Optical function of the finite-thickness corrugated pellicle of euglenoids.

We explore the electromagnetic response of the pellicle of selected species of euglenoids. These microorganisms are bounded by a typical surface pellicle formed by S-shaped overlapping bands that resemble a corrugated film. We investigate the role played by this structure in the protection of the cell against UV radiation. By considering the pellicle as a periodically corrugated film of finite thickness, we applied the C-method to compute the reflectance spectra. The far-field results revealed reflectance peaks with a Q-factor larger than 103 in the UV region for all the illumination conditions investigated. The resonant behavior responsible for this enhancement has also been illustrated by near-field computations performed by a photonic simulation method. These results confirm that the corrugated pellicle of euglenoids shields the cell from harmful UV radiation and open up new possibilities for the design of highly UV-reflective surfaces.

Ultrastructural alterations in Phacus brachykentron (Euglenophyta) due to excess of organic matter in the culture medium.

Morphological and ultrastructural changes induced by exposure to excess of organic matter were analyzed in Phacus brachykentron (Pochm.). The cells were isolated from sites in Matanza River, Buenos Aires, Argentina, which have a high degree of organic matter contamination coming from waste waters discharges of the meat industry. Master strains were cultured on soil water medium and a toxicity bioassay was performed. As a result of the enriched medium, several morphological and ultrastructural cellular alterations were observed by optical, scanning, and transmission electron microscopy. Among these, we can point out changes in cell dimensions, remarkable widening of some pellicle bands, increased number and volume of paramylon grains, displacement of the nucleus from the central to the lateral position, some chloroplasts with their thylakoids disordered, and cell lysis. The response to organic enrichment was very fast, i.e. during the 48h of the bioassay. Therefore, any significant increase of organic matter would rapidly affect wild euglenoids. Our results suggest that the alterations observed, such as the presence of large intracellular paramylon bodies or the deformation of euglenoid cells in natural samples, have the potential to be used as environmental bioindicators.

Ultrastructure and molecular phylogenetic position of Heteronema scaphurum: a eukaryovorous euglenid with a cytoproct.

Euglenids comprise a distinct clade of flagellates with diverse modes of nutrition, including phagotrophy, osmotrophy and phototrophy. Much of the previous research on euglenids has focused on phototrophic species because of their ecological abundance and significance as indicators for the health of aquatic ecosystems. Although largely understudied, phagotrophic species probably represent the majority of euglenid diversity. Phagotrophic euglenids tend to be either bacterivorous or eukaryovorous and use an elaborate feeding apparatus for capturing prey cells. We characterized the ultrastructure and molecular phylogenetic position of Heteronema scaphurum, a eukaryovorous euglenid collected in freshwater. This species was equipped with a distinct cytoproct through which waste products were eliminated in the form of faecal pellets; a cytoproct has not been reported in any other member of the Euglenida. Heteronema scaphurum also had a novel predatory mode of feeding. The euglenid ensnared and corralled several green algal prey cells (i.e. Chlamydomonas) with hook-like flagella covered in mucous before engulfing the bundle of prey cells whole. Molecular phylogenetic analyses inferred from small subunit rDNA sequences placed this species with other eukaryovorous euglenids, which was consistent with ultrastructural features associated with the feeding apparatus, flagellar apparatus, extrusomes, and pellicle.

Trachelomonas (Euglenophyta) from a eutrophic reservoir in Central Mexico.

