In situ ultrastructures of two evolutionarily distant apicomplexan rhoptry secretion systems.

Parasites of the phylum Apicomplexa cause important diseases including malaria, cryptosporidiosis and toxoplasmosis. These intracellular pathogens inject the contents of an essential organelle, the rhoptry, into host cells to facilitate invasion and infection. However, the structure and mechanism of this eukaryotic secretion system remain elusive. Here, using cryo-electron tomography and subtomogram averaging, we report the conserved architecture of the rhoptry secretion system in the invasive stages of two evolutionarily distant apicomplexans, Cryptosporidium parvum and Toxoplasma gondii. In both species, we identify helical filaments, which appear to shape and compartmentalize the rhoptries, and an apical vesicle (AV), which facilitates docking of the rhoptry tip at the parasite's apical region with the help of an elaborate ultrastructure named the rhoptry secretory apparatus (RSA); the RSA anchors the AV at the parasite plasma membrane. Depletion of T. gondii Nd9, a protein required for rhoptry secretion, disrupts the RSA ultrastructure and AV-anchoring. Moreover, T. gondii contains a line of AV-like vesicles, which interact with a pair of microtubules and accumulate towards the AV, leading to a working model for AV-reloading and discharging of multiple rhoptries. Together, our analyses provide an ultrastructural framework to understand how these important parasites deliver effectors into host cells.

Oral combination of eugenol oleate and miltefosine induce immune response during experimental visceral leishmaniasis through nitric oxide generation with advanced cytokine demand.

Conventional therapy of visceral leishmaniasis (VL) remains challenging with the pitfall of toxicity, drug resistance, and expensive. Hence, urgent need for an alternative approach is essential. In this study, we evaluated the potential of combination therapy with eugenol oleate and miltefosine in Leishmania donovani infected macrophages and in the BALB/c mouse model. The interactions between eugenol oleate and miltefosine were found to be additive against promastigotes and amastigotes with xΣFIC 1.13 and 0.68, respectively. Significantly (p < 0.001) decreased arginase activity, increased nitrite generation, improved pro-inflammatory cytokines, and phosphorylated p38MAPK were observed after combination therapy with eugenol oleate and miltefosine. >80% parasite clearance in splenic and hepatic tissue with concomitant nitrite generation, and anti-VL cytokines productions were observed after orally administered miltefosine (5 mg/kg body weight) and eugenol oleate (15 mg/kg body weight) in L. donovani-infected BALB/c mice. Altogether, this study suggested the possibility of an oral combination of miltefosine with eugenol oleate against visceral leishmaniasis.

Spermatological characters in the Lepocreadioidea, with first data on Holorchis pycnoporus (Aephnidiogenidae), a parasite of the Striped seabream Lithognathus mormyrus (Sparidae) from the Gulf of Gabes (Tunisia).

The ultrastructural characteristics of the mature spermatozoon of Holorchis pycnoporus (Digenea, Lepocreadioidea, Aephnidiogenidae) are described by means of transmission electron microscopy (TEM). Live worms were collected from the digestive tract of the Striped seabream Lithognathus mormyrus (Teleostei, Sparidae), off the Gulf of Gabès at La Chebba (Tunisia). The ultrastructural study reveals that the male gamete of H. pycnoporus is a filiform cell tapered at both extremities and exhibiting the type III of the digenean spermatozoon proposed by Bakhoum et al. (2017a), characterized by the presence of (1) two axonemes with the 9 + '1' pattern of the Trepaxonemata, (2) external ornamentation of the plasma membrane located in a posterior part of the anterior region of the spermatozoon and associated with cortical microtubules, (3) two bundles of parallel cortical microtubules with maximum number located in the middle part of the spermatozoon, and (4) generally two mitochondria. Moreover, H. pycnoporus shares a set of ultrastructural characteristics with the studied Aephnidiogenidae such as: (1) two 9+'1' axonemes of different lengths, (2) an anterior electron-dense material, (3) mitochondrion/a, (4) an external ornamentation of the plasma membrane associated with cortical microtubules, and (5) two bundles of parallel cortical microtubules with their maximum number (around 24 microtubules) located in the middle or posterior part of the spermatozoon. In the Aephnidiogenidae, the mature spermatozoon exhibits a similar ultrastructural pattern. Some differences are observed, particularly the location of maximum number of cortical microtubules and the number of mitochondria. The presence of the anterolateral electron-dense material is the major particularity in species belonging to the Lepocreadioidea. This anterior dense material could be a synapomorphy for the superfamily and an ultrastructural argument supporting the monophyletic status of the Lepocreadioidea (Bray and Cribb, 2012).

