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Article

Phylogeny of Trachelomonas and Strombomonas (Euglenaceae) Based on Morphological and Molecular Data

by
Xiaodie Jiang
1,2,
Xi Chen
1,
Wanting Pang
1,* and
Quanxi Wang
1,*
1
College of Life Sciences, Shanghai Normal University, Shanghai 200234, China
2
School of Environmental and Geographical Sciences, Shanghai Normal University, Shanghai 200234, China
*
Authors to whom correspondence should be addressed.
Diversity 2022, 14(8), 623; https://doi.org/10.3390/d14080623
Submission received: 9 June 2022 / Revised: 29 July 2022 / Accepted: 3 August 2022 / Published: 5 August 2022
(This article belongs to the Special Issue Diversity and Ecology of Algae in China)
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Abstract

:
The classification of Trachelomonas and Strombomonas is based on the morphology of loricae, which may not reflect phylogenetic relationships. There are different views on the relationship between the two genera. It is thus important for researchers to classify the two genera based on more characteristics besides loricae, such as protoplast and molecular data, and to establish a natural classification system. In this study, 29 strains (13 species) of Trachelomonas and 12 strains (eight species) of Strombomonas were collected from various locations in China. The morphological characteristics of Trachelomonas and Strombomonas were observed in the field and culture conditions, and the sequences of nuclear SSU rDNA, nuclear LSU rDNA, plastid-encoded LSU rDNA and plastid-encoded LSU rDNA of strains were obtained. We constructed a phylogenetic tree by combining four gene sequences with maximum likelihood and Bayesian methods. Based on the development of the loricae, it was found that the shape of the loricae and the presence or absence of the collar were relatively stable in culture, while the length of the collar presence or absence of spines and the color of the loricae changed. The phylogenetic tree showed that Trachelomonas and Strombomonas were sister branches, which supported Trachelomonas and Strombomonas as independent genera. Taxa in the genus Strombomonas sorted into two clades and Trachelomonas sorted into five strongly supported clades. Key morphological features could be attributed to each of the clades.

1. Introduction

In 1833, Ehrenberg established the genus Trachelomonas based on cells surrounded by a lorica [1]. Deflandre (1926) improved the taxonomic system of the genus based on the lorica shape and ornamentation [2]. This system included two sections (Rotundatae and Caudatae) and seven subsections (Sphaericae, Ellipticae, Ampulliformes, Saccatae, Longisetae, Colliferae, Accuminatae). Later, Deflandre (1930) separated the subsection Saccatae, which has distinctive collar, possession of a tailpiece, lack of ornamentation, and the ability aggregate particles on the surface of the lorica from the genus Trachelomonas and established a new genus, Strombomonas. However, some authors suggested the taxonomic positioning of organisms that are presently classified as Strombomonas be reconsidered [3]. The classification of Trachelomonas and Strombomonas has been heavily dependent on the morphology of loricae. Only Pringsheim classified 13 Trachelomonas species into 6 groups by combining the morphology of loricae and protoplasts [4]. Singh [5], Rosowski et al. [6], Barnes et al. [7], and Wang Quanxi et al. [8] suggested both loricae and protoplasts characteristics be considered in the systematic classification of Trachelomonas and Strombomonas.
With the development of molecular biology, the understanding of the phylogeny relationships between Trachelomonas and Strombomonas also changed. The monophyly of these two lorica-bearing genera could not be confirmed initially [9]. A single monophyletic loricate clade was supported based on SSU and LSU rDNA data of three species. Marin et al. (2003) revised the classification of euglenoids and reclassified Strombomonas into Trachelomonas [10]. However, only thirty Trachelomonas strains and one Strombomonas strain were used for the SSU rDNA analysis. Subsequently, Triemer et al. (2006) sequenced SSU and LSU rDNA of eight Strombomonas and 25 Trachelomonas strains, and combined them in a multigene phylogenetic analysis. Trachelomonas and Strombomonas were confirmed to be two independent genera [11]. This was also supported by other studies [12,13,14,15,16,17]. These analyses added 11 definite species and a few undetermined species (T. sp.). However, the position of many taxa on the tree depends on the number of species and the genes used [18].
It is difficult to construct phylogenetic studies of loricate genera only based on loricae morphology. In addition, many features of loricae are unstable, which also causes difficulties for the classification of Trachelomonas and Strombomonas [12]. At present, it is necessary to find out stable morphological characteristics of the two genera. In AlgaeBase, there are 374 accepted species of Trachelomonas, but molecular data are available for only 22 species in NCBI [19]. For Strombomonas, molecular data are available for eight of 89 species. This data gap makes it difficult to establish phylogenetic relationships between these two genera and within each of them. Previous Chinese studies on Trachelomonas and Strombomonas were based only on morphology. A total of 106 species of Trachelomonas and 30 species of Strombomonas have been reported from China [20,21]. Shi et al. divided Trachelomonas into nine groups (Volvocinae, Curtae, Oblongae, Cylindricae, Ovoideae, Hispidae, Pyriformes, Caudatae, Scabrae) based on the shape and ornamentation of loricae for facilitating identification. Almost no molecular data from Chinese strains have been published so far except for the taxa T. subplanctonica, S. borystheniensis, S. fluviatilis, S. gibberosa, S. maxima and S. triquetra [21,22]. A total of 13 species of Trachelomonas and eight species of Strombomonas were collected from China. Lorica and protoplast of isolated and cultured strains were observed and measured to explore the stability of traits. The nuclear SSU rDNA (nSSU rDNA), nuclear LSU rDNA (nLSU rDNA), plastid-encoded SSU rDNA (cpSSU rDNA) and plastid-encoded LSU rDNA (cpLSU rDNA) of each species were obtained and combined in a multigene phylogenetic analysis. A phylogenetic tree was constructed by combining four genes and using maximum likelihood and Bayesian methods to assess the relationship between the two genera and clarify the phylogenetic characteristics of each lineage.