This study provides valuable information on the ultrastructure and environmental conditions of the Trachelomonas Ehr. (Euglenophyceae) genus in the Guadalupe Dam, a eutrophic reservoir located in the suburbs of Mexico City, which receives a considerable volume of wastewaters. Specimens were collected at surface level between November 2005 and May 2006. Using LM and SEM twelve taxa from phytoplankton were identified of which, 9 are new records for Mexico. The reservoir is warm monomictic, with basic pH values (7.4-10.1), a high concentration of chlorophyll a(18-101 microg l(-1), a permanent anoxic bottom, specific conductivity (K25) of 205 to 290 microS cm(-1), N-NO3, 0.19-1.2 mg l(-1) and P-PO4 0.22-1.6 mg l(-1). Water temperature was 15.6-23.0 degrees C. Most of the Trachelomonas species were found during the dry season, when concentrations of organic matter, nitrogen and phosphorus as well as the temperature were the highest. Higher species richness was also associated with the warmer months. This research contributes to increase our knowledge on Trachelomonas in Mexico and constitutes the first detailed description of lorica ultrastructure of 12 taxa that grow in a body of water with high concentration of nutrients and a moderate amount of mineral contents.

Evolution of distorted pellicle patterns in rigid photosynthetic euglenids (phacus dujardin).

Members of the euglenid genus Phacus are morphologically differentiated from other photosynthetic species by the presence of a rigid cytoskeleton (pellicle) and predominantly dorsoventrally flattened, leaf-shaped cells. In order to better understand the evolutionary history of this lineage, we used scanning electron microscopy to examine patterns of pellicle strips in Phacus acuminatus, Phacus longicauda var. tortus, Phacus triqueter, Phacus segretii, Phacus pleuronectes, Phacus similis, Phacus pusillus, Phacus orbicularis, Phacus warszewiczii, and Discoplastis spathirhyncha, a putative close relative of Phacus and Lepocinclis. Our observations showed that while the earliest diverging species in our analyses, namely P. warszewiczii, has three whorls of exponential reduction, most members of Phacus have clustered patterns of posterior strip reduction that are bilaterally symmetrical distortions of the radially symmetrical "whorled" patterns found in other photosynthetic euglenids. Comparative morphology, interpreted within the context of molecular phylogenetic analyses of combined nuclear small subunit rDNA and partial nuclear large subunit rDNA sequences, demonstrates that clustered patterns of posterior strip reduction arose after the divergence of Phacus from other photosynthetic euglenids and are the result of developmental processes that govern individual strip length. Clustered patterns of pellicle strips in Phacus do not appear to be adaptively significant themselves; they evolved in association with the origin of cell flattening and cell rigidity, which may be adaptations to a planktonic lifestyle.

Ultrastructure and molecular phylogeny of Calkinsia aureus: cellular identity of a novel clade of deep-sea euglenozoans with epibiotic bacteria.

BACKGROUND: The Euglenozoa is a large group of eukaryotic flagellates with diverse modes of nutrition. The group consists of three main subclades - euglenids, kinetoplastids and diplonemids--that have been confirmed with both molecular phylogenetic analyses and a combination of shared ultrastructural characteristics. Several poorly understood lineages of putative euglenozoans live in anoxic environments, such as Calkinsia aureus, and have yet to be characterized at the molecular and ultrastructural levels. Improved understanding of these lineages is expected to shed considerable light onto the ultrastructure of prokaryote-eukaryote symbioses and the associated cellular innovations found within the Euglenozoa and beyond. RESULTS: We collected Calkinsia aureus from core samples taken from the low-oxygen seafloor of the Santa Barbara Basin (580 - 592 m depth), California. These biflagellates were distinctively orange in color and covered with a dense array of elongated epibiotic bacteria. Serial TEM sections through individually prepared cells demonstrated that C. aureus shares derived ultrastructural features with other members of the Euglenozoa (e.g. the same paraxonemal rods, microtubular root system and extrusomes). However, C. aureus also possessed several novel ultrastructural systems, such as modified mitochondria (i.e. hydrogenosome-like), an "extrusomal pocket", a highly organized extracellular matrix beneath epibiotic bacteria and a complex flagellar transition zone. Molecular phylogenies inferred from SSU rDNA sequences demonstrated that C. aureus grouped strongly within the Euglenozoa and with several environmental sequences taken from low-oxygen sediments in various locations around the world. CONCLUSION: Calkinsia aureus possesses all of the synapomorphies for the Euglenozoa, but lacks traits that are specific to any of the three previously recognized euglenozoan subgroups. Molecular phylogenetic analyses of C. aureus demonstrate that this lineage is a member of a novel euglenozoan subclade consisting of uncharacterized cells living in low-oxygen environments. Our ultrastructural description of C. aureus establishes the cellular identity of a fourth group of euglenozoans, referred to as the "Symbiontida".