Spermatological characteristics of Sclerodistomoides pacificus (Digenea, Sclerodistomoididae) a parasite of the flying fish Cheilopogon pinnatibarbatus (Teleostei, Exocoetidae).

Sclerodistomoides pacificus is the only species described now in Sclerodistomoididae. We present in this paper the first ultrastructural data of the mature spermatozoon of a species from the genus Sclerodistomoides. Adult specimens of S. pacificus (Digenea: Hemiuroidea: Sclerodistomoididae), were parasites of the gall-bladder of the teleost fish Cheilopogon pinnatibarbatus captured in the Atlantic Ocean, near Dakar (Senegal). The male gamete is a filiform cell which exhibits a similar ultrastructural organization to that reported in most species belonging to the Hemiuroidea with two axonemes of the 9 + '1' pattern of trepaxonematans, a nucleus, a mitochondrion, external ornamentation of the plasma membrane not associated with cortical microtubules and located in the anterior region of the spermatozoon, and parallel cortical microtubules disposed in one side of the spermatozoon. However, the present study allowed describing for the first time a moniliform mitochondrion in the Hemiuroidea. The presence of a moniliform mitochondrion and the absence of filamentous external ornamentation described in other Hemiuridae: Lecithochirium microstomum, L. musculus and Hemiurus appendiculatus are a good tool for phylogenetic purposes in the Hemiuroidea. Moreover, spermatological organisation and model are discussed in context with those of previous studies in the Hemiuroidea.

Division and Adaptation to Host Environment of Apicomplexan Parasites Depend on Apicoplast Lipid Metabolic Plasticity and Host Organelle Remodeling.

Apicomplexan parasites are unicellular eukaryotic pathogens that must obtain and combine lipids from both host cell scavenging and de novo synthesis to maintain parasite propagation and survival within their human host. Major questions on the role and regulation of each lipid source upon fluctuating host nutritional conditions remain unanswered. Characterization of an apicoplast acyltransferase, TgATS2, shows that the apicoplast provides (lyso)phosphatidic acid, required for the recruitment of a critical dynamin (TgDrpC) during parasite cytokinesis. Disruption of TgATS2 also leads parasites to shift metabolic lipid acquisition from de novo synthesis toward host scavenging. We show that both lipid scavenging and de novo synthesis pathways in wild-type parasites exhibit major metabolic and cellular plasticity upon sensing host lipid-deprived environments through concomitant (1) upregulation of de novo fatty acid synthesis capacities in the apicoplast and (2) parasite-driven host remodeling to generate multi-membrane-bound structures from host organelles that are imported toward the parasite.

Taxonomic traits in the microstructure of flowers of parasitic Orobanche picridis with particular emphasis on secretory structures.

Orobanche picridis is an obligate root parasite devoid of chlorophyll in aboveground organs, which infects various Picris species. Given the high level of phenotypic variability of the species, the considerable limitation of the number of taxonomically relevant traits (mainly in terms of generative elements), and the low morphological variation between species, Orobanche is regarded as one of the taxonomically most problematic genera. This study aimed to analyse the taxonomic traits of O. picridis flowers with the use of stereoscopic and bright-field microscopy as well as fluorescence, scanning, and transmission electron microscopy. The micromorphology of sepals, petals, stamens, and pistils was described. For the first time, the anatomy of parasitic Orobanche nectaries and the ultrastructure of nectaries and glandular trichomes were presented. Special attention was paid to the distribution and types of glandular and non-glandular trichomes as well as the types of metabolites contained in these structures. It was demonstrated that the nectary gland was located at the base of the gynoecium and nectar was secreted through modified nectarostomata. The secretory parenchyma cells contained nuclei, large amyloplasts with starch granules, mitochondria, and high content of endoplasmic reticulum profiles. Nectar was transported via symplastic and apoplastic routes. The results of histochemical assays and fluorescence tests revealed the presence of four groups of metabolites, i.e. polyphenols (tannins, flavonoids), lipids (acidic and neutral lipids, essential oil, sesquiterpenes, steroids), polysaccharides (acidic and neutral polysaccharides), and alkaloids, in the trichomes located on perianth elements and stamens.

