bluuebunny/crossref_metadata_2025 huggingface dataset:
Transcription Rates and Transcript Stabilities of Macronuclear Genes in Vegetative Euplotes crassus Cells
Authors: BRANDT ANDREAS , KLEIN ALBRECHT
Published: May 2007

ABSTRACT. In hypotrichous ciliates such as Euplotes crassus genes in the transcriptionally active macronucleus are present on individual minichromosomes which occur in gene‐specific copy numbers. This different degree of gene amplification can be understood as a means to preset the expression potential of the respective genetic information. In addition, the actual steady state transcript amounts are governed by the transcription rates and transcript stabilities. To establish the relative effects of these three parameters the copy numbers of genes transcribed by the three different polymerases were determined. The transcript levels of growing or starving vegetative cells were then determined, and nuclear run‐on assays were performed to determine the transcription rates of the genes in the different nutritional states. A weak correlation between the gene copy numbers and transcription rates was found. The transcripts of genes synthesized by RNA polymerase II exhibited different stabilities upon starvation of the cells, compared to the supposedly stable ribosomal 5S and 26S RNA. Refeeding of the cells after starvation also resulted in a differential response with respect to the accumulation of the transcripts of different genes transcribed by RNA polymerase II, which can be interpreted in the context of the gene functions.
Calcium and Hydrogen Ion Concentrations in the Parasitophorous Vacuoles of Epithelial Cells Infected with the Microsporidian Encephalitozoon hellem
Authors: LEITCH GORDON J. , SCANLON MARY , VISVESVARA GOVINDA S. , WALLACE SARA
Published: May 2007

ABSTRACT. Microsporidia of the genus Encephalitozoon undergo merogony and sporogony in a parasitophorous vacuole within the host cell. Cultured green monkey kidney cells infected with Encephalitozoon hellem were loaded with the fluorescent dyes fura‐2 or BCECF in order to measure intracellular concentrations of calcium and hydrogen ions respectively. Both the parasitophorous vacuole calcium concentration and pH values resembled those of the host cell cytoplasm in infected cells. Calcein entered the parasitophorous vacuole but not other host cell vacuoles or parasite stages within the parasitophorous vacuole. The lack of a pH or calcium concentration gradient across the parasitophorous vacuole membrane and the permeability of this membrane to a large anion such as calcein suggest that the vacuole membrane surrounding E. hellem resembles that surrounding some other intracellular parasites such as Toxoplasma gondii. A potential role is discussed for the parasitophorous vacuole calcium concentration in germination in situ.
Ultrastructure of the Parabasalid Protist Holomastigotoides
Authors: LINGLE WILMA L. , SALISBURY JEFFREY L.
Published: May 2007

ABSTRACT. The ultrastructure of two species of Holomastigotoides is presented. The basic unit of organization of these large cells is the flagellar band. Each flagellar band consists of a row of flagellar basal bodies linked by three fiber systems. The number of flagellar bands is species dependent. The flagellar bands originate at the cell apex and are arranged in parallel spirals of increasing gyre, thus defining the conical shape of the cell. In the cell apex a striated root called a parabasal fiber is juxtaposed with the basal bodies of each flagellar band. Linear extensions of two parabasal fibers function as the spindle poles for the persistent extra‐nuclear spindle. The nucleus is in close contact with the spindle poles and spindle microtubules. Parallel sheets of microtubules which constitute axostyles are nucleated along the underside of the parabasal fibers. The axostyles extend away from the cell apex, with many reaching the basal region of the cell. Some of the axostyles follow the spiral pattern of the flagellar bands. Numerous Golgi bodies are spaced regularly along the flagellar bands. Together the parabasal fiber, axostyles and Golgi bodies associated with a flagellar band are termed a parabasal complex.
Heterovesicula cowani N. G., N. Sp. (Heterovesiculidae N. Fam.), a Microsporidian Parasite of Mormon Crickets, Anabrus simplex Haldeman, 1852 (Orthoptera: Tettigoniidae)
Authors: LANGE CARLOS E. , MACVEAN CHARLES M. , HENRY JOHN E. , STREETT DOUGLAS A.
Published: May 2007

ABSTRACT. Heterovesicula cowani, n. g., n. sp., is a dimorphic microsporidium described from the adipose tissue of the Mormon cricket, Anabrus simplex Haldeman. Proliferation of the microsporidium is by karyokinesis of uninucleate and binucleate cells to form binucleate and tetranucleate cells, respectively. These cells will undergo binary fission (merogony). Ultimately, the meronts undergo karyokinesis without subsequent cytokinesis producing spherical multinucleate plasmodia that are transitional to 2 types of sporogony. Transitional to disporoblastic sporogony, a fragile interfacial envelope delaminates from the plasmodium with morphogenesis to a monfiliform plasmodium consisting of fusiform binucleate diplokaryotic sporonts. These undergo karyokinesis to form tetranucleate diplokaryotic sporonts that undergo cytokinesis during disintegration of the plasmodium into isolated binucleate sporonts. Transitional to octosporoblastic sporogony, multinucleate plasmodia disintegrate into short monofiliform plasmodia of diplokaryotic sporonts which then segregate while undergoing gradual nuclear dissociation (haplosis by nuclear dissociation). These undergo two sequences of karyokinesis and subsequent multiple fission to form eight uninucleate (haploid) sporoblasts in a fusiform arrangement within a persistent envelope. Binucleate spores are ovocylindrical, about 5.4 × 1.7 μm (fresh), with an isofilar polar filament singly coiled about 11 turns. Uninucleate spores are ovoid to slightly pyriform, 4.0 × 1.7 μm (fresh), with an isofilar filament singly coiled about 9 turns. A new family, Heterovesculidae, is proposed for the new genus.
Composition of Pneumocystis carinii Neutral Lipids and Identification of Coenzyme Q10 as the Major Ubiquinone Homolog
Authors: ELLIS JAYNE E. , WYDER MICHAEL A. , ZHOU LINDA , GUPTA ARUN , RUDNEY HARRY , KANESHIRO EDNA S.
Published: May 2007

