Summary
The architecture of eukaryotic cells is underpinned by complex arrrays of microtubules that stem from an organizing center, referred to as the MTOC. With few exceptions, MTOCs consist of two ...basal bodies that anchor flagellar axonemes and different configurations of microtubular roots. Variations in the structure of this cytoskeletal system, also referred to as the ‘flagellar apparatus’, reflect phylogenetic relationships and provide compelling evidence for inferring the overall tree of eukaryotes. However, reconstructions and subsequent comparisons of the flagellar apparatus are challenging, because these studies require sophisticated microscopy, spatial reasoning and detailed terminology. In an attempt to understand the unifying features of MTOCs and broad patterns of cytoskeletal homology across the tree of eukaryotes, we present a comprehensive overview of the eukaryotic flagellar apparatus within a modern molecular phylogenetic context. Specifically, we used the known cytoskeletal diversity within major groups of eukaryotes to infer the unifying features (ancestral states) for the flagellar apparatus in the Plantae, Opisthokonta, Amoebozoa, Stramenopiles, Alveolata, Rhizaria, Excavata, Cryptophyta, Haptophyta, Apusozoa, Breviata and Collodictyonidae. We then mapped these data onto the tree of eukaryotes in order to trace broad patterns of trait changes during the evolutionary history of the flagellar apparatus. This synthesis suggests that: (i) the most recent ancestor of all eukaryotes already had a complex flagellar apparatus, (ii) homologous traits associated with the flagellar apparatus have a punctate distribution across the tree of eukaryotes, and (iii) streamlining (trait losses) of the ancestral flagellar apparatus occurred several times independently in eukaryotes.
Morphostasis of traits in different species is necessary for reconstructing the evolutionary history of complex characters. Studies that place these species into a molecular phylogenetic context test ...hypotheses about the transitional stages that link divergent character states. For instance, the transition from a phagotrophic mode of nutrition to a phototrophic lifestyle has occurred several times independently across the tree of eukaryotes; one of these events took place within the Euglenida, a large group of flagellates with diverse modes of nutrition. Phototrophic euglenids form a clade that is nested within lineages of phagotrophic euglenids and that originated through a secondary endosymbiosis with green algae. Although it is clear that phototrophic euglenids evolved from phagotrophic ancestors, the morphological disparity between species representing these different nutritional modes remains substantial.
We cultivated a novel marine euglenid, Rapaza viridis n. gen. et sp. ("green grasper"), and a green alga, Tetraselmis sp., from the same environment. Cells of R. viridis were comprehensively characterized with light microscopy, SEM, TEM, and molecular phylogenetic analysis of small subunit rDNA sequences. Ultrastructural and behavioral observations demonstrated that this isolate habitually consumes a specific strain of Tetraselmis prey cells and possesses a functional chloroplast that is homologous with other phototrophic euglenids. A novel feeding apparatus consisting of a reduced rod of microtubules facilitated this first and only example of mixotrophy among euglenids. R. viridis also possessed a robust photoreception apparatus, two flagella of unequal length, euglenoid movement, and a pellicle consisting of 16 strips and one (square-shaped) whorl of posterior strip reduction. The molecular phylogenetic data demonstrated that R. viridis branches as the nearest sister lineage to phototrophic euglenids.
The unusual combination of features in R. viridis combined with its molecular phylogenetic position completely conforms to the expected transitional stage that occurred during the early evolution of phototrophic euglenids from phagotrophic ancestors. The marine mixotrophic mode of nutrition, the preference for green algal prey cells, the structure of the feeding apparatus, and the organization of the pellicle are outstanding examples of morphostasis that clarify pivotal stages in the evolutionary history of this diverse group of microbial eukaryotes.
Kleptoplasts (kP) are distinct among photosynthetic organelles in eukaryotes (i.e., plastids) because they are routinely sequestered from prey algal cells and function only temporarily in the new ...host cell. Therefore, the hosts of kleptoplasts benefit from photosynthesis without constitutive photoendosymbiosis. Here, we report that the euglenozoan
has only kleptoplasts derived from a specific strain of green alga,
sp., but no canonical plastids like those found in its sister group, the Euglenophyceae.
showed a dynamic change in the accumulation of cytosolic polysaccharides in response to light-dark cycles, and
C isotopic labeling of ambient bicarbonate demonstrated that these polysaccharides originate in situ via photosynthesis; these data indicate that the kleptoplasts of
are functionally active. We also identified 276 sequences encoding putative plastid-targeting proteins and 35 sequences of presumed kleptoplast transporters in the transcriptome of
. These genes originated in a wide range of algae other than
sp., the source of the kleptoplasts, suggesting a long history of repeated horizontal gene transfer events from different algal prey cells. Many of the kleptoplast proteins, as well as the protein-targeting system, in
were shared with members of the Euglenophyceae, providing evidence that the early evolutionary stages in the green alga-derived secondary plastids of euglenophytes also involved kleptoplasty.
