Polar Filament Research Articles

Myxobolus jialingensis n. sp. (Myxozoa: Myxobolidae) infecting urinary bladder and hepatopancreas of yellowhead catfish Tachysurus fulvidraco from China.

In the present study, we described a novel myxosporean species, Myxobolus jialingensis n. sp. (Myxozoa: Myxobolidae), which infected the urinary bladder and hepatopancreas of yellowhead catfish Tachysurus fulvidraco in China. The mature spores of M. jialingensis n. sp. were pyriform with the length of 15.8 ± 0.7 (15.4-17.0) μm and width of 8.0 ± 0.3 (7.8-8.9) μm. Two pyriform polar capsules were slightly unequal in size: the larger polar capsule was 7.4 ± 0.3 (6.7-8.0) μm in length and 3.1 ± 0.2 (2.8-3.6) μm in width; and the smaller polar capsule measured 7.3 ± 0.3 (6.6-8.1) μm in length and 3.3 ± 0.2 (2.9-3.6) μm in width. The polar capsules were directed toward the apex of the spore, packing seven to eight spirals of the polar filaments. The small subunit ribosomal RNA gene (18S rDNA) sequence of M. jialingensis n. sp. was unique among all myxozoans, and the highest similarity was 96.1% with M. voremkhai. Phylogenetic analysis based on 18S rDNA sequences revealed that myxosporeans infecting the close host affinity (belonging to the same order) had close phylogenetic relationship and, some myxosporeans infecting the same host order might have multiple origins.

The Initiation of a Solar Streamer Blowout Coronal Mass Ejection Arising from the Streamer Flank

Abstract Streamer blowout (SBO) coronal mass ejections (CMEs) represent a particular class of CMEs that are characterized by a gradual swelling of the overlying streamer and a slow CME containing a flux-rope structure. SBO CMEs arising from the streamer flank fall into a special category of SBO CMEs involving three lower arches under the higher streamer arcade. However, the initiation mechanism for this special category of SBO CMEs remains elusive, due to the observational limitations. Here we report critical observations of a SBO CME associated with the eruption of a polar crown filament that originated from the streamer flank. The filament slowly rose toward the solar equator with the writhing motion, and underwent a sudden acceleration before its eruption. Interestingly, during the rising, the filament fields experienced gradual external reconnections, which is evidenced by the dip-shaped bottom of the enveloping flux-rope structure changing from a smooth concave, the slow inflows (∼1.8 km s−1) from both the filament fields and the coronal loops beneath, and the persistent brightenings around the interface between the filament fields and the coronal loops beneath. The newly formed lower loops at the filament source and the Y-shaped structure in the stretched tail fields indicate the internal reconnections for the filament eruption. The clear signatures of external and internal reconnections shed light on the initiation mechanisms of SBO CMEs.

Molecular and morphological characterisation of a novel microsporidian species, Tubulinosema suzukii, infecting Drosophila suzukii (Diptera: Drosophilidae)

A microsporidium showing morphological characteristics typical of a Tubulinosema species was discovered in Drosophila suzukii. All developmental stages were diplokaryotic and grew in direct contact with the host cell cytoplasm. Spores from fresh preparations were ovoid to slightly pyriform and measured 4.29 × 2.47 μm in wet mount preparations. The spore wall consisted of a 125 nm thick endospore covered by a double layered exospore of 39 nm and 18 nm. The polar filament measured 67 μm in length, was slightly anisofilar and was arranged in ten coils in one or rarely two rows. The two posterior coils were 95 nm in diameter while the anterior coils were 115 nm in diameter. Early developmental stages were surrounded by electron-dense, 35.3 nm diameter, surface ornaments scattered over the membrane. Tubular elements with diameters of approximately 75 nm were seen attaching to the periphery of meronts and sporonts. Tissues infected included fat body, midgut and muscle. A 1915 bp rDNA fragment, covering the small subunit (SSU), the internal transcribed spacer (ITS) and the 5′ end of the large subunit ribosomal DNA, was amplified by PCR and sequenced. Phylogenetic analyses of the SSU rDNA fragment revealed closest relationship to Tubulinosema pampeana (Host: Bombus atratus, South America) and Tubulinosema loxostegi (Host: Loxostege sticticalis, ubiquitous), but using the complete dataset of SSU-ITS-LSU rDNA genes revealed T. hippodamiae (Host: Hippodamiae convergens) as the most closely related species. Based on the morphological and genetic features a new species, Tubulinosema suzukii sp. nov., is proposed for this microsporidium isolated from D. suzukii.

