cryptic diversity │ molecular phylogeny │ morphology │ ultrastructure │ ecology
Traditional view of the euglenoids relationships was based especially on the number of flagella and the nutrition. Alternatively, another morphological features were designed by Leedale (1967) and euglenoids were divided into six main orders: Euglenales, Eutreptiales and Euglenamorphales which contain mainly photosynthetic genera and some colourless species. The rest of colourless genera are from the orders Rhabdomonadales (osmotrophic genera), Sphenomonadales (osmotrophic or phagotrophic genera) and Heteronematales (phagotrophic genera with feeding apparatus). Series of molecular studies show that relationships between euglenoids are not as clear as it seems according to morphologically based taxonomic studies. Euglenoids are close relatives to the kinetoplastids and diplonemids. Together these organisms form a group Euglenozoa (Triemer & Farmer, 2007, Cavalier-Smith, 2016) and current knowledge of the evolutionary relationships of euglenoids is shown in the Figure 1. Photoautotrophic euglenoids are a monophyletic group with the basal mixotrophic genus Rapaza (Karnkowska et al., 2015, Kim et al., 2015). Photoautotrophic euglenoids, together with some colourless groups (Peranemids, Anisonemids, Aphagea group and Neometanema), belong to the bigger group Spirocuta with spiral pellicle strips arrangement as synapomorphy (Cavalier-Smith, 2016). Phylogenetic relationships of the colourless groups seem to be a little bit complicated as is visible in the Figure 1A, in which some genera are polyphyletic (e.g. Astasia and Distigma). The position of several groups is not clear and some genera still miss molecular data for the evaluation of their position in this tree (e.g. Urceolus and Sphenomonas).
Figure 1. Phylogenetic relationships within euglenoids and related organisms. (A) Current knowledge of the evolutionary tree of euglenoids, based primarily on SSU rRNA gene phylogenies, photoautotrophic taxa are shown in green, primary osmotrophs in blue, “typical” phagotrophic taxa in black, and symbiontids in orange, taxa shown in gray are important taxa whose positions are inferred from morphological information (no molecular data are currently available), double lines on a branch represent paraphyletic groups (according to Leander et al., 2017, modified and simplified); (B) Schematic diagram of phylogenetic relationship of the phototrophic euglenoids, based on multigenes (nSSU, nLSU, cpSSU, hsp90 and psbO) Bayeasian analysis (according to Karnkowska et al., 2015, modified and simplified).
Can be ubiquite species cryptic?
Which relationships between morphologically similar species creating complexes?
Which genera need to be revised?
Photosyntetic euglenoids form monophyletic lineage divided into two orders: Eutreptiales including marine and brackish genera – Eutreptiella and Eutreptia – with characteristic two or four flagella (Bicudo & Menezes, 2016, Yamaguchi et al., 2012) and Euglenales with the dominance of freshwater species belonging to the common algal communities, e.g. plankton, mephyton and neuston (Wołowski & Hindák, 2005). In this order, Kim et al. (2010) established new family Phacaceae including genera Discoplastis, Lepocinclis and Phacus, the rest of euglenales genera belong to the family Euglenaceae (name designed by Dujardin, 1841, confirmed by Silva, 1980). A common feature of genera from Phacaceae family is the presence of small discoid chloroplasts without pyrenoids and 32 pellicle strips (Kim et al., 2010, Leander et al., 2001, 2007). Phacus and Lepocinclis have similarities in pellicle strips arrangement and of dimorphic paramylon grains (Monfilis et al., 2011). Discoplastis seem to be a monophyletic sister group to the Phacus and Lepocinclis (Karnkowska et al., 2015, Triemer et al., 2006). Two Phacus species (P. warszewiczii and P. limnophila) branched independently from other species of the genus and they are characterized by morphological differences: P. warszewiczii has longitudinally twisted cell body having here curved ridges; P. limnophila has elongated spindles shape and two long, straight, rod-shaped paramylon grains (Kim et al., 2015).
In the Euglenaceae group, several lineages have developed – Euglenaformis proxima as basal organisms of this group, Trachelomonas-Strombomonas-Colacium lineage, Monomorphina-Cryptoglena-Euglenaria lineage and Euglena-lineage (Karnkowska et al., 2015).
The group including loricate genera Trachelomonas and Strombomonas together with Colacium seems to be monophylethic. This relationship is not supported only by the molecular data (Karnkowska et al., 2015, Ciugulea et al., 2008), but there is also morphological synapomorphisms: 40-48 pellicle strips (Leander et al., 2001, 2007) and the production of extracellular mucosal structures – loricae and stalks (see above). Divergency between Trachelomonas and Strombomonas was studied by Deflandre (1930), who established Strombomonas as a new independent genus based on the differences of lorica morphology. According to the results of Marin et al. (2003), these genera were synonymous, but the work of Ciugulea et al. (2008) supported the hypothesis about independence of these genera and their sister position.
