Phylogenetic relationships of coexisting Heterocypris (Crustacea, Ostracoda) lineages with different reproductive modes from Lampedusa Island (Italy) Valeria Rossi a, * , Andrea Gandolfi b , Francesca Baraldi a,b , Carlo Bellavere a , Paolo Menozzi a a Department of Environmental Sciences, University of Parma, Viale G.P.Usberti 11/a Campus, 43100 Parma, Italy b Natural Resources Department, IASMA Research Centre, San Michele a/Adige, Trento, Italy Received 25 October 2006; revised 2 April 2007; accepted 12 April 2007 Available online 3 May 2007 Abstract Coexisting bisexual and unisexual populations of individuals belonging to the genus Heterocypris are found in ephemeral freshwater ponds on the island of Lampedusa (Pelagie Islands, Italy). Different reproductive modes were associated with a peculiar morphological trait: a lamella hyalina on the posterior margin of the left valve was observed in amphimictic females, a feature missing in apomictic females. In order to clarify the phylogenetic relationships among taxa with different morphological traits and reproductive modes, we used four polymorphic enzyme loci (GPI, ICD2, MPI and PGM) and mitochondrial DNA 16S ribosomal sequences. We identified three main evolutionary units that showed a combination of morphological and reproductive characteristics: (1) amphimictic females of H. barbara with a lamella hyalina, according to the typical feature of the species, and apomictic females of H. barbara without lamella that are sympatric in one temporary pond; (2) apomictic females of H. incongruens without a lamella, as typical of the species; (3) apomictic females without a lamella, living in sympatry with H. barbara, but characterised by a high genetic diversity from both H. incongruens and H. barbara. We discuss the possible origin of apomictic lineages as a result of independent transition episodes to apomixis from different sexual ancestors. Time of divergence reflected the genetic differentiation within and among multiple ancestors and different possible routes to parthenogenesis. Ó 2007 Elsevier Inc. All rights reserved. Keywords: Evolution of sex; Mitochondrial DNA; Allozymes; Heterocypris incongruens; H. barbara; Temporary ponds; Ostracods 1. Introduction The origin of asexual organisms that naturally coexist with sexual relatives may hold the key to comprehending the maintenance of sex and recombination, a long-standing problem in evolutionary biology (Bell, 1982; Maynard Smith, 1986; Stearns, 1987). Most ways in which partheno- genetic lineages could arise (spontaneous loss of sex, conta- gious origin, interspecific and intraspecific hybridisation) and persist may be demonstrable if sexual and parthenoge- netic lineages coexist (Simon et al., 2003). In hermaphro- dites, in organisms with environmental sex determination and cyclical parthenogens, and in gynogenetic and hybridogenetic sperm dependent asexual forms, mixed populations are common and coexistence may be necessary (Booj and Guldemond, 1984; Hebert, 1987; Spolsky et al., 1992a; Vrijenhoek and Pfeiler, 1997; Weinzierl et al., 1999; Ciros-Perez et al., 2002; Simon et al., 2002; Pongratz et al., 2003; Schley et al., 2004). In some cases biochemical and molecular tools were used to disentangle age and origin of asexual lineages and to evaluate how past and actual gene flow between parthenogenetic lineages and close sex- ual relatives may explain persistence and genetic variability of clonal lineages (Innes and Hebert, 1988; Quattro et al., 1055-7903/$ - see front matter Ó 2007 Elsevier Inc. All rights reserved. doi:10.1016/j.ympev.2007.04.013 * Corresponding author. Fax: +39 0521 905402. E-mail address: valeria.rossi@unipr.it (V. Rossi). www.elsevier.com/locate/ympev Molecular Phylogenetics and Evolution 44 (2007) 1273–1283