2. Materials and Methods

2.1. Strains Collection, Cultivation and Morphological Study

During 2019–2021, we collected 13 Trachelomonas species and eight Strombomonas species from various sites in China (Table S1). Phytoplankton algae were collected with a 20 μm plankton net. Individual euglenoid cells were isolated with capillary pipettes under a Nikon Ts2 inverted microscope (NIKON, Tokyo, Japan). All strains were cultured in AF–6 medium [23] and were maintained at 22–25 °C under conditions of a 14:10 light: dark cycle at 3000 lux photons from cool white fluorescent tubes. Species (environmental samples and laboratory cultures of strains) were observed and identified using an Axio Imager A2 microscope (Carl Zeiss Inc., Hallbergmoos, Germany) and photographed with a microscope appendant camera (DP72, Olympus, Tokyo, Japan). More than 50 specimens for each strain were measured with ImageJ software [24]. The measurement results were analyzed by constructing box plot with SPSS v21.0 (Statistical Product and Service Solutions, Chicago, IL, USA) [25]. Excel 2016 was used to conduct one-way analysis of variance for the measured data.

2.2. DNA Extraction, Amplification and Sequencing

The total DNA of strains was extracted using Plant Genomic DNA Kit (Tiangen Biotech Co., Beijing, China). We amplified and sequenced the nSSU rDNA, nLSU rDNA, cpSSU rDNA and cpLSU rDNA from our strains. Polymerase chain reactions (PCR) were performed using published primers, reaction mixes and amplification conditions [13,26,27,28]. The PCR products were purified using a SanPrep column DNA gel purification kit (Sangon, Shanghai, China), and and sequenced in BGI Tech Corporation (Shanghai, China).