Several lineages of euglenozoans are enveloped with epibiotic bacteria and live in low oxygen and anoxic marine sediments, such as Bihospites bacati and Calkinsia aureus. A combination of shared ...ultrastructural traits and molecular phylogenetic inferences demonstrate that these lineages belong to a clade called the “Symbiontida.” Bihospites and Calkinsia possess all of the synapomorphies for the Euglenozoa plus several novel traits. Bihospites has a distinctive cell surface organization reminiscent of the pellicle strips in euglenids, a robust C-shaped feeding apparatus that encircles the nucleus, and a diverse community of epibiotic bacteria. Calkinsia has a novel “extrusomal pocket” and a thick (orange) extracellular matrix beneath a uniform layer of epibiotic bacteria. Despite the absence of molecular phylogenetic data, similar ultrastructural traits in Postgaardi mariagerensis and its epibiotic bacteria strongly suggest that this species is also a member of the Symbiontida. Molecular phylogenetic trees inferred from small subunit (SSU) ribosomal DNA sequences have shown that Bihospites and Calkinsia group strongly with a diverse set of environmental DNA sequences (eDNA) generated from low-oxygen marine samples collected at different depths from different locations around the world. These data demonstrate a diverse array of symbiontids that have yet to be characterized at the genomic, cellular, and behavior levels, which underscores how poorly we currently understand the biology and ecology of the group. Moreover, current data suggest that the communities of epibiotic bacteria associated with Bihospites, Calkinsia, and Postgaardi co-evolved with their hosts and are metabolically integrated with modified mitochondria positioned immediately beneath the host's plasma membrane. No symbiontid species has ever been cultivated, so improved knowledge about these eukaryotic organisms and their intimate relationships with bacteria in low oxygen environments will likely be achieved using culture-independent approaches, such as isolated-cell metagenomics.
Broad comparisons of microtubular root systems (the flagellar apparatus) demonstrate independent patterns of cytoskeletal streamlining in parasitic protists relative to their free-living ancestors.
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•Overview of the flagellar apparatus in six major lineages of parasitic protists.•Identification of broad patterns of cytoskeletal homology with free-living ancestors.•Streamlining of the ancestral cytoskeleton occurred independently in parasitic lineages.
The microtubular cytoskeleton of most single-celled eukaryotes radiates from an organizing center called the flagellar apparatus, which is essential for locomotion, feeding and reproduction. The structure of the flagellar apparatus tends to be conserved within diverse clades of eukaryotes, and modifications of this overall structure distinguish different clades from each other. Understanding the unity and diversity of the flagellar apparatus provides important insights into the evolutionary history of the eukaryotic cell. Diversification of the flagellar apparatus is particularly apparent during the multiple independent transitions to parasitic lifestyles from free-living ancestors. However, our understanding of these evolutionary transitions is hampered by the lack of detailed comparisons of the microtubular root systems in different lineages of parasitic microbial eukaryotes and those of their closest free-living relatives. Here we help to establish this comparative context by examining the unity and diversity of the flagellar apparatus in six major clades containing both free-living lineages and endobiotic (parasitic and symbiotic) microbial eukaryotes: stramenopiles (e.g., Phytophthora), fornicates (e.g., Giardia), parabasalids (e.g., Trichomonas), preaxostylids (e.g., Monocercomonoides), kinetoplastids (e.g., Trypanosoma), and apicomplexans (e.g., Plasmodium). These comparisons enabled us to address some broader patterns associated with the evolution of parasitism, including a general trend toward a more streamlined flagellar apparatus.
Recent culture-independent surveys of eukaryotic small-subunit ribosomal DNA (SSU rDNA) from many environments have unveiled unexpectedly high diversity of microbial eukaryotes (microeukaryotes) at ...various taxonomic levels. However, such surveys were most probably biased by various technical difficulties, resulting in underestimation of microeukaryotic diversity. In the present study on oxygen-depleted sediment from a deep-sea methane cold seep of Sagami Bay, Japan, we surveyed the diversity of eukaryotic rDNA in raw sediment samples and in two enrichment cultures. More than half of all clones recovered from the raw sediment samples were of the basidiomycetous fungus Cryptococcus curvatus. Among other clones, phylotypes of eukaryotic parasites, such as Apicomplexa, Ichthyosporea, and Phytomyxea, were identified. On the other hand, we observed a marked difference in phylotype composition in the enrichment samples. Several phylotypes belonging to heterotrophic stramenopiles were frequently found in one enrichment culture, while a phylotype of Excavata previously detected at a deep-sea hydrothermal vent dominated the other. We successfully established a clonal culture of this excavate flagellate. Since these phylotypes were not identified in the raw sediment samples, the approach incorporating a cultivation step successfully found at least a fraction of the “hidden” microeukaryotic diversity in the environment examined.
The plastid genomes of the green algal order Chlamydomonadales tend to expand their non-coding regions, but this phenomenon is poorly understood. Here we shed new light on organellar genome evolution ...in Chlamydomonadales by studying a previously unknown non-photosynthetic lineage. We established cultures of two new Polytoma-like flagellates, defined their basic characteristics and phylogenetic position, and obtained complete organellar genome sequences and a transcriptome assembly for one of them.