The lipids of purified preparations of Pneumocystis carinii carinii freshly isolated from infected rats were analyzed and compared with those of whole lungs from normal and methylprednisolone‐immunosuppressed uninfected rats. In this study, the neutral lipid fraction was examined in detail; the relative concentrations of individual classes making up this fraction were quantified. Of particular interest was the nature of the organism's ubiquinone (coenzyme Q, CoQ) fraction because atovaquone, a hydroxynaphtho‐quinone (566C80) analog of ubiquinone, is efficacious in the treatment of P. carinii pneumonia. The ubiquinone concentration in both P. carinii and lung tissues was relatively low compared to that present in rat heart and liver tissues. Two homologs were identified in the organism: CoQ10 was the predominant homolog with lesser amounts of CoQ9 present. In contrast, the lungs of normal and immunosuppressed uninfected rats had CoQ9 and lesser amounts of CoQ8, but no detectable CoQ10. Furthermore, radiolabeled mevalonic acid was incorporated in vitro into the ubiquinone fraction of P. carinii indicating that the organism has the de novo branch of the isoprenoid biosynthetic pathway leading to polyprenyl formation. Hence, it was concluded that CoQ10 (if not both CoQ110 and CoQ9) in P. carinii as not scavenged from the host but was synthesized by the organism. Although lung tissues contained substantial free fatty acids, the organism was enriched in these lipids. The high concentration of free fatty acids and relatively low level of triglycerides in P. carinii suggest that fatty acids may represent major carbon sources for ATP production by the organism.
An Arrayed Bacteriophage P1 Genomic Library of Pneumocystis carinii
Authors: METCHEVA IVELINA S. , STEDMAN TIMOTHY T. , BUCK GREGORY A.
Published: May 2007

We have constructed an arrayed, large insert, multiple coverage genomic library of Pneumocystis carinii DNA using the bacteriophage P1 cloning system. The library consists of ∽4800 independent clones with an average insert size of ∽55 kbp individually arrayed in 50 microtiter plates, and is readily screened on ten or fewer microtiter plate‐sized filters using a high density colony replicating device. Screening of the library for unique P. carinii sequences detected an average of 4–5 positive clones for each, consistent with a several‐fold coverage of the ∽10‐mbp P. carinii genome. Restriction and hybridization analyses demonstrated that the P1 clones in this library are quite stable and contain few, if any, chimeric inserts. Thus, this arrayed, large insert library off. carinii genomic DNA will be a valuable tool in the future genetic dissection of this important pathogen.
Effect of Cobalamin Deficiency on the Biosynthesis of Phosphatidylcholine in Euglena gracilis
Authors: INUI HIROSHI , OHYA OSAMU , ISEGAWA YUJI , KITAOKA SHOZABURO , MIYATAKE KAZUTAKA , NAKANO YOSHIHISA
Published: May 2007

Euglena gracilis requires cobalamin (Cbl) as an essential growth factor. Phosphatidylcholine (PC) synthesis was greatly reduced by Cbl deficiency. Rapid cell division occurred after Cbl was replenished, and PC was actively synthesized during the cell divisions. When the deficient cells were given methionine (a precursor for the choline moiety), active synthesis of PC occurred even without the Cbl supplement, although cell division was not induced. As methionine synthase in Euglena requires methylcobalamin as a coenzyme, decrease in methionine synthesis may account for reduced PC synthesis under Cbl‐deficient conditions. Phosphatidyleth‐anolamine and phosphatidylserine synthesis were also suppressed, commensurate with decrease of PC synthesis, under Cbl deficiency, even though Cbl is not thought to participate in their synthesis. In contrast, a lot of triglyceride and wax ester accumulated in Cbl‐deficient cells. Moreover, Cbl depletion altered fatty acid composition, notably due to increased proportion of odd‐numbered fatty acids
The Contribution of Ciliated Protozoa to Plankton and Benthos Biomass in a European Ricefield
Authors: MADONI PAOLO
Published: May 2007

The densities and biomass of ciliates inhabiting the water‐sediment interface and the water column of an experimental ricefield were investigated throughout four annual cycles. Ciliate abundance and biomass were higher at the water‐sediment interface than in the water column. In both sites, large ciliates (> 105μm3) contributed the higher biomass values, but the highest densities were found in the intermediate size class (104‐105μm3). The prorodontid Coleps hirtus dominated the ciliate assemblage and usually comprised > 50% of total ciliate density. Blooms of C. hirtus, occurred in June in the water column and in July at the sediment surface. During the four cycles of rice cultivation, the average daily values for production of the entire ciliate community was 69.6 mgC/m2/d, and the net production efficiency (K2) was 72.0%. The estimated production values in the present study are high if compared to production measured for ciliates in other freshwater ecosystems.
Long‐Term Storage of Infective Microsporidian Spores in Liquid Nitrogen
Authors: MADDOX JOSEPH V. , SOLTER LEELLEN F.
Published: May 2007