A new Myxobolus (Cnidaria: Myxosporea) infecting the ornamental catfish Corydoras schwartzi from the Purus River in Brazil

A new microscopic cnidarian is described, infecting five of thirty (16.6%) specimens of Corydoras schwartzi caught in the Purus River, State of Amazonas, Brazil. Histological analysis showed that cyst development occurred in the serosa layer of the intestine. Mature myxospores are ovoid in body shape in frontal view, 22.4 ± 0.3 μm in total length and 16.3 ± 0.1 μm in width. Internally, two aubergine-shaped, elongate symmetrical polar capsules occupy more than half the length of the spore, 14.3 ± 0.2 μm in length and 6.5 ± 0.1 μm in width. Ultrastructural analysis provided evidence of five polar filament coils inside the polar capsule and binucleated sporoplasm containing a moderate number of sporoplasmosomes. The valvogenic cells abutting each other form a sutural ridge and frequently a thin layer of homogeneous material separates the cells. The outer surface of the myxospore valves is smooth, with no evidence of formation of ridges in the valves. Immature myxospores in various stages of development were observed. This study is the first report of a myxosporean parasitizing C. schwartzi and the first report of a myxosporean infection in the intestine of an ornamental fish from South America.

Insights into myxozoan composition and physiology revealed by histochemical properties of myxospores.

Myxozoa (phylum Cnidaria) are a diverse group of metazoan parasites that predominately infect fish. Little is known regarding the composition and physiology of their myxospore life stage. The objective of this work was to investigate the composition of myxospores and extrasporogonic stages of nine myxozoan species infecting various teleost fish using histochemical staining techniques. Thirty histochemical stains were applied to formalin-fixed, paraffin-embedded tissues processed routinely for light microscopic evaluation. The polar capsules were the most consistent stain target across the taxa examined. Polar capsule staining with Alizarin red, von Kossa and methyl green-pyronin suggests the presence of intracapsular calcium and phosphate, which may contribute to polar filament discharge or pathogenesis of host invasion. The shell valves and suture lines of most myxozoans were stained with Luna and phosphotungstic acid haematoxylin stains, consistent with the presence of chitin and microfibrils, respectively. Vacuoles were consistently highlighted by diastase-susceptible periodic acid-Schiff and Grocott's methenamine silver staining, indicating glycogen. Other histochemical stains exhibited inconsistent staining across the taxa, suggesting differences in myxospore composition potentially reflective of physiologic variations and tissue tropisms. This work provides some information on conserved features and taxa-associated composition of myxospores and lends insight into myxozoan physiology and host-parasite interactions.

Novel myxosporean species parasitizing an economically important fish from the Amazon basin.

