Phylogenetic relationships of species within the tribe Labiostrongylinea (Nematoda: Cloacinidae) from Australian marsupials based on ribosomal DNA spacer sequence data Neil B. Chilton a, b, ⁎, Florence Huby-Chilton a , Ian Beveridge a , Lesley R. Smales c, d , Robin B. Gasser a , Ross H. Andrews e a Department of Veterinary Science, University of Melbourne, Werribee, Victoria 3030, Australia b Department of Biology, University of Saskatoon, Saskatchewan, Canada S7N 5E2 c School of Biological and Environmental Science, Central Queensland University, Rockhampton, Queensland 4702, Australia d South Australian Museum, Adelaide, South Australia 5000, Australia e Department of Parasitology, Liver Fluke and Cholangiocarcinoma Research Centre, Khon Kaen University, Khon Kaen 40002, Thailand abstract article info Article history: Received 6 January 2011 Received in revised form 6 June 2011 Accepted 11 June 2011 Available online 17 June 2011 Keywords: Nematodes Labiostrongylinea Phylogenetic relationships Ribosomal DNA Internal transcribed spacers Macropodid and potoroid marsupials Parasitic nematodes of the tribe Labiostrongylinea (Family Cloacinidae) occur in the stomachs of a wide variety of potoroid and macropodid marsupials in Australia, Papua Indonesia and Papua New Guinea. The aim of the present study was to infer the evolutionary relationships of the five genera of labiostrongyline nematodes that occur in Australian potoroids and macropodids using sequence data of the nuclear first and second internal transcribed spacers of ribosomal DNA. The phylogenetic analyses resulted in the separation of the Labiostrongylinea into two major groups reflecting coevolution between hosts and parasites. Two nematode species belonging to the genus Potorostrongylus formed a sister group to the remaining species of the Labiostrongylinea. This genus occurs exclusively in potoroid marsupials, which are considered to be basal to the macropodid marsupials. The second major group included species of Labiostrongylus, Labiosimplex, Labiomultiplex and Parazoniolaimus, all of which occur in macropodids. These species formed two distinct clades, one predominating in the host genera Thylogale and Onychogalea, and the second in the genus Macropus, which includes the more recent macropodids. However, there is also evidence of colonisation by both nematode clades of relatively unrelated hosts. In addition, genetic differences among individuals of Lm. eugenii from geographically isolated populations of M. eugenii, and among Ls. longispicularis from different subspecies of M. robustus suggest the existence of sibling species that may have arisen by allopatric speciation. The broad coevolutionary relationship between the labiostrongyline nematodes and their marsupial hosts therefore represents a mixture of potential cospeciation and colonisation events. © 2011 Elsevier Ireland Ltd. All rights reserved. 1. Introduction Coevolution between parasites and their hosts is a well-recognised phenomenon [1] even if the relative roles of cospeciation and colonisation in this process are debatable. The cloacinine nematodes (Strongylida: Cloacininae) that occur in the sacculated forestomachs of kangaroos and wallabies (Macropodidae) and rat-kangaroos (Potoroidae) are a diverse assemblage of parasites, comprising 36 genera and more than 255 species arranged in six tribes: Cloacininea, Coronostrongylinea, Labiostrongylinea, Macropostrongylinea, Pharyngos- trongylinea and Zoniolaiminea [2]. Cloacinine nematodes have clearly coevolved with their macropodoid hosts (Macropodidae + Potoroidae) as sacculated forestomachs have evolved in macropodoids over the last 30 million years [3–5]. However, the mechanisms involved in the evolution of the numerous groups of cloacinine nematodes are far from clear. Beveridge and Chilton [2] examined the phylogenetic relationships of eight relatively small genera of the subfamily Cloacininae using cladistic analyses of morphological characters. They concluded that, although there was some evidence of cospeciation in the genera examined, colonisation appeared to be the more important mechanism of diversification within this nematode radiation [2]. Their conclusions are potentially limited by the fact that they examined genera comprising a relatively small number of species, and the analyses were conducted exclusively on morphological characters. Using a larger genus or series of closely related genera combined with molecular techniques may provide more significant insights into the evolution of this diverse group of nematodes. The Labiostrongylinea contains at least 35 species [2,6] that are distinguished from members of the other tribes by differences in the morphology of lips, buccal capsule and oesophagus [7]. The type genus, Labiostrongylus, was subdivided by Smales [8,9] into three new Parasitology International 60 (2011) 381–387 ⁎ Corresponding author at: Department of Biology, University of Saskatoon, Saskatchewan, Canada S7N 5E2. Tel.: +1 306 9664407; fax: +1 306 966446. E-mail address: neil.chilton@usask.ca (N.B. Chilton). 1383-5769/$ – see front matter © 2011 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.parint.2011.06.012 Contents lists available at ScienceDirect Parasitology International journal homepage: www.elsevier.com/locate/parint