Thirty‐one species of microsporidia, isolated from insects and stored in liquid nitrogen for up to 25 yr, were infectious when removed from liquid nitrogen. The natural hosts of all of these microsporidia were terrestrial insects, representing six different insect orders: Coleoptera, Diptera, Hemiptera, Hymenoptera, Lepidoptera, and Orthoptera. All microsporidia from terrestrial insects that were tested survived storage in liquid nitrogen, while Nosema algerae, a microsporidium from aquatic mosquito hosts did not survive freezing in liquid nitrogen. A Nosema species from the alfalfa weevil, Hypera postica, lost some infectivity in a water storage medium after 25 yr in liquid nitrogen. Liquid nitrogen storage of microsporidian spores in 50% and 100% glycerol media reduced loss of infectivity and is recommended for extended storage of microsporidia from terrestrial insect hosts.
Insights into the Evolution of Nuclear Dualism in the Ciliates Revealed by Phylogenetic Analysis of rRNA Sequences
Authors: HAMMERSCHMIDT BRIGITTE , SCHLEGEL MARTIN , LYNN DENIS H. , LEIPE DETLEF D. , SOGIN MITCHELL L. , RAIKOV IGOR B.
Published: May 2007

The small subunit rRNA gene sequences of the karyorelictean ciliates, Loxodes striatus and Protocruzia sp., and the heterotrichian ciliates, Climacostomum virens and Eufolliculina uhligi, were used to test the evolution of nuclear dualism in the Phylum Ciliophora. Phylogenies derived using a least squares distance method, neighbour joining, and maximum parsimony demonstrate that the karyorelictean ciliates sensu Small and Lynn, 1985 do not form a monophyletic group. However, Loxodes and the heterotrich ciliates form the first branch in the ciliate lineage, and Protocruzia branches, in distance methods, basal to the spirotrich lineage. It is proposed that Protocruzia be removed from the Class Karyorelictea, and placed in closer taxonomic association with the spirotrich lineage. The distribution of nuclear division types along the phylogenetic tree is consistent with the notion that macronuclei incapable of division represent a derived rather than a primitive or “karyorelictid” character trait.
Analyses of 22S Dynein Binding to Tetrahymena Axonemes Lacking Outer Dynein Arms
Authors: SULLIVAN JEANELL , LUDMANN SUSAN A. , HAMASAKI TOSHIKAZU , PENNOCK DAVID G.
Published: May 2007

ABSTRACT. Tetrahymena thermophila mutants homozygous for the oad mutation become nonmotile when grown at the restrictive temperature of 39° C. Axonemes isolated from nonmotile oad mutants (oad 39° C axonemes) lack approximately 90% of their outer dynein arms and are deficient in 22S dynein. Here we report that oad 39° C axonemes contain 40% of the 22S dynein heavy chains that wild‐type axonemes contain and that oad axonemes do not undergo ATP‐induced microtubule sliding in vitro. Wild‐type 22S dynein will bind to the outer arm position in oad axonemes and restore ATP‐induced microtubule sliding in those axonemes. Unlike wild‐type 22S dynein, oad 22S dynein does not bind to the outer arm position in oad axonemes. These data indicate that the oad mutation affects some component of the outer arm dynein itself rather than the outer arm dynein binding site. These data also indicate that oad axonemes can be used to assay outer dynein arm function.
Comparative Development of Two Microsporidian Species: Enterocytozoon bieneusi and Enterocytozoon salmonis, Reported in AIDS Patients and Salmonid Fish, Respectively
Authors: DESPORTES‐LIVAGE ISABELLE , CHILMONCZYK STEFAN , HEDRICK RONALD , OMBROUCK CÉCILE , MONGE DANIELLE , MAIGA IBRAHIM , GENTILINI MARC
Published: May 2007

ABSTRACT. Enterocytozoon bieneusi and Enterocytozoon salmonis are reported in HIV‐infected patients and in salmonid fish, respectively. Both species share the early development of the extrusion apparatus of the spores, which is completed prior to fission of the sporogonic syncytium into sporoblasts, and the early synthesis of polar tube constituents, but they differ in other developmental and sporogenetic processes. Enterocytozoon bieneusi develops in direct contact with the cytoplasm of epithelial cells whereas E. salmonis occurs only in the nucleus of leucocytes and epithelioid cells. Sporogonic nuclei, which are scattered throughout the sporont in E. bieneusi, are located in the periphery in E. salmonis. The multilamellar structures associated with the nuclear envelopes and the endoplasmic reticulum cisternae are specific for E. bieneusi. Additionally, the evolution of the polar tube precursors proceeds differently in the two parasites. In E. bieneusi, they transform into electron‐dense bodies associated with a reticulum and polar tubes derive from these structures according to a process similar to that reported in other microsporidia. In E. salmonis, polar tube precursors fuse directly at their ends and the polar tubes appear to be formed by the assemblage of these fused precursors with a material previously synthesized in the vicinity of nuclei. In conclusion, both species appear to be less closely related than was supposed in earlier descriptions.
Molecular Characterization of the D Surface Protein Gene Subfamily in Paramecium tetraurelia
Authors: BREUER MARION , SCHULTE GERALD , SCHWEGMANN KLAUS J. , SCHMIDT HELMUT J.
Published: May 2007

ABSTRACT. When Paramecium tetraurelia expresses the D serotype, detectable by serum tests, high molecular mRNA could be isolated, which corresponds to the molecular mass of the D surface protein. Using this D specific mRNA as a probe for screenings in different genomic libraries a subfamily of five very similar genes was found, named α‐51D, γ1‐51D, γ2‐51D, δ‐51D and ε‐51D. Each of them is about 8‐kb long, they show regions of identity to each other, and there is no evidence that any are defective genes or pseudogenes. Up to now serotype D is the only known serotype showing this phenomenon. Another novel feature is that two of the D isogenes are closely linked. The sequence for the entire coding region of the α‐51D gene has been determined, as well as the upstream and downstream noncoding regions. Its deduced amino acid sequence shows the same characteristic cysteine periodicity displayed by all other immobilization antigen (i‐ag) genes from Paramecium. However, in contrast to most other such genes, tandem repeats are missing from the 7599‐bp long coding region of the α‐51D gene. When the sequences of the type 51D genes are compared to each other, the similarity is very high and extends to coding as well as to noncoding regions. Similarity within noncoding regions is usually only observed for allelic i‐ag genes. We conclude that the type D genes constitute a family of isogenes that are nonallelic. They contain slightly different consensus sequences with possible functions as regulatory regions.
Investigating the Comparative Biology of the Heterokonts with Nucleic Acids1
Authors: WEE JAMES L. , HINCHEY JEANETTE M. , NGUYEN KIM X. , KORES PAUL , HURLEY DAVID L.
Published: May 2007