We discovered a novel deeply diverged chlamydomonadalean lineage that has no close photosynthetic relatives and represents an independent case of photosynthesis loss. To accommodate these organisms, we establish the new genus Leontynka, with two species (L. pallida and L. elongata) distinguishable through both their morphological and molecular characteristics. Notable features of the colourless plastid of L. pallida deduced from the plastid genome (plastome) sequence and transcriptome assembly include the retention of ATP synthase, thylakoid-associated proteins, the carotenoid biosynthesis pathway, and a plastoquinone-based electron transport chain, the latter two modules having an obvious functional link to the eyespot present in Leontynka. Most strikingly, the ~362 kbp plastome of L. pallida is by far the largest among the non-photosynthetic eukaryotes investigated to date due to an extreme proliferation of sequence repeats. These repeats are also present in coding sequences, with one repeat type found in the exons of 11 out of 34 protein-coding genes, with up to 36 copies per gene, thus affecting the encoded proteins. The mitochondrial genome of L. pallida is likewise exceptionally large, with its >104 kbp surpassed only by the mitogenome of Haematococcus lacustris among all members of Chlamydomonadales hitherto studied. It is also bloated with repeats, though entirely different from those in the L. pallida plastome, which contrasts with the situation in H. lacustris where both the organellar genomes have accumulated related repeats. Furthermore, the L. pallida mitogenome exhibits an extremely high GC content in both coding and non-coding regions and, strikingly, a high number of predicted G-quadruplexes.
With its unprecedented combination of plastid and mitochondrial genome characteristics, Leontynka pushes the frontiers of organellar genome diversity and is an interesting model for studying organellar genome evolution.
Oxymonads are a group of flagellates living as gut symbionts of insects or vertebrates. They have several unique features, one of them being the absence of mitochondria. Diversity of this group is ...seriously understudied, which is particularly true for small species from the family Polymastigidae. We isolated 34 strains of oxymonads with Polymastigidae-like morphology from 24 host species and unused cesspits and sequenced the SSU rRNA gene. Our strains formed two clades in the phylogenetic tree with Streblomastix strix branching between them. This topology was also supported by a three-gene phylogenetic analysis. Despite considerable genetic differences between the clades, light and electron microscopy revealed only subtle differences. The larger clade is considered genus Monocercomonoides and the isolates belonging here were classified into three new species (including the first potentially free-living species), two previously described species, and three unclassified lineages. The smaller clade, here described as Blattamonas gen. nov., consists of three newly described species. Concomitantly with the description of Blattamonas, we elevate the Monocercomonoides subgenus Brachymonas to the genus level. Our study shows that, despite their conserved morphology, the molecular diversity of Polymastigidae-like oxymonads is broad and represents a substantial part of the diversity of oxymonads.
Two monospecific genera of marine benthic dinoflagellates, Adenoides and Pseudadenoides, have unusual thecal tabulation patterns (lack of cingular plates in the former; and no precingular plates and ...a complete posterior intercalary plate series in the latter) and are thus difficult to place within a phylogenetic framework. Although both genera share morphological similarities, they have not formed sister taxa in previous molecular phylogenetic analyses. We discovered and characterized a new species of Pseudadenoides, P. polypyrenoides sp. nov., at both the ultrastructural and molecular phylogenetic levels. Molecular phylogenetic analyses of SSU and LSU rDNA sequences demonstrated a close relationship between P. polypyrenoides sp. nov. and Pseudadenoides kofoidii, and Adenoides and Pseudadenoides formed sister taxa in phylogenetic trees inferred from LSU rDNA sequences. Comparisons of morphological traits, such as the apical pore complex (APC), demonstrated similarities between Adenoides, Pseudadenoides and several planktonic genera (e.g. Heterocapsa, Azadinium and Amphidoma). Molecular phylogenetic analyses of SSU and LSU rDNA sequences also demonstrated an undescribed species within Adenoides.
The Fornicata (Excavata) is a group of microbial eukaryotes consisting of both free-living lineages (e.g., Carpediemonas) and parasitic lineages (e.g. Giardia and Retortamonas) that share several ...molecular and ultrastructural traits. Carpediemonas-like organisms (CLOs) are free-living lineages that diverged early within the Fornicata, making them important for inferring the early evolutionary history of the group. Molecular phylogenetic analyses of free-living fornicates, including sequences from environmental PCR surveys, have demonstrated that CLOs form six different lineages. Representatives from five of these lineages have been studied at the ultrastructural level. The sixth lineage has been labeled “CL2” but has yet to be described with ultrastructural data. Improved understanding of CL2 is expected to help elucidate character evolution within the Fornicata. Therefore, we comprehensively characterized CL2 (NY0171) in order to understand the ultrastructural traits in this lineage, especially the organization of the microtubular root system (i.e., the flagellar apparatus). CL2 shared several morphological features with other fornicates, including reduced mitochondria and an arched B fiber bridging flagellar roots 1 and 2. The molecular phylogenetic position combined with some distinctive ultrastructural traits (e.g., a curved ventral groove) in CL2 required us to establish a new genus and species, Aduncisulcus paluster gen. et sp. nov.
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