2.3. Phylogenetic Analyses

The sequences were submitted to the BLAST search program of the National Center for Biotechnology Information (NCBI) to find closely related sequences. All sequences were downloaded from GenBank and aligned using the Clustal W [29] option in the BioEdit v7.2.1 (BioEdit Sequence Alignment Editor, Scotts Valley, CA, USA) sequence analysis software [30]. Very short sequences (<200 bp) were excluded from the alignment. Based on Neighbor–joining phylogenetic analyses, the identical and near identical sequences were excluded and one or a few sequences of each species were retained. Table S2 lists the 73 taxa with GenBank accession numbers used in this study. Sequences of the Colacium genus were used to root the trees. Sequences of four genes were aligned separately. A combined alignment was generated (5431 aligned sites; cpSSU Rdna = 1–854; nSSU rDNA = 855–2713; cpLSU rDNA = 2714–3758; nLSU rDNA = 3759–5431) by using PhyloSuite v1.2.2 (A desktop platform for streamlined molecular sequence data management and state of the art evolutionary phylogenetics studies, Wuhan, China) [31]. The best-fit model for the concatenated alignments were selected using ModelFinder v1.6.8 (Fast and Accurate Model Selection, Canberra, Australia) [32] with all algorithm and AIC criterion (for ML: cpSSU rDNA, nSSU rDNA = GTR + F + R4; nLSU rDNA = TIM2 + F + R4; cpLSU rDNA = TVM + F + I + G4; for BI: cpSSU rDNA, nSSU rDNA, cpLSU rDNA, nLSU rDNA = GTR + F + I + G4). ML phylogenies were inferred using IQ-TREE v1.6.8 (Efficient Tree Reconstruction, Canberra, Australia) [33] with 5000 ultrafast bootstraps (under Edge-linked partition model), as well as the Shimodaira-Hasegawa-like approximate likelihood-ratio test [34]. Bayesian Inference phylogenies were inferred using MrBayes v3.2.6 (Bayesian Inference of Phylogeny, Stockholm, Sweden) [35] with 2 parallel runs, 10,000,000 generations, of which the initial 25% of sampled data were discarded as burn-in (under partition model). Figtree v1.4.2 (Produce Figures of Phylogenetic Trees, Edinburgh, UK, Britain) was used to edit all resulting phylogenetic trees.

3. Result

3.1. Morphological Characteristics

The morphological characteristics of 13 species of Trachelomonas and eight species of Strombomonas varied (Figure 1, Figure 2, Figure 3 and Figure 4 and Table 1). Among these species, T. cervicula is a new record for China.
  • Trachelomonas cervicula A.Stokes (Figure 1A–C)
Description: loricae spherical without collar, surface smooth, diameter 20–29 µm; cells with metabolic movements, chloroplasts discoid without pyrenoid, range 5–10; paramylon grains rod-shaped granules, numerous; stigma obvious.
Reference strain: Deposited as FACHB-3562 in the Freshwater Algae Culture Collection at the Institute of Hydrobiology Chinese Academy of Science, Wuhan, Hubei Province, China.
Collected location: 31°14′91″ N 121°37′75″ E; phytoplankton, Shanghai.

3.2. Stability of Loricae Morphology

In measuring the length and width of the loricae of five strains of Trachelomonas (Figure S1), we found the length of T. planctonica var. oblonga and T. subplanctonica decreasing significantly before and after cultivation (Figure S1A). In comparison, there was no significant change in the loricae width of T. hispida, T. lefevrei and T. planctonica var. oblonga after culture (Figure S1B). For the length-width ratio of the loricae of five strains, only T. subplanctonica had a significant difference (Figure S1E).
The collar morphology in cultured specimens is different from those in the field. In cultured samples, the collars of T. armata (Figure 1G,H), T. hispida (Figure 3G–I), T. planctonica var. oblonga (Figure 1M,N) and T. subplanctonica (Figure 2D,E) became shorter. In measuring the length of the collar, we found that the collar length of loricae in the cultured samples was significantly reduced compared with that in the field sample (Figure S1C). There was no significant difference in the collar width of T. hispida, T. planctonica var. oblonga, T. lefevrei, but the collar width of T. similis and T. subplanctonica decreased significant (Figure S1D). Comparing the collar length-width ratio of five strains, only T. lefevrei had no significant difference (Figure S1F). The collar of T. similis became short and straight (Figure 1P,Q). And the teeth on the collar edge of T. lefevrei (Figure 1J,K) were not present in cultured specimens.
In the field samples, the loricae color of T. subplanctonica varied from colorless, yellowish brown to reddish brown. The same situation also occurred in all other strains. The spines on the loricae surface did not grow in culture for most species. The cone spines of T. armata (Figure 1G,H), T. hispida (Figure 3G–I), T. cf. sydneyensis var. minima (Figure 3J,K) and Trachelomonas sp. (Figure 2M,N and Figure 3D,E) did not grow (Figure 2M,N). The rod spines on the surface of the loricae of T. cf. bacillifera var. minima still grew in culture (Figure 2P,Q).