During the last decade investigations of heterokont protists utilizing molecular methods have challenged established biosystematic concepts. Most investigations emphasized the chloroplast genome or sequences from nuclear‐encoded, ribosomal genes. Refinement of DNA isolation protocols, advent of “universal primers” and the polymerase chain reaction, automated sequencing and increased accessibility of DNA sequence databases have expanded data‐gathering efficiency and increased dataset sizes. Because independent datasets have been easier to obtain, the testing of specific phylogenetic hypotheses has been facilitated, altering relationship concepts, primarily at phylum/class levels, and perceptions of cellular evolution. New approaches have emphasized ecological studies and extended studies to genus/species levels and poorly investigated genomes. This paper reviews studies documenting these impacts and identifies some current limitations. Additionally, new DNA sequence data from our laboratory on nuclear‐encoded rDNA internal transcribed spacers and the chloroplast‐encoded psb A gene suggest that these regions will provide taxonomic resolution for the Synu‐rophyceae, at the class/order level and subspecies level, respectively.
P. carinii DNA Detected by ITSs Nested PCR in Serum and PBMC of AIDS Patients with PCP
Authors: ATZORI CHIARA , AGOSTONI FIORENZA , ANGELI ELENA , ORLANDO GIOVANNA , CARGNEL ANTONIETTA
Published: May 2007

SUMMARY Pneumocystis carinii pneumonia (PCP) is usually diagnosed by examination of BAL, a sample often unpleasant to be collected from immunocompromised host affected by acute respiratory disease. We studied by the Internal Transcribed Spacers (ITSs) nested PCR the presence of P. carinii DNA in serum and Peripheral Blood Mononuclear Cells (PBMC) during acute episodes of PCP to test blood as a possible noninvasive diagnostic tool.
Leishmania mexicana amazonensis: Differential Display Analysis and Cloning of mRNAs from Attenuated and Infective Forms
Authors: HEARD PAMELA L. , LEWIS CLARENCE S. , CHAUDHURI GAUTAM
Published: May 2007

The virulence of Leishmania mexicana is determined by the concerted action of several parasite molecules. These cells lose their infectivity to host macrophages after prolonged cultivation in axenic growth media. Both virulent and attenuated variants of the parasite cells were cloned. The differential display reverse transcription‐polymerase chain reaction technique was employed to understand whether this natural attenuation of the parasite cells is accompanied by differential expression of selected genes in those cells. Twelve different dinucleotide‐anchored oligo(dT) antisense primers were used to make cDNAs from poly(A)+ mRNAs isolated from a clonal population of virulent and avirulent cells following a protocol optimized for Leishmania mRNAs. Those cDNAs were subjected to amplifications using each of the three different arbitrary decanucleotide primers and the corresponding anchored oligo(dT) primer. This procedure revealed four virulent‐specific cDNA probes and one avirulent‐specific cDNA probe. Differential expressions of these genes were confirmed by northern hybridization using the cloned cDNA probes. These results indicate that differential expression of genes may be the key in determining the molecular basis of leishmanial virulence.
Effect of Platelet‐Activating Factor on the Process of Cellular Differentiation of Herpetomonas muscarum muscarum
Authors: LOPES ANGELA H. C. S. , DUTRA PATRICIA M. L. , RODRIGUES CLAUDIA O. , SOARES MAURILIO J. , ANGLUSTER JAYME , CORDEIRO RENATO S. B.
Published: May 2007

ABSTRACT. The effects of platelet‐activating factor (PAF), at doses ranging from 10−6 M to 10−10 M, on cell growth and on cell differentiation of Herpetomonas muscarum muscarum were investigated. Cell differentiation was evaluated by both light and electron microscopy. At the concentrations used, PAF did not interfere with the protozoan growth. However, parasites grown in the presence of PAF (10−6 M) were significantly more differentiated than those grown in the absence of PAF, since the first day of culture. On the first two days of culture, PAF doses ranging from 10−10 M to 10−7 M, did not significantly interfere with the differentiation of these parasites, although after the third day of culture, all PAF doses used significantly increased the protozoan differentiation. Specific PAF receptor antagonists totally abrogated (WEB 2086 and WEB 2170)or significantly decreased (BN 52021) PAF effect on cell differentiation. These findings indicate PAF triggers the process of cell differentiation in Herpetomonas muscarum muscarum and suggest these parasites have receptors for PAF.
The Plaque Matrix (PQM) and Tubules at the Surface of Intramuscular Parasite, Trachipleistophora hominis
Authors: WEIDNER EARL , CANNING ELIZABETH U. , HOLLISTER WAFAA S.
Published: May 2007