Who Needs a Contractile Actomyosin Ring? The Plethora of Alternative Ways to Divide a Protozoan Parasite.

Cytokinesis, or the division of the cytoplasm, following the end of mitosis or meiosis, is accomplished in animal cells, fungi, and amoebae, by the constriction of an actomyosin contractile ring, comprising filamentous actin, myosin II, and associated proteins. However, despite this being the best-studied mode of cytokinesis, it is restricted to the Opisthokonta and Amoebozoa, since members of other evolutionary supergroups lack myosin II and must, therefore, employ different mechanisms. In particular, parasitic protozoa, many of which cause significant morbidity and mortality in humans and animals as well as considerable economic losses, employ a wide diversity of mechanisms to divide, few, if any, of which involve myosin II. In some cases, cell division is not only myosin II-independent, but actin-independent too. Mechanisms employed range from primitive mechanical cell rupture (cytofission), to motility- and/or microtubule remodeling-dependent mechanisms, to budding involving the constriction of divergent contractile rings, to hijacking host cell division machinery, with some species able to utilize multiple mechanisms. Here, I review current knowledge of cytokinesis mechanisms and their molecular control in mammalian-infective parasitic protozoa from the Excavata, Alveolata, and Amoebozoa supergroups, highlighting their often-underappreciated diversity and complexity. Billions of people and animals across the world are at risk from these pathogens, for which vaccines and/or optimal treatments are often not available. Exploiting the divergent cell division machinery in these parasites may provide new avenues for the treatment of protozoal disease.

A lipid-binding protein mediates rhoptry discharge and invasion in Plasmodium falciparum and Toxoplasma gondii parasites.

Members of the Apicomplexa phylum, including Plasmodium and Toxoplasma, have two types of secretory organelles (micronemes and rhoptries) whose sequential release is essential for invasion and the intracellular lifestyle of these eukaryotes. During invasion, rhoptries inject an array of invasion and virulence factors into the cytoplasm of the host cell, but the molecular mechanism mediating rhoptry exocytosis is unknown. Here we identify a set of parasite specific proteins, termed rhoptry apical surface proteins (RASP) that cap the extremity of the rhoptry. Depletion of RASP2 results in loss of rhoptry secretion and completely blocks parasite invasion and therefore parasite proliferation in both Toxoplasma and Plasmodium. Recombinant RASP2 binds charged lipids and likely contributes to assembling the machinery that docks/primes the rhoptry to the plasma membrane prior to fusion. This study provides important mechanistic insight into a parasite specific exocytic pathway, essential for the establishment of infection.

Cyclic AMP signalling controls key components of malaria parasite host cell invasion machinery.

Cyclic AMP (cAMP) is an important signalling molecule across evolution, but its role in malaria parasites is poorly understood. We have investigated the role of cAMP in asexual blood stage development of Plasmodium falciparum through conditional disruption of adenylyl cyclase beta (ACß) and its downstream effector, cAMP-dependent protein kinase (PKA). We show that both production of cAMP and activity of PKA are critical for erythrocyte invasion, whilst key developmental steps that precede invasion still take place in the absence of cAMP-dependent signalling. We also show that another parasite protein with putative cyclic nucleotide binding sites, Plasmodium falciparum EPAC (PfEpac), does not play an essential role in blood stages. We identify and quantify numerous sites, phosphorylation of which is dependent on cAMP signalling, and we provide mechanistic insight as to how cAMP-dependent phosphorylation of the cytoplasmic domain of the essential invasion adhesin apical membrane antigen 1 (AMA1) regulates erythrocyte invasion.

An ImageJ tool for simplified post-treatment of TEM phase contrast images (SPCI).

Tools for taking advantage of phase-contrast in transmission electron microscopy are of great interest for both biological and material sciences studies as shown by the recent use of phase plates and the development of holography. Nevertheless, these tools most often require highly qualified experts and access to advanced equipment that can only be considered after preliminary investigations. Here we propose to address this issue by the development of an ImageJ plugin that allow the retrieval of a phase image by simple numerical treatment applied to two defocused images. This treatment based on Tikhonov regularization requires the adjustment of a single parameter. Moreover, it is possible to use this approach on one-image. Although in that case the retrieved image gives only qualitative information, it is able to enhance the image contrast appropriately. This can be of interest for specimens producing low contrast images under the electron microscopes, such as some frozen hydrated biological samples or those sensible to electron radiation unsuitable for holographic studies.