This paper provides morphological and phylogenetic analyses of two new myxobolid species found infecting Piaractus brachypomus from the Amazon basin. The fish were caught in the Tapajós River, in the municipality of Santarém, in the state of Pará, Brazil. The plasmodial development of Henneguya brachypomus n. sp. occurred in the gill lamellae while Myxobolus pirapitingae n. sp. developed in the pyloric cecum. Morphological analyses did not identify inflammatory infiltrate for either species, but H. brachypomus n. sp. induced stretching of the epithelium, compression of the adjacent tissues, and displacement and deformation of the neighboring lamellae. The mature myxospores of H. brachypomus n. sp. were ellipsoid, with a length of 11.7-13.8μm, a width of 4.0-4.6μm, and a thickness of 3.5-4.3μm. The polar capsules were elongated, with a length of 5.6-7.3μm and a width of 1.3-2.0μm, and each contained a polar filament with 8-9 coils. The caudal process was 40.5-48.1μm long and the total length of the myxospore was 52.4-61.6μm. Myxobolus pirapitingae n. sp. exhibited rounded mature myxospores measuring 10.0-11.1μm in length, 7.0-7.6μm in width, and 5.4-6.3μm in thickness. The polar capsules were of equal size and occupied less than half the myxospore, measuring 3.5-4.0μm in length and 2.0-2.6μm in width, with each containing a polar filament with 6-7 coils. Phylogenetic analysis based on partial small subunit ribosomal DNA (ssrDNA) sequences showed that H. brachypomus n. sp. clustered as a sister species of Henneguya piaractus, while M. pirapitingae n. sp. was grouped in a sub-clade together with Myxobolus matosi and Myxobolus colossomatis.

Narcinecotyle longifilamentus n. gen., n. sp. (Monogenea: Hexabothriidae), gill parasite of the numbfish Narcine entemedor (Torpediniformes: Narcinidae) from the Mexican Pacific coast.

A new genus and species of Hexabothriidae Price, 1942 is described from specimens infecting the gill of the numbfish Narcine entemedor Jordan and Starks from the Pacific coast of Mexico. In addition to the new taxon described here, species of 8 genera also display symmetrical haptors and have eggs with two polar filaments. However, they differ in the combination of the following features: distal portion of the male copulatory organ tubular, unarmed and proximally dilated confining an internal coiled duct, as well as dorsal origin of haptoral appendix. Based on molecular data derived from 3 loci, the mitochondrial Cytochrome C oxidase subunit 1 as well as the nuclear ribosomal 18S and 28S, the new species was found nested within Hexabothriidae together with the other 4 genera with representatives in GenBank. This is the first species of Hexabothriidae reported from a species of Narcinidae.

A new lineage of crayfish-infecting Microsporidia: The Cambaraspora floridanus n. gen. n. sp. (Glugeida: Glugeidae) complex from Floridian freshwaters (USA)

Crayfish are a vital ecological asset in their native range but can be highly damaging as invasive species. Knowledge of their diseases, including high levels of research on Aphanomyces astaci (crayfish plague), show that disease plays a vital role during crayfish invasions. Microsporidian diseases in crayfish are less studied but are thought to have important links to crayfish health and invasion dynamics.In this study we provide a systematic description of a novel microsporidian parasite from the Floridian crayfish, Procambarus paeninsulanus, with additional genetic identification from related Microsporidia from Procambarus fallax, Cambarellus shufeldtii and Cambarellus blacki. This novel microsporidium from P. paeninsulanus is described in a new genus, Cambaraspora, and species, Cambaraspora floridanus, and represents a novel member of the Clade V Microsporidia within the Glugeidae. The parasite develops in the muscle tissue of P. paeninsulanus, within a sporophorous vesicle, and produces a spore with 19–21 turns of the polar filament measuring 6.136 ± 0.84 µm in length and 2.12 ± 0.23 µm in width. The muscle-infecting nature of the parasite suggests that it is horizontally transmitted.Genetic data for the 18S of the parasite from all hosts confirms its assignment to Clade V and reveal it to be a relative of multiple fish-infecting parasites. It shows closest genetic relationship to Glugea plecoglossi, but branches alongside multiple microsporidia from fish, crustaceans and eDNA isolates.The information presented here suggests that this novel parasite may have the potential to infect piscine hosts and is a likely mortality driver in the P. paeninsulanus population. Its potential as a control agent or wildlife disease invasion threat is explored, as well as the placement of this novel microsporidium within the Glugeidae.