Ploeotids Represent Much of the Phylogenetic Diversity of Euglenids.

Ploeotids are an assemblage of rigid phagotrophic euglenids that have 10-12 pellicular strips and glide on their posterior flagellum. Molecular phylogenies place them as a poorly resolved, likely paraphyletic assemblage outside the Spirocuta clade of flexible euglenids, which includes the well-known phototrophs and primary osmotrophs. Here, we report SSU rRNA gene sequences from 38 ploeotids, using both single-cell and culture-based methods. Several contain group I or non-canonical introns. Our phylogenetic analyses place ploeotids in 8 distinct clades: Olkasia n. gen., Hemiolia n. gen., Liburna n. gen., Lentomonas, Decastava, Keelungia, Ploeotiidae, and Entosiphon. Ploeotia vitrea, the type of Ploeotia, is closely related to P. oblonga and Serpenomonas costata, but not to Lentomonas. Ploeotia cf. vitrea sensu Lax and Simpson 2013 is not related to P. vitrea and has a different pellicle strip architecture (as imaged by scanning electron microscopy): it instead represents a novel genus and species, Olkasia polycarbonata. We also describe new genera, Hemiolia and Liburna, for the morphospecies Anisonema trepidum and A. glaciale. A recent system proposing 13 suprafamilial taxa that include ploeotids is not supported by our phylogenies. The exact relationships between ploeotid groups remain unresolved and multigene phylogenetics or phylogenomics are needed to address this uncertainty.

Epiplasts: Membrane Skeletons and Epiplastin Proteins in Euglenids, Glaucophytes, Cryptophytes, Ciliates, Dinoflagellates, and Apicomplexans.

Animals and amoebae assemble actin/spectrin-based plasma membrane skeletons, forming what is often called the cell cortex, whereas euglenids and alveolates (ciliates, dinoflagellates, and apicomplexans) have been shown to assemble a thin, viscoelastic, actin/spectrin-free membrane skeleton, here called the epiplast. Epiplasts include a class of proteins, here called the epiplastins, with a head/medial/tail domain organization, whose medial domains have been characterized in previous studies by their low-complexity amino acid composition. We have identified two additional features of the medial domains: a strong enrichment of acid/base amino acid dyads and a predicted ß-strand/random coil secondary structure. These features have served to identify members in two additional unicellular eukaryotic radiations-the glaucophytes and cryptophytes-as well as additional members in the alveolates and euglenids. We have analyzed the amino acid composition and domain structure of 219 epiplastin sequences and have used quick-freeze deep-etch electron microscopy to visualize the epiplasts of glaucophytes and cryptophytes. We define epiplastins as proteins encoded in organisms that assemble epiplasts, but epiplastin-like proteins, of unknown function, are also encoded in Insecta, Basidiomycetes, and Caulobacter genomes. We discuss the diverse cellular traits that are supported by epiplasts and propose evolutionary scenarios that are consonant with their distribution in extant eukaryotes.IMPORTANCE Membrane skeletons associate with the inner surface of the plasma membrane to provide support for the fragile lipid bilayer and an elastic framework for the cell itself. Several radiations, including animals, organize such skeletons using actin/spectrin proteins, but four major radiations of eukaryotic unicellular organisms, including disease-causing parasites such as Plasmodium, have been known to construct an alternative and essential skeleton (the epiplast) using a class of proteins that we term epiplastins. We have identified epiplastins in two additional radiations and present images of their epiplasts using electron microscopy. We analyze the sequences and secondary structure of 219 epiplastins and present an in-depth overview and analysis of their known and posited roles in cellular organization and parasite infection. An understanding of epiplast assembly may suggest therapeutic approaches to combat infectious agents such as Plasmodium as well as approaches to the engineering of useful viscoelastic biofilms.