3.3. Phylogenetic Analysis and Morphological Characteristics of the Clades

We sequenced nSSU rDNA, nLSU rDNA, cpSSU rDNA and cpLSU rDNA sequences from all Trachelomonas and Strombomonas strains. The topology was similar for the phylogenetic trees obtained by both Maximum likelihood and Bayesian inference, in terms of the location of the analyzed strains (Figure 5). In the phylogenetic trees, both Trachelomonas and Strombomonas are monophyletic groups and sister branches of each other (1.00/100). All strains are separated into seven clades (A–H).
All Strombomonas species fall into the A-B clades. S. gibberosa was independent of other Strombomonas species. The lorica of S. gibberosa has long collar and stout caudal tail. The middle of the lorica is wide-rhomboidal. S. gibberosa has 7–9 chloroplasts with haplopyrenoids. Clade A was a relatively low supported clade (0.66/88) of six species, including S. cf. borystheniensis strain SHNUQ6C3. S. cf. borystheniensis is closely related to S. borystheniensis and S. ovalis, and clustered into a subclade (1.00/100). S. triquetra, S. balvayi and S. eurystoma clustered into a subclade (1.00/98). Loricae of species in Clade A have short collars, the posterior end rounded or have a short caudate process. All these species have five or more chloroplasts with haplopyrenoids or diplopyrenoid. Clade B contains four species (1.00/100), with S. fluviatilis and S. maxima clustered into a subclade (1.00/100), S. verrucosa and S. acuminata clustered into another subclade (1.00/100). These four species have fusiform to broadly elliptic loricae which have long collars and long tails except S. verrucosa. Each species had 6–17 chloroplasts with haplopyrenoids.
All Trachelomonas species fall into the C-G clades. Clade C and clade D both contain four species that have strong support (1.00/100). The species of Clade C have round loricae without collars and spines. The cells have more than five chloroplasts with haplopyrenoids. Clade D consists of four species with strong support (1.00/100). All species have rectangular loricae with collar or annular thickenings and without spines. Each cell had 8–10 chloroplasts with haplopyrenoids. Clade E contains three closely related species (1.00/97). Loricae of species in clade E are oval, with annular thickenings or short collars and with spines. The cells had more than ten chloroplasts with haplopyrenoids. Clade F contains nine species (1.00/97). T. lefevrei, T. planctonica, T. planctonica var. oblonga, T. similis and T. cf. crebea are clustered into a subclade with strong support (1.00/100). T. subplanctonica, T. bernardinensis and T. undulaticollum clustered into another subclade with low support (0.59/-). T. conradii and T. scabra clustered into a subclade with strong support (1.00/97). These nine species have round or oval loricae with short collars or annular thickenings and are without spines. The cells have 7–18 chloroplasts with haplopyrenoids. Clade G contains 11 species (1.00/100). T. ellipsoidalis is independent of other species of Trachelomonas. In addition to T. ellipsoidalis, other species in the G clade clustered into a subclade with strong support (1.00/100). Species in clade G have the greatest morphological diversity of all the group identified herein. Loricae are rectangular to round, elliptic or nearly round, with short collars or annular thickenings, and possess spines. Each cell had chloroplasts with haplopyrenoids or diplopyrenoid.

4. Discussion

4.1. Stable Morphological Characteristics

The morphologic identification of loricate genera is based on the shape, color, ornamentation of loricae and collar shape. However, these characteristics may change under culture conditions. Poniewozik et al. found that the loricae shape of T. hispida changed as the loricae aged [36]. We measured lorica sizes of five Trachlemonas species, and found that lorica shape did not change significantly after being cultivated (Figure 1, Figure 2 and Figure 3 and Figure S1E). The collar became shorter and smaller but was not lost. Poniewozik et al. showed that the loricae surface could not stably form spines [36]. The cone spines on the loricae surface in our cultured strains did not grow steadily. Moreover, the loricae color was light to dark during different periods of development both in culture samples and field samples [21,36]. We also found this phenomenon in this study. It might have been a result of different rates of incorporating and releasing microelements in and out of the lorica structure. [36,37]. Therefore, we thought the shape of lorica and whether the lorica has a collar are more reliable in identifying species. And spines are also important characteristic of those species in the field samples.