Morphological and Molecular Characterisation of Myxidium kudoi Meglitsch, 1937 from the Blue Catfish Ictalurus furcatus, Valenciennes in Oklahoma, USA

Myxidium kudoi Meglitsch, 1937 has been described from the type host, blue catfish Ictalurus furcatus, with no additional host records or molecular data available for this species. To provide molecular data and a novel host locality for this species and carry out phylogenetic analyses to infer the evolutionary relationship of the species to other members of the family Myxidiidae for which DNA sequence data is available. These data were collected using myxospores from the gallbladder of a blue catfish, Ictalurus furcatus collected from Lake Texoma, Oklahoma, USA. Myxospores were morphologically consistent with the only other account of this species and not any other Myxidium species described from siluriform fishes. Myxospores were oblong with rounded ends and were 10.8-12.6 (11.6 ± 0.5) µm in length and 4.7-6.6 (5.7 ± 0.5) µm in width. Polar capsules were subspherical and 2.7-3.9 (3.4 ± 0.3) µm in length and 2.4-3.5 (3.1 ± 0.3) µm in diameter, with each capsule containing a polar filament with 3-4 coils. Molecular data consisted of a 2918-bp sequence of the partial 18S, complete ITS1, 5.8S, ITS2, and partial 28S ribosomal rRNA regions as well as a 2455-bp sequence of partial 28S ribosomal RNA. The partial 18S and 28S data was used in a concatenated Bayesian phylogenetic analysis to further infer the evolutionary relationships of the Myxidiidae. Additionally, the partial 18S data was used in a separate phylogenetic analysis. The present work reports novel morphological and molecular data for Myxidium kudoi as well as a novel locality of occurrence for this species. In concatenated phylogenetic analysis using 18S and 28S data and other molecular data from Myxozoa, M. kudoi grouped with other freshwater Myxidiidae. In the single-locus, 18S analysis, M. kudoi grouped with Myxidium rhodei from Rutilus rutilus and Myxidium amazonense from Corydoras melini, the only other Myxidium species of catfish for which molecular data are available.

Ultrastructural description and phylogeny of a novel microsporidian, Glugea eda n. sp. from the striated fusilier, Caesio striata, in the Red Sea off Saudi Arabia

Glugea eda n. sp. is described from the mesenteries of the striated fusilier, Caesio striata, collected from the Red Sea coast off Yanbu' al Bahr, Saudi Arabia. Numerous blackish xenomas, ranged from 3 to 5 mm, were found in the body cavity associated with the mesenteries. Mature spores are monomorphic, ellipsoidal with an average size of 5(4–6) μm in length and 2.2 (2–3) μm in width. Observations of the ultrastructure revealed that the development was asynchronous and that the nuclei were isolated throughout the life cycle with uninucleate meronts. Sporoblasts were uninucleated and existed together with sporonts in a fully formed parasitophorous vacuole. The polar filament of the mature spore was isofilar with 24–28 coils, arranged in three rows. Phylogenetic analysis placed the current microsporidia within the clade grouping Glugea species and close to the species described from the Red Sea and Arabian Gulf. The morphometric and molecular comparison with other members of the genus Glugea evidenced the taxonomic novelty of the present form, suggesting that it should be considered as a new species. To the best of our knowledge, the parasite here described represents the first occurrence of microsporidian infection in the fish of the family Caesionidae.

Coelozoic parasite of the family Ceratomyxidae (Myxozoa, Bivalvulida) described from motile vermiform plasmodia found in Hemiodus unimaculatus Bloch, 1794.