ABSTRACT. Surface plaque matrix (PQM) and a tubular arrangement of filaments border Trachipleistophora hominis parasites during growth within host muscle. The PQM at the parasite surface forms a network of processes which can be associated with filamentous tubules. Peroxidase tracer delineated the PQM and showed apparent connections with the tubules. The tubules at the interface of T. hominis‐infected cells are structurally similar to the extrasporular tubules of the microsporidian, Ameson michaelis. The extrasporular tubules of A. michaelis and the proteins from T. hominis‐infected muscle reacted to keratin antibodies, K8.13, K4 and K13. Conversely, antibodies produced to T. hominis‐infected muscle, reacted with the extrasporular tubular proteins of A. michaelis. The PQM and tubular elements are thought to play an important role in affecting molecular traffic between the host and parasite.
Three New Limax Amoebae Isolated from Marine Surface Sediments: Vahlkampfia caledonica N. Sp., Saccamoeba marina N. Sp., and Hartmannella vacuolata N. Sp.
Authors: ANDERSON O. ROGER , ROGERSON ANDREW , HANNAH FIONA
Published: May 2007

ABSTRACT. Three new limax amoebae, isolated from marine, surface sediment samples are described using light microscopic and fine structural features. One species, characterized by eruptive locomotion typical of the family Vahlkampfiidae, is assigned the name Vahlkampfia caledonica (47.4 ± 16.0 μm × 12.1 ± 3.2 μm). The other two monopodial species move with steady locomotion characteristic of the family Hartmannellidae. One is a Saccamoeba with a distinct posterior bulbous uriod, vaculoes containing prominent crystals, glycocalyx with cup‐like components, and spherical nucleus with central nucleolus. It is assigned the name Saccamoeba marina (72.5 ± 14.9 ±μm × 20. 7 ± 4. 5 μM). The other hartmannellid limax amoeba moves by steady locomotion and has a rather constant monopodial from, lacks a uroid, but has occasional trailing masses of cytoplasm, contains cup‐like structures in the glycocalyx, and is characterized μm). Few limax amoebae have been described from marine environment and these data provide additional evidence that limax amoebae may be more abundant in marine sediments that realized previously.
Opportunistic Properties of Nosema algerae (Microspora), a Mosquito Parasite, in Immunocompromised Mice
Authors: TRAMMER THOMAS , DOMBROWSKI FRANK , DOEHRING MARTINA , MAIER WALTER A. , SEITZ HANNS M.
Published: May 2007

ABSTRACT. In the last ten years microspordia have been recognized as opportunistic pathogens in AIDS patients. The sources of infection and the mechanisms of transmission of these organisms in humans are mostly uncertain. Transmission of invertebrte microsporidia to mammals is normally considered impossible, temperature being a limiting factor for development. Mice treated with cortisone acetate and with cyclosporin A, respectively, as well as athymic mice were injected intravenously, intranasally, perorally and subcutaneously with spores of Nosema algerae, a microsporidian species of culicine mosquitoes. No infection could be detected in tissue samples of cortisone acetate and cyclosporin A treated mice. However, the experimental inoculation of spores into the tail and foot of athymic mice caused severe infection in skeletal muscles and the connective tissue. In some tails, nerve tissue and bone marrow were also infected. Vegetative stages and spores were seen in direct contact to host cell cytoplasma. For the first time the prolonged and progressive development of an invertebrate microsporidium in a mammalian host is shown. The possibility of invertebrate microsporidia as a source of human microsporidiosis should now be taken into consideration.
Ultrastructural Features of Spindle Microtubule Organization During the Neclear Division of Encephalitozoon hellem
Authors: SACCHI LUCIANO , BIGLIARDI ELISA , LANZARINI PAOLO , CORONA SILVIA , GATTI SIMONETTA , SCAGLIA MASSIMO
Published: May 2007

Ultrastructural studies were carried out to describe the nuclear division cycel of a strain of Enephalitozoon hellem isolated from an Italina AIDS patient. The nuclear division occurs during the porliferative vegetative phase and it is characterized by the intranuclear mitosis and by the lack of centrioles. The spindle termini are electron dense spindle plaques (ESPs), resembling to the spindle pole bodies (SPBs) of Sacchoaromycetes. The ESPs are bifacial organella forming microtubules on both nucleoplasmic and cytoplasmic faces. In the outer layer of the spindle plaque are present vesicular elements lined by a double membrance of unknown function. The peculiar morphological feacture of E. hellem ESPs indicate that bothe intranclear spindle and cytoplasmic microtubules are involved in the nuclear division.
Evidence for Reversible Tyrosine Protein Phosphorylation in the Okadaic Acid‐Producing Marine Dinoflagellate Prorocentrum lima
Authors: DAWSON JOHN F. , OSTERGAARD HANNE L. , KLIX HEIDE , BOLAND MARION P. , HOLMES CHARLES F. B.
Published: May 2007

ABSTRACT. The protist Prorocentrum lima, a primary producer of the tumour promoter okadaic acid, is a member of the dinoflagellate class of marine microorganisms. Herein, we have identified and characterized a protein tyrosine kinase (designated PLIK 1A) in P. lima that autophosphorylates almost exclusively on tyrosine residues. PLIK 1A was shown to have an approximate molecular mass of 38 kDa by SDS‐PAGE and a native molecular mass within the range of 47–55 kDa by Superdex‐75 gel filtration. Phosphoamino acid analysis of autophosphorylated PLIK 1A revealed the presence of phosphotyrosine and autophosphorylated PLJK 1A reacted with monoclonal anti‐phosphotyrosine antibodies in a Western immunoblot. In addition, two protein tyrosine phosphatases were identified in P. lima that had apparent molecular masses within the ranges of 150–168 kDa and 73–82 kDa as determined by Superdex‐200 gel filtration. These P. lima phosphatases, termed PLPTP‐I and PLPTP‐II, efficiently dephosphorylated tyrosine phosphorylated myelin basic protein. owever, only PLPTP‐I was capable of dephosphorylating the tyrosine phosphorylated substrate angiotensin. Both PLPTP‐I and PLPTP‐II were able to dephosphorylate tyrosine autophosphorylated PLIK 1A. These data provide the first evidence for reversible tyrosine protein phosphorylation in P. lima by protein tyrosine kinases and phosphatases
Germination of Nosema algerae (Microspora) Spores: Conditional Inhibition by D2O, Ethanol and Hg2+ Suggests Dependence of Water Influx upon Membrane Hydration and Specific Transmembrane Pathways
Authors: FRIXIONE EUGENIO , RUIZ LOURDES , CERBÓN JORGE , UNDEEN ALBERT H.
Published: May 2007