4.2. The Relationship between Phylogenetic Tree and Morphological Classification

Strombomonas was separated from Trachelomonas based on its rough and less ornamented lorica surface. Although early molecular data reclassified Strombomonas into Trachelomonas [10], subsequent studies recovered strongly supported monophyletic Strombomonas and Trachelomonas clades [11,12,13,14,15,16,17]. Our molecular data analysis supported the separation of these two genera. Brosnan et al. [38] also supports the separation of Trachelomonas and Strombomonas as distinct genera using two sets of morphological characters, loricae development and pellicle strip reduction. Lorica development in Strombomonas occurred from the anterior of the cell to the posterior. But for Trachelomonas, a layer of mucilage was excreted over the entire protoplast, followed by creation of the collar at the anterior end. In our cultivation process, this phenomenon was also found. The loricae tails of cultured Strombomonas were often missing (Figure 3B,E,J), while those of Trachelomonas were intact (Figure 1, Figure 2 and Figure 3). The protoplasts of Trachelomonas and Strombomonas species were very similar, and the chloroplasts mostly contained haplopyrenoids or diplopyrenoids. Therefore, our results supported that Trachelomonas and Stromomonas were independent based on the morphological data of loricae and protoplasts as well as molecular data. However, the systematic classification status of those species which are morphologically intermediate between the two genera need to be analyzed based on more morphology and molecular data.
The morphology of loricae is the only characteristic in the classification and identification of loricate genera [2,39,40]. Huber-Pestalozzi once divided Trachelomonas into two sections (Rotundatae, Caudatae) and Strombomonas into four sections (Rotundatae, Complanatae, Prismaticae, Intermediae) according to the morphology and ornamentations of loricae [40]. Shi et al. [20] thought that the classification of the two genera by Huber-Pestalozzi lacked protoplast characteristics and was artificial and they did not systematically classify the two genera.
The study of Ciugulea et al. has shown that the molecular phylogeny was inconsistent with Huber-Pestalozzi’s classification but was consistent with Pringsheim’s [4] classification. [12]. This result is only partly supported by our study. The six groups classified by Pringsheim are interlaced in our phylogenetic tree. Pringsheim placed T. cervicula in group six, this species fell in clade C of our study. Clade C also included T. volvocinopsis which was placed to group two in Pringsheim. Pringsheim placed T. pertyi in group three. It fell into clade G, which also contains T. zorensis and T. hispida of group one. It may be because Pringsheim [4] only cultivated 13 species, and the classification was using not only the shape of the loricae, but also the color of the loricae and the number of chloroplasts, which were not stable morphological characteristics. It was clear from the phylogeny that classification based on protoplast morphology alone is not sufficient and often misleading.
Our results on the loricate taxa were similar with Ciugulea et al. [12] and Kim et al. [18]. In our study, seven clades with well-defined phylogenetic positions were presented. For Strombomonas, the clade relationships and composition were more similar to the results found in the study of Ciugulea et al. [12]. All species of Strombomonas were divided into Clade A-B. Clade A corresponded to clade H and clade G in Ciugulea et al. [12]. Our clade B corresponded to clade F in Ciugulea et al. [12]. For Trachelomonas, the clade relationships and composition obtained herein were more similar with the study of Kim et al. [18]. Different from the results of Ciugulea et al., we did not recover the sister taxon relationship between clades E and G in this study. Taxa of Trachelomonas included in this study divided into Clade C-G. Clade C corresponded to clade D in Ciugulea et al. [12] and to a subclade of clade F2 in Kim et al. [18]. Clade C contained sequences of T. cervicula not present before. Clade D corresponded to clade C in Ciugulea et al. [12] and to another subclade of clade F2 in Kim et al. [18]. Clade E corresponded to clade B in Ciugulea et al. [12] and to clade F3 in Kim et al. [18] but added T. armata strain SHNUS50B3. Clade F corresponded to clade E in Ciugulea et al. [12] and to clade F4 in Kim et al. [18]. In clade F, there were five species (T. planctonica var. oblonga, T. subplanctonica, T. undulaticollum) not present in Ciugulea et al. [12] and Kim et al. [18]. The morphological characteristics of these species were consistent with those concluded by Ciugulea et al. [12]. Clade G corresponded to clade A in Ciugulea et al. [12] and to clade F1 in Kim et al. [18]. In this clade, T. cf. bacillifera var. minima, T. cf. sydneyensis var. minima were not present in previous studies of Ciugulea et al. [12] and Kim et al. [18]. After adding our additional species, the morphological characteristics of this clade became diversified, with the shape of loricae not only oval, but also round and rectangular to round, and the chloroplasts are with haplopyrenoids or diplopyrenoid.
The pyrenoid nucleus was not easily observed. We think that it is necessary to examine protoplasts of species, although protoplast characteristics are rare and not suitable for observation. The phylogeny of loricate genera should be established based on the morphology of loricae and protoplast.