Myxozoans of the family Ceratomyxidae are common coelozoic parasites of marine, anadromous, and freshwater fish, and may also be found, less frequently, parasitizing the tissue of these hosts. The diversity and ecology of the freshwater species of the genus Ceratomyxa have been poorly investigated, leading to a knowledge gap that restricts the understanding of the distribution and prevalence of this group of parasites. In the present study, parasites were found inside vermiform plasmodia, characterised by oscillatory movements in the characiform species Hemiodus unimaculatus. The crescent-shaped and elongated spores, perpendicular to the suture line, have a mean length of 28.9 ± 2.7μm and width of 2.6 ± 0.1μm, with two symmetrical oval polar capsules, 1.9 ± 0.3μm in length and 1.7 ± 0.2μm in width, containing polar filaments with three or four coils, located near the central suture, with symmetrical lateral elongations 14.3 ± 1.1μm in length and binucleate amoeboid sporoplasm. The integrated comparative analysis of the morphological characteristics and partial SSU rRNA sequences supported the identification of a new species of coelozoic Ceratomyxa, found in the gallbladder of H. unimaculatus, from the Tocantins basin, in the municipalities of Estreito and Imperatriz in eastern Brazilian Amazonia.The new species was denominated Ceratomyxa fonsecai n. sp.

Neohexostoma gymnosardae n. sp. (Monogenea, Hexostomatidae), a gill parasite of Gymnosarda unicolor (Valenciennes) (Teleostei, Scombridae) in the South China Sea.

Parasite biodiversity of fish in coral reefs of the South China Sea is still incompletely explored. We describe here a new species of Neohexostoma (Monogenea: Hexostomatidae) from the gill filaments of the dogtooth tuna Gymnosarda unicolor (Scombridae), collected off Yongshu Reef, South China Sea. Neohexostoma gymnosardae n. sp. is distinguished from its congeners by the following features: (i) haptor clearly marked from body proper by a strongly constricted peduncle, divided in its posterior margin into two symmetrical lobes, (ii) vagina armed with scattered small blunt spines, (iii) eggs tied by their long polar filaments, (vi) esophagus with several lateral diverticula, (v) intestinal ceca unfused and extending into the haptor. We present an analysis of the relationships of this monogenean based on partial 28S rDNA sequences. An identification key for species of Neohexostoma is provided. This is the first member of the genus Neohexostoma known to parasitize a species of Gymnosarda.

Henneguya sacacaensis n. sp. (Myxozoa: Myxosporea) parasitizing gills of the acará bicudo Satanoperca jurupari (Osteichthyes: Cichlidae) in eastern Amazon.

This study describes Henneguya sacacaensis n. sp. in specimens of the Osteichthyes Satanoperca jurupari (Heckel, 1840), collected in the Rio Curiaú Environmental Protection Area in the city of Macapá, state of Amapá Brazil. Using optical microscopy and molecular analysis, these cyst-shaped parasites were analyzed. The gills of 57.14% of the analyzed S. jurupari contained hundreds of spores. The cysts found on the gill lamellae were oval-shaped and whitish. The Henneguya spores had an average length of 46.5 (41.3-56.92) µm. The fusiform body of the Henneguya measured 16.5 (13.16-20.01) µm long and 5.1 (3.91-6.12) µm in width, the two polar capsules had a taper of 3.83 (3.4-4.32) µm and a width of 1.68 (1.4-1.99) µm, and the tail measured 30 (22.47-41.67) µm in length, containing a polar filament coiled seven to nine times. Morphogical and phylogenetic analysis allowed the preposition of a new species, Henneguya sacacaensis n. sp, that belongs to the family Myxobolidae and the genus Henneguya.

Ultrastructural and molecular characterization of Nosema alticae sp. nov. (Microsporidia: Nosematidae), pathogen of the flea beetle, Altica hampei Allard, 1867 (Coleoptera: Chrysomelidae)

In this study, the first microsporidian pathogen from Altica hampei (Coleoptera: Chrysomelidae) is described based on light microscopy, ultrastructural characteristics and comparative 16S SSU rDNA analysis. All developmental stages of the microsporidium are diplokaryotic and in direct contact with the host cell cytoplasm. Giemsa-stained mature spores are oval in shape and measured 3.82 ± 0.35 μm in length and 2.54 ± 0.27 μm in width. The polar filament of the binucleate spores is isofilar with 12–14 coils. Coils are 140.28 ± 4.88 nm (135.59–147.06; n = 36) in diameter and consist of six concentric layers of different electron density and thickness. The spores have a relatively thick (161.72 ± 29.19 nm) trilaminar spore wall. Morphological, ultrastructural and molecular features indicate that the described microsporidium belongs to the genus Nosema and is named Nosema alticae sp. nov.