Genus Euglena, according to the traditional morphology-based concept, seemed to be polyphyletic and it was supported by several studies using molecular methods (Karnkowska-Ishikawa et al., 2011, 2012, 2013, Kosmala et al., 2005, 2009, Marin et al., 2003, Wang & Chen, 2004, Zakryś et al., 2001, 2002, 2004). As the results of these studies showed, several new genera from this genus were separated, e.g. Discoplastis (Triemer et al., 2006), Euglenaformis (Bennet et al., 2014) and Euglenaria (Linton et al., 2010), and several species were moved to another genera, e.g. Euglena acus, E. oxyuris, E. spirogyra, E. tripteris to the genus Lepocinclis (Marin et al., 2003) and Euglena limnophila to the genus Phacus (Linton et al., 2010). As well as two species of Euglena that fall outside Euglena clade: E. archaeoplastidiata and E. velata (Karnkowska-Ishikawa et al., 2012, Kim et al. 2010, 2015, Kim & Shin, 2008).
Common features for the monophyletic lineage of Monomorphina and Cryptoglena genera are rigid pellicle from small number (15-16) of broad pellicle strips and cell containing only one plastid (Leander et al., 2001, 2007).
Genus Euglenaria seems to be monophyletic with some morphological features (e.g. lobate plastids with diplopyrenoids) similar to those other Euglena species, but there is a distinction in molecular signatures in nuclear SSU rDNA sequences (Linton et al., 2010, Karnkowska-Ishikawa et al., 2012). According to Leander et al. (2017), the phylogenetic position of that Euglenaria-lineage is not well resolved.
The solution to these taxonomic problems came with methods of molecular phylogenetics at the turn of the 20th and the 21st centuries. Zakryś et al. (2002) resolved problem of Euglena geniculata and E. myxocylindracea, which are according to authors‘ results genetically and morphologically identical; Marin et al. (2003) studied euglenoids phylogeny based on SSU rDNA sequences, authors renewed the genus Monomorphina, which was a sub-part of the genus Phacus and made changes in the taxonomic position of common worldwide members of the genus Euglena (E. acus, E. oxyuris, E. spirogyra, E. tripteris) to the genus Lepocinclis; Shin & Triemer (2004) focused on a type of species of the genus Euglena – E. viridis; Zakryś et al. (2004) studied isolates of the common Euglena agilis using combination of ITS2 of extrachromosomal rDNA and the chloroplast SSU rDNA sequences. During the last ten years, several revisions were made in the number of euglenoid genera, e.g. Euglena sensu lato (Bennet et al., 2014, Karnkowska-Ishikawa et al., 2011, 2012, 2013, Kosmala et al., 2009, Linton et al., 2010, Triemer et al., 2006), Lepocinclis (Kosmala et al., 2005), Monomorphina (Kosmala et al., 2007b, Nudelman et al., 2005) and Phacus (Karnkowska-Ishikawa, et al., 2010, Kosmala et al., 2007a, Łukomska-Kowalczyk, 2015). The question about cryptic species diversity is studied and discussed in several works, Kim et al. (2013a, b) described quite a high number of cryptic species in the genus Monomorphina and Cryptoglena; Kim & Shin (2014) studied the cryptic diversity in the genus Phacus with the description of seven new species and the work about morphological and genetic diversity of Euglena deses group with an emphasis on cryptic species was published by Kim et al. (2016).
The effort to understand the taxonomy and phylogenetic relationships of euglenoids also brings new methods. Bennet & Triemer (2012) designed new methods for obtaining nuclear gene sequences from filed samples. Authors use this method successfully in the study of position of Lepocinclis horridus (formerly Phacus horridus) and Lepocinclis helicoideus (formerly Euglena helicoideus). This method could solve the problem with the complicated or impossible cultivation of some species and the inability to work with cultures of these organisms. Łukomska-Kowalczyk et al. (2016) focused on the use of barcoding in the photosynthetic euglenoids and, as a result of their studies, authors suggested two molecular markers (COI and 18S rDNA) as potential DNA barcodes. Despite a significant progress in this area, there is still a relatively high number of species complexes together with some genera, e.g. Colacium, Strombomonas and Trachelomonas, where molecular data is insufficient or fully missing – these problems bring tasks and challenges for further researches.
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