ABSTRACT. The germination of microsporidian spores under conditions expected to affect water flow across the plasma membrane‐wall complex was studied by assessing their responses to in vitro stimulation with Na+ or K+. Partial or full substitution of common water with D2O, which more effectively coats ions and electrostatically‐charged cell surfaces with relatively stable hydration layers, delayed and inhibited spore germination in a concentration‐dependent manner; yet, preincubation in 100% D2O did not change the normal response to standard stimulation. Water structure‐breaking conditions, such as an increase in temperature (within the 15° C to 40° C range) or in ionic strength (2‐ to 10‐fold normal), opposed the inhibition by D2O and allowed significant stimulation by Li+, the monovalent cation with the largest hydration diameter and a usually weak stimulant action on the spores. Ethanol, known to reduce water permeation across cell membranes and phospholipid bilayers, also caused a powerful and dose‐dependent (1% to 4% v/v) inhibition of spore germination, but pretreatment with ethanol did not affect the normal response. HgCl2, an inhibitor of specific water channels, blocked spore germination at just 250 μM in the normal stimulation solution irrespective of the temperature, and permitted only a delayed response in high salt stimulation solutions. However, the inhibition by Hg2+ was abolished by the simultaneous presence of 2‐mercaptoethanol in the medium. These results suggest (1) that spore germination is keenly dependent upon the hydration states of both the plasma membrane‐wall complex and the stimulant ions, and (2) that osmotic water flows into the spores through specific transmembrane pathways with critical sulfhydryl groups, i.e. analogous to the water channels that facilitate water movements across the plasma membranes of highly permeable cells.
Pneumocystis carinii of the Common Shrew, Sorex araneus, Shows a Discrete Phenotype
Authors: LAAKKONEN JUHA , SUKURA ANTTI
Published: May 2007

ABSTRACT. We carried out an immunohistological and morphological study on Pneumocystis carinii originating from the common shrew, Sorex araneus. Immunologic properties were studied by applying two commercially available immunofluorescence staining kits with differing developmental form specificity to a lung homogenate. The cyst form‐specific staining kit reacted with cysts originating from S. araneus. Ultrastructurally this particular antigen epitope specifically deposited on the electron‐lucent middle layer of the cyst pellicle. The immunohistochemical staining kit reacting with both cyst and trophozoite forms from human and rat origin did not react with any developmental forms of P. carinii originating from S. araneus. Both kits demonstrated P. carinii in the lung homogenate of a field vole, Microtus agrestis. In morphologic examination, the methenamine silver‐stained cyst forms of P. carinii from S. araneus and from M. agrestis differed in size from each other and from those originating from laboratory rats. Ultrastructurally P. carinii from S. aruneus did not differ from organisms of rat origin.
Stomatogenesis of the Ciliate Genus Strombidinopsis with an Improved Description of S. spiniferum and S. acuminatum
Authors: Dale Torbjørn , Lynn Denis H.
Published: May 2007

Protargol‐stained cultured samples of Strombidinopsis spiniferum and S. acuminatum revealed a kind of enantiotropic mode of cell division. Stomatogenesis may be divided into eight stages. Stage 1 was the appearance of an anterior anarchic field of a few kinetosomes, probably originating apokinetally near a somatic kinety on the dorsal side of the cell. In Stage 2, the anarchic field becomes denser and larger by kinetosomal replication. The kinetosomes become aligned as oral polykinetids (OPk's) in Stage 3. In Stage 4, the paroral (PO) emerges when most of the anarchic field has transformed into OPk's. In Stage 5. all OPk's are apparently formed, and the anlage resembles a barrel, lacking a few staves. In Stage 6, the barrel rotates approximately 90° transversely to the cell's axis, concomitantly the outer end of the barrel splays open. In Stage 7, the somatic kineties start to divide, the anlage has opened to the cell surface, and the part with the internal OPk's spirals into the oral cavity while the OPk's form a closed circle. At this stage, one or two large macronuclei were seen preparing to divide. In Stage 8, the adult or interphase stage, the cytopharynx is formed. Based on our results, the original redescriptions of S. spinifierum and S. acuminatum must be modified in three respects: the cells have a PO; the OPk's are composed of three rows of kinetosomes (uncertain for S. acuminatum), not two as originally reported; and the number of internal OPk's is not always 3, but ranges from 3‐4 in S. spiniferum and 3‐5 in S. acuminatum.
Brachiola vesicularum, N. G., N. Sp., a New Microsporidium Associated with AIDS and Myositis
Authors: Call Ann , Takvorian Peter M. , Lewin Sharon , Rendel Michael , Sian Corazon S. , Wittner Murray , Tanowitz Herbert B. , Keohane Elaine , Weiss Louis M.
Published: May 2007