Two's company, three's a crowd: co-occurring pollinators and parasite species in Breynia oblongifolia (Phyllanthaceae).

BACKGROUND: Obligate pollination mutualisms (OPMs) are specialized interactions in which female pollinators transport pollen between the male and female flowers of a single plant species and then lay eggs into those same flowers. The pollinator offspring hatch and feed upon some or all of the developing ovules pollinated by their mothers. Strong trait matching between plants and their pollinators in OPMs is expected to result in reciprocal partner specificity i.e., a single pollinator species using a single plant species and vice versa, and strict co-speciation. These issues have been studied extensively in figs and fig wasps, but little in the more recently discovered co-diversification of Epicephala moths and their Phyllanthaceae hosts. OPMs involving Epicephala moths are believed occur in approximately 500 species of Phyllanthaceae, making it the second largest OPM group after the Ficus radiation (> 750 species). In this study, we used a mixture of DNA barcoding, genital morphology and behavioral observations to determine the number of Epicephala moth species inhabiting the fruits of Breynia oblongifolia, their geographic distribution, pollinating behavior and phylogenetic relationships. RESULTS: We found that B. oblongifolia hosts two species of pollinator that co-occurred at all study sites, violating the assumption of reciprocal specificity. Male and female genital morphologies both differed considerably between the two moth species. In particular, females differed in the shape of their ovipositors, eggs and oviposition sites. Phylogenetic analyses indicated that the two Epicephala spp. on B. oblongifolia likely co-exist due to a host switch. In addition, we discovered that Breynia fruits are also often inhabited by a third moth, an undescribed species of Herpystis, which is a non-pollinating seed parasite. CONCLUSIONS: Our study reveals new complexity in interactions between Phyllantheae and Epicephala pollinators and highlights that host switching, co-speciation and non-pollinating seed parasites can shape species interactions in OPMs. Our finding that co-occurring Epicephala species have contrasting oviposition modes parallels other studies and suggests that such traits are important in Epicephala species coexistence.

Changes in parasite-chaetognath species assemblages in the Mexican Central Pacific before and during El Niño 1997-1998.

We investigated the seasonal and interannual changes in diversity, abundance, and prevalence of chaetognaths and their parasites collected monthly during 1996-1998 in the Mexican Central Pacific. We tested the hypothesis of a positive relationship between abundance and species richness of chaetognaths and their parasites, and investigated the influence of the 1997-1998 El Niño event on this host-parasite interaction. Of the 9 chaetognath species collected in the present study, only 7 were found to be parasitized. Of 78154 chaetognath specimens collected, 790 were parasitized (1% prevalence) with at least 1 type of epibiont (cysts, perhaps protists) and 6 types of endoparasites: protists (apicomplexans, dinoflagellates, and ciliates), digeneans, cestodes, acanthocephalans, nematodes, and other unidentified endoparasites. Cysts, digeneans, and cestodes were the most abundant parasites. Mean intensity ranged from 1-4 endoparasites and from 1-21 epibionts host-1. Zonosagitta bedoti and Flaccisagitta enflata were the most abundant chaetognath species and had the highest parasite diversity. Mesosagitta minima and Parasagitta euneritica had the highest parasite prevalence (>2%). A 2-way cluster analysis defined sampling month groups as before, during, and after the 1997-1998 El Niño. The highest abundances of chaetognaths and parasites were associated with a high thermal stratification index, salinity, and mixed layer depth. We conclude that there is a positive, non-linear correlation between the abundance of chaetognaths and their parasites. Although El Niño decreased the abundance and diversity of chaetognaths throughout the time series, the abundance and diversity of their parasites were not significantly different among hydro-climatic periods, suggesting that host abundance must decrease orders of magnitude to influence host availability for parasites.

Cellular dissection of malaria parasite invasion of human erythrocytes using viable Plasmodium knowlesi merozoites.