Unusual mitochondrial genome structures throughout the Euglenozoa.

Mitochondrial DNA of Kinetoplastea is composed of different chromosomes, the maxicircle (bearing 'regular' genes) and numerous minicircles (specifying guide RNAs involved in RNA editing). In trypanosomes [Kinetoplastea], DNA circles are compacted into a single dense body, the kinetoplast. This report addresses the question whether multi-chromosome mitochondrial genomes and compacted chromosome organization are restricted to Kinetoplastea or rather occur throughout Euglenozoa, i.e., Kinetoplastea, Euglenida and Diplonemea. To this end, we investigated the diplonemid Rhynchopus euleeides and the euglenids Petalomonas cantuscygni, Peranema trichophorum and Entosiphon sulcatum, using light and electron microscopy and molecular techniques. Our findings together with previously published data show that multi-chromosome mitochondrial genomes prevail across Euglenozoa, while kinetoplast-like mtDNA packaging is confined to trypanosomes.

Did trypanosomatid parasites have photosynthetic ancestors?

Some molecular phylogenies of plastid-like genes suggest that chloroplasts (the structures responsible for photosynthesis in plants and algae) might have been secondarily lost in trypanosomatid parasites. Chloroplasts are present in some euglenids, which are closely related to trypanosomatids, and it has been argued that chloroplasts arose early in the diversification of the lineage Euglenozoa, to which trypanosomatids and euglenids belong (plastids-early hypothesis). This article reviews how euglenid ultrastructural systems are functionally integrated and phylogenetically correlated. I argue that chloroplast acquisition profoundly altered the structure of certain euglenids, and that the complete absence of these modifications in other euglenozoans is most consistent with their never having had a chloroplast. Ultrastructural evidence suggests that chloroplasts arose relatively recently within a specific subgroup of euglenids and that trypanosomatids are not secondarily non-photosynthetic (plastids-recent hypothesis).

The Glu-modification of alpha-tubulin in the feeding apparatus of the primitive flagellate Entosiphon sulcatum is only apparent after detergent treatment.

Using specific monoclonal antibodies, we investigated the distribution of post-translational modified Tyr- and Glu-tubulins during interphase of the primitive flagellate Entosiphon sulcatum. Immunofluorescence studies of simultaneously permeabilized and fixed cells revealed that microtubular structures comprising Ca(2+)-labile subpellicular and flagellar MTs and Ca(2+)-stable MTs in the siphon complex (feeding organelle) reacted surprisingly unorthodox with antibodies against Tyr- and Glu-tubulin: Unexpectedly, the siphon complex consisting of Ca(2+)-stable MTs appeared exclusively Tyr-positive, whereas the Ca(2+)-labile subpellicular and flagellar MTs reacted with the Glu- as well as with the Tyr-antibody. That the siphon MTs were indeed Ca(2+)-stable and all other MTs had become solubilized, was verified by EM-observation. This surprising result contrasting considerably with the permanent nature of the siphon complex, was reconsidered after preceding lysis and extraction procedures. Depending on the type of detergent used and on extraction times applied, the MTs of the siphon complex now always showed also Glu-positivity, indicating the presence of detyrosinated alpha-tubulin as a biochemical marker of stabilized MTs. Since saponin, irrespective of subsequent extraction times, always produced a Glu-positive reaction and ultrastructural analysis never gave compelling evidence for a drastic MAP-removal, we conclude that the Glu-epitope became freely accessible due to conformational changes in the tubulin polymeres.

[Potentially toxic and harmful phytoflagellates from the Mexican Pacific coasts]. / Fitoflagelados potencialmente tóxicos y nocivos de costas del Pacífico Mexicano.