5. Conclusions

We collected 13 species of Trachelomonas and eight species of Strombomonas in China. We isolated and cultured specimens from field samples to observe the stability of loricae, and supplemented the protoplast characteristics of species. We extracted DNA, amplificated and sequenced four genes from the cultured samples. A phylogenetic tree was constructed based on Bayesian and maximum likelihood methods. The results showed that the shape of the lorica, whether the lorica has a collar and spines could be used to identify species. Our data support Trachelomonas and Strombomonas as separate genera. We define the phylogenetic location of each clade. Species of each clade identified with molecular methods share common morphological characteristics.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/d14080623/s1, Table S1: The collection localities of strains in this study, Table S2: Relevant sequence information downloaded from GenBank in this study. Figure S1: Box plot display of loricae length, loricae width, collar length, collar width, loricae length/loricae width and collar length/collar width of field samples and cultured samples.

Author Contributions

Conceptualization, X.J. and X.C.; methodology, X.J. and X.C.; formal analysis, X.J. and X.C.; Investigation, X.J. and X.C.; data curation, X.J. and X.C.; writing original draft preparation, X.J., W.P. and Q.W.; writing review and editing, X.J., W.P. and Q.W.; supervision, Q.W.; Project administration, W.P.; funding acquisition, W.P. and Q.W. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by National Natural Science Foundation of China (No. 32070227).

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

Data can be found within the manuscript and Supplementary Materials.

Acknowledgments

We are grateful to Shuran Wan for her kind help in field work. Thanks to John Patrick Kociolek of the University of Colorado for his extensive English revision of our manuscript. Thanks to the four reviewers for their valuable comments.

Conflicts of Interest

The authors declare no conflict of interest.