Brachiola vesicularum, n. g., n. sp., is a new microsporidium associated with AIDS and myositis. Biopsied muscle tissue, examined by light and electron microscopy, revealed the presence of organisms developing in direct contact with muscle cell cytoplasm and fibers. No other tissue types were infected. All parasite stages contain diplokaryotic nuclei and all cell division is by binary fission. Sporogony is disporoblastic, producing 2.9 times 2 μm diplokaryotic spores containing 8‐10 coils of the polar filament arranged in one to three rows, usually two. Additionally, this microsporidium produces electron‐dense extracellular secretions and vesiculotubular appendages similar to Nosema algerae. However, the production of protoplasmic extensions which may branch and terminate in extensive vesiculotubular structures is unique to this parasite. Additionally, unlike Nosema algerae, its development occurred at warm blooded host temperature (37‐38° C) and unlike Nosema connori, which disseminates to all tissue types, B. vesicularum infected only muscle cells. Thus, a new genus and species is proposed. Because of the similarities with the genus Nosema, this new genus is placed in the family Nosematidae. Successful clearing of this infection (both clinically and histologically) resulted from treatment with albendazole and itraconozole.
Cytoskeleton Structure and Composition in Choanoflagellates
Authors: Karpov Serglei A. , Leadbeater Barry S. C.
Published: May 2007

The structure and composition of the cytoskeleton has been studied in Monosiga ovata (Protozoa: Order Choanofiagellida Kent 1880) using a combination of methods in association with light and electron microscopy. Supplementary observations are included for Desmarella moniliformis. The basal body of the single anterior flagellum is subtended proximally and at right angles by a second, non‐flagellar basal body. The edges of the two basal bodies are connected by a fibrillar bridge. A long, narrow, striated, fibrillar rootlet extends posteriorly from the lower edge of the non‐flagellar basal body towards the Golgi apparatus. It is associated throughout most of its length with the surface of a flattened sac. Rootlet microtubules pass radially from a ring of electron dense material which encircles the distal end of the flagellar basal body. These microtubules extend outwards for about one‐third of the length of the cell. Within each collar tentacle is a longitudinal bundle of microfilaments composed of actin as illustrated by rhodamine‐phalloidin staining for fluorescence microscopy. The base of each microfilament bundle is associated with one or more rootlet microtubules by fine fibrillar bridges. The attachment between microtubules and tentacle microfilaments is further demonstrated by their coordinated displacement when the cytoskeleton becomes dislodged. The role of the cytoskeleton in maintaining the position of the collar tentacles during interphase and cell division is discussed.
Roots1
Authors: Salisbury Jeffrey L.
Published: May 2007

ABSTRACTMany unicellular eukaryotic organisms possess complex fiber systems that organize and anchor the flagellar basal apparatus in the cell [20, 24]. In 1978 we first published the observation that one of these fiber systems, the striated flagellar root of the quadriflagellate green algaTetraselmis subcordiformis(=Platymonas subcordiformis), is a contractile organelle [31]. We subsequently found that striated flagellar roots are composed, in part, of the Ca2+‐binding protein centrin [30]. Since that time, centrin has been found to be a ubiquitous component of the flagellar basal apparatus, basal bodies and centrioles, and centrosomes and mitotic spindle poles of eukaryotic cells (for general reviews see [28, 34]). While we have learned a great deal about centrin from other organisms, our earliest success in understanding the biology of centrin was in large part due to the extraordinary extent to whichTetraselmiscells have elaborated their centrin‐based organelles. In this paper, I will return attention to several unanswered questions concerning Tetraselmis striated flagellar root behavior and I will suggest several new directions that students may wish to pursue in order to tease fresh insights from this fascinating organism.
The Evolutionary History of the Genus Acanthamoeba and the Identification of Eight New 18S rRNA Gene Sequence Types
Authors: Stothard Diane R. , Schroeder‐Diedrich Jill M. , Awwad Mohammad H. , Gast Rebecca J. , Ledee Dolena R. , Rodriguez‐Zaragoza Salvador , Dean Chantal L. , Fuerst Paul A. , Byers Thomas J.
Published: May 2007

ABSTRACT The 18S rRNA gene (Rns) phylogeny of Acanthamoeba is being investigated as a basis for improvements in the nomenclature and taxonomy of the genus. We previously analyzed Rns sequences from 18 isolates from morphological groups 2 and 3 and found that they fell into four distinct evolutionary lineages we called sequence types T1‐T4. Here, we analyzed sequences from 53 isolates representing 16 species and including 35 new strains. Eight additional lineages (sequence types T5‐T12) were identified. Four of the 12 sequence types included strains from more than one nominal species. Thus, sequence types could be equated with species in some cases or with complexes of closely related species in others. The largest complex, sequence type T4, which contained six closely related nominal species, included 24 of 25 keratitis isolates. Rns sequence variation was insufficient for full phylogenetic resolution of branching orders within this complex, but the mixing of species observed at terminal nodes confirmed that traditional classification of isolates has been inconsistent. One solution to this problem would be to equate sequence types and single species. Alternatively, additional molecular information will be required to reliably differentiate species within the complexes. Three sequence types of morphological group 1 species represented the earliest divergence in the history of the genus and, based on their genetic distinctiveness, are candidates for reclassification as one or more novel genera.
Identification and Characterization of an Endo/exonuclease in Pneumocystis carinii that is Inhibited by Dicationic Diarylfurans with Efficacy Against Pneumocystis Pneumonia
Authors: Hildebrandt Ellen , Boykin David W. , Kumar Arvind , Tidwell Richard R. , Dykstra Christine C.
Published: May 2007

