Edaphochlamys debaryana (Goroschankin) Pröschold & Darienko 2018

Pröschold, Thomas, Darienko, Tatyana, Krienitz, Lothar & Coleman, Annette W., 2018, Chlamydomonas schloesseri sp. nov. (Chlamydophyceae, Chlorophyta) revealed by morphology, autolysin cross experiments, and multiple gene analyses, Phytotaxa 362 (1), pp. 21-38 : 34

Edaphochlamys debaryana (Goroschankin) Pröschold & Darienko comb. nov.

Basionym: Chlamydomonas debaryana Goroschankin (1891) , Bull. Soc. Imp. Nat. Moscou, N.S. 5: 106–108, fig. 9–12.

Comment: All strains designated as C. debaryana were almost identical in morphology and SSU and ITS rDNA sequences. Only the strain SAG 70.81 differed from C. debaryana as shown in Fig. 3 . This strain could be preliminary identified as C. cf. latifrons ; however, this needs further studies.

As demonstrated in our study, unicellular genera Chlamydomonas and Edaphochlamys are closely related to the colonial genera of the Goniaceae , Tetrabaenaceae , and Volvocaceae . The Goniaceae include the genera Gonium and Astrephomene ( Nozaki & Kuroiwa, 1992) , the Tetrabaenaceae the genera Basichlamys and Tetrabaena ( Nozaki & Ito, 1994) . Both families were considered as intermediate families between unicellular taxa such as Chlamydomonas and the Volvocaceae , which comprises the colonial genera Pandorina , Volvulina , Yamagishiella , Eudorina , Platydorina , Colemanosphaera , Pleodorina , and Volvox ( Nozaki et al., 2000, Nozaki, 2003, Nozaki et al., 2014). However, in contrast to the phylogenies using chloroplast genes, where Chlamydomonas and Vitreochlamys species were often at the base of the Volvocales sensu stricto, the phylogenetic analyses of SSU and ITS rDNA sequences always demonstrated that the unicellular taxa are distributed among the colonial lineages ( Nakada et al., 2016; this study). Our study here revealed five lineages ( Chlamydomonas , Edaphochlamys , Tetrabaenaceae , Goniaceae , and Volvocaceae ) among the Core-Reinhardtinia, four of them are highly supported in bootstrap and Bayesian analyses. Only the family Goniaceae (* in Figs 3–4 ) was not supported in our analyses. The genera Chlamydomonas and Edaphochlamys were topologically sisters of the families Goniaceae / Volvocaceae and Tetrabaenaceae , respectively. These results were partially confirmed by the activity tests of gamete lytic enzymes (GLE) derived from Chlamydomonas reinhardtii . In these tests the GLE dissolved not only the cell walls of several Chlamydomonas species such as C. reinhardtii , C. incerta , and partly C. debaryana (only SAG 26.72, but no reaction by SAG 4.72 and SAG 14.72), it also degraded those of the colonial genera Gonium , Astrephomene , Basichlamys (= Gonium sacculiferum ), and Tetrabaena (= Gonium sociale ; Matsuda et al., 1987, Matsuda, 1988). All these data indicated the close relationship of these taxa. In contrast, the gamete autolysin had no influence on the other colonial Volvocaceae .