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Figure 1. Morphology of Trachelomonas strains in this study. Each species includes: morphology of lorica in the field (left), morphology of lorica in culture (middle), Protoplast morphology (right). (A–C) T. cervicula strain SHNUS39D1; (D–F) T. playfairii strain SHNUN14B1; (G–I) T. armata strain SHNUS50B3; (J–L) T. lefevrei strain SHNUCC; (M–O) T. planctonica var. oblonga strain SHNUS2C1; (P–R) T. similis strain SHNUS41C3. Species are all in the same scale, scale bar 20 μm.
Figure 1. Morphology of Trachelomonas strains in this study. Each species includes: morphology of lorica in the field (left), morphology of lorica in culture (middle), Protoplast morphology (right). (A–C) T. cervicula strain SHNUS39D1; (D–F) T. playfairii strain SHNUN14B1; (G–I) T. armata strain SHNUS50B3; (J–L) T. lefevrei strain SHNUCC; (M–O) T. planctonica var. oblonga strain SHNUS2C1; (P–R) T. similis strain SHNUS41C3. Species are all in the same scale, scale bar 20 μm.
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Figure 2. Morphology of Trachelomonas strains in this study. Each species includes: the morphology of lorica in the field (left), morphology of lorica in culture (middle), protoplast morphology (right); (A–C) T. cf. crebea strain SHNUS39B2; (D–F) T. subplanctonica strain SHNUS17C2; (G–I) T. bernardinensis strain SHNUWeijian; (J–L) T. undulaticollum strain SHNUS35D5; (M–O) Trachelomonas sp. strain SHNUC4C3; and (P–R) T. cf. bacillifera var. minima strain SHNUQ4B2. Species are all in the same scale, scale bar 20 μm.
Figure 2. Morphology of Trachelomonas strains in this study. Each species includes: the morphology of lorica in the field (left), morphology of lorica in culture (middle), protoplast morphology (right); (A–C) T. cf. crebea strain SHNUS39B2; (D–F) T. subplanctonica strain SHNUS17C2; (G–I) T. bernardinensis strain SHNUWeijian; (J–L) T. undulaticollum strain SHNUS35D5; (M–O) Trachelomonas sp. strain SHNUC4C3; and (P–R) T. cf. bacillifera var. minima strain SHNUQ4B2. Species are all in the same scale, scale bar 20 μm.
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Figure 3. Morphology of Trachelomonas strains in this study: (A–L) each species includes: morphology of lorica in the field (left), morphology of lorica in culture (middle), protoplast morphology (right); (A–C) Trachelomonas sp. strain SHNUS12C2; (D–F) Trachelomonas sp. strain SHNUS26A3; (G–I) T. hispida strain SHNUQ1D6; (J–L) T. cf. sydneyensis var. minima strain SHNUC1D1; (M,N) Morphology of Trachelomonas sp. strain SHNUS25D1 in culture; (O,P) morphology of Trachelomonas sp. strain SHNUN16B1 in culture; (Q) morphology of Trachelomonas sp. strain SHNUS30A1 in culture; and (R) protoplast morphology of Trachelomonas sp.strain SHNUC2D. Species are all in the same scale, scale bar 20 μm.
Figure 3. Morphology of Trachelomonas strains in this study: (A–L) each species includes: morphology of lorica in the field (left), morphology of lorica in culture (middle), protoplast morphology (right); (A–C) Trachelomonas sp. strain SHNUS12C2; (D–F) Trachelomonas sp. strain SHNUS26A3; (G–I) T. hispida strain SHNUQ1D6; (J–L) T. cf. sydneyensis var. minima strain SHNUC1D1; (M,N) Morphology of Trachelomonas sp. strain SHNUS25D1 in culture; (O,P) morphology of Trachelomonas sp. strain SHNUN16B1 in culture; (Q) morphology of Trachelomonas sp. strain SHNUS30A1 in culture; and (R) protoplast morphology of Trachelomonas sp.strain SHNUC2D. Species are all in the same scale, scale bar 20 μm.
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Figure 4. Morphology of Strombomonas strains in this study: (A–I) each species includes: morphology of lorica in the field (left), morphology of lorica in culture (middle), protoplast morphology (right); (A–C) S. maxima strain SHNUS6A3; (D–F) S. fluviatilis strain SHNUQ6B2; (G–I) S. borystheniensis strain SHNUQ6C2; (J,K) Morphology of S. verrucosa strain SHNUS28C1; (L) protoplast morphology of S. ovalis strain SHNUS49C6; (M–R) each species including: morphology of lorica in the field (left), protoplast morphology (right); (M,N) S. gibberosa strain SHNUQ6A1; (O,P) S. triquetra strain SHNUQ6A4; and (Q,R) S. cf. borystheniensis strain SHNUQ6C3. Species are all in the same scale, scale bar 20 μm.
Figure 4. Morphology of Strombomonas strains in this study: (A–I) each species includes: morphology of lorica in the field (left), morphology of lorica in culture (middle), protoplast morphology (right); (A–C) S. maxima strain SHNUS6A3; (D–F) S. fluviatilis strain SHNUQ6B2; (G–I) S. borystheniensis strain SHNUQ6C2; (J,K) Morphology of S. verrucosa strain SHNUS28C1; (L) protoplast morphology of S. ovalis strain SHNUS49C6; (M–R) each species including: morphology of lorica in the field (left), protoplast morphology (right); (M,N) S. gibberosa strain SHNUQ6A1; (O,P) S. triquetra strain SHNUQ6A4; and (Q,R) S. cf. borystheniensis strain SHNUQ6C3. Species are all in the same scale, scale bar 20 μm.