The phytoflagellates are a heterogeneous group of autotrophic, heterotrophic and mixothrophic flagellates of trophic importance in several ecosystems. As in the rest of Latin America, the phytoflagellates that occur in the Mexican Pacific coasts are virtually unknown except for a few records. Their study require complicated collection and analysis methods, a probable cause for the scarce knowledge of this group in tropical and subtropical areas. Material recently collected from various localities along the Mexican Pacific coasts was used to study phytoflagellates, including toxic and potentially toxic species. Plankton samples were treated by gravity and pump filtration, using different methods for fixation and analysis. The phyla Euglenophyta, Heterokontophyta and Haptophyta were found. They occur as plankton in oceanic and shallow coastal waters.

Ultrastructure and molecular phylogenetic position of Neometanema parovale sp. nov. (Neometanema gen. nov.), a Marine phagotrophic euglenid with skidding motility.

Heteronema is a commonly encountered genus of phagotrophic euglenids that contains very different morphotypes, including elongate gliding species and ovoid skidding forms. We report the first ultrastructural and sequence data from a culture of an ovoid skidding heteronemid, KM051. Cells were 8-23.5 µm long with 22 pellicular strips and a fibrous extracellular layer. The tubular extrusomes had dense centre sections. The feeding apparatus was barely visible by light microscopy, but included two microtubule-supported rods. The flagella had hollow, inflated transition zones, heteromorphic paraxonemal rods, and sheaths of flagellar hairs. The posterior flagellum bore a knob that, unusually, sat >2 µm distal to the flagellar base. No ultrastructural features were uniquely shared by KM051 and the elongate, gliding species Heteronema scaphurum. Conversely, the pellicular microtubule array resembles that in deep-branching primary osmotrophs (Aphagea). 18S ribosomal DNA (18S rDNA) phylogenies showed that KM051 is related to a recently obtained Heteronema c.f. exaratum sequence. These skidding heteronemids are not closely related to H. scaphurum, and instead are closely related to Dinema, Anisonema and specifically, Aphagea. The skidding species in Heteronema are transferred to Neometanema gen. nov. (along with most species of Metanema Klebs, 1893), with KM051 described as Neometanema parovale sp. nov.

On Keelungia pulex nov. gen. et nov. sp., a heterotrophic euglenoid flagellate that lacks pellicular plates (Euglenophyceae, Euglenida).

Keelungia pulex nov. gen. et nov. sp. is described from coastal waters of NE Taiwan. The new species is heterotrophic and feeds on bacteria. Cells are oblong-ovoid, biflagellate and glide along the sides of the flask. Each cell is approximately 8-11µm long, and one of the smallest euglenoid flagellates presently known. Keelungia lacks pellicular plates and in this respect resembles diplonemids and Symbiontida, which are thought to be among the basal groups of Euglenozoa. SEM showed the presence of 10 evenly spaced longitudinal striae in the cell surface, but the striae are difficult to see in the light microscope. TEM showed each stria to comprise a double set of very low longitudinal ridges separated by a shallow furrow, and supported by ca 5 microtubules beneath the plasmalemma, unlike the situation in diplonemids and Symbiontida. The cell surface was further subtended by an extensive system of rough cisternae of endoplasmic reticulum. Keelungia pulex is phylogenetically related to species of Ploeotia and to Lentomonas applanata, but differs in details of the feeding apparatus and in the absence of pellicular plates. Sequencing of SSU rDNA indicates that Ploeotia, Keelungia and Entosiphon form a clade near the base of the euglenoid phylogenetic tree.

Reconciling the bizarre inheritance of microtubules in complex (euglenid) microeukaryotes.

