater snails belonging to the genus Parafossarulus, and some nidi were estimated to be present due to the introduction of these snails. Thus, the number of clonorchiasis cases increased in recreational areas near Kronshtadtka as a result of the water storage formation http://dx.doi.org/10.1016/j.ijpara.2014.06.009 0020-7519/Ó 2014 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved. ⇑ Corresponding author. Tel.: +7 (423) 231 0410; fax: +7 (423) 231 0193. E-mail address: chelomina@ibss.dvo.ru (G.N. Chelomina). International Journal for Parasitology 44 (2014) 795–810 Contents lists available at ScienceDirect International Journal for Parasitology journal homepage: www.elsevier.com/locate/ijpara