Aquatic Botany 134 (2016) 113–119 Contents lists available at ScienceDirect Aquatic Botany journal homepage: www.elsevier.com/locate/aquabot Molecular evaluation of the validity of the morphological characters of three Swedish Chara sections: Chara, Grovesia, and Desvauxia (Charales, Charophyceae) Petra Nowak a,∗ , Hendrik Schubert a , Ralf Schaible b a University of Rostock, Biosciences, Ecology, Albert-Einstein-Str. 3, 18059 Rostock, Germany b Max Planck Institute for Demographic Research, Konrad Zuse Str. 1, 18057 Rostock, Germany a r t i c l e i n f o Article history: Received 29 September 2015 Received in revised form 2 August 2016 Accepted 2 August 2016 Available online 3 August 2016 Keywords: Chara Morphology Phylogeny Species concept Taxonomy a b s t r a c t Charophytes are aquatic macrophytes with a complex morphology that belong to the Streptophytina phylum. The high degree of morphological plasticity in the genus Chara has led researchers to propose two competing species concepts, a micro- and macrospecies concept, the relevance of which we tested by phylogenetic analyses. We conducted a phylogenetic survey based on the phylogenetic relationships between 14 species of the genus Chara using molecular data from two nuclear (18S rDNA, nrITS-1) and three chloroplast (rbcL, atpB, matK) markers. As the combined molecular data indicated that the overall level of genetic divergence between the investigated species was, at ≤3%, relatively low, a robust species delineation was difficult to achieve. The sequence dataset of 6214 base pairs did not separate individuals of closely related taxa with distinct morphological characters; e.g., the interspecific genetic variability for C. contraria and C. filiformis or C. rudis and C. hispida was zero. In contrast, the molecular data revealed clear differences between C. vulgaris and C. contraria; species that are distinguished only by their relative levels of development of primary and secondary cortex cell rows. The low levels of genetic variability between as well as within the taxa studied here suggest that there has been a relatively recent, but still incomplete, speciation of Chara species. © 2016 Elsevier B.V. All rights reserved. 1. Introduction Charophytes are a group of morphologically highly differenti- ated algae that belong to the Streptophyta phylum (Mattox and Stewart, 1985). As they are assumed to be closely related to the ancestors of land plants (Laurin-Lemay et al., 2012), these organ- isms can serve as models for the development of acclimation mechanisms that allow for survival in terrestrial habitats (Braun et al., 2007). In addition, charophytes are of interest to researchers because of their potential applications, including as bioindicators of environmental quality, for which a reliable determination of charophytes by robust characters is essential (e.g., Krause, 1997; Steinhardt et al., 2009). The taxonomy of charophyte algae relies on an almost purely phenetic species concept; i.e., the classification is based on overall similarities in the morphological characters of the organisms. For charophytes (and many other algae groups), the absence of mor- ∗ Corresponding author. E-mail address: petra.nowak@uni-rostock.de (P. Nowak). phological traits with strong links to reproduction make it difficult to validate the traits used. Because there is no direct connection between the vegeta- tive morphological characters used for species delineation and reproduction (biological species concept), there are two compet- ing concepts of the level for “species”-ranked taxonomic units for charophytes: a microspecies concept (e.g., Krause, 1997) that posits the existence of approximately 30 distinct European Chara species, and a macrospecies concept (Wood and Imahori, 1965) that recognises only 18 distinct species of genus Chara globally. The macrospecies tend to be polymorphic, with each macrospecies encompassing a number of subspecies, varieties, and forms—most of which are raised to the species level in the microspecies concept. The advocates of both concepts use vegetative traits almost exclu- sively as diagnostic characters (especially the number of cortex cells and the number, shape, and length of stipulodes and spines), and differ only in their conclusions regarding the taxonomic rele- vance of these traits. In contrast to the microspecies concept the macrospecies concept rejects the assumption that the reproduc- tion mode (especially dioecism and monoecism) is a character that should be used for species delineation. This difference was http://dx.doi.org/10.1016/j.aquabot.2016.08.001 0304-3770/© 2016 Elsevier B.V. All rights reserved.