A new myxozoan species Henneguya unitaeniata sp. nov. (Cnidaria: Myxosporea) on gills of Hoplerythrinus unitaeniatus from Mato Grosso State, Brazil.

On the basis of morphological and molecular analyses, a new myxozoan parasite is described from the gills of the fish Hoplerythrinus unitaeniatus, collected in the municipality of Nova Xavantina, Mato Grosso State, Brazil. Plasmodia of Henneguya unitaeniata sp. nov. were oval and whitish and were found surrounded by collagen fibers forming plasmodia wall between gill filaments on the gill arch. The spores were ellipsoidal with two similar polar capsules. Morphometric analysis showed a total spore mean length of 23.8 ± 1.5 μm, spore body mean length of 14.5 ± 0.7 μm, caudal appendage mean length of 10.3 ± 1.4 μm, thickness mean length of 4.3 ± 0.3 μm, polar capsule mean length of 4.2 ± 0.5 μm, polar capsule mean width of 1.8 ± 0.3 μm, spore mean width of 4.8 ± 0.4 μm, and 4-5 polar filament coils. Phylogenetic analysis showed Henneguya unitaeniata sp. nov. as a basal species in a subclade formed by myxozoans that parasitize bryconid fishes.

Ortholinea scatophagi (Myxosporea: Ortholineidae), a novel myxosporean infecting the spotted scat, Scatophagus argus (Linnaeus 1766) from southwest coast of India

A new species of myxosporean, Ortholinea scatophagi n. sp. infecting the urinary bladder of the spotted scat, Scatophagus argus (Linnaeus 1766) is described. O. scatophagi n. sp. is characterized by spherical myxospores with a slightly flattened anterior end and equal spore valves with extra sutural ridges on its surface; measured 7.34 ± 0.67 μm in length, 6.90 ± 0.71 μm in width and 6.48 ± 0.37 μm in thickness. Two polar capsules, equal, spherical to oval in shape, arranged diametrically opposite and measured 2.59 ± 0.42 μm in length and 2.24 ± 0.35 μm in width. Polar filaments, 21.84 ± 2.86 μm long, with four to five coils. Scanning electron microscopy revealed the presence of extra sutural ridges on spore surface. Pansporoblasts spherical to irregular in shape, measured 31.08 ± 2.67 μm in length and 13.88 ± 5.40 μm in width; Monosporic, disporic and polysporic plasmodial stages were observed; plasmodia spherical or irregular in shape with granular cytoplasm containing refractile granules. The species was compared with 23 existing nominal species of Ortholinea, based on morphology and morphometry. Molecular analysis resulted in a 1773 bp long SSU rDNA sequence (GenBank accession number MN 310514). In phylogenetic analyses the present parasite clustered with other members of Ortholinea, under the freshwater urinary clade. Considering the morphologic, morphometric and molecular differences with previously described species of Ortholinea, and differences in host and geographic locations, the present species is treated as new and the name Ortholinea scatophagi n. sp.is proposed.

Chromospheric Synoptic Maps of Polar Crown Filaments

Polar crown filaments form above the polarity inversion line between the old magnetic flux of the previous cycle and the new magnetic flux of the current cycle. Studying their appearance and their properties can lead to a better understanding of the solar cycle. We use full-disk data of the Chromospheric Telescope (ChroTel) at the Observatorio del Teide, Tenerife, Spain, which were taken in three different chromospheric absorption lines (H-alpha 6563A, CaII-K 3933A, and HeI 10830A), and we create synoptic maps. In addition, the spectroscopic HeI data allow us to compute Doppler velocities and to create synoptic Doppler maps. ChroTel data cover the rising and decaying phase of Solar Cycle 24 on about 1000 days between 2012 and 2018. Based on these data, we automatically extract polar crown filaments with image-processing tools and study their properties. We compare contrast maps of polar crown filaments with those of quiet-Sun filaments. Furthermore, we present a super-synoptic map summarizing the entire ChroTel database. In summary, we provide statistical properties, i.e. number and location of filaments, area, and tilt angle for both the maximum and declining phase of Solar Cycle 24. This demonstrates that ChroTel provides a promising data set to study the solar cycle.