ABSTRACT Dicationic diarylfurans and dicationic carbazoles are under development as therapeutic agents against opportunistic infections. While their ability to bind to the minor groove of DNA has been established, the complete mechanism of action has not. We demonstrate here that an effective diarylfuran, 2,5‐bis[4‐(N‐isopropylguanyl)phenyl]furan. inhibits an endo/exonuclease activity present in Pneumocystis carinii, Cryptococcus neoformans, Candida albicans, and Saccharomyces cerevisiae. This activity was purified from the particulate fraction of P. carinii. The enzyme requires Mg++ or Mn++, and shows preferences for single‐ over double stranded DNA and for AT‐rich over GC‐rich domains. A panel of 12 dicationic diarylfurans and eight dicationic carbazoles, previously synthesized, were evaluated for inhibition of the purified nuclease and for efficacy against Pneumocystis pneumonia in rats. Among the diarylfurans, potency of nuclease inhibition, in vivo antimicrobial activity, and DNA binding strength were all strongly correlated (p < 0.001). These findings suggest that one target for antimicrobial action of the diarylfurans may be a nucleolytic or other event requiring unpairing of DNA strands. Dicationic carbazoles which were strong nuclease inhibitors all displayed anti‐Pneumocystis activity in vivo, but there were also noninhibitory carbazoles with in vivo efficacy.
Graviresponses of Gliding and Swimming Loxodes Using Step Transition to Weightlessness
Authors: Machemer‐Röhnisch Sigrun , Bräucker Richard , Machemer Hans
Published: May 2007

Cells of Loxodes striatus were adjusted to defined culturing, experimental solution, O2‐supply, temperature, and state of equilibration to be subjected to step‐type transition of acceleration from normal gravity (1 g) to the weightless condition (μg) during free fall in a 500‐m drop shaft. Cellular locomotion inside a vertical experimental chamber was recorded preceding transition and during 10 s of μg. Cell tracks from video records were used to separate cells gliding along a solid surface from free swimmers, and to determine gravitaxis and gravikinesis of gliding and swimming cells. With O2 concentrations ≥ 40% air saturation, gliders and swimmers showed a positive gravitaxis. In μg gravitaxis of gliders relaxed within 5 s, whereas gravitaxis relaxation of swimmers was not completed even after 10 s. Rates of horizontal gliders (319 μm/s) exceeded those of horizontal swimmers (275 μm/s). Relaxation of gravikinesis was incomplete after 10 s of μg. Analysis of the locomotion rates during the g‐step transition revealed that gliders sediment more slowly than swimmers (14 versus 45 μm/s). The gravikinesis of gliders cancelled sedimentation effects during upward and downward locomotion tending to maintain cells at a predetermined level inside sediments of a freshwater habitat. At ≥ 40% air saturation, gravikinesis of swimmers augmented the speed of the majority of cells during gravitaxis, which favours fast vertical migrations of Loxodes.
Grazing Behavior of Heterotrophic NanoFlagellates Observed with a High Speed VTR System
Authors: Ishigaki Tetsuji , Terazaki Makoto
Published: May 2007

The grazing patterns of heterotrophic nanoflagellates (Cafeteria roenbergensis and Actinomonas mirabilis) were recorded by high speed video micrography. Experiments were carried out at 10, 15 and 25° C at a salinity of 36 psu and at 20° C when the salinity was either 20 or 36 psu. Bacteria entrained in a stream of water generated by flagellar beating were propelled toward the cell body (phase 1). Each bacterium destined for ingestion made contact with the anterior end of the flagellum of Cafeteria or was captured with an actinopod of Actinomonas (phase 2). The captured bacterium was drawn close to the cell body (phase 3) and ingested (phase 4). Cafeteria was observed during 38 grazing events and appeared to have two different patterns of ingestion, “enclosing” and “pocket phagocytosis.” The mean time for food capture was 0.10 s and the subsequent ingestion was complete on average in 1.59 s. No further bacteria were captured until ingestion was complete. Actinomonas was observed during 27 grazing events in which capture of panicles on an actinopod was followed by transport to the cell body and subsequent “enclosing phagocytosis.” The mean time to complete grazing was 39.16 s, which is about 25 times longer than for Cafeteria; the phase of transport to the cell body occupied most of this time. During the grazing cycle, Actinomonas captured other bacteria on the actinopodia, but sometimes released them. Grazing time in Cafeteria was positively correlated with temperature but not correlated with salinity. Grazing behavior in Actinomonas was correlated with neither temperature nor salinity.
Adhesion of Plasmodium gallinaceum Ookinetes to the Aedes aegypti Midgut: Sites of Parasite Attachment and Morphological Changes in the Ookinete
Authors: Zieler Helge , Garon Claude F. , Fischer Elizabeth R. , Shahabuddin Mohammed
Published: May 2007

Plasmodium gallinaceum ookinetes adhered to Aedes aegypti midgut epithelia when purified ookinetes and isolated midguts were combined in vitro. Ookinetes preferentially bound to the microvillated luminal surface of the midgut, and they seemed to interact with three types of structures on the midgut surface. First, they adhered lo and migrated through a network‐like matrix, which we have termed microvilli‐associated network, that covers the surface of the microvilli. This network forms on the luminal midgut surface in response to blood or protein meals. Second, the ookinetes bound directly to the microvilli on the surface of the midgut and were occasionally found immersed in the thick microvillar layer. Third, the ookinetes associated with accumulations of vesicular structures found interspersed between the microvillated cells of the midgut. The origin of these vesicular structures is unknown, but they correlated with the surface of midgut cells invaded by ookinetes as observed by TEM. After binding to the midgut. ookinetes underwent extensive morphological changes: they frequently developed one or more annular constrictions, and their surface roughened considerably, suggesting that midgut components remain bound to the parasite surface. Our observations suggest that, in a natural infection, the ookinete interacts in a sequential manner with specific components of the midgut surface. Initial binding to the midgut surface may activate the ookinete and cause morphological changes in preparation for invasion of the midgut cells.