Plasmodium knowlesi, a zoonotic parasite causing severe-to-lethal malaria disease in humans, has only recently been adapted to continuous culture with human red blood cells (RBCs). In comparison with the most virulent human malaria, Plasmodium falciparum, there are, however, few cellular tools available to study its biology, in particular direct investigation of RBC invasion by blood-stage P. knowlesi merozoites. This leaves our current understanding of biological differences across pathogenic Plasmodium spp. incomplete. Here, we report a robust method for isolating viable and invasive P. knowlesi merozoites to high purity and yield. Using this approach, we present detailed comparative dissection of merozoite invasion (using a variety of microscopy platforms) and direct assessment of kinetic differences between knowlesi and falciparum merozoites. We go on to assess the inhibitory potential of molecules targeting discrete steps of invasion in either species via a quantitative invasion inhibition assay, identifying a class of polysulfonate polymer able to efficiently inhibit invasion in both, providing a foundation for pan-Plasmodium merozoite inhibitor development. Given the close evolutionary relationship between P. knowlesi and P. vivax, the second leading cause of malaria-related morbidity, this study paves the way for inter-specific dissection of invasion by all three major pathogenic malaria species.

Balantioides coli: morphological and ultrastructural characteristics of pig and non-human primate isolates.

Balantioides coli is a ciliated protozoon that inhabits the intestine of pigs, non-human primates and humans. Light microscopy studies have described over 50 species of the genus Balantioides but their validity is in doubt. Due to the limited information about this genus, this study is aimed to identify morphological characteristics of Balantioides coli isolated using fluorescence microscopy and both scanning (SEM) and transmission electron microscopy (TEM). Trophozoites isolated from the feces of pig and macaque were washed and subjected to centrifugation. These cells were fixed with paraformaldehyde for immunofluorescence. Other aliquots of these trophozoites were fixed with glutaraldehyde, post fixed with osmium tetroxide and processed for SEM and TEM. Immunofluorescence studies revealed microtubules with a longitudinal distribution to the main axis of the parasite and in the constitution of cilia. SEM demonstrated a high concentration of cilia covering the oral apparatus and a poor presence of such structures in cytopyge. TEM revealed in the plasma membrane, several associated structures were observed to delineate the cellular cortex and mucocysts. The cytoskeleton of the oral region was observed in detail and had an organization pattern consisting of microtubules, which formed files and nematodesmal networks. Organelles such as hydrogenosomes like and peroxisomes were observed close to the cortex. Macronuclei were observed, but structures that were consistent with micronuclei were not identified. Ultrastructural morphological analysis of isolates confirms its similarity to Balantioides coli. In this study were identified structures that had not yet been described, such as hydrogenosomes like and cytoskeletal structures.

First paleoparasitological study of micromammal coprolites from the holocene of the Somuncurá Plateau Protected Natural Area (Patagonia Argentina).

The Somuncurá Plateau is a Protected Natural Area located in the middle of the northern extra-Andean arid Patagonia. Inhabited by at least 20 small mammal species, is the place with the uppermost species richness in Patagonia. The aim of this study was to examine the parasite remains from micromammal coprolites collected in association with a bone sequence recovered at the east of the Somuncurá Plateau (site "Alero Las Lechuzas"). Coprolites came from the four temporal units previously defined: unit I (4790 ±â€¯100 yrs. 14C B.P.), unit II, unit III (7840 ±â€¯120 yrs. 14C B.P.) and unit IV. Each coprolite was processed, rehydrated, homogenized, processed by spontaneous sedimentation and examined using a light microscope. Coprolites and eggs were described, measured and photographed. Samples were positive for two nematode species: Helminthoxys caudatus Freitas, Lent & Almeida, 1937 (Oxyurida, Oxyuridae) and Trichuris spp. (Trichinellida: Trichuridae). This is the first paleoparasitological study developed for the Somuncurá Plateau Protected Area. Moreover, this is the first time that the genus Helminthoxys is reported from ancient times worldwide. Coprolites were attributed to the mountain cavy Microcavia australis (Rodentia, Caviidae).The presence of H. caudatus for the Middle Holocene of northern Patagonia contributes to the study of the history of the histricomorphs and pinworms relationships.

Functional insights into pathogen biology from 3D electron microscopy.

In recent years, novel imaging approaches revolutionised our understanding of the cellular and molecular biology of microorganisms. These include advances in fluorescent probes, dynamic live cell imaging, superresolution light and electron microscopy. Currently, a major transition in the experimental approach shifts electron microscopy studies from a complementary technique to a method of choice for structural and functional analysis. Here we review functional insights into the molecular architecture of viruses, bacteria and parasites as well as interactions with their respective host cells gained from studies using cryogenic electron tomography and related methodologies.

Role of Polyamines in Parasite Cell Architecture and Function.