Polar tube structure and three polar tube proteins identified from Nosema pernyi

Microsporidian spores contain a single polar filament that is coiled around the interior of the spore. Upon germination the polar tube (post-germination polar filament) is ejected by inversion into a host cell. The sporoplasm flows through the polar tube, directly infecting the cytoplasm of the cell. Various species of microsporidia display differences in the number of coils in the polar filament and in the amino acid sequence of the polar tube proteins (PTPs). Nosema pernyi is a lethal pathogen that causes microsporidiosis in the Chinese oak silkworm, Antheraea pernyi. In this study, we identified three PTPs in N. pernyi using RT-PCR and LC-MS/MS. Polar tube protein 3 was localized in the polar tube using immuno-histochemical staining and an immunofluorescence assay. Co-immunoprecipitation data and LC-MS/MS analysis revealed that some potential proteins, like immune related proteins in A. pernyi may interact with PTP3.

A microtubule RELION-based pipeline for cryo-EM image processing

Microtubules are polar filaments built from αβ-tubulin heterodimers that exhibit a range of architectures in vitro and in vivo. Tubulin heterodimers are arranged helically in the microtubule wall but many physiologically relevant architectures exhibit a break in helical symmetry known as the seam. Noisy 2D cryo-electron microscopy projection images of pseudo-helical microtubules therefore depict distinct but highly similar views owing to the high structural similarity of α- and β-tubulin. The determination of the αβ-tubulin register and seam location during image processing is essential for alignment accuracy that enables determination of biologically relevant structures. Here we present a pipeline designed for image processing and high-resolution reconstruction of cryo-electron microscopy microtubule datasets, based in the popular and user-friendly RELION image-processing package, Microtubule RELION-based Pipeline (MiRP). The pipeline uses a combination of supervised classification and prior knowledge about geometric lattice constraints in microtubules to accurately determine microtubule architecture and seam location. The presented method is fast and semi-automated, producing near-atomic resolution reconstructions with test datasets that contain a range of microtubule architectures and binding proteins.

Unusual dynamics of the divergent malaria parasite PfAct1 actin filament

Gliding motility and host cell invasion by the apicomplexan parasite Plasmodium falciparum (Pf), the causative agent of malaria, is powered by a macromolecular complex called the glideosome that lies between the parasite plasma membrane and the inner membrane complex. The glideosome core consists of a single-headed class XIV myosin PfMyoA and a divergent actin PfAct1. Here we use total internal reflection fluorescence microscopy to visualize growth of individual unstabilized PfAct1 filaments as a function of time, an approach not previously used with this actin isoform. Although PfAct1 was thought to be incapable of forming long filaments, filaments grew as long as 30 µm. Polymerization occurs via a nucleation-elongation mechanism, but with an ∼4 µM critical concentration, an order-of-magnitude higher than for skeletal actin. Protomers disassembled from both the barbed and pointed ends of the actin filament with similar fast kinetics of 10 to 15 subunits/s. Rapid treadmilling, where the barbed end of the filament grows and the pointed end shrinks while maintaining an approximately constant filament length, was visualized near the critical concentration. Once ATP has been hydrolyzed to ADP, the filament becomes very unstable, resulting in total dissolution in <40 min. Dynamics at the filament ends are suppressed in the presence of inorganic phosphate or more efficiently by BeFX A chimeric PfAct1 with a mammalian actin D-loop forms a more stable filament. These unusual dynamic properties distinguish PfAct1 from more canonical actins, and likely contribute to the difficultly in visualizing PfAct1 filaments in the parasite.