Acta Tropica 98 (2006) 25–33 Paragonimus heterotremus Chen and Hsia (1964), in Vietnam: A molecular identification and relationships of isolates from different hosts and geographical origins Thanh H. Le a,∗ , Nguyen Van De b , David Blair c , Donald P. McManus d , Hideto Kino e , Takeshi Agatsuma f a Institute of Biotechnology, Vietnam b Institute for Malariology, Parasitology and Entomology, Vietnam c James Cook University, Australia d Queensland Institute of Medical Research, Australia e Hamamatsu University School of Medicine, Japan f Kochi Medical School, Japan Received 29 November 2005; received in revised form 25 December 2005; accepted 11 January 2006 Available online 15 March 2006 Abstract Paragonimus heterotremus Chen and Hsia (1964), and paragonimiasis caused by this species is a newly detected disease in Vietnam. Twelve samples of Paragonimus (Platyhelminthes: Trematoda: Digenea: Paragonimidae) from different life-stages (eggs, miracidia, metacercariae, adults from natural and experimental hosts) and host species (crab, dog, cat and human) were collected in different geographical locations in Vietnam. DNA sequences were obtained from each for partial mitochondrial cytochrome c oxidase subunit 1 (cox1) (387 bp) and the entire second ribosomal internal transcribed spacer (ITS-2) (361 bp). The ITS-2 sequences were identical among all specimens, including those previously reported in GenBank. For cox1, there were sequence differences between specimens from Vietnam (four provinces, different locations) and those from Guangxi (China) and Saraburi (Thailand). Phylogenetic trees inferred from cox1 and ITS-2 sequences using sequence data for 15 P. heterotremus and for other Paragonimus spp. revealed that all P. heterotremus originating from Vietnam, Thailand and China form a distinct group. This information also confirms the identity of the Vietnamese specimens as P. heterotremus. © 2006 Published by Elsevier B.V. Keywords: Mitochondrial DNA; Paragonimus heterotremus; Cytochrome c oxidase subunit 1 (cox1); Internal transcribed spacer-2 (ITS-2); Metac- ercariae; Identification; Phylogeny 1. Introduction Paragonimus heterotremus Chen and Hsia (1964) (synonym Paragonimus tuanshanensis Chung, Ho, ∗ Corresponding author. Tel.: +84 4 7567297; fax: +84 4 8363144. E-mail address: im-ibt@hn.vnn.vn (T.H. Le). Cheng and Tsao, 1964) (see Chen, 1965) is one of over 40 recognized species in the genus Paragonimus Braun, 1899 (see Blair et al., 1999a). This species is reported to occur in southern regions of China as well as Thai- land, Laos and Vietnam (Chen and Hsia, 1964; Miyazaki and Vajrasthira, 1966; Kino et al., 1995; Blair et al., 1999a; De et al., 2003; Doanh et al., 2005; Waikagul and Yoonuan, 2005), and recently, in Manipur, India (Singh, 0001-706X/$ – see front matter © 2006 Published by Elsevier B.V. doi:10.1016/j.actatropica.2006.01.008