However, these results raised the question of why the phylogenetic analyses using nuclear and plastid-coding genes showed different tree topologies. Possible explanations are: (1) The phylogenies of nuclear and plastid-coding genes were based on different data sets. Unfortunately, no congruent alignments including sequences of the same strains have been available until now. (2) Nuclear and chloroplast genes of unicellular and colonial taxa have different evolutionary rates. Whereas the nuclear genes appear to evolve at similar rates, the plastid-coding genes evolved differently among unicellular and colonial species. To address these questions further, we created small congruent datasets (ITS and rbc L) of representatives of all lineages and analyzed them separately ( Fig. 4 ). The analyses of both datasets clearly showed different tree topologies. The ITS phylogeny revealed five clades, which represented the three families (the Goniaceae is only weakly supported) and the two genera Chlamydomonas and Edaphochlamys . In contrast to highly or moderately supported lineages using ITS, the rbc L phylogeny did not show this topology and most lineages were not supported by bootstrap and Bayesian analyses. This clearly demonstrated that rbc L is too conserved to achieve a robust phylogenetic resolution. For better understanding of the plastid-coding genes, we re-analyzed the datasets used by Nakada et al. (2016) separately and concatenated (see Figs S1 ; Supplemental Material). The third codon positions were excluded from these analyses because these positions were saturated. The tree topologies using rbc L, atp B, psa A, psa B, and psb C were very different and showed no or only weak support for genera and families. These results were probably caused by different evolutionary rates of each gene and low genetic variations among the plastid-coding genes. The concatenated dataset of all genes also revealed high support for only some of the genera and lineages. As Wang et al. (2014) and references therein highlighted, if the tree topologies of single genes differ significantly, then the gene sequences should not be combined in a concatenated dataset. This resulted in incongruent species trees, which is caused by different genetic history of the selected genes. As consequence of our study, the nuclear SSU and ITS sequences combined in a concatenated dataset produce a much better resolution and therefore should be preferred for taxonomy of the volvocalean algae.

Figures

FIGURE 1. Morphology and phenotypic plasticity of Chlamydomonas schloesseri sp. nov. A–J. vegetative cells, arrow in F. marked the eyespot; K–O. different sporangia; scale bar = 10 μm.

FIGURE 3. Molecular phylogeny of Chlamydomonas and representatives belonging to the Tetrabaenaceae, Goniaceae and Volvocaceae based on SSU and ITS rDNA sequence comparisons. The phylogenetic trees shown were inferred using the maximum likelihood method based on the data sets (2688 aligned positions of 75 taxa) using PAUP 4.0b10. For the analyses the best model was calculated by Modeltest 3.7. The setting of the best model was given as follows: GTR+I+G (base frequencies:A 0.2516, C 0.2232, G 0.2514, T 0.2738; rate matrix A-C 1.3107, A-G 2.3565, A-U 2.4706, C-G 0.4123, C-U 4.2284, G-U 1.0000) with the proportion of invariable sites (I = 0.6048) and gamma shape parameter (G = 0.3904). The branches in bold are highly supported in all analyses (Bayesian values> 0.95 calculated with PHASE and MrBayes; bootstrap values> 70% calculated with PAUP using maximum likelihood, neighbor-joining, maximum parsimony and RAxML using maximum likelihood). The Goniaceae are only moderated supported in bootstrap and Bayesian analyses, which is indicated by an asterisk.

FIGURE 4. Molecular phylogeny of Chlamydomonas and representatives belonging to the Tetrabaenaceae and Goniaceae based on rbcL and ITS rDNA sequence comparisons. The phylogenetic trees shown were inferred using the maximum likelihood method based on the data sets (29 taxa: 915 aligned positions for ITS, 1128 for rbcL) using PAUP 4.0b10. For the analyses the best model was calculated by Modeltest 3.7. The setting of the best model was given as follows: (ITS) GTR+I+G (base frequencies: A 0.2634, C 0.2225, G 0.2192, T 0.2949; rate matrixA-C 1.2503, A-G 2.1014,A-U 2.0643, C-G 0.3957, C-U 3.3009, G-U 1.0000) with the proportion of invariable sites (I = 0.2682) and gamma shape parameter (G = 0.6858); (rbcL) GTR+I+G (base frequencies: A 0.2777, C 0.1688, G 0.2048, T 0.3487; rate matrix A-C 0.2771, A-G 3.5238, A-U 4.0693, C-G 0.6646, C-U 7.4921, G-U 1.0000) with the proportion of invariable sites (I = 0.5523) and gamma shape parameter (G = 0.6480). The branches in bold are highly supported in all analyses (Bayesian values> 0.95 calculated with PHASE and MrBayes; bootstrap values> 70% calculated with PAUP using maximum likelihood, neighbor-joining, maximum parsimony and RAxML using maximum likelihood). The Goniaceae are only weakly supported in bootstrap and Bayesian analyses, which is indicated by an asterisk.