THE SPECIES EUGLENA DESES (EUGLENACEAE) REVISITED: NEW MORPHOLOGICAL AND MOLECULAR DATA 1 Anna Karnkowska-Ishikawa, Rafal Milanowski, and Bo _ zena Zakrys´ 2 Department of Plant Systematics and Geography, Faculty of Biology, University of Warsaw, Al. Ujazdowskie 4, PL-00-478 Warszawa, Poland For this study, we have examined the literature and the morphological diversity, as well as analyzed the nuclear SSU rDNA sequences of two very common and cosmopolitan species formerly known as Euglena deses Ehrenb. and Euglena intermedia (G. A. Klebs) F. Schmitz. Our studies have shown that there is evidence for distinguishing only one species (E. deses). Here, we define new diagnostic features for E. deses, namely, periplast ornamenta- tion (the presence of small papillae—discovered for the first time in this species) and the lateral location of the anterior canal opening, from which the flagel- lum emerges. We also designate the epitype and emend the diagnosis for E. deses. Key index words: Euglena adhaerens ; Euglena deses ; Euglena satelles ; Euglenea; Euglenida; phylogeny; SSU rDNA; taxonomical revision Abbreviations: BA, Bayesian analysis; bs, nonpara- metric bootstrap; ML, maximum likelihood; MP, maximum parsimony; NJ, neighbor joining; pp, posterior probability E. deses has occupied researchers since long ago, both because it is one of the first green euglenoid species described by Ehrenberg (1835, 1838) and because it is very commonly encountered through- out the world. According to Pringsheim (1956), E. deses is nearly as common as Euglena viridis. After years of multiple studies, the result is a literature composed of numerous descriptions of taxa with morphology similar to that of E. deses, with the rela- tionships between the taxa and their distinguishing criteria still being unclear: E. deses var. intermedia G. A. Klebs 1883, E. intermedia (G. A. Klebs) F. Schmitz 1884, E. intermedia var. klebsii Lemmermann 1910, E. deses var. tenuis Lemmermann 1910, E. deses var. grac- ilis Playfair 1921, E. deses var. minuta Playfair 1921, Euglena sima Wermel 1924, Euglena klebsii (Lem- merm.) Mainx 1927, E. intermedia var. brevis F. E. Fritsch et M. F. Rich 1930, Euglena satelles Braslavska- Spectorova 1937, Euglena adhaerens Matvienko 1938, E. deses var. carterae E. G. Pringsheim 1953, E. interme- dia fo. major T. G. Popova 1955, E. deses fo. mesnili E. G. Pringsheim 1956, E. deses fo. klebsii (Lem- merm.) T. G. Popova 1966, E. deses fo. major T. G. Popova 1966, E. deses var. digrana Zakrys´ 1986, and E. intermedia var. acidophila Z. X. Shi 1989. The most recent research, undertaken to clarify the issues contended in the literature for over a century, took place relatively not long ago. It concerned the ultrastructure of the cells of cultivated strains repre- senting taxa morphologically similar to E. deses (Zak- rys´ et al. 2001, Kusel-Fetzmann and Weidinger 2008). The studies have revealed the presence of pyrenoids (an important diagnostic feature) in all species within this group, which we have labeled with the working name ‘‘E. deses group.’’ Moreover, in the last of these recent research articles, the authors took note of a new feature, which may be of diagnostic value: the lat- eral or apical location of the opening to the anterior canal, from which the flagellum emerges (Kusel-Fetz- mann and Weidinger 2008). This finding has inspired us to take on the research presented here, through which we compare the current morphological diag- nostic features with the results of phylogenetic analy- ses carried out for nuclear SSU rDNA sequences. MATERIALS AND METHODS Euglenoid strains and culture conditions. The strains used in this study are described in Table S1 (in the supplementary material). All strains were cultivated in a liquid soil-water medium, enriched by a small piece of garden pea (medium 3c; Schlo ¨sser 1994), under identical conditions, in a growth chamber maintained at 17°C and 16:8 light:dark, ca. 27 lmol photons Æ m )2 Æ s )1 provided by cool-white fluorescent tubes (Philips, Amsterdam, the Netherlands). LM observations. Observation of morphological features (cell size and shape, shape and number of large paramylon grains, periplast ornamentation, chloroplast size) were per- formed using a light microscope (Nikon Eclipse E-600 with Nomarski contrast; Nikon, Tokyo, Japan), equipped with the software for image recording and processing. Photographic documentation was made using the digital camera, Nikon DX-1200, connected to a microscope. Cultures were sampled every 2 weeks, for periods of 3–4 months. Such sampling enabled us to observe all the cells during their developmental stages, from the young (immediately after division) to the steadily aging to the old. Biometric studies. The LUCIA measurement program (Lab- oratory Imaging s. r. o., Prague, Czech Republic) was used to perform biometric studies. Three parameters were measured 1 Received 24 June 2010. Accepted 2 November 2010. 2 Author for correspondence: e-mail zakrys@biol.uw.edu.pl. J. Phycol. 47, 653–661 (2011) Ó 2011 Phycological Society of America DOI: 10.1111/j.1529-8817.2011.00982.x 653