Conservation of taxonomically difficult species: the case of the Australian orchid, Microtis angusii Nicola S. Flanagan 1,3, *, Rod Peakall 1 , Mark A. Clements 2 & J. Tupac Otero 2 1 School of Botany and Zoology, The Australian National University, Canberra, ACT 0200, Australia; 2 Centre for Plant Biodiversity Research, GPO Box 1600 Canberra, ACT 2601, Australia; 3 Present address: Biochemistry Department, University College Cork, Cork, Ireland (*Corresponding author: Phone: +353-21- 4904124; Fax: +353-21-4904259; E-mail: nicflanagan@fastmail.fm) Received 18 May 2005; accepted 3 January 2006 Key words: AFLP, clonality, ITS, practical outcomes, species diagnosis Abstract As species are the common currency for conservation efforts, their accurate description is essential for efficient preservation of biological diversity. The genus Microtis (Orchidaceae) is typified by a paucity of consistent morphological characters, confounding taxonomic attempts. We report the results of a study of the conservation genetics of the recently discovered, endangered Australian orchid species M. angusii (Jones). This species was known only from one small population, with identification of further populations hampered by taxonomic difficulties. We used a combination of 122 AFLP markers and DNA sequence variation in the ribosomal ITS gene region to investigate the population genetic structure of the type population of M. angusii. Six further putative M. angusii populations were also analysed with these markers. Two of these populations showed high genetic affinity to M. angusii, bearing identical ITS sequences. Both the type and a second population were invariable across all AFLP loci. The third population, 3 km distant, showed minor genetic differentiation. These two new populations warrant immediate protection. Phylogenetic relationships between M. angusii and close relatives revealed its genetic affiliation to an unidentified, more distant pop- ulation, and to the species M. unifolia. Given the propensity in Microtis for both selfing and clonality, mechanisms that both reduce within population variability and promote divergence between isolated pop- ulations, we recommend an extended study of both the genetic structure and breeding systems in the M. angusii/M. unifolia group, in order to ensure that the protection provided is both adequate and justified. Introduction Accurate species-level taxonomic assessment is essential for the effective conservation of biological diversity. Typically, species are a natural taxonomic rank that forms the basis for both conservation assessments and management (Mace 2004). The assessment of habitats for prioritizing conservation efforts requires accurate lists of the constituent species, while effective and efficient conservation of individual threatened species requires that their taxonomic status be unambiguous. This widespread use of species as the ‘unit’ of conservation is largely a consequence of their unique status in the taxo- nomic hierarchy. Species are often seen as having an objective reality, with gene flow, and thus the evolutionary potential to generate further diversity, being mainly restricted to within species (Hey 2001). Despite considerable discussion in the literature regarding the concept of a species, practical limi- tations mean that most species are still described through the traditional assessment of morphological characters. However, it is now widely recognized that for biological diversity to persist, it is essential to Conservation Genetics (2006) 7:847–859 Ó Springer 2006 DOI 10.1007/s10592-006-9119-8