We introduce a hypothetical model that explains how surface microtubules in euglenids are generated, integrated and inherited with the flagellar apparatus from generation to generation. The Euglenida is a very diverse group of single-celled eukaryotes unified by a complex cell surface called the "pellicle", consisting of proteinaceous strips that run along the longitudinal axis of the cell and articulate with one another along their lateral margins. The strips are positioned beneath the plasma membrane and are reinforced with subtending microtubules. Euglenids reproduce asexually, and the two daughter cells inherit pellicle strips and associate microtubules from the parent cell in a semi-conservative pattern. In preparation for cell division, nascent pellicle strips develop from the anterior end of the cell and elongate toward the posterior end between two parent (mature) strips, so that the total number of pellicle strips and underlying microtubules is doubled in the predivisional cell. Each daughter cell inherits an alternating pattern of strips consisting of half of the nascent strips and half of the parent (mature) strips. This observation combined with the fact that the microtubules underlying the strips are linked to the flagellar apparatus created a cytoskeletal riddle: how do microtubules associated with an alternating pattern of nascent strips and mature strips maintain their physical relationship to the flagellar apparatus when the parent cell divides? The model of microtubular inheritance articulated here incorporates known patterns of cytoskeletal semi-conservatism and two new inferences: (1) a multigenerational "pellicle microtubule organizing center" (pMTOC) extends from the dorsal root of the flagellar apparatus, encircles the flagellar pocket, and underpins the microtubules of the pellicle; and (2) prior to cytokinesis, nascent pellicle microtubules fall within one of two "left/right" constellations that are linked to one of the two new dorsal basal bodies.

Description of Rhynchopus euleeides n. sp. (Diplonemea), a free-living marine euglenozoan.

We describe Rhynchopus euleeides n. sp., using light and electron microscopy. This free-living flagellate, which was isolated earlier from a marine habitat, can be grown axenically in a rich medium based on modified seawater. In the trophic stage, cells are predominantly elliptical and laterally flattened, but frequently change their shape (metaboly). Gliding is the predominant manner of locomotion. The two flagella, which are typically concealed in their pocket, are short stubs of unequal length, have conventional axonemes, but apparently lack a paraxonemal rod. Swarmer cells, which form only occasionally, are smaller in size and carry two conspicuous flagella of more than 2 times the body length. Cells are decorated with a prominent apical papillum. Both the flagellar pocket and the adjacent feeding apparatus seem to merge together into a single sub-apical opening. The mitochondrion, which is most likely single, is located peripherally. It is reticulated in shape and contains only a few lamellar cristae. Mitochondrial DNA is abundant and evenly distributed throughout the organelle. Morphological synapomorphies confirm the affiliation of the species with the genus Rhynchopus (Diplonemea, Euglenozoa). We discuss the characters that distinguish Rhynchopus from Diplonema corroborating the validity of the two genera.

Food vacuole contents in the ciliate, Balantidium jocularum (Balantididae), a symbiont in the intestine of the surgeonfish, Naso tonganus (Acanthuridae).

During the past 16 years, the ciliate Balantidium jocularum has been collected from the intestines of many specimens of its fish host, Naso tonganus, all collected from the Great Barrier Reef near Lizard Island, Australia. Ciliates for this study of food consumption were isolated in 1988, 1989, 2003, and 2005. Nineteen specimens of B. jocularum were examined in the transmission electron microscope to determine the contents of both food vacuoles and a putative discharging cytoproct vacuole. Food vacuoles contained rod-shaped bacteria, tightly coiled spirilliform bacteria, and one or more euglenid flagellates. In several balantidia of somewhat different form than the type species of B. jocularum, the large bacterium, Epulopiscium fishelsoni, was observed in light microscope protargol preparations. Some putative phagolysosomes retained spirilliform bacteria that were apparently intact, and others contained partially digested flagellates. Food in a single discharging cytoproct vacuole consisted of normal appearing spirilliform bacteria, some other bacteria, and no flagellates. The results argue for non-selective ingestion of food and selective digestion; hence, somewhat inefficient food processing.

Dinoflagellate, Euglenid, or Cercomonad? The ultrastructure and molecular phylogenetic position of Protaspis grandis n. sp.

Protaspis is an enigmatic genus of marine phagotrophic biflagellates that have been tentatively classified with several different groups of eukaryotes, including dinoflagellates, euglenids, and cercomonads. This uncertainty led us to investigate the phylogenetic position of Protaspis grandis n. sp. with ultrastructural and small subunit (SSU) rDNA sequence data. Our results demonstrated that the cells were dorsoventrally flattened, shaped like elongated ovals with parallel lateral sides, 32.5-55.0 mum long and 20.0-35.0 mum wide. Moreover, two heterodynamic flagella emerged through funnels that were positioned subapically, each within a depression and separated by a distinctive protrusion. A complex multilayered wall surrounded the cell. Like dinoflagellates and euglenids, the nucleus contained permanently condensed chromosomes and a large nucleolus throughout the cell cycle. Pseudopodia containing numerous mitochondria with tubular cristae emerged from a ventral furrow through a longitudinal slit that was positioned posterior to the protrusion and flagellar apparatus. Batteries of extrusomes were present within the cytoplasm and had ejection sites through pores in the cell wall. The SSU rDNA phylogeny demonstrated a very close relationship between the benthic P. grandis n. sp. and the planktonic Cryothecomonas longipes. These ultrastructural and molecular phylogenetic data for Protaspis indicated that the current taxonomy of Protaspis and Crythecomonas is in need of re-evaluation. The composition and identity of Protaspis is reviewed and suggestions for future taxonomic changes are presented. Problems within the genus Cryothecomonas are highlighted as well, and the missing data needed to resolve ambiguities between the two genera are clarified.

Unique mitochondrial genome structure in diplonemids, the sister group of kinetoplastids.

Kinetoplastid flagellates are characterized by uniquely massed mitochondrial DNAs (mtDNAs), the kinetoplasts. Kinetoplastids of the trypanosomatid group possess two types of mtDNA molecules: maxicircles bearing protein and mitoribosomal genes and minicircles specifying guide RNAs, which mediate uridine insertion/deletion RNA editing. These circles are interlocked with one another to form dense networks. Whether these peculiar mtDNA features are restricted to kinetoplastids or prevail throughout Euglenozoa (euglenids, diplonemids, and kinetoplastids) is unknown. Here, we describe the mitochondrial genome and the mitochondrial ultrastructure of Diplonema papillatum, a member of the diplonemid flagellates, the sister group of kinetoplastids. Fluorescence and electron microscopy show a single mitochondrion per cell with an ultrastructure atypical for Euglenozoa. In addition, DNA is evenly distributed throughout the organelle rather than compacted. Molecular and electron microscopy studies distinguish numerous 6- and 7-kbp-sized mitochondrial chromosomes of monomeric circular topology and relaxed conformation in vivo. Remarkably, the cox1 gene (and probably other mitochondrial genes) is fragmented, with separate gene pieces encoded on different chromosomes. Generation of the contiguous cox1 mRNA requires trans-splicing, the precise mechanism of which remains to be determined. Taken together, the mitochondrial gene/genome structure of Diplonema is not only different from that of kinetoplastids but unique among eukaryotes as a whole.

Comparative morphology of the euglenid pellicle. I. Patterns of strips and pores.

In anticipation that improved knowledge of euglenid morphology will provide robust apomorphy-based definitions for clades, transmission and scanning electron microscopy were used to reveal novel morphological patterns associated with the euglenid pellicle. In some taxa, the number of pellicle strips around the cell periphery reduces as discrete whorls at the anterior and posterior ends of the cell. The number of whorls at either end varies between selected euglenid taxa but is invariant within a taxon. The pattern of strip reduction associated with these whorls is shown to have at least three evolutionarily linked states: exponential, pseudoexponential, and linear. Two general equations describe these states near the posterior end of euglenid cells. Exponential patterns of strip reduction near the anterior end are described by a third equation. In addition, several euglenid taxa were found to possess conspicuous pellicle pores. These pores are arranged in discrete rows that follow the articulation zones between adjacent strips. The number of strips between rows of pores varies between taxa and displays a series of consecutive character states that differ by a power of two. The patterns of pores may not only have phylogenetical and taxonomical value but may provide morphological markers for following strip maturation during cytoskeletal reproduction.