In the absence of accessible, effective vaccines, the fight against parasitic disease relies mostly on chemotherapy. Nevertheless, the considerable side effects, high costs and growing number of refractory cases comprise substantial drawbacks. Thus, the search for new antiparasitic compounds remains a high priority. The polyamine biosynthesis, conversion and transport pathways offer different targets for selective chemotherapy. Polyamine analogues and other antagonists may provide tools in the search for new lead compounds. Light and electron microscopy techniques may encompass valuable approaches to elucidate the possible mechanisms of action of different antiparasitic compounds, allowing the identification of subcellular target compartments, presumably establishing the basis for a more rational drug design and/or planning of therapeutic strategies.

Syk inhibitors interfere with erythrocyte membrane modification during P falciparum growth and suppress parasite egress.

Band 3 (also known as the anion exchanger, SLCA1, AE1) constitutes the major attachment site of the spectrin-based cytoskeleton to the erythrocyte's lipid bilayer and thereby contributes critically to the stability of the red cell membrane. During the intraerythrocytic stage of Plasmodium falciparum's lifecycle, band 3 becomes tyrosine phosphorylated in response to oxidative stress, leading to a decrease in its affinity for the spectrin/actin cytoskeleton and causing global membrane destabilization. Because this membrane weakening is hypothesized to facilitate parasite egress and the consequent dissemination of released merozoites throughout the bloodstream, we decided to explore which tyrosine kinase inhibitors might block the kinase-induced membrane destabilization. We demonstrate here that multiple Syk kinase inhibitors both prevent parasite-induced band 3 tyrosine phosphorylation and inhibit parasite-promoted membrane destabilization. We also show that the same Syk kinase inhibitors suppress merozoite egress near the end of the parasite's intraerythrocytic lifecycle. Because the entrapped merozoites die when prevented from escaping their host erythrocytes and because some Syk inhibitors have displayed long-term safety in human clinical trials, we suggest Syk kinase inhibitors constitute a promising class of antimalarial drugs that can suppress parasitemia by inhibiting a host target that cannot be mutated by the parasite to evolve drug resistance.

The unusual eyes of Xenos peckii (Strepsiptera: Xenidae) have green- and UV--sensitive photoreceptors.

The highly specialized evolution of Strepsiptera has produced one of the most unusual eyes among mature insects, perhaps in line with their extremely complex and challenging life cycle. This relatively rare insect order is one of the few for which it has been unclear what spectral classes of photoreceptors any of its members may possess, an even more apt question given the nocturnal evolution of the group. To address this question, we performed electroretinograms on adult male Xenos peckii: we measured spectral responses to equi-quantal monochromatic light flashes of different wavelengths, and established VlogI relationships to calculate spectral sensitivities. Based on opsin template fits, we found maximal spectral sensitivity (λmax) in the green domain at 539 nm. Application of a green light to 'bleach' green receptors revealed that a UV peak was contributed to by an independent UV opsin with a λmax of 346 nm. Transcriptomics and a phylogenetic analysis including 50 other opsin sequences further confirmed the presence of these two opsin classes. While these findings do not necessarily indicate that these unorthodox insects have color vision, they raise the possibility that UV vision plays an important role in the ability of X. peckii males to find the very cryptic strepsipteran females that are situated within their wasp hosts.

The Cytoskeleton of Parabasalian Parasites Comprises Proteins that Share Properties Common to Intermediate Filament Proteins.

Certain protist lineages bear cytoskeletal structures that are germane to them and define their individual group. Trichomonadida are excavate parasites united by a unique cytoskeletal framework, which includes tubulin-based structures such as the pelta and axostyle, but also other filaments such as the striated costa whose protein composition remains unknown. We determined the proteome of the detergent-resistant cytoskeleton of Tetratrichomonas gallinarum. 203 proteins with homology to Trichomonas vaginalis were identified, which contain significantly more long coiled-coil regions than control protein sets. Five candidates were shown to associate with previously described cytoskeletal structures including the costa and the expression of a single T. vaginalis protein in T. gallinarum induced the formation of accumulated, striated filaments. Our data suggests that filament-forming proteins of protists other than actin and tubulin share common structural properties with metazoan intermediate filament proteins, while not being homologous. These filament-forming proteins might have evolved many times independently in eukaryotes, or simultaneously in a common ancestor but with different evolutionary trajectories downstream in different phyla. The broad variety of filament-forming proteins uncovered, and with no homologs outside of the Trichomonadida, once more highlights the diverse nature of eukaryotic proteins with the ability to form unique cytoskeletal filaments.