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Figure 5. Bayesian phylogenetic tree based on combined nuclear SSU, nuclear LSU plastid SSU and plastid LSU rDNA sequences. Support values for all analyses are shown on branches as follows: Bayesian posterior probabilities (BA)/SH aLRT values/maximum likelihood bootstrap values (ML). The pp values < 0.50, SH aLRT values < 50 or rbs values < 50 was marked with a hyphen (-). Strain collected from China in red.
Figure 5. Bayesian phylogenetic tree based on combined nuclear SSU, nuclear LSU plastid SSU and plastid LSU rDNA sequences. Support values for all analyses are shown on branches as follows: Bayesian posterior probabilities (BA)/SH aLRT values/maximum likelihood bootstrap values (ML). The pp values < 0.50, SH aLRT values < 50 or rbs values < 50 was marked with a hyphen (-). Strain collected from China in red.
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Table
Table 1. Morphological characters of Trachelomonas and Strombomonas strains in this study.
Table 1. Morphological characters of Trachelomonas and Strombomonas strains in this study.
TaxonIsolate/StrainLength (µm)Width (µm)Number of ChloroplastsPyrenoidLoricae Shape
T. armata (Ehrenberg) SteinSHNUS50B332–4915–2210–15HaplopyrenoidOvoid with short collar, posterior end with long cone thorns
T. bernardinensis VischerSHNUWeijian31–5818–296–10HaplopyrenoidFusiform with collar, collar with toothed edge, a visible process at the posterior end
T. cervicula A.StokesSHNUS39D120–2920–295–10NoneSpherical without collar, surface smooth
T. cf. bacillifera var. minimaSHNUQ4B224–2520–218–10HaplopyrenoidElliptical without collar, surface with rod spines
T. cf. crebeaSHNUS39B220–3720–278–10DiplopyrenoidElliptical with collar, surface with granular process
T. cf. sydneyensis var. minimaSHNUC1D123–2619–2010–12HaplopyrenoidElliptical without collar, surface with short conical spines
T. hispida (Perty) SteinSHNUQ1D630–3322–247–10DiplopyrenoidOblong without collar, surface with short cone spines
T. lefevrei DeflandreSHNUCC27–3120–249–10HaplopyrenoidElliptical with collar, collar with toothed edge
T. planctonica var. oblonga SvirenkoSHNUS2C120–2317–2010–20HaplopyrenoidElliptical with collar, collar with toothed edge
T. similis StokesSHNUS41C320–2316–175–8HaplopyrenoidWide elliptical with oblique collar, surface punctured
T. playfairii DeflandreSHNUN14B119–2616–208–10HaplopyrenoidElliptical with oblique collar, collar with toothed edge
Trachelomonas sp.SHNUS26A316–1913–167–10HaplopyrenoidElliptical without collar, anterior end with annular thickenings, surface with short spines
Trachelomonas sp.SHNUC4C321–2321–235–8DiplopyrenoidSpherical without collar, surface with short conical spines
Trachelomonas sp.SHNUS12C220–2217–2010–12HaplopyrenoidElliptical without collar, surface with fine spines
T. subplanctonica Jiang & PangSHNUS17C223–3419–306–10HaplopyrenoidEllipsoidal with collar, collar with toothed edge, surface punctured
T. undulaticollum ShiSHNUS35D528–3020–228–10HaplopyrenoidElliptical with long collar
S. borystheniensis (Roll) PopovaSHNUQ6C222–3019–206–11DiplopyrenoidWide ellipsoidal with a wide short collar, without tail
S. cf. borystheniensisSHNUQ6C327–3818–255–7HaplopyrenoidEllipsoidal with collar, without tail
S. gibberosa (Playfair) DeflandreSHNUQ6A140–5321–357–9HaplopyrenoidMiddle wide-rhomboidal, loricae with a wide short collar and a long tail
S. fluviatilis (Lemmermann) DeflandreSHNUQ6B253–7121–289–13HaplopyrenoidFusiform with straight collar and a long tail
S. maxima (Skvortsov) DeflandreSHNUS6A370–10928–438–17HaplopyrenoidBroadly ovate with straight collar and a long tail
S. triquetra (Playfair) DeflandreSHNUQ6A434–4320–216–10HaplopyrenoidInverted triangle with short collar and a short caudate process
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Jiang, X.; Chen, X.; Pang, W.; Wang, Q. Phylogeny of Trachelomonas and Strombomonas (Euglenaceae) Based on Morphological and Molecular Data. Diversity 2022, 14, 623. https://doi.org/10.3390/d14080623

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Jiang X, Chen X, Pang W, Wang Q. Phylogeny of Trachelomonas and Strombomonas (Euglenaceae) Based on Morphological and Molecular Data. Diversity. 2022; 14(8):623. https://doi.org/10.3390/d14080623

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Jiang, Xiaodie, Xi Chen, Wanting Pang, and Quanxi Wang. 2022. "Phylogeny of Trachelomonas and Strombomonas (Euglenaceae) Based on Morphological and Molecular Data" Diversity 14, no. 8: 623. https://doi.org/10